chiniofon and roquinimex

Prostate cancer is the mostly commonly diagnosed non-skin cancer in males. The culmination of the last 70 years of clinical drug development has documented that androgen ablation plus taxane-based systemic chemotherapy enhances survival, but is not curative, in metastatic prostate cancer. To effect curative therapy, additional drugs must be developed that enhance the response when combined with androgen ablation/taxane therapy.. The history of the discovery and development of tasquinimod as a second-generation oral quinoline-3-carboxamide analogue for prostate cancer will be presented.. The mechanism for such anticancer efficacy is via tasquinimod's ability to inhibit the 'angiogenic switch' within cancer sites required for their continuous lethal growth.. Tasquinimod is a novel inhibitor of tumor angiogenesis that enhances the therapeutic anticancer response when combined with other standard-of-care modalities (radiation, androgen ablation, and/or taxane-based chemotherapies) in experimental animal models, but does not inhibit normal wound healing. It has successfully completed clinical Phase II testing in humans and will shortly enter registration Phase III evaluation for the treatment of metastatic prostate cancer.

Topics: Administration, Oral; Angiogenesis Inhibitors; Animals; Clinical Trials as Topic; Drug Evaluation, Preclinical; Humans; Hydroxyquinolines; Male; Prostatic Neoplasms; Quinolines; Quinolones

NK cells are a subset of mononuclear cells which have long been suspected of playing an immunoregulatory role in the prevention of autoimmune diseases. Here, we briefly discuss the characteristics of NK cells--particularly what is known of their functional capabilities--and summarise the major findings from studies of NK cells in human and animals susceptible to three major autoimmune diseases: multiple sclerosis, systemic lupus erythematosus and type 1 (autoimmune) diabetes mellitus. In each case, we present the evidence for an association between disease and deficiencies in NK cells. The prospect of clinical interventions that stimulate NK cell activity are discussed and the current status described.

Topics: Animals; Autoimmune Diseases; Cell Differentiation; Cytokines; Diabetes Mellitus, Type 1; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Humans; Hydroxyquinolines; Interferons; Killer Cells, Natural; Lupus Erythematosus, Systemic; Mice; Multiple Sclerosis; Rats; Self Tolerance

Linomide (roquinimex, LS 2616) is a quinoline-3-carboxamide with pleiotropic immune modulating capacity and it has therapeutic effects in several experimental animal models of autoimmune diseases. Linomide has been evaluated in clinical trials for multiple sclerosis, and was indeed shown to have disease inhibitory effects. However, due to unexpected side effects recorded in patients treated with Linomide, premature termination of clinical trials was required. The basic mechanism(s) of action of Linomide in inducing beneficial effects in autoimmune diseases is still elusive. Some experimental evidence indicates that Linomide influences the regulation of the cytokine profile, resulting in the inhibition of autoimmune and inflammation pathologies. This review focuses on Linomide applied in models for autoimmune and inflammation pathologies of the central and the peripheral nervous system, and summarises its very encouraging disease inhibitory effects and their potential pharmacological basis. The beneficial effects recorded with Linomide in both experimental and clinical trials emphasise the possible value of substances with Linomide-like activity for clinical use in autoimmune and inflammation pathologies in the near future.

Topics: Adjuvants, Immunologic; Animals; Autoimmune Diseases; Central Nervous System; Cytokines; Humans; Hydroxyquinolines; Immunity, Cellular; Peripheral Nervous System

Insulin-dependent diabetes mellitus (IDDM) is considered to be an autoimmune disorder characterized by destruction of the pancreatic beta-cells by auto-reacting lymphocytes. An attractive therapeutic approach to this disease would be to abrogate the autoimmune process at an early stage, thus preserving a critical mass of pancreatic beta-cells necessary for maintenance of normal glucose tolerance. Linomide (quinoline-3-carboxamide, Roquinimex, LS 2616), is a novel, orally absorbed, immunomodulatory drug that has been shown to be effective in various models of autoimmunity without causing non-specific immunosuppression. In this review, we describe the efficacy of Linomide for ameliorating the autoimmune process and diabetes in the non-obese diabetic (NOD) model of IDDM when administered at early stages of the disease. We also show that advanced disease in the NOD mouse can be treated effectively by combining Linomide with therapeutic modalities designed to increase pancreatic beta-cell mass. Subsequent clinical studies have shown that Linomide preserves beta-cell function in individuals with new-onset IDDM. Based on these data, Linomide or derivatives thereof might be useful for treatment of human IDDM.

Topics: Adjuvants, Immunologic; Animals; Autoimmune Diseases; Diabetes Mellitus, Type 1; Hydroxyquinolines; Mice; Mice, Inbred NOD

The last years have produced a plethora of new information including extensive studies, retrospective analysis and new perspectives on data interpretation on multiple sclerosis (MS) treatment. Considering how difficult it is to study a disease such MS with its variability, unpredictability and duration, it seems hard to resemble definite results from this experience. However, corticosteroids have been the mainstay of treatment for the management of acute relapses, showing the capacity to shorten the duration of relapses, accelerate the recovery. At present, interferon beta is generally considered to be the treatment of choice for patients with relapsing remitting disease. Glatiramer acetate is still not available in many parts of Europe, but its results demonstrate a reduction of relapses in 30% of cases. Most European experts only consider as alternative treatment the immunosuppressive drugs, chosen if patients demonstrate unacceptable side effects of interferon or clearly do not respond. Very different and even more confusing data still come from experimental trial in secondary progressive MS, where the target of treatment is to slow the progression of disability. Different drugs (methotrexate, mitoxantrone, linomide, steroids and even interferons) are employed, but the results are still debated. Future therapies are being derived from constantly changing and evolving concept of MS immunopathogenesis: therefore many experimental and clinical trials use anti-integrin antibodies or insulin growth factors, metallo-proteinase inhibitors or T-cell vaccination. Some of the above treatment may have a chance of producing the gaining control of the disease without much inner toxicity.

Topics: Adjuvants, Immunologic; Adrenal Cortex Hormones; Azathioprine; Clinical Trials as Topic; Forecasting; Glatiramer Acetate; Humans; Hydroxyquinolines; Immunoglobulins, Intravenous; Immunosuppressive Agents; Interferons; Methotrexate; Multiple Sclerosis; Peptides; Polymers; Recurrence; Retrospective Studies

Topics: Adjuvants, Immunologic; Administration, Oral; Adolescent; Adult; Double-Blind Method; Female; Humans; Hydroxyquinolines; Male; Middle Aged; Multiple Sclerosis

Topics: Animals; Cell Movement; Graft Rejection; Graft vs Host Reaction; Hydroxyquinolines; Immunosuppressive Agents; Intestine, Small; Rats

Topics: Adjuvants, Immunologic; Animals; Apoptosis; Cell Cycle; Humans; Hydroxyquinolines; Lymphocyte Activation; Signal Transduction; T-Lymphocytes

Topics: Adjuvants, Immunologic; Androgens; Animals; Antibodies, Anti-Idiotypic; Antiphospholipid Syndrome; Aspirin; Bone Marrow Transplantation; Bromocriptine; Cyclosporine; Disease Models, Animal; Estrogens; Ethylenes; Heparin; Hydroxyquinolines; Immunoglobulins, Intravenous; Immunosuppression Therapy; Immunosuppressive Agents; Lupus Erythematosus, Systemic; Lymphocyte Transfusion; Mice; T-Lymphocytes, Regulatory

Programmed cell death (PCD) is a necessary process that helps to regulate the lifespan of lymphocytes and maintain the compartmental balance of lymphoid organs. In addition, PCD is required for the generation and maintenance of self-tolerance. Strategies that inhibit PCD cause profound alterations in the (patho)physiology of the immune system. Here, Guido Kroemer and Carlos Martínez-A. discuss the multiplicity of PCD-inducing pathways, which have been revealed through the use of PCD-inhibitory agents, and analyse the levels at which these agents act.

Topics: Animals; Apoptosis; Cycloheximide; Cyclosporine; Humans; Hydroxyquinolines; Lymphocytes; Mifepristone; Proto-Oncogenes; Tretinoin

It is now known that syngeneic transplantation, T lymphocyte depletion and absence of graft-versus-host disease all increase the risk of relapse following allogeneic transplantation for the myeloid leukemias, both acute and chronic. Leukemia-specific immune responses appear to play a major role in the therapy of the myeloid leukemias. In recent years attempts have been made to better characterize and effectively utilize these antileukemic immune responses, concentrating on clinical states of minimal residual disease. This review will discuss the role of such immunotherapy following autologous bone marrow transplantation for myeloid leukemias, and will focus on recent experience and ongoing clinical trials using the novel immunomodulator Linomide.

Topics: Adjuvants, Immunologic; Humans; Hydroxyquinolines; Immunotherapy; Interferons; Interleukin-2; Killer Cells, Natural; Leukemia, Myeloid; T-Lymphocytes

Topics: Adjuvants, Immunologic; Animals; Diabetes Mellitus, Type 1; Encephalomyelitis, Autoimmune, Experimental; Female; Humans; Hydroxyquinolines; Mice; Mice, Inbred NOD

Topics: Animals; Disease Models, Animal; Graft Rejection; Graft Survival; Guanidines; Heart Transplantation; Hydroxyquinolines; Immunosuppressive Agents; Rats; Transplantation, Heterologous; Transplantation, Homologous

Topics: Acute Disease; Bone Marrow Transplantation; Graft vs Host Disease; Granulocyte-Macrophage Colony-Stimulating Factor; Humans; Hydroxyquinolines; Immunotherapy; Interferons; Interleukin-2; Leukemia; Macrophages; T-Lymphocytes

Topics: Bone Marrow Transplantation; Combined Modality Therapy; Forecasting; Graft vs Host Disease; Humans; Hydroxyquinolines; Immunologic Factors; Immunotherapy, Adoptive; Interleukin-2; Killer Cells, Natural; Leukemia

Due to an unexpected increase in serious cardiovascular events in MS patients treated with Linomide, a synthetic immunomodulator, two phase-III multinational relapsing remitting (RR) and secondary progressive (SP) MS trials had to be discontinued. MRI results of 413 patients who participated for at least 3 months were analysed. Patients received placebo, 2.5 or 5 mg Linomide. Scans were performed at pre-enrolment, month 3 and termination. The number and volume of enhancing lesions (ELV), and the number of active scans were evaluated. At month 3, the decrease in the number of enhancing lesions in the placebo group was 11%, compared with 15% in the 2.5 mg group (P=0.027) and 23% in the 5 mg group (P=0.057). Using the percentage of active scans as outcome parameter, the odds ratio for improvement between placebo and 2.5 mg group was 1.62 (P=0.14); between placebo and 5 mg Linomide group 3.58 (P=0.003). At termination, a rebound effect was noted in the 2.5 mg group (P=0.01). Analysis of the ELV showed no significant difference between placebo and treatment groups. Although Linomide has unacceptable side effects, it seems to have a modest effect on MS disease activity, as measured by MRI. Multiple Sclerosis (2000) 6, 99 - 104

Topics: Adjuvants, Immunologic; Adolescent; Adult; Cardiovascular Diseases; Humans; Hydroxyquinolines; Magnetic Resonance Imaging; Middle Aged; Multiple Sclerosis, Chronic Progressive; Multiple Sclerosis, Relapsing-Remitting; Treatment Outcome

Topics: Adjuvants, Immunologic; Adult; Aged; Disability Evaluation; Dose-Response Relationship, Drug; Double-Blind Method; Female; Humans; Hydroxyquinolines; Male; Middle Aged; Multiple Sclerosis, Chronic Progressive; Multiple Sclerosis, Relapsing-Remitting; Treatment Failure

To determine whether linomide (roquinimex) is better than placebo in slowing the time to confirmed clinical worsening in patients with relapsing-remitting (RR) and secondary progressive (SP) MS.. In this 27-center, randomized, double-blind, placebo-controlled, multiple-dose, phase III trial, 715 patients with active RRMS (n = 90) or SPMS (n = 625) were randomized to receive either linomide (1.0, 2.5, or 7.5 mg orally daily) or placebo. Patients were evaluated at 3-month intervals clinically and with MRI. The planned primary outcome was the time to the development of "confirmed" clinical worsening (increase of >/= 1.0 Expanded Disability Status Scale [EDSS] score for an enrollment EDSS score /= 0.5 point for an enrollment EDSS score of >/= 5.5) not associated with an acute relapse.. The trial was terminated 1 month after it became fully enrolled due to unanticipated serious cardiopulmonary toxicities (pericarditis, pleural effusion, myocardial infarction, and possible pulmonary embolism), pancreatitis, and death. Notable arthralgia, myalgia, bursitis, and facial and peripheral edema were common adverse events. The high dose of linomide (7.5 mg) was not well tolerated. The trial was too brief to determine unequivocal clinical benefits. Trends suggested an unconfirmed early effect on change in EDSS score at 6 months for the medium dose (2.5 mg daily).. MS patients may be more prone to develop important linomide treatment-related adverse events than other previously studied patients. However, linomide may be a potentially more toxic drug than was suspected from observations made in smaller studies for other indications. Phase III trials may identify infrequent and important toxicities that may not be anticipated by phase I and II trials.

Topics: Adjuvants, Immunologic; Adult; Adverse Drug Reaction Reporting Systems; Aged; Disability Evaluation; Dose-Response Relationship, Drug; Double-Blind Method; Female; Humans; Hydroxyquinolines; Male; Middle Aged; Multiple Sclerosis, Chronic Progressive; Multiple Sclerosis, Relapsing-Remitting; Treatment Failure

To determine the safety and efficacy of roquinimex (linomide) in the management of relapsing-remitting and secondary progressive MS as monitored by MRI.. Preclinical studies and several short term randomized trials of linomide suggested clinical and MRI-measured benefits with acceptable risk for closely followed MS patients.. The North American Linomide Trial formally screened 853 individuals for relapsing or secondary progressive, clinically definite MS; recent disease activity or progression; and an Expanded Disability Status Scale score at entry of 3.0 to 6.5 inclusive. MRI was obtained on 811 subjects at pre-enrollment, 718 cases at enrollment, and then at three monthly intervals until the trial was prematurely terminated for unacceptable toxicity.. Enhancement was found on 40.2% of 718 entry scans. Statistically robust correlations were found between clinical demographic data and several entry MRI measures including CSF volume, a reflection of brain atrophy. Assessment of the effect of treatment on MRI-measured disease was limited by early trial termination. However, active treatment for 3 months reduced the proportion of patients with one or more enhancements. An exploratory analysis suggested that 2.5 mg was the most active of three doses tested in limiting the total volume of enhanced tissue, the proportion of MRI-defined lesions designated as "black holes," and by a novel MRI composite disease measure.. The short term signature of the effect of linomide on MRI-measured aspects of the disease suggests that safer drugs of this class might be useful in the management of MS. The use of a composite index of the heterogeneous nature of the pathology of MS as captured by MRI may have merit as an outcome measure in clinical trials.

Topics: Adjuvants, Immunologic; Adult; Adverse Drug Reaction Reporting Systems; Aged; Atrophy; Brain; Double-Blind Method; Female; Humans; Hydroxyquinolines; Magnetic Resonance Imaging; Male; Middle Aged; Multiple Sclerosis, Chronic Progressive; Multiple Sclerosis, Relapsing-Remitting; Treatment Failure

Roquinimex, Linomide, a quinoline derivative with pleiotropic immunomodulatory activity, has previously been shown to enhance natural killer (NK) cell number and activity after ABMT in patients with AML. In this study 278 AML patients in remission were randomized to receive Roquinimex 0.2 mg/kg body weight or placebo twice weekly for 2 years following ABMT. Out of 139 patients in each group, 109 Roquinimex patients and 108 placebo patients were in their first CR. Median age at inclusion was 41 years for Roquinimex patients and 39 years for placebo patients. Twelve patients in each group had their marrow purged prior to reinfusion. Relapse and death were study endpoints. Surviving patients were followed for 2.6 to 6. 9 years. The total number of relapses was 60 in the Roquinimex group and 63 in the placebo group (not significant). Leukemia-free and overall survivals were similar in the two groups. Recovery of platelet counts was significantly delayed in the Roquinimex group as compared to placebo. No other significant differences regarding toxicity parameters were recorded. In conclusion, previous findings on NK cells could not be confirmed and the study showed no benefit for Roquinimex over placebo regarding relapse or survival following ABMT for AML in remission.

Topics: Adolescent; Adult; Aged; Angiogenesis Inhibitors; Bone Marrow Purging; Bone Marrow Transplantation; Child; Combined Modality Therapy; Disease-Free Survival; Female; Follow-Up Studies; Humans; Hydroxyquinolines; Killer Cells, Natural; Leukemia, Myeloid, Acute; Male; Middle Aged; Placebos; Recurrence; Survival Rate; Time Factors; Transplantation, Autologous

The absorption and disposition of roquinimex (Linomide) were studied in four male and two female healthy volunteers. The subjects received a single oral aqueous solution of 14C-labelled roquinimex, about 0.1 mg/kg, after an overnight fast. Blood samples were taken and urine and faeces were collected for 10 days after dosing. The plasma, urine and faeces concentrations of roquinimex and metabolites were determined by high-performance liquid chromatography (HPLC) with radiochemical detection. The metabolites were identified by HPLC-mass spectroscopy (MS). The plasma concentration-time profiles of roquinimex exhibited a rapid absorption followed by a bi-exponential disposition. A secondary peak was observed between 6 and 8 h, indicating enterohepatic circulation (EHC) of roquinimex. The terminal disposition half-life was estimated as 27 h. The primary metabolic pathways of roquinimex were hydroxylation, demethylation and conjugation. The major compound in plasma was roquinimex; metabolites were only occasionally detected. In urine and faeces, roquinimex accounted for 2% of the dose and conjugated and hydroxylated metabolites each accounted for about 30% of the dose. A model was derived for the plasma concentrations of roquinimex and the amount of urinary excreted roquinimex to take into account EHC. This model improved the goodness-of-fit according to common goodness-of-fit criteria. The values of the pharmacokinetic parameters were similar using compartmental and non-compartmental methods, indicating that the contribution of EHC of roquinimex is of minor importance in the evaluation of the pharmacokinetics of roquinimex.

Topics: Adjuvants, Immunologic; Administration, Oral; Adult; Chromatography, High Pressure Liquid; Enterohepatic Circulation; Female; Half-Life; Humans; Hydroxyquinolines; Intestinal Absorption; Male; Middle Aged; Models, Biological

The reproducibility of an automated method for estimating the volume of white matter abnormalities on brain magnetic resonance (MR) images of multiple sclerosis (MS) patients was evaluated. Twenty MS patients underwent MR imaging twice within 30 minutes. Measurement variability is introduced mainly by MRI acquisition and image registration procedures, which demonstrate significantly worse reproducibility than the image segmentation. The correction of partial volume artifacts is essential for sensitive measurements of overall lesion burden. The average lesion volume difference (bias) between two MR exams of the same MS patient (N = 20) was 0.05 cm3, with a 95% confidence interval between -0.17 and +0.28 cm3, suggesting that the proposed measurement system is suitable for clinical follow-up trials, even in relatively small patient cohorts. The limits of agreement for lesion volume were between -1.3 and +1.5 cm3, implying that in individual patients changes in lesion load need to be at least this large to be detected reliably. This automated method for estimating lesion burden is a reliable tool for the evaluation of MS progression and exacerbation in patient cohorts and potentially also in individual patients.

Topics: Adjuvants, Immunologic; Adult; Analysis of Variance; Brain; Double-Blind Method; Female; Follow-Up Studies; Humans; Hydroxyquinolines; Magnetic Resonance Imaging; Multiple Sclerosis; Observer Variation; Reproducibility of Results; Time Factors

Roquinimex, Linomide, is a quinoline derivative with pleiotropic immunomodulatory activities which has been shown to enhance NK function. As part of a phase III placebo-controlled multicenter study patients were randomized to receive Roquinimex, 0.2 mg/kg body weight, or a placebo twice weekly for a duration of 2 yr following autologous bone marrow transplantation for acute myeloid leukemia in remission. At Arhus University Hospital 7 patients were randomized to receive the active drug and 6 to receive the placebo. Surviving patients were followed for 2 yr with immunological monitoring of their natural immune effector cells (NK- and LAK cell activity). Peripheral heparinized blood samples were obtained twice before the onset of conditioning therapy and at several time points after ABMT, and whole blood samples were analyzed by flow cytometry for the detection of leukocyte differentiation antigens as well as by 4 h 51Cr release assays for cytotoxicity. In contrast to previous experience with Linomide, in the present study we found that at 36 wk or later time points Linomide patients exhibited a significant suppression of circulating natural effector cell number and activity when compared with the control group. These observations underline the need for further exploration into novel and manageable immunostimulators.

Topics: Acute Disease; Adjuvants, Immunologic; Adult; Bone Marrow Transplantation; Cytotoxicity, Immunologic; Female; Humans; Hydroxyquinolines; Killer Cells, Lymphokine-Activated; Killer Cells, Natural; Leukemia, Myeloid; Leukocytes; Male; Middle Aged; Transplantation, Autologous; Treatment Outcome

Unmanipulated autologous bone marrow transplant (ABMT) offers patients with chronic myelogenous leukemia (CML) a long-term survival of 10%, at best. Immunotherapy has a role in the myeloid leukemias, and there is increasing evidence that of all hematopoietic neoplasms, CML may be the most susceptible to immune regulation. Roquinimex is known to enhance T cell, NK cell and macrophage activity. A phase II study was initiated in March 1992 to evaluate the role of roquinimex in Ph chromosome-positive CML post ABMT. Patients were conditioned with busulfan/ cyclophosphamide followed by reinfusion of unmanipulated Ph-positive bone marrow stem cells (>1 x 108 NBC/kg). When engraftment of neutrophils (ANC) reached 100/microl, patients received oral roquinimex twice weekly, escalating to a maximal dose of 0.2 mg/kg in 2 weeks. Seventeen patients have entered the study; 11 in first chronic phase (CP1); two in second chronic phase (CP2) and four in accelerated phase (AP). All required significant myelosuppressive therapy prior to ABMT to maintain stable blood counts and most had also received prior interferon therapy. All patients survived the transplant. Subsequent toxicity consisted mainly of musculoskeletal aches and peripheral edema. Additionally, specific skin changes were observed including graft-versus-host-like disease and eccrine sweat gland necrosis. Eight out of 17 patients are alive 28-60 months post ABMT. Of the nine patients who died, two were in CP2 and three in AP. All patients in CP1 went into a complete hematological remission post ABMT and seven of the 11 patients had at least a major cytogenetic response (greater than 65% Ph-negative metaphases) at 1 year or beyond and four of the 11 patients had a complete cytogenetic response at 2 years or beyond. Cytogenetic response post transplant often developed over time and did not simply represent post ABMT engraftment with Ph-negative cells. The clinical and cytogenetic data in these patients are encouraging and suggest that roquinimex may have significant activity when given post ABMT to patients with Ph-positive CML.

Topics: Adult; Aged; Antineoplastic Agents; Bone Marrow Transplantation; Combined Modality Therapy; Humans; Hydroxyquinolines; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Middle Aged; Patient Selection; Survival Analysis; Transplantation, Autologous

In order to investigate the bioavailability and the rate-limiting step of the absorption of roquinimex, an oral solution and a tablet formulation (Linomide(R)) were given to healthy volunteers. The study was conducted as a randomized three-period crossover study in seven male and seven female healthy volunteers. The subjects received an intravenous infusion, an oral solution and an oral tablet formulation, each of 5 mg (about 0.07 mg kg(-1)), as single doses after an overnight fast on three occasions, with a wash-out period of 3 weeks in between. Venous blood samples were taken over 7 days and the plasma concentrations of roquinimex were determined by high-performance liquid chromatography (HPLC) with ultraviolet (UV)-detection. The pharmacokinetics of roquinimex was characterized by a low plasma clearance, 4.9 mL h(-1) kg(-1) and a small volume of distribution, 0.22 L kg(-1). The oral bioavailability of the drug was complete for both the solution and the tablet formulation. The absorption rate was faster for the solution than for the tablet. The disposition of roquinimex was biphasic, with a terminal disposition half-life of 32 h. Between 4 and 8 hours after dosing, a secondary plasma peak was observed, indicating enterohepatic circulation of the drug. No major sex differences were shown in the pharmacokinetics of roquinimex. In conclusion, dissolution rate-limited absorption of roquinimex was shown, which demonstrates that disintegration and dissolution of the tablet play a major role in the absorption process of roquinimex. Despite the delayed absorption after administration of the tablet, the extent of absorption was complete.

Topics: Adjuvants, Immunologic; Adult; Biological Availability; Blood Proteins; Cross-Over Studies; Female; Half-Life; Humans; Hydroxyquinolines; Intestinal Absorption; Male; Protein Binding; Solubility

The quinoline-3-carboxamide, linomide, protects non-obese diabetic mice from diabetes. The effects of linomide on insulin needs and beta cell function were studied in recent juvenile Type I diabetes in a double-blind trial. Patients with recent onset diabetes were randomly assigned to treatment with a fixed dose of 2.5 mg linomide (42 patients) or placebo (21 patients) for 1 year, in addition to insulin and diet. Glycated haemoglobin was 10-15% lower at 9 months (p = 0.003) and 12 months (p < 0.05) in the linomide group. The insulin dose was 32-40% smaller in the linomide group at 3 (p < 0.03), 6 (p < 0.02), 9 (p < 0.001) and 12 months (p = 0.01). Insulin doses correlated negatively with C peptide values (p = 0.001-0.002). The trend for higher C peptide values in the linomide group did not reach significance. In a post hoc subgroup analysis performed in 40 patients (25 from the linomide group and 15 from the placebo group) who still had detectable residual beta cell function at entry, linomide was associated with 45-59% higher C peptide value at 6 months (p < 0.05), 9 months (p < 0.05) and 12 months (p < 0.05). The main adverse effects of linomide were mild transitory anaemia (45 vs 10% in the linomide and placebo groups), thrombocytopenia (24 vs 10%), and mild joint discomfort (45 vs 5%) with no clinical signs. In conclusion, low-dose linomide reduced the insulin needs in patients with juvenile Type I diabetes of recent onset and improved beta cell function in patients who still had detectable beta cell function at entry. These results support further clinical and experimental studies to define the effects of linomide in Type I diabetes provided the safety of linomide is reliably established.

Topics: Adjuvants, Immunologic; Adolescent; Adult; Antineoplastic Agents; C-Peptide; Child; Diabetes Mellitus, Type 1; Double-Blind Method; Female; Glycated Hemoglobin; Humans; Hydroxyquinolines; Hypoglycemic Agents; Insulin; Islets of Langerhans; Male

Thirty-one patients with advanced renal carcinoma or malignant melanoma were treated in the first feasibility study of alpha-interferon (Roferon) and the new oral immunomodulating agent, Linomide. Linomide 5 mg or 10 mg p.o. daily was given for 2 weeks; alpha-interferon was then added at 3 MU s.c. three times weekly, escalating in each patient by 3 MU per week, if tolerable, up to 12 MJ. The combination was poorly tolerated with nausea, vomiting, somnolence and myalgia commonly reported. Adverse events accounted for treatment withdrawal in ten patients and contributed to withdrawal in four other patients. Treatment with Linomide alone in the first 2 weeks led to a significant increase in white blood cells, neutrophils and platelets. When alpha-interferon was added, the platelet count decreased significantly over the following 6 weeks. Nineteen patients had white cell phenotype and function measured. After 2 weeks of 5 mg Linomide, a transient but significant decrease in the absolute number of activated T-helper cells (CD4+DR+) was observed. No changes in natural killer (NK) cell number or activity were observed. Twenty-two patients were evaluable for response. One with metastatic renal cell carcinoma had a complete response and six had stable disease. This study does not support the use of the combination because significant toxicity was seen without the anticipated immunological benefits.

Topics: Adjuvants, Immunologic; Adult; Aged; Antineoplastic Combined Chemotherapy Protocols; Dose-Response Relationship, Drug; Feasibility Studies; Humans; Hydroxyquinolines; Interferon-alpha; Kidney Neoplasms; Male; Melanoma; Middle Aged; Treatment Outcome

A Phase II clinical trial was undertaken using roquinimex (Linomide) in patients with myelodysplastic syndromes (MDS). Roquinimex is an orally active drug with immunostimulating activities demonstrated in vitro and clinically. Seventeen patients with MDS were enrolled in the study. Eligibility was limited to cytopenic patients with <20% marrow blasts. The drug was given orally twice weekly for 12 weeks with frequent monitoring of clinical, hematologic, and immunologic parameters. An increase in CD8+ and CD56+/CD3- cells was detected by 3 weeks. There was, however, no augmentation of natural killer or lymphokine-activated killer cell activity; progenitor cells were unchanged. Four patients had improvement in neutrophil counts, and two patients had improvement in platelet counts. Despite this improvement, the responses were transient or not maintained after discontinuation of therapy. One patient with RAEB, who was red cell transfusion dependent, experienced a complete remission that has persisted 14 months after completion of therapy. Adverse events developed in >25% of patients and included arthralgia, fever, headache, and myalgia. These side effects led to early withdrawal of therapy in five patients. These findings suggest that roquinimex may be of occasional benefit to patients with myelodysplastic syndromes.

Topics: Adjuvants, Immunologic; Aged; Female; Hematopoiesis; Humans; Hydroxyquinolines; Male; Middle Aged; Myelodysplastic Syndromes; Prospective Studies

Following a previous EORTC GU-Group study, in which linomide showed some activity in poor prognosis patients, this study was initiated to determine the effect of linomide in more favourable patients. 35 patients with metastatic renal cell carcinoma with good prognostic factors, i.e. good performance status, prior nephrectomy, no prior systemic therapy, and no liver, bone or brain metastases, were treated with linomide, a quinoline derivative with immunomodulating properties, at a dose of 10 mg daily, after an initial dose escalation during the first 4 weeks of treatment. In 29 evaluable patients, no responses were observed (95% confidence interval 0-10%). Best overall response was no change in 9 patients, for a median duration of 4 months. Linomide in this schedule was poorly tolerated, with 17% (6 patients) of patients being withdrawn and 23% (8 patients) having dose reductions due to adverse events, mostly influenza-like symptoms of myalgia, arthralgia and fatigue. Several cases of pericarditis and neuropathy were observed. In spite of selection of favourable prognosis patients and an optimal daily dosing schedule, linomide was not an effective treatment in renal cell carcinoma. In view of toxicity and lack of efficacy, there is no rationale in further testing the drug in this disease.

Topics: Antineoplastic Agents; Carcinoma, Renal Cell; Female; Humans; Hydroxyquinolines; Kidney Neoplasms; Male; Prognosis

The aim of this study was to determine the objective tumour response rate and duration of response and toxicity of linomide (Roquinimex) treatment in patients with disseminated renal cell carcinoma, pretreated or not pretreated with immunotherapy. From March 1991 to July 1992, 72 patients with metastatic and progressive renal cell cancer were entered of whom 9 (12%) were not evaluable for response. Linomide was given orally, twice weekly, 5 mg during the first week with dose escalation to 10 mg during the second week and 15 mg thereafter. Treatment was continued until disease progression or unacceptable toxicity. No haematological toxicity but slight anaemia was observed. A significant WBC (white blood cell count) increase (P < 0.0001, paired T-test) was found during treatment. The most often reported non-haematological side-effects were: flu-like syndrome (54%, grade III-IV 7%), nausea/vomiting (41% and 3%, respectively) and neurotoxicity (34% and 2%). Most side-effects were of mild or moderate intensity (WHO grade 1 or 2). The objective overall response rate was 4%: 1 CR and 2 PRs. Stable disease was reported for 28 patients (40%). The duration of response was 17, 22 and 30 (CR) months. Median time to progression was 5 months. Linomide at the given dose and schedule is well tolerated, but has limited antitumour activity in metastatic renal cell carcinoma.

Topics: Adult; Aged; Antineoplastic Agents; Carcinoma, Renal Cell; Female; Humans; Hydroxyquinolines; Immunotherapy; Kidney Neoplasms; Male; Middle Aged

Roquinimex (Linomide) has been demonstrated to suppress tumor growth in animal models. The effect is at least in part related to enhanced numbers and activity of natural killer (NK) cells. In this clinical pilot study, roquinimex was given at increasing doses (0.05 mg/kg to 0.6 mg/kg) to 13 patients (performance status 0-3) with various malignant disorders. Immunology parameters were followed and side effects were observed during the study. The plasma pharmacokinetics of roquinimex was studied at the 0.2 mg/kg dose level. The clinical side effects were dominated by musculoskeletal discomfort, nausea, and pain. No significant hematological or biochemical toxicity was observed. Pharmacokinetic analysis at the 0.2 mg/kg dose level revealed a Cmax of 4.0 mumol/L at tmax of 1.2 hr and an elimination half-life of 42 hr. Increased numbers of phenotypic NK cells, activated T (DR+CD4+) cells, and monocytes were observed after administration of roquinimex compared with pretreatment values. Roquinimex seems to be an active immunomodulator with manageable toxicity. Further exploration of therapeutic efficacy is warranted.

Topics: Adjuvants, Immunologic; Adult; Aged; Antineoplastic Agents; Female; Humans; Hydroxyquinolines; Killer Cells, Natural; Male; Metabolic Clearance Rate; Middle Aged; Neoplasms; Pilot Projects; T-Lymphocytes

Linomide (quinoline-3-carboxamide) is a synthetic immunomodulator that increases the natural killer cell activity. We previously demonstrated that linomide effectively inhibited the clinical and histopathologic signs of acute and chronic relapsing experimental autoimmune encephalomyelitis. We report a double-blind, placebo-controlled study to evaluate tolerability and to obtain preliminary indications of the clinical efficacy of linomide on secondary progressive MS. Thirty patients suffering from clinically definite and laboratory-supported secondary progressive MS, with an expanded disability status scale (EDSS) of 3.0 to 7.0, were included in this study. Patients were treated daily with linomide (2.5 mg) or placebo orally and were followed up for side effects and changes in their neurologic status; monthly MRI scans were taken throughout the treatment period. Twenty-four patients completed at least 6 months of treatment. Mild to moderate side effects, including muscle pains, arthralgia, and edema, were present in 11 of the 15 patients receiving placebo and in 13 of the 15 patients treated with linomide. At 24 weeks, the mean shift in EDSS was +0.272 +/- 0.156 in the placebo group versus -0.166 +/- 0.167 in the linomide group (p = 0.0451). The percentage of patients with evidence of "activity" on their MRI (new, enlarging, or new gadolinium diethylenetriaminepentaacetic acid [Gd-DTPA]-enhancing lesions) throughout the treatment period was 75% in the placebo group and 33% in the linomide group (p = 0.0205). The mean total number of new Gd-DTPA-enhancing lesions per MRI scan for the same period was 0.42 +/- 0.143 in the placebo group and 0.19 +/- 0.114 in the linomide group (p = 0.0387). In this study, linomide proved to be safe and well tolerated in patients with secondary progressive MS. In addition, our results indicate that linomide tends to inhibit the progression of the disease, especially preventing the appearance of new active lesions in the MRI scans. Based on these results, two multicenter phase III trials are currently under way in the United States and in Europe and Australia.

Topics: Adjuvants, Immunologic; Adult; Double-Blind Method; Female; Humans; Hydroxyquinolines; Magnetic Resonance Imaging; Male; Middle Aged; Multiple Sclerosis; Pilot Projects

Topics: Adjuvants, Immunologic; Administration, Oral; Adolescent; Adult; Double-Blind Method; Female; Humans; Hydroxyquinolines; Male; Middle Aged; Multiple Sclerosis

The synthetic immunomodulator Linomide, a quinoline-3-carboxamide, has a profound inhibitory influence in several experimental autoimmune diseases, including acute and chronic experimental allergic encephalomyelitis. In a double-blind trial, 31 patients with relapsing-remitting multiple sclerosis were randomized to oral doses of 2.5 mg Linomide or placebo once a day for six months. Fourteen patients receiving Linomide and 14 receiving placebo completed the trial, and the results were based on this population. The mean number of active (new and enlarged T2 weighted) lesions per monthly MRI scan was 1.37 in the patients receiving Linomide and 4.22 in the patients receiving placebo (p = 0.043). The percentage of scans with active MRI lesions was lower in the Linomide-treated group (p = 0.0064). When neurologic deficit was assessed by the Regional Functional Scoring System (RFSS), the Linomide group showed an improvement of 1% of the maximal RFSS range and the placebo group a deterioration of 0.2% (p = 0.14). There were three patients with relapses in the Linomide-treated group and six in the placebo group (p = 0.22). A slightly decreased proportion of natural killer cells in cerebrospinal fluid and peripheral blood was noted in the Linomide group. A severe adverse event of pleuropericarditis occurred in one of the Linomide-treated patients. The most frequent adverse event was musculoskeletal pain, of mild to severe degree, which tended to diminish after three months on Linomide therapy.

Topics: Adjuvants, Immunologic; Adolescent; Adult; Double-Blind Method; Female; Humans; Hydroxyquinolines; Magnetic Resonance Imaging; Male; Middle Aged; Multiple Sclerosis; Recurrence

Recently, the synthetic immunomodulator Linomide (quinoline-3-carboxamide, LS 2616) was reported to prevent IDDM and insulitis in NOD mice. The mechanism for this protective effect is not known. The cytokine interleukin 1 (IL-1) may be a pathogenetic factor in the initial destruction of the beta-cells leading to IDDM. This study was undertaken to investigate the influence of Linomide on IL-1beta induced diabetogenic and hormonal changes in the rat in vivo, and on IL-1beta mediated synthesis of NO and inhibition of insulin secretion in isolated islets of Langerhans ex vivo. Normal male Wistar Kyoto rats received 4.0 microg/kg of recombinant human IL-1beta (rhIL-1beta) i.p. daily for 5 days with or without Linomide (8-9 mg/kg/day) in the drinking water. Litters of neonatal Wistar rats were pretreated for 3 days with injections of 10 mg/kg of Linomide i.p., and pancreatic islets of Langerhans were isolated for ex vivo studies. Linomide alone caused significant hypercorticosteronemia, hypoglucagonemia, lymphopenia and neutrophilia. Linomide had no effect on IL-1beta induced hyperglycemia, hyperglucagonemia, lymphopenia, neutrocytosis, or hypercorticosteronemia on day three and hypocorticosteronemia on day five. Further, Linomide did not prevent rhIL-1beta mediated reduction in insulin secretion or increase in NO synthesis ex vivo. In conclusion, Linomide does not seem to exert its protective effect on IDDM development via inhibition of interleukin 1 action on islet insulin release or NO production, but the increase in plasma corticosterone may contribute to the understanding of the immunomodulatory effects of Linomide.

Topics: Adjuvants, Immunologic; Animals; Blood Cell Count; Corticosterone; Glucagon; Humans; Hydroxyquinolines; Insulin Antagonists; Interleukin-1; Islets of Langerhans; Male; Random Allocation; Rats; Rats, Inbred WKY

Linomide, a synthetic quinoline carboxamide, has the ability to stimulate various lymphocyte subpopulations. We have shown its inhibitory effect on the clinical and histological signs of acute and chronic relapsing EAE. In these models linomide induces suppression of lymphocyte response to antigens, production of autoantibody, antigen presentation to specific T-cell lines and Mac-I expression, and induces activation of NK and suppressor-inducer cells. We have subsequently shown its inhibitory effect on clinical and MRI signs of patients with secondary progressive multiple sclerosis. Results of a double blind, placebo controlled, short term pilot study with p.o. linomide, showed a significant effect on the clinical disability scale (EDSS) (P = 0.045) and on the mean total number of new lesions in serial monthly MRI scans (P = 0.021). The increase of CD45Ra, CD8 and CD16 positive cells in linomide treated patients may indicate the importance of suppressor-inducer, suppressor and NK cells for the inhibition of the autoimmune response in the disease.

Topics: Adjuvants, Immunologic; Adult; Disease Progression; Double-Blind Method; Encephalomyelitis, Autoimmune, Experimental; Female; Humans; Hydroxyquinolines; Lymphocytes; Macrophages; Magnetic Resonance Imaging; Male; Middle Aged; Multiple Sclerosis; Pilot Projects

Topics: Adjuvants, Immunologic; Double-Blind Method; Humans; Hydroxyquinolines; Multiple Sclerosis; Recurrence; Remission Induction; Treatment Outcome

Quinoline-3-carboxamide (Linomide) is a novel, synthetic immunomodulator acting via immunologic and non-immunologic mechanisms. It has shown efficacy against various malignancies, experimental autoimmune encephalomyelitis, and septic shock in animal models and has been investigated for clinical use in minimal residual myeloid leukemia with promising results. Interleukin-2 has shown considerable efficacy in palliative anti-tumor-treatment of advanced renal cell cancer, revealing remission rates of up to 40% in combination therapy regimens. Linomide is reported to exhibit synergistic effects with interleukin-2. Here we report on a clinical phase I/II study examining tolerance and efficacy of a combination therapy schedule of SQ interleukin-2 and PO Linomide in advanced renal cell cancer. Seventeen patients received 10 IU/m2 interleukin-2 per week for 8 weeks, resting interleukin-2 for another 8 weeks. In week 5 they started 5 mg Linomide daily, continued with 10 mg from week 7 to 16. No objective remissions were observed. Among 15 patients evaluable for response, 10 (66.7%) were progredient during the study. Three patients died during the observation period, including two not evaluable for response. Median survival was 4.0 months, median progression-free survival 2.5 months with a Kaplan-Meier estimate of 3.63 months. Fever, reduced general condition, nausea/vomiting, dyspnea, anorexia, chills and hypotension were the most common side effects, reaching WHO grade 3 in 6 and grade 4 in 2 cases. In summary, Linomide in combination with interleukin-2 provides no advantages in efficacy or toxicity over other therapy regimens employing interleukin-2.

Topics: Adjuvants, Immunologic; Adult; Aged; Carcinoma, Renal Cell; Female; Humans; Hydroxyquinolines; Interleukin-2; Kidney Neoplasms; Male; Middle Aged

Patients with chronic myelogenous leukemia (CML) can be cured with allogeneic bone marrow transplantation. Over the past decade, it has become clear that immunological mechanisms, in the form of graft-versus-leukemia, constitute an integral part of this therapy. Because of limitations imposed by a lack of suitable donors, age, and toxicity, only a minority of patients can be offered allogeneic bone marrow transplantation (BMT). Recently, attempts have been made to employ autologous bone marrow transplantation (ABMT) for the therapy of CML using a variety of pre- and post-transplantation manipulations. This report describes the rationale for an ongoing clinical trial using the immunomodulator roquinimex (Linomide), following autologous bone marrow transplantation, in an attempt to stimulate the immunological responses thought to be critical for successful therapy in CML.

Topics: Adjuvants, Immunologic; Adolescent; Adult; Aged; Bone Marrow Transplantation; Combined Modality Therapy; Female; Humans; Hydroxyquinolines; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Male; Middle Aged; Transplantation, Autologous

Immunostimulatory therapy is at present considered after autologous bone marrow transplantation (ABMT) in order to mimic the allogeneic graft-versus-leukemia effect and thereby reduce the relapse rate. In a pilot study, five adults with acute myeloid leukemia were treated with the new immunomodulator Linomide post-ABMT. Linomide (0.3 mg/kg/week orally) was given in cycles of three weeks followed by three weeks of rest for up to six months. During treatment periods cyclic increases of CD56+CD3- and CD16+ NK cells were observed in parallel with enhanced cytotoxic activity of patient cells against both the NK-sensitive K562 and NK-resistant Daudi cell lines. A cyclic increase of CD14+ monocytic cells was also recorded. The proliferative responses of patient cells to PHA and allogeneic cells (MLC) were enhanced during Linomide therapy. The in vitro production of TNF alpha, IFN gamma, and IL-1 followed the same cyclic increase during treatment periods. Side effects were generally mild, and no harmful effects on engraftment were seen. Linomide therapy after ABMT thus induces a broad immunostimulation that offers a potential benefit with regard to leukemia-free survival.

Topics: Adjuvants, Immunologic; Antigens, CD; Bone Marrow Transplantation; Cytokines; Female; Humans; Hydroxyquinolines; Killer Cells, Natural; Leukemia, Myeloid, Acute; Lymphocyte Activation; Male; Middle Aged; Monocytes; Pilot Projects; T-Lymphocytes

According to WHO, in 2017, about 90.5 million people suffered from cancer and about 8.8 million deaths occurred due to disease. Although the chemotherapeutic agents have decreased the mortality among the cancer patients but high toxicity and non-specific targets are still major drawbacks. Many researchers have identified linomide, a 4-hydroxy-2-quinolone derivative, as a lead molecule for the development of anticancer agents. With this background, we thought of the following objective.. The objective of this research work involves the synthesis of a series of N-(2-(4- hydroxy-2-oxo-1-phenyl-1,2-dihydroquinolin-3-yl)-2-oxoethyl)-N-alkyl substituted benzene sulfonamides IVa-d (1-3) by replacing the anilide moiety at the third position of linomide with sulfamoylacyl and also N-methyl by N-phenyl functionality. To perform in silico anticancer activity by using Molegro Virtual Docker (MVD-2013, 6.0) software and in vitro anticancer activity by MTT assay.. The starting material 4-hydroxy-1-phenylquinolin-2(1H)-one was treated with N-bromosuccinamide to yield compound II. Condensation of compound II with primary amines resulted in compounds IIIa-d, which, on coupling with substituted aromatic sulfonyl chlorides yield the title compounds IVa-d (1-3).. All the synthesized compounds were satisfactorily characterized by spectral data. The results of docking revealed that the synthesized compounds exhibited well-conserved hydrogen bonds with one or more amino acid residues in the active pocket of EGFRK tyrosine kinase domain (PDB ID: 1m17). The MolDock Score of compound IVd-1 (-115.503) was the highest amongst those tested. The in vitro anticancer activity results showed that compound IVc-1 (R= - (CH2) 2-CH3 ; R'= -H) and IV d-1 (R= -CH2-C6H5; R'= -H) were found to be most potent against K562 cell line with an IC50 of 0.451 μM/ml and 0.455 μM/ml respectively. Compound IVd-1 also showed better potency against A549 cell line with IC50 value of 0.704 μM/ml.. The results of in silico and in vitro anticancer activity are in agreement with each other. Compound IV d-1 was found to be most active of the series.

Topics: A549 Cells; Antineoplastic Agents; Catalytic Domain; Cell Proliferation; Crystallography, X-Ray; Drug Design; Drug Screening Assays, Antitumor; ErbB Receptors; Humans; Hydrogen Bonding; Hydroxyquinolines; Inhibitory Concentration 50; K562 Cells; Molecular Docking Simulation; Neoplasms; Structure-Activity Relationship

Transferring parental splenocytes into unirradiated F1 mice induces a chronic graft-versus-host disease (GVHD), characterized by the production of Th2 cytokines and immunocomplex-mediated glomerulonephritis resembling systemic lupus erythematosus (SLE). The effects of H1521, a new derivative of 4-hydroxyquinoline-3-carboxamide, were investigated in chronic GVHD lupus model. H1521 was administered to chronic GVHD mice for 10 weeks. Nephritic symptoms were monitored and cytokine expression in the spleen was detected. To clarify the direct effect of H1521 on CD4(+) T cell, CD4(+) T cells were isolated and co-cultured with H1521 under neutral and Th1 or Th2 driving conditions in vitro. H1521 (32 mg/kg) reduced the incidence of proteinuria by 50% in chronic GVHD mice. Ameliorated lupus symptoms and improved renal histopathology damage were also observed. Administration of H1521 had little impact on Th1 cytokine IL-2 and IFN-gamma or Th2 cytokine IL-4 and IL-10 mRNA expression. In contrast, severely deficient IFN-gamma production by concanavalin A-stimulated spleen cells in chronic GVHD mice was completely restored by H1521. In accordance with this, decreased T-bet mRNA expression became normalized with H1521 (32 mg/kg) treatment. In addition, in vitro studies demonstrated that H1521 preferentially favored Th1 differentiation in CD4(+) T cell and promoted IFN-gamma secretion in Th1 differential CD4(+) T cell. However, IL-4 secretion in naive or Th2 differential CD4(+) T cell was unaffected by H1521. In conclusion, H1521 can induce Th1 cytokine profile in CD4(+) T cells and has possible therapeutic value in Th2-predominant immune diseases.

Topics: Adjuvants, Immunologic; Animals; Chronic Disease; Cytokines; Disease Models, Animal; Female; Graft vs Host Disease; Hydroxyquinolines; Lupus Erythematosus, Systemic; Mice; Mice, Inbred Strains; Th1 Cells; Th2 Cells

Roquinimex is an immunomodulator that can effectively inhibit the development of several autoimmune diseases in animal models, but the mechanism is still unknown. In this study, we investigated the effect of roquinimex on chronic graft-versus-host disease (GVHD) in mice, a well-established model for human systemic lupus erythematosus (SLE). Oral administration of roquinimex significantly suppressed the development of proteinuria and ameliorated nephritis symptoms in chronic GVHD mice. In addition, renal histopathology and immunohistochemistry studies revealed reduced glomerulonephritis and decreased IgG deposition in chronic GVHD mice treated with roquinimex. Chronic GVHD is characterized by a predominance of Th2 cytokines, and proinflammatory cytokines that also play an important role in the pathology of tissue damage. Therefore, we focused on the effect of roquinimex on cytokine production. Chronic GVHD mouse splenocytes exhibited severely reduced interferon (IFN)-gamma production in response to Concanavalin (Con A) stimulation and an overt Th2 skewness. Roquinimex treatment, however, induced IFN-gamma production and restored the Th1/Th2 cytokine balance, although only a minimal effect of roquinimex on interleukin (IL)-4 secretion was observed. The production of the proinflammatory cytokines TNF-alpha and IL-1 beta by peritoneal macrophages from lipopolysaccharide (LPS)-treated GVHD mice was significantly inhibited by roquinimex treatment. These data suggested that the beneficial effect of roquinimex on lupus might, at least in part, result from a restoration of Th1/Th2 cytokine balance and inhibition of inflammatory cytokine production.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Antinuclear; Blood Urea Nitrogen; Cells, Cultured; Cholesterol; Chronic Disease; Creatinine; Cytokines; Enzyme-Linked Immunosorbent Assay; Female; Graft vs Host Disease; Hydroxyquinolines; Immunoglobulin G; Inflammation Mediators; Kidney; Macrophages; Male; Mice; Mice, Inbred DBA; Proteinuria; Spleen; Th1 Cells; Th2 Cells; Triglycerides

Linomide is an immunomodulator that can effectively inhibit the development of several autoimmune diseases in animal models. Previously, linomide was shown to influence macrophage function, although the mechanism was elusive. In this study, we investigated the effect of linomide on the macrophage inflammatory cytokine, tumor necrosis factor-alpha (TNF-alpha), production induced by lipopolysaccharide (LPS) in vitro on the murine macrophage cell line, RAW264.7. Linomide exposure reduced LPS-evoked TNF-alpha production in a dose-dependent manner. Gel shift and reporter gene analyses revealed linomide inhibited LPS-induced NF-kappaB binding to the NF-kappaB consensus oligonucleotide and NF-kappaB-mediated reporter gene expression. Immunoblot analysis showed that linomide inhibited phosphorylation of p38 kinase and c-jun N terminal kinase (JNK) in LPS-stimulated RAW264.7 cells. Taken together, these results suggest that linomide inhibits TNF-alpha production by suppressing the activation of NF-kappaB and mitogen-activated protein kinase (MAPK), which might, at least in part, contribute to the beneficial effects of linomide in the treatment of autoimmune diseases.

Topics: Animals; Cell Line; Enzyme Activation; Hydroxyquinolines; JNK Mitogen-Activated Protein Kinases; Lipopolysaccharides; Macrophages; Mice; NF-kappa B; p38 Mitogen-Activated Protein Kinases; Tumor Necrosis Factor-alpha

Replicating, neuroattenuated gamma(1)34.5-deleted herpes simplex virus (HSV)-vectors are tools for experimental therapy of gliomas and autoimmune diseases. Immunomodulative treatment with Linomide (quinoline-3-carboxamide) has earlier been shown to facilitate some virus infections and reduce autoimmunity. Now we aimed at elucidating the safety of immunomodulatory therapy during infection of mice with HSV vectors. We focused on immunological and virological changes in the nervous system. BALB/c mice were infected intranasally with the HSV-1 recombinant viruses R3616, R3659 and R8306 (with mouse IL-4 transgene) and either treated with Linomide or left untreated as control groups. Treatment with Linomide was started 7 days before infection. Virological analysis consisted of viral culture and PCR for HSV DNA. Cytokine responses were studied with quantitative RT-PCR and EIA. Immunomodulatory treatment did not change the clinical course of infections. The expression of IL-4 and IL-10 in brains increased in Linomide-treated mice, particularly in infection with R8306. The expression of IL-23p19 was decreased in brains in Linomide-treated, vector-infected mice, in comparison with nontreated but virus-infected animals. Immunomodulatory treatment did not increase the viral load in brains in any of the mouse groups infected with R3616, R3659 or R8306. Immunomodulative treatment with Linomide did not compromise the safety of replicating HSV-vectors, not even the one with IL-4 transgene, suggesting that combination of immunomodulation with virotherapy may be beneficial in the treatment of certain diseases of the central nervous system. Further investigations are needed to elucidate the effects of immunomodulatory therapy in order to improve vector survival and efficacy of gene therapy.

Topics: Adjuvants, Immunologic; Animals; Brain; Cytokines; Female; Gene Expression Regulation; Genetic Therapy; Genetic Vectors; Herpes Simplex; Herpesvirus 1, Human; Hydroxyquinolines; Immunotherapy; Mice; Mice, Inbred BALB C; RNA, Messenger; Spleen; Th2 Cells; Trigeminal Ganglion

Linomide, Figure 1, produces robust and consistent in vivo growth inhibition of prostate cancer models via its anti-angiogenic activity and inhibition of autoimmune encephalomyelitis models of multiple sclerosis (MS). MS clinical trials were discontinued because of unacceptable toxicity, due to dose-dependent induction of proinflammation.. Therefore, linomide analogs were initially screened to determine their in vivo potency to inhibit growth of the Dunning R-3327 AT-1 rat prostate cancer model in rats and their potency to inhibit angiogenesis in a Matrigel assay in mice.. Based upon its superior potency (i.e., 30- to 60-fold more potent than linomide) in these assays and its lack of a proinflammation in the Beagle-dog, ABR-215050 (tasquinimod), Figure 1, was characterized for dose-response ability to inhibit the growth of a series of four additional human and rodent prostate cancer models in mice. Pharmacokinetic analysis following oral dosing documented that blood and tumor tissue levels of ABR-215050 as low as 0.5-1 microM are therapeutically effective. This efficacy is correlated with inhibition of angiogenesis in a variety of assays (endothelial capillary tube formation, aortic ring assay, chorioallantoic membrane assay, real-time tumor blood flow and PO(2) measurements, tumor blood vessel density, and tumor hypoxic and apoptotic fractions).. Based upon its robust and consistent anti-angiogenic activity and thus tumor growth, ABR-215050 has entered clinical trials for the treatment of prostate cancer.

Topics: Angiogenesis Inhibitors; Animals; Cell Growth Processes; Cell Line, Tumor; Dogs; Humans; Hydroxyquinolines; Male; Mice; Neovascularization, Pathologic; Prostatic Neoplasms; Quinolines; Quinolones; Rats; Xenograft Model Antitumor Assays

Linomide (quinoline-3-carboxamide) is an immunomodulator with anti-inflammatory effects in rodents with autoimmune diseases. Its mode of action still remains to be elucidated. We hypothesized that an investigation of T cell interactions with the extracellular matrix (ECM), composed of glycoproteins such as fibronectin (FN) and laminin (LN), might provide better understanding of their in vivo mode of action in extravascular inflammatory sites. We examined the effect of Linomide on T cell adhesion to intact ECM, and separately to LN, and FN, and on the release and production of tumor necrosis factor (TNFalpha) and nitrogen oxide (NO) in relation to adhesive molecules in non-obese diabetic (NOD) female spleen cells, focusing on intracellular adhesion molecule-1 (ICAM-1) and CD44. NOD female mice that developed spontaneous autoimmune insulitis, which destroys pancreatic islets and subsequently leads to insulin-deficient diabetes mellitus, were studied. Linomide, given in the drinking water or added to tissue cultures in vitro, inhibited the beta1 integrin-mediated adhesion of T cells to ECM, FN and LN, as well as the production and release of TNFalpha and NO, which play a major role in the induction and propagation of T cell-mediated insulitis. In addition, exposure of T cells to Linomide resulted in increased expression of CD44 and ICAM-1 molecules on spleen cells of Linomide-treated mice; such an increase in adhesion molecule expression may lead to more effective arrest of T cell migration in vivo. The regulation of T-cell adhesion, adhesion receptor expression, and inhibition of TNFalpha and NO secretion by Linomide may explain its beneficial role and provide a new tool for suppressing self-reactive T cell-dependent autoimmune diseases.

Topics: Adjuvants, Immunologic; Administration, Oral; Animals; Cell Adhesion; Extracellular Matrix; Female; Hyaluronan Receptors; Hydroxyquinolines; Intercellular Adhesion Molecule-1; Mice; Mice, Inbred NOD; Nitric Oxide; Spleen; T-Lymphocytes; Tumor Necrosis Factor-alpha

A series of novel quinoline-3-carboxamide derivatives were synthesized and evaluated for their immunomodulatory activity. The compounds were tested in vitro for effects on spleen lymphocyte proliferation and TNF-alpha production by macrophage. Three compounds showed immunomodulatory profiles similar to and more potent than those of linomide and FR137316 and were selected for further pharmacological studies in vivo.

Topics: Animals; Cell Proliferation; Cells, Cultured; Drug Design; Drug Screening Assays, Antitumor; Hydroxyquinolines; Immunologic Factors; Lymphocytes; Macrophages; Quinolines; Rats; Spleen; Tumor Necrosis Factor-alpha

Roquinimex-related 3-quinolinecarboxamide derivatives were prepared and evaluated for treatment of autoimmune disorders. The compounds were tested in mice for their inhibitory effects on disease development in the acute experimental autoimmune encephalomyelitis model and selected compounds in the beagle dog for induction of proinflammatory reaction. Structure-activity relationships are discussed. Compound 8c, laquinimod, showed improved potency and superior toxicological profile compared to the lead compound roquinimex (1b, Linomide) and was selected for clinical studies (currently in phase II).

Topics: Adjuvants, Immunologic; Animals; Autoimmune Diseases; Dogs; Encephalomyelitis, Autoimmune, Experimental; Female; Hydroxyquinolines; Mice; Mice, Inbred C57BL; Quinolones; Structure-Activity Relationship

Herpes simplex virus (HSV) is a common neurotropic virus which infects epithelial cells and subsequently the trigeminal ganglia (TG) and brain tissue. We studied how immunomodulation with roquinimex (Linomide) affects the course of corneal HSV infection in BALB/c mice. BALB/c mice have also been used in a model for HSV-based vectors in treating an autoimmune disease of the central nervous system (CNS). We addressed the questions of how immunomodulation affects the local as well as the systemic immune response and whether roquinimex could facilitate the spread of HSV to the CNS. The cytokine response in the brain and TG was studied using a quantitative rapid real-time RT-PCR method. We were interested in whether immunomodulation affects the expression of the recently described Th1-cytokine IL-23p19 in the brain and TG. The expression of IL-23 mRNA was decreased in brains of roquinimex-treated BALB/c mice. Also the expression of IL-12p35 and IFN-gamma mRNAs decreased. No significant changes were seen in IL-4 and IL-10 mRNA expression. The cytokine response was also studied using supernatants of stimulated splenocytes by EIA. Roquinimex treatment suppressed the production of IFN-gamma and also the production of IL-10 in HSV-infected BALB/c mice.

Topics: Adjuvants, Immunologic; Animals; Brain; Cytokines; Female; Gene Expression Regulation; Herpesvirus 1, Human; Hydroxyquinolines; Interferon-gamma; Interleukin-10; Interleukin-23; Interleukin-23 Subunit p19; Interleukins; Keratitis, Herpetic; Mice; RNA, Messenger

The immunomodulator Linomide has been shown to protect against septic liver injury by reducing hepatic accumulation of leukocytes although the detailed anti-inflammatory mechanisms remain elusive. This study examined the effect of Linomide on the production of CXC chemokines, including macrophage inflammatory protein-2 (MIP-2) and cytokine-induced neutrophil chemoattractant (KC) and interleukin-10 (IL-10) in lipopolysaccharide (LPS)/d-galactosamine (Gal)-induced liver injury in mice. It was found that pretreatment with 300 mg kg(-1) of Linomide markedly suppressed leukocyte recruitment, perfusion failure, and hepatocellular damage and apoptosis in the liver of endotoxemic mice. Administration of Linomide inhibited endotoxin-induced gene expression of MIP-2 and KC and significantly reduced the hepatic production of MIP-2 and KC by 63 and 80%, respectively. Moreover, it was found that Linomide increased the liver content of IL-10 by more than three-fold in endotoxemic mice. The protective effect of Linomide against endotoxin-induced inflammation and liver injury was abolished in IL-10-deficient mice, suggesting that the beneficial effect of Linomide is dependent on the function of IL-10. Taken together, these novel findings suggest that the protective effect of Linomide is mediated via local upregulation of IL-10, which in turn decreases the generation of CXC chemokines and pathological recruitment of leukocytes in the liver of endotoxemic mice.

Topics: Alanine Transaminase; Animals; Apoptosis; Aspartate Aminotransferases; Chemical and Drug Induced Liver Injury; Chemokine CXCL1; Chemokines, CXC; Endotoxins; Enzyme-Linked Immunosorbent Assay; Hydroxyquinolines; Immunologic Factors; Intercellular Signaling Peptides and Proteins; Interleukin-10; Interleukin-16; Leukocytes; Liver; Liver Function Tests; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Reverse Transcriptase Polymerase Chain Reaction

Linomide is an immunomodulator which has been shown to potently inhibit autoimmunity in several animal models for human autoimmune diseases, including type I diabetes in the nonobese diabetic (NOD) mouse. In this study, we investigate the basis for Linomide's protective effects in the NOD mouse by immunohistochemical and RT-PCR analysis of the phenotype and cytokine expression by cells infiltrating the islets of Langerhans in the pancreas. Linomide treatment was found to reduce the infiltration of T cells, B cells, dendritic cells (DC) and MHC class II(+) cells into the islets, but did not reduce macrophage (MPhi) infiltration. This was seen following Linomide treatment at 3-5, 4-8 and 14-24 weeks of age and thus appears to be independent of the stage of the autoreactive process and the extent of insulitis. The reduced insulitis may be due to reduced expression of adhesion molecules since decreased numbers of islet-associated blood vessels expressing CD106 and MAdCAM-1 were detected following Linomide treatment. Furthermore, short term Linomide treatment (3 or 7 days), which did not alter the number of infiltrating cells, was found to inhibit the production of TNF-alpha which is known to induce the expression of CD106 and MAdCAM-1. These results suggest that the reduced insulitis observed in Linomide-treated animals is secondary to a functional modulation of infiltrating cells.

Topics: Aging; Animals; B-Lymphocytes; Cell Movement; Cytokines; Dendritic Cells; Female; Gene Expression Regulation; Hydroxyquinolines; Insulin; Islets of Langerhans; Macrophages; Mice; Mice, Inbred NOD; T-Lymphocytes; Tumor Necrosis Factor-alpha

The synthesis and anti-angiogenic activities of linomide and its analogues are reported. Three of the analogues are 3.3-69 times more potent than linomide at inhibiting blood vessel formation in the CAM angiogenesis assay. These compounds possessed considerable anti-proliferative activity against isolated HUVEC cells with no activity against epithelial-derived prostate tumor cells.

Topics: Allantoin; Angiogenesis Inhibitors; Animals; Cell Division; Chick Embryo; Endothelium, Vascular; Humans; Hydroxyquinolines; Male; Membranes; Muscle, Smooth, Vascular; Prostatic Neoplasms; Regional Blood Flow; Structure-Activity Relationship; Tumor Cells, Cultured

Roquinimex is a modulator of the immune system and has been shown to attenuate induction of several inflammatory and autoimmune diseases. The objective of the present study was to determine the efficacy of roquinimex in a model of murine colitis.. For this purpose, Balb/c mice were exposed to 5% dextran sodium sulfate (DSS) in the drinking water for five to six days. Roquinimex (300 mg kg(-1) day(-1)) was administered by subcutaneous (s.c.) injection 3 days prior to and throughout the treatment period with DSS. In separate experiments, 300 mg kg(-1) day(-1) of roquinimex was given therapeutically after initiation of DSS challenge.. DSS provoked clinical signs of colitis, reduced crypt height (CH) and increased mucosal damage score (MDS) as analyzed by histology. In addition, challenge with DSS increased the colonic content of myeloperoxidase (MPO). Prophylactic administration of DSS-treated mice with roquinimex significantly reduced clinical signs of colitis, MDS and the CH-reduction. Moreover, in roquinimex treated animals, the MPO activity was significantly reduced by more than 50% compared to DSS control mice. Notably, therapeutic administration of roquinimex in DSS-treated mice also significantly inhibited the MDS, CH-reduction and MPO activity.. These findings suggest that roquinimex strongly inhibits murine colitis and may provide a novel pharmacological approach to treat inflammatory bowel disease.

Topics: Adjuvants, Immunologic; Animals; Colitis; Colon; Dextran Sulfate; Hydroxyquinolines; Injections, Subcutaneous; Male; Mice; Mice, Inbred BALB C; Peroxidase

Linomide prevents the development of autoimmune insulitis and insulin-deficient diabetes mellitus in female NOD mice. Linomide prevents development of autoimmune manifestations in other experimentally induced and spontaneous autoimmune diseases as well, but the mechanism of action is unknown. The present report summarizes our investigations on the effect of Linomide on different functional T cell subsets in NOD mice analyzed according to their cytokine profile. Supernatants from cultured splenocytes and peritoneal cells taken from Linomide-treated mice contained lower levels of TNFalpha, IL-1 beta, IFN gamma and IL-12 versus higher levels of IL-4, IL-6 and IL-10 in comparison with supernatants from cultures of untreated mice. Our results suggest that regulation of autoimmunity following oral Linomide administration in NOD mice induces a shift from Th(1) to Th(2) phenotype response, thereby preventing the development of diabetes by active cytokine-induced immunoregulation of T cell subsets, including downregulation of Th(1) and upregulation of Th(2).

Topics: Adjuvants, Immunologic; Animals; Autoimmunity; Concanavalin A; Cytokines; Diabetes Mellitus, Type 1; Female; Hydroxyquinolines; Inflammation; Mice; Mice, Inbred NOD; Spleen; Th1 Cells; Th2 Cells

A new orally active drug, laquinimod (ABR-215062), was shown to completely inhibit the development of murine acute experimental autoimmune encephalomyelitis (EAE). Furthermore, leukocyte infiltration into the central nervous system (CNS) was abolished in the laquinimod-treated animals. By direct comparison based on dose and total exposure, laquinimod was approximately 20 times more potent than the immunomodulator roquinimex. Laquinimod also had clear therapeutic effect when given after clinical onset in a chronic relapsing EAE model. It therefore represents a new orally active immunoregulatory drug without general immunosuppressive properties for the treatment of the autoimmune disease multiple sclerosis.

Topics: Adjuvants, Immunologic; Animals; Anti-Inflammatory Agents; B-Lymphocytes; Cells, Cultured; Central Nervous System; Chemotaxis, Leukocyte; Dexamethasone; Dose-Response Relationship, Drug; Encephalomyelitis, Autoimmune, Experimental; Female; Hydroxyquinolines; Immunosuppression Therapy; Mice; Mice, Inbred C57BL; Multiple Sclerosis; Neuroimmunomodulation; Quinolones; T-Lymphocytes

A significant reduction of the pan T lymphocytes as well as CD4+ and CD8 subsets of cells in the gut mucosa of the septic rats has previously been demonstrated. In contrast, the populations of major histocompatibility complex (MHC) class II-positive cells and macrophages increased. The aim of this study was to evaluate if the immunomodulator Linomide influenced the immune cell distribution in the small intestinal mucosa in sepsis and, furthermore, if these changes coincide with changes in the concentration of tumor necrosis factor-alpha (TNF-alpha) in plasma or ascites. Polymicrobial sepsis was induced in rats by cecal ligation and puncture (CLP). Three different experimental groups were used: CLP, Linomide p.o. + CLP, and Linomide i.p.+ CLP, with adequate controls. Specimens were taken from the small bowel for immunohistologic staining and grading of mucosal injury. The following monoclonal antibodies were used: W3/25, OX8, R73, OX6, and ED1. All slides were examined by one "blinded" examiner. Mucosal injury was graded from 0 to 5. The immunostained tissues were also analyzed by an automatic color-based image system. All controls had a normal appearance of the mucosa (grade 0-1), whereas the septic animals had a median grade of III (II-IV) mucosal injury. Linomide i.p. + CLP decreased mucosal damage to median I (0-IV, P < 0.05). Linomide had no effects on the immune cell distribution in controls. In CLP rats, a significant reduction in both CD4+ and CD8+ T lymphocytes as well as an increased number of macrophages and MHC class II-positive cells was seen in the villi as compared with sham-operated controls (P < 0.05). Linomide attenuated these changes for CD8+ and T lymphocytes and macrophages. Sepsis caused increased concentrations of TNF-alpha in portal blood and ascites 3 h from CLP induction. This increase was attenuated by Linomide.

Topics: Adjuvants, Immunologic; Animals; Ascitic Fluid; Disease Models, Animal; Hydroxyquinolines; Immunohistochemistry; Intestinal Mucosa; Intestine, Small; Male; Rats; Rats, Wistar; Sepsis; Tumor Necrosis Factor-alpha

Linomide (quinoline-3-carboxamide) is an immunomodulator with diverse effects on the immune system. Its beneficial effects on experimental autoimmune disease models have been linked to downregulation of Th1 cytokines and altered macrophage functions. We studied this effect of downregulation of Th1-type of immune response on Semliki Forest A7 virus infection in experimental autoimmune encephalomyelitis (EAE) susceptible Th1-prone SJL mice and in EAE-resistant Th2-prone BALB/c mice. We aimed at addressing the target-cell population of Linomide responsible for this Th1 downregulation. Treatment with Linomide led to increased virus infection in brain and this effect coincided with decreased production of IL-12 and IFN-gamma from stimulated spleen cells in SJL mice. In contrast, IL-12 and IFN-gamma expression were increased in Linomide-treated BALB/c mice. Treatment of infected SJL mice resulted in decreased percentage of CD11b+ and CD11c+ cells. Thus, the target cell population of Linomide may be antigen-presenting cells (APC) which are considered as candidates for regulatory cells of Th1/Th2 balance.

Topics: Adjuvants, Immunologic; Alphavirus Infections; Animals; Antigen-Presenting Cells; Brain; CD11b Antigen; Encephalomyelitis, Autoimmune, Experimental; Hydroxyquinolines; Interferon-gamma; Interleukin-12; Mice; Mice, Inbred BALB C; RNA, Viral; Semliki forest virus; Spleen; Th1 Cells; Th2 Cells; Viral Load

The effect of linomide, an immunomodulatory drug, on natural killer (NK) cells and T cell-dependent immune responses following syngeneic or allogeneic bone marrow transplantation (BMT) was investigated in BALB/c mice inoculated with B-cell leukemia (BCL1). Linomide given in the drinking water had no impact on graft survival or graft versus leukemia (GVL) effects. Although linomide regulates anti-self reactivity in mice with experimental and spontaneous autoimmune disorders, the anti-tumor effects induced by allogeneic donor lymphocytes were not affected. This indicates that different mechanisms regulate anti-self and anti-leukemia effects. Alternatively, linomide might affect the homing of self-reactive lymphocytes to specific target organs in autoimmune disorders, although the homing process may not be relevant to the control of leukemia by alloreactive lymphocytes.

Topics: Adjuvants, Immunologic; Animals; Bone Marrow Transplantation; Dose-Response Relationship, Drug; Female; Graft vs Host Disease; Graft vs Leukemia Effect; Hydroxyquinolines; Killer Cells, Natural; Leukemia, B-Cell; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL

Linomide is an immunomodulator that ameliorates several autoimmune and inflammatory diseases. We assessed the effect of Linomide on microvascular perfusion failure, leukocyte recruitment and hepatocellular injury induced by tumor necrosis factor alpha (TNF-alpha) and D-Galactosamine (Gal).. After 3 days of Linomide pretreatment (1, 10 and 100 mg/kg/day), rats were challenged with TNF-alpha/Gal for 24 h. Microvascular perfusion, leukocyte-endothelium interactions in hepatic postsinusoidal venules and leukocyte sequestration in sinusoids were evaluated using intravital microscopy. Liver enzymes were measured spectrophotometrically.. Challenge with TNF-alpha/Gal significantly reduced sinusoidal perfusion, and increased leukocyte rolling, adhesion and liver enzymes. Interestingly, pretreatment with Linomide (10 and 100 mg/kg/day) significantly reduced TNF-alpha/Gal-induced leukocyte rolling by 65 and 63%, and leukocyte adhesion by 87 and 84%, respectively. Moreover, Linomide (10 and 100 mg/kg/day) decreased sinusoidal sequestration of leukocytes by 71 and 51%, and markedly improved sinusoidal perfusion. Moreover, Linomide reduced aspartate aminotransferase by 87-97%, and alanine aminotransferase by 79-96%. However, Linomide had no protective effect when administered concomitantly with TNF-alpha/Gal.. These data demonstrate a dose-dependent inhibitory effect of Linomide on perfusion failure, leukocyte recruitment and hepatocellular injury provoked by TNF-alpha. Indeed, these findings suggest that Linomide may be an effective substance for protection of the liver in sepsis.

Topics: Adjuvants, Immunologic; Animals; Antineoplastic Agents; Cell Adhesion; Cell Communication; Chemical and Drug Induced Liver Injury; Endothelium, Vascular; Galactosamine; Hydroxyquinolines; Leukocytes; Liver; Liver Diseases; Male; Microcirculation; Rats; Rats, Sprague-Dawley; Sepsis; Tumor Necrosis Factor-alpha

The therapeutic effects of ABR-215062, which is a new immunoregulator derived from Linomide, have been evaluated in experimental autoimmune neuritis (EAN), a CD4(+) T cell-mediated animal model of Guillain-Barré syndrome in man. In previous studies, we reported that Linomide suppressed the clinical EAN and myelin antigen-reactive T and B cell responses. Here EAN induced in Lewis rats by inoculation with peripheral nerve myelin P0 protein peptide 180-199 and Freund's complete adjuvant was strongly suppressed by ABR-215062 administered daily subcutaneously from the day of inoculation. ABR-215062 dose-dependently reduced the incidence of EAN, ameliorated clinical signs and inhibited P0 peptide 180-199-specific T cell responses as well as also the decreased inflammation and demyelination in the peripheral nerves. The suppression of clinical EAN was associated with inhibition of the inflammatory cytokines IFN-gamma and TNF-alpha, as well as the enhancement of anti-inflammatory cytokine IL-4 in lymph node cells and periphery nerve tissues, respectively, in a dose-dependent manner. These effects indicate that ABR-215062 may mediate its effects by regulation of Th1/Th2 cytokine balance and suggest that ABR-215062 is potentially a new chemical entity for effective treatment of autoimmune diseases.

Topics: Adjuvants, Immunologic; Animals; Cell Movement; Dose-Response Relationship, Drug; Dose-Response Relationship, Immunologic; Hydroxyquinolines; Injections, Subcutaneous; Interferon-gamma; Male; Neuritis, Autoimmune, Experimental; Peripheral Nerves; Quinolones; Rats; Rats, Inbred Lew; Th1 Cells; Th2 Cells; Tumor Necrosis Factor-alpha

A significant correlation exists between disability and the volume of black holes (BHL VOL), defined as hypointense lesions on T1-weighted cranial magnetic resonance imaging. A consistent correlation has also been reported between urinary myelin basic protein-like material (MBPLM) and the transition toward secondary progression (SP) from relapsing-remitting (RR) multiple sclerosis (MS).. To improve the management of MS through a noninvasive and cost-effective test for monitoring disease activity or disease status.. From 662 patients with MS (86 with RR MS, 259 with SP MS without continued attacks, and 317 with SP MS with continued attacks), 24-hour urine samples were obtained at enrollment in the phase 3 Linomide (roquinimex) drug study. The urine specimens were analyzed for MBPLM and correlated with clinical features and findings on cranial magnetic resonance imaging.. Significant but weak correlations existed between urinary MBPLM and BHL VOL in all patients with MS (r = 0.114, P =.003; n = 662), patients with SP MS without attacks (r = 0.185, P =.003; n = 259), and all patients with SP MS (r = 0.122, P =.003; n = 576). No significant correlations were detected in the RR MS group or any of the disease groups or subgroups whose Expanded Disability Status Scale score was 5.0 or lower. In subgroup analysis, the most significant correlation was detected between urinary MBPLM after adjustment for creatinine and BHL VOL in patients with SP MS with an Expanded Disability Status Scale score of 5.5 or higher but without continued relapses (r = 0.417, P<.001; n = 138).. In patients with advanced SP MS, urinary MBPLM may possibly serve as an indicator of failed remission and axonal damage. Urinary MBPLM correlates with disease status in MS, especially the transition of RR MS to SP MS with advancing disability.

Topics: Adjuvants, Immunologic; Axons; Brain; Cost-Benefit Analysis; Cross-Sectional Studies; Disability Evaluation; Disease Progression; Female; Humans; Hydroxyquinolines; Magnetic Resonance Imaging; Male; Multiple Sclerosis; Multiple Sclerosis, Chronic Progressive; Multiple Sclerosis, Relapsing-Remitting; Myelin Basic Protein; Predictive Value of Tests; Randomized Controlled Trials as Topic; Severity of Illness Index

A modified procedure for the synthesis of Linomide is described. The synthesized drug was characterized and assessed for its in vivo antiangiogenic activity. In a murine angiogenesis assay Linomide treatment inhibited new blood vessel formation as documented by reduced microvessel area and blood volume.

Topics: Angiogenesis Inhibitors; Animals; Hydroxyquinolines; Mice

Linomide is a potent immunomodulator and has been reported to prevent type 1 diabetes mellitus in non-obese diabetic (NOD) mice and to reduce the incidence of other autoimmune diseases in animal models. The mechanisms of action seem to involve antigen expression by down regulation of macrophage activity and to antagonise the activation of Th1 cells during the cellular immune response. With the purpose to investigate the effect of Linomide on the incidence of spontaneous autoimmune thyroiditis (AIT) in female NOD mice we administered Linomide in drinking water (100 mg/kg/day) to NOD mice from 5th to 19th week of age. The mice were sacrificed at the end of week 19. None of the mice developed diabetes during the study period. The incidence of thyroiditis was evaluated on paraffin HE-stained sections and graduated on a scale from 0 to 4. Thirty-two percent of 37 mice treated with Linomide developed thyroiditis compared to 45% of 22 controls (p=0.31, chi2 =1.00). Among the mice who developed thyroiditis no difference in the degree of thyroiditis was found. Therefore no beneficial effect of Linomide on the incidence of spontaneous AIT in NOD mice could be demonstrated.

Topics: Adjuvants, Immunologic; Administration, Oral; Animals; Cell Movement; Diabetes Mellitus, Type 1; Female; Hydroxyquinolines; Incidence; Lymphocytes; Mice; Mice, Inbred NOD; Thyroid Gland; Thyroiditis, Autoimmune

Wallerian degeneration after peripheral nerve transection leads to the phagocytosis of degenerated myelin and axon components by macrophages. These phagocytes are recruited from the systemic circulation and Wallerian degeneration may therefore be used as a model for myelin removal by hematogenous macrophages, a feature that is also a hallmark of demyelinating diseases of the central and peripheral nervous system. The immunomodulator linomide has been shown to be effective in the treatment of experimental demyelinating diseases although the exact mode of its action is not yet defined. The present study investigated the effect of linomide on monocyte invasion and myelin phagocytosis after sciatic nerve transection. Linomide had a dual effect in Wallerian degeneration. Monocyte migration from the circulation to the damaged nervous system was significantly reduced. Additionally, the myelin phagocytic capacity of macrophages was impaired, finally resulting in a significant delay in the removal of myelin. The present experiments may provide an explanation for the effects of linomide during the course of demyelinating diseases of the nervous system.

Topics: Adjuvants, Immunologic; Animals; Cell Migration Inhibition; Hydroxyquinolines; Injections, Subcutaneous; Macrophages; Mice; Mice, Inbred C57BL; Myelin Sheath; Peripheral Nerve Injuries; Peripheral Nerves; Phagocytosis; Sciatic Nerve; Wallerian Degeneration

It has been demonstrated that quinoline-3-carboxamide, linomide, inhibited angiogenesis and reduced the volume of tumors grown from human hormone-resistant prostate cancer cell lines after subcutaneous implantation in mice. However, subcutaneous xenograft models may not mimic human conditions due to the absence of prostatic stromal cells at the ectopic site. Therefore, we investigated the influence of linomide on local tumor growth and metastasis of the human hormone-resistant prostate cancer cell line PC-3 in an orthotopic model.. In 30 athymic nude mice, 5x10(5) PC-3 cells were injected into the dorsal prostate after surgical exposure. After 7 days, group 1 (n = 15 mice) received linomide 100 mg/kg/day in the drinking water (per os). The other 15 mice (group 2) served as controls. All mice were sacrificed after 38 days followed by macroscopical and histological evaluation of local tumor growth and metastasis. Microvessel density was determined by immunohistochemical staining for von Willebrand factor as well as silver staining followed by morphometric analysis in an area of highest vessel density.. In the control group, local tumorigenicity and locoregional lymph node metastasis was 100%. The mean weight of the local tumor was 894 mg (395- 1,261 mg). The mean transversal diameter of the lymph node metastases was 4.0 mm (1.5-5. 4 mm). In the treatment group, local tumor growth and lymph node metastasis was 100% with a mean local tumor weight of 869 mg (232-1, 131 mg) and a mean lymph node metastasis diameter of 4.6 mm (1.3-5.9 mm). Microvessel density of the local tumor in the control and treatment group did not differ significantly.. Contrary to the results reported in subcutaneous animal models, linomide per os has no effect on net tumor growth and metastasis after orthotopic implantation of the human hormone-resistant prostate cancer cell line PC-3 in nude mice.

Topics: Angiogenesis Inhibitors; Animals; Cell Division; Humans; Hydroxyquinolines; Male; Mice; Mice, Nude; Neoplasm Metastasis; Neoplasm Transplantation; Prostatic Neoplasms; Tumor Cells, Cultured

1. In vitro studies with roquinimex, an immuno-modulator, in liver microsomes from mouse and rat were conducted to evaluate the primary metabolism and compare the metabolite pattern as well as the rate of metabolism with the in vivo pharmacokinetics of the compound in these two species. 2. In the presence of NADPH, roquinimex was metabolized to six primary metabolites (R1-6) by liver microsomes from mouse and rat. The formation of these metabolites was qualitatively similar in both species, and was greatly enhanced by pretreatment with PCN, an inducer of cytochrome P4503A. 3. The identification of the R1-6 demonstrated that roquinimex had been hydroxylated and demethylated. Hydroxylation at different sites of the quinoline moiety was the dominating reaction in both species. 4. Comparison of the resulting microsomal intrinsic clearance of 0.3 micromol mg(-1) protein min(-1) in mouse liver microsomes, versus 0.03 micromol mg(-1) protein min(-1) in rat liver microsomes demonstrated that the mouse possesses about a 10-fold greater metabolic capacity for roquinimex than the rat. 5. The in vivo pharmacokinetics of roquinimex demonstrated a 7-fold higher clearance in mouse than in the rat (82 ml h(-1) kg(-1) in mouse, 10.6 ml h(-1) kg(-1) in rat), which is in concordance with the in vitro findings.

Topics: Adjuvants, Immunologic; Animals; Aryl Hydrocarbon Hydroxylases; Cytochrome P-450 CYP3A; Cytochrome P-450 Enzyme System; Hydroxyquinolines; Mice; Microsomes, Liver; NADP; Oxidoreductases, N-Demethylating; Rats; Species Specificity

Roquinimex (Linomide) is an immunotherapeutic agent used in conjunction with autologous bone marrow transplantation (ABMT) for treatment of acute and chronic myelogenous leukemia (AML and CML). This agent may induce graft-versus-host reactions (GVHR) as well as graft-versus-leukemia (GVL) effects.. We documented the incidence of acute cutaneous GVHR associated with roquinimex immunotherapy. The presence or absence of autologous GVHR was also correlated with a potential GVL effect in patients with CML treated with ABMT and subsequent roquinimex immunotherapy in the period after the transplant.. Fifteen patients undergoing bone marrow transplantation and roquinimex immunotherapy for CML were followed up, and clinicopathologic data were analyzed.. Acute cutaneous GVHRs were observed in 6 of 15 patients (40%) treated with roquinimex. Ten of 11 evaluable patients receiving roquinimex exhibited eccrine sweat gland necrosis (ESGN) (90.9%), which was independent of the acute GVHR. Neither bone marrow engraftment status nor the survival rates of patients with and without GVHR was significantly different.. Roquinimex immunotherapy enhances the incidence of GVHR and was associated with a high rate of ESGN in patients with CML who were undergoing ABMT. There was no significant association between ESGN and acute GVHR. Acute autologous GVHR caused by roquinimex did not correlate with a GVL effect in our study of 15 patients with CML.

Topics: Adjuvants, Immunologic; Adult; Bone Marrow Transplantation; Female; Graft vs Host Disease; Graft vs Leukemia Effect; Humans; Hydroxyquinolines; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Male; Necrosis; Sweat Gland Diseases; Transplantation, Autologous

1. Roquinimex, a novel immunomodulator, is metabolized in liver microsomes from mouse and rat via cytochrome P450s to four hydroxylated and two demethylated metabolites (R1-6). The study investigated which cytochrome P450 enzyme(s) is responsible for the metabolism of roquinimex in man. 2. Enzyme kinetic analysis demonstrated an apparent Km = 1.28-7.00 mM and Vmax = 50-159 pmol x mg(-1) microsomal protein x min(-1) for the primary metabolites in human liver microsomes. The sum of Cl(int) for the primary pathways was 0.167 microl x mg(-1) microsomal protein x min(-1). 3. A correlation between the formation rate of R1-6 and 6beta-hydroxylation of testosterone was obtained within a panel of liver microsomes from 11 individuals (r2 = 0.72-0.97). Furthermore, significant inhibition (>90%) of roquinimex primary metabolism was demonstrated by ketoconazole and troleandomycin, specific inhibitors of CYP3A4 as well as with anti-CYP3A4 antibodies. Moreover, a similar metabolite pattern was produced from roquinimex by incubation with cDNA-expressed CYP3A4 as by human liver microsomes. 4. In conclusion, these data indicate a major role for CYP3A4 in the formation of roquinimex primary metabolites in human liver microsomes.

Topics: Antibodies; Cytochrome P-450 CYP3A; Cytochrome P-450 Enzyme Inhibitors; Cytochrome P-450 Enzyme System; DNA, Complementary; Enzyme Inhibitors; Female; Gene Expression; Humans; Hydroxylation; Hydroxyquinolines; Ketoconazole; Kinetics; Male; Microsomes, Liver; Mixed Function Oxygenases; Testosterone; Troleandomycin

The immunomodulator Linomide (roquinimex) ameliorates the development of numerous inflammatory and immunological diseases, including sepsis, arthritis, and encephalomyelitis. However, the mechanism underlying this protective effect of Linomide remains unclear. In this study, we wanted to evaluate the effect of Linomide treatment on the different steps in the extravasation process of leukocytes stimulated by tumor necrosis factor alpha (TNF-alpha) in vivo. For this purpose, we used intravital microscopy in the mouse cremaster muscle microcirculation. We found that pretreatment with Linomide dose-dependently (3-300 mg/kg) reduced TNF-alpha-induced leukocyte adhesion and tissue recruitment. Notably, at 300 mg/kg response to TNF-alpha was nearly abolished, i.e. leukocyte adhesion was decreased by 83% and recruitment by 86%. In fact, the anti-inflammatory effect of this dose of Linomide corresponded in magnitude to the potency of 10 mg/kg of dexamethasone. Moreover, administration of Linomide did not alter the systemic leukocyte counts. On the other hand, 1-10 mg/kg of dexamethasone decreased the circulating number of mononuclear leukocytes by 77%. Taken together, our novel findings demonstrate that Linomide is a potent inhibitor of leukocyte adhesion and recruitment in cytokine-activated tissues. These data may help explain the documented protection provided by Linomide in inflammatory diseases characterized by cytokine and leukocyte accumulation.

Topics: Adjuvants, Immunologic; Animals; Cell Adhesion; Cell Movement; Dexamethasone; Dose-Response Relationship, Drug; Glucocorticoids; Hydroxyquinolines; Leukocyte Count; Leukocytes; Male; Mice; Microcirculation; Microscopy, Video; Muscle, Skeletal; P-Selectin; RNA, Messenger; Tumor Necrosis Factor-alpha

Linomide is a potent immunomodulator that has been shown to inhibit autoimmunity in several animal models of autoimmune disease, including experimental autoimmune encephalomyelitis (EAE). Linomide's mechanism of action is unknown, however, it has been suggested to modulate the function of antigen presenting cells (APC) and that this may account for the inhibition of autoimmune disease. In this study we have been able to show that Linomide treatment of SJL/N mice upregulates the cell surface expression of several activation markers on macrophages and B cells. Thus, we found the following markers, expressed as a % of control, to be significantly upregulated following Linomide treatment; MHC class II (260%), Ly-6A/E (520%), CD11a (280%), CD54 (190%) and CD80 (200%) on macrophages and Ly-6A/E (250%) and CD11a (150%) on B cells. The duration and dosage of Linomide required to obtain these effects is similar to those required for EAE inhibition. Several Linomide analogues were made by the introduction of structural modifications into the Linomide molecule, resulting in a number of compounds with varying effects on EAE. We found a linear relationship between the compound's ability to inhibit EAE and its ability to upregulate MHC class II on macrophages (p<0.001), such that compounds which were able to inhibit EAE also upregulated MHC class II expression, whereas those that did not inhibit EAE were unable to do so. These results suggest that drug-mediated activation of distinct APC functions may be protective in autoimmunity.

Topics: Adjuvants, Immunologic; Animals; Antigens, Differentiation; Antigens, Differentiation, B-Lymphocyte; Antigens, Ly; B-Lymphocytes; Dose-Response Relationship, Drug; Encephalomyelitis, Autoimmune, Experimental; Female; Genes, MHC Class II; Hydroxyquinolines; Macrophage Activation; Macrophages, Peritoneal; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Inbred NOD; Up-Regulation

Linomide is a synthetic immunomodulator which was shown to protect animals against a wide range of experimental autoimmune diseases. In this study we have investigated the effects of Linomide on the thymus in an effort to elucidate the mechanisms by which this immunomodulator suppresses autoimmune reactivity. Normal or adrenalectomized SJL/J mice were treated orally for 10 days with linomide (80 mg/kg/day). Thymocytes were tested by FACS for the analysis of the CD4 and CD8 markers and TCR expression on their surface. Thymuses from these animals were examined for size and cellularity and immunohistopathologically for the detection of apoptosis and for the expression of the markers CD4 and CD8. A significant reduction in the thymus size and cellularity was observed in mice treated with Linomide, starting from day 3 after treatment, accompanied by an enhanced apoptotic death of cortical thymocytes, which was first noted on day 1 of treatment and peaked on day 3. FACS analysis and immunohistochemistry revealed a significant depletion of the CD4(+)/CD8(+) (double positive) cells with a parallel relative increase of the more mature, medullar, single positive, lymphocytes. These effects on the thymus were not mediated through a corticosteroid-dependent pathway, and were also observed in adrenalectomized and Linomide-treated animals. These observations may be of importance for the clarification of the role of thymus in autoimmunity and the possible ways for immune intervention with immunomodulators like Linomide at this level.

Topics: Adjuvants, Immunologic; Adrenal Cortex Hormones; Animals; Apoptosis; Atrophy; Autoimmunity; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Female; Hydroxyquinolines; Mice; Mice, Inbred Strains; Thymus Gland

Linomide (quinoline-3-carboxamide) is a synthetic immunomodulator that suppresses several experimental autoimmune diseases. Here we report the effects of Linomide on experimental autoimmune neuritis (EAN), a CD4+ T cell-mediated animal model of acute Guillain-Barré syndrome (GBS) in humans. EAN induced in Lewis rats by inoculation with bovine peripheral nervous system (PNS) myelin and Freund's complete adjuvant was strongly suppressed by Linomide administered daily subcutaneously from the day of inoculation. Linomide dose-dependently delayed the interval between immunization and onset of clinical EAN, as well as the severity of EAN symptoms. These clinical effects were associated with dose-dependent down-modulation of PNS antigen-induced T and B cell responses and with suppression of the proinflammatory cytokines IL-12, interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha) mRNA. In PNS sections, Linomide suppressed IL-12 and TNF-alpha, and up-regulated IL-10 mRNA expression. These findings suggest that Linomide could be useful in certain T cell-dependent autoimmune diseases.

Topics: Adjuvants, Immunologic; Animals; Antigens; B-Lymphocytes; Cytokines; Dose-Response Relationship, Immunologic; Hydroxyquinolines; Immunity, Cellular; Interferon-gamma; Lymph Nodes; Male; Myelin Sheath; Neuritis, Autoimmune, Experimental; Rats; Rats, Inbred Lew; RNA, Messenger; T-Lymphocytes; Tumor Necrosis Factor-alpha

The aim of the present investigation was to study the possible synergistic effect between cyclosporin A (CsA) and antithymocyte globulin (ATG), using the potent immunostimulator, Linomide. DA rats were transplanted with a PVG/c heart to the neck vessels, and the recipients were treated for 10 days with oral CsA (10mg/kg), oral Linomide (160mg/kg) and/or ATG, which was given as a single dose of either 0.02 ml, 0.1 ml or 0.2 ml prior to transplantation. Rats given a combination of ATG and Linomide or CsA and Linomide were used as controls. Synergism between CsA and ATG was tested using the two immunosuppressive drugs given in combination in order to override the challenge of Linomide. CsA or ATG treatment alone resulted in rats with long-term surviving grafts. Addition of Linomide to CsA-treated recipients was followed by early graft rejection. Similarily, Linomide triggered rejection in rats given a low dose of ATG and in recipients given a high dose of ATG if Linomide treatment was prolonged to 21 days. The combination of ATG, CsA and 21 days of Linomide resulted in a significantly prolonged graft survival compared with either ATG + Linomide or CsA + Linomide. These findings demonstrate the synergistic capacity of ATG and CsA in combined immunosuppressive therapy.

Topics: Adjuvants, Immunologic; Animals; Antilymphocyte Serum; Cyclosporine; Dose-Response Relationship, Immunologic; Drug Administration Schedule; Drug Synergism; Drug Therapy, Combination; Female; Graft Survival; Heart Transplantation; Hydroxyquinolines; Immune Tolerance; Intubation, Gastrointestinal; Male; Rats; Rats, Inbred Strains

Five years after the identification of the von Hippel-Lindau (VHL) gene, physicians, scientists and concerned VHL family members met to review the current state of knowledge on the diagnosis and treatment of VHL and to summarize the latest information on the biochemistry of the VHL protein (pVHL). The NIH and University of Pennsylvania groups reported the detection of germ-line mutations in 100% (93 of 93) of VHL families studied. Several studies determined the frequency of VHL germ-line mutations in individuals with a single manifestation of VHL without a family history of VHL. National groups to improve the diagnosis and treatment of individuals with VHL disease have been established in Great Britain, Denmark, France, Holland, Italy, Japan, Poland, and the United States. Evidence for the existence of genes that modify the expression of VHL was presented. The VHL protein appears to have several distinct functions: (a) down-regulation of hypoxia-inducible mRNAs; (b) proper assembly of the extracellular fibronectin matrix; (c) regulation of exit from the cell cycle; and (d) regulation of expression of carbonic anhydrases 9 and 12.

Topics: Animals; Carcinoma, Renal Cell; Central Nervous System Neoplasms; Cystadenoma, Papillary; Disease Models, Animal; DNA Mutational Analysis; Exons; Genes, Lethal; Genetic Testing; Genotype; Hemangioblastoma; Hemangioma; Humans; Hydroxyquinolines; Kidney Neoplasms; Ligases; Mice; Mice, Knockout; Mice, Nude; Neoplasms; Neovascularization, Pathologic; Nephrectomy; Pancreatic Neoplasms; Paraganglioma; Phenotype; Polymerase Chain Reaction; Proteins; Radiosurgery; Retinal Neoplasms; Trophoblasts; Tumor Suppressor Proteins; Ubiquitin-Protein Ligases; von Hippel-Lindau Disease; Von Hippel-Lindau Tumor Suppressor Protein

The drug Linomide is an immunomodulator showing marked down-regulation of several experimental autoimmune diseases. In this study, its effect on three different experimental models of thyroid disease and on spontaneous infiltration of salivary glands (sialoadenitis), was investigated. Although very effective at preventing thyroid infiltrates in mice immunized with mouse thyroglobulin and complete Freund's adjuvant and in spontaneous models of thyroiditis and sialoadenitis, it completely failed to modify experimental autoimmune thyroiditis (EAT) induced in mice immunized with mouse thyroglobulin and lipopolysaccharide. There was no significant shift in the observed isotypes of anti-mouse thyroglobulin antibodies and only anti-mouse thyroglobulin antibodies in the spontaneous model were completely down-modulated by the drug. One surprising fact to emerge was that Linomide-treated donor mice, although protected from thyroid lesions themselves, were still able to transfer EAT showing that they must have been effectively primed while being treated with Linomide. It is possible that the drug down modulated EAT by interfering with the trafficking of primed effector cells.

Topics: Adjuvants, Immunologic; Animals; Autoimmune Diseases; Disease Models, Animal; Drug Administration Schedule; Female; Freund's Adjuvant; Hydroxyquinolines; Lipopolysaccharides; Lymph Nodes; Male; Mice; Mice, Inbred CBA; Mice, Inbred NOD; Sialadenitis; Thyroglobulin; Thyroiditis, Autoimmune

Topics: Animals; Antineoplastic Agents; Drug Synergism; Humans; Hydroxyquinolines; Interleukin-12; Macrophages; Male; Melanoma, Experimental; Mice; Neovascularization, Pathologic; Prostatic Neoplasms

Since prostate cancer (PC) development involves a combination of genetic predisposition and promotional mechanisms, especially the metabolic conversion of testosterone to 5alpha dihydrotestosterone (DHT) by 5alpha reductase, how do mechanisms in man relate to prostate-seminal vesicle (P-SV) tumor development in Lobund-Wistar (L-W) rats? The disease in man and in L-W rats shares developmental mechanisms and characteristics to the extent that prevention of P-SV tumors in L-W rats could be predictive of similar results in man. The epidemiology of PC in man and P-SV tumors in L-W rats indicates that both are hormone-related diseases based on genetic predisposition, high production of androgens (which are activated to DHT by 5alpha reductase), and early development of androgen-dependent and metastasizing late androgen-independent stages of adenocarcinomas, all after long latency periods.. L-W rats at risk of developing spontaneous or induced P-SV tumors were subjected to putative antitumor agents or procedures. These included dietary restriction, testosterone ablation, soybean-derived isoflavones, antiangiogenic linomide, tamoxifen, and a vitamin D analogue.. L-W rats subjected to 1) early onset of dietary restriction manifested suppression of spontaneous and induced development of P-SV tumors; 2) testosterone-ablation by nonesterified DHT (NE-DHT) suppressed early onset of induced P-SV tumors and to a lesser extent late onset of spontaneous tumors; 3) diets containing soy protein isolate (high isoflavones) manifested marginal suppressive effects against induced P-SV tumors, but in 12-month-old rats, the development of spontaneous tumors was reduced in incidence; 4) early administrations of antiangiogenic linomide suppressed development of induced P-SV tumors and of transplanted prostate adenocarcinoma III (PA-III) tumors, but linomide had little antitumor effect against large advanced stage tumors; and 5) tamoxifen and vitamin D analogue suppressed development of P-SV tumors. Results in conditions 1-3 were negative when tested against PA-III tumors.. Developing stages of P-SV tumors were prevented in L-W rats with autochthonous spontaneous and induced tumors, but most of the agents tested were of no therapeutic benefit against advanced-stage and transplanted PA-III tumors. However, early administrations of antiangiogenic linomide suppressed early growth of induced and transplanted PA-III tumors.

Topics: Animals; Anticarcinogenic Agents; Antineoplastic Agents; Antineoplastic Agents, Hormonal; Disease Susceptibility; Estrogens, Non-Steroidal; Glycine max; Hydroxyquinolines; Isoflavones; Male; Neovascularization, Pathologic; Phytoestrogens; Plant Preparations; Prostatic Neoplasms; Rats; Rats, Wistar; Tamoxifen; Testosterone; Vitamin D

Linomide is a synthetic immunomodulator that has been shown to protect animals against a wide range of spontaneously developing or induced autoimmune diseases. We have previously reported that Linomide blocks both the clinical and the histopathological manifestations of experimental autoimmune encephalomyelitis (EAE) in various animal models. In this study, in an effort to elucidate the mechanisms by which Linomide suppresses EAE, and autoimmunity in general, we investigated the in vivo effects of this drug on the TH1/TH2 lymphocyte balance, which is important for the induction or inhibition of autoireactivity. Naive SJL/J mice were treated orally for 15 days with Linomide (80 mg/kg/day). Spleen cells were obtained at various time points during the treatment period and were stimulated in vitro with concanavalin A. Interleukins IL-4, IL-10 and IL-12, transforming growth factor-beta (TGFbeta) and interferon-gamma (IFNgamma) cytokine production was evaluated both by means of detection of the cytokines in the medium (by ELISA technique) and by detection of the cytokine mRNA production, using a semiquantitative reverse transcriptase polymerase chain reaction method. A significant upregulation of IL-4, IL-10 and TGFbeta was observed following treatment with Linomide, which peaked at day 10 (IL-10) or day 15 (IL-4). On the other hand, IL-12 and IFNgamma production were either unchanged or decreased. It seems therefore that Linomide induces in vivo a shift towards TH2 lymphocytes which may be one of the mechanisms of downregulation of the autoimmune reactivity in EAE. Our observations indicate that downregulation of TH1 cytokines (especially IL-12) and enhancement of TH2 cytokine production may play an important role in the control of T-cell-mediated autoimmunity. These data may contribute to the design of new immunomodulating treatments for a group of autoimmune diseases.

Topics: Adjuvants, Immunologic; Animals; Autoimmunity; Cytokines; Down-Regulation; Encephalomyelitis, Autoimmune, Experimental; Enzyme-Linked Immunosorbent Assay; Female; Hydroxyquinolines; Interleukin-10; Interleukin-4; Mice; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Th2 Cells

In murine severe experimental autoimmune encephalomyelitis (EAE), an animal model for the human disease multiple sclerosis (MS), we tested the efficacy of a 5-halo-6-phenyl pyrimidinone compound, bropirimine (PNU-54461). We observed that the compound is active in suppressing EAE when administered orally, a significant pharmacological advantage compared to some current therapies for the treatment of MS. Furthermore, bropirimine was most efficacious when dosing was begun 5-10 days after injection of myelin basic protein, the protein isolated from the central nervous system and used for inducing EAE in our model. This is a period of time following the initial immunological events leading to the disease, when large-scale leukocyte infiltration into the central nervous system begins. Following oral dosing, bropirimine peaked in the blood within 3 h and was cleared to undetectable concentrations within 16-18 h. Despite the pharmacokinetics in the blood, bropirimine was fully efficacious when dosed orally every two or three days. Surprisingly, bropirimine treatment did not result in a statistically significant decrease in leukocyte infiltration into the lower spinal cord, unless the compound was dosed daily at a high concentration. We also observed the concentration and time course of alpha-interferon in blood following oral dosing of bropirimine. The kinetics of interferon in the blood are similar to, but clearly distinguishable from, the pharmacokinetics of bropirimine in the blood. It is not clear whether or not the induction of interferon plays a key role in the efficacy of bropirimine. Nevertheless, the results using bropirimine in EAE suggest that the compound may be useful for the treatment of multiple sclerosis.

Topics: Adjuvants, Immunologic; Animals; Cytosine; Dose-Response Relationship, Drug; Encephalomyelitis, Autoimmune, Experimental; Female; Hydroxyquinolines; Immunohistochemistry; Interferons; Mice; Multiple Sclerosis

The role of lymphocytes and their subpopulations in lung fibrosis is as yet unclear.. To define the role of immunomodulation in bleomycin-induced inflammatory fibrotic lung injury, by testing the effect of two known Th1 inhibitors: linomide and pentoxifylline.. C57BL/6 mice were treated by a single intratracheal instillation of 0.06 mg bleomycin in 0.01 ml saline or saline alone. Treatment groups included: (1) intratracheal bleomycin and daily treatment with linomide or pentoxifylline; (2) intratracheal bleomycin and daily water; (3) intratracheal saline and daily linomide or pentoxifylline; (4) intratracheal saline and daily water. Linomide and pentoxifylline were available per os in the drinking water from 1 day prior to intratracheal instillation. Animals were studied 14 days after intratracheal instillation. Lung injury was evaluated by total and differential cell count in bronchoalveolar lavage fluid, by a semiquantitative morphological index of lung injury and a quantitative image analysis of cellularity, fibrosis fraction and alveolar wall area fraction, and by biochemical analysis of lung hydroxyproline content.. Linomide or pentoxifylline did not cause any lung injury in saline-treated control mice. Overt signs of lung injury were apparent in bleomycin-treated mice. These changes were not affected by daily treatment with linomide or pentoxifylline, which were given in the highest tolerable dose.. This study does not support the use of linomide or pentoxifylline to prevent or ameliorate lung fibrosis and may suggest that drug-induced differentiation of T lymphocytes into Th1/th2 subpopulations does not affect the evolution of bleomycin-induced lung injury.

Topics: Adjuvants, Immunologic; Animals; Antibiotics, Antineoplastic; Bleomycin; Hydroxyquinolines; Male; Mice; Mice, Inbred C57BL; Pentoxifylline; Pulmonary Fibrosis; Th1 Cells; Th2 Cells

When applied in rodent transplant models most immunosuppressive drugs yield adequate graft protection for as long as the drug is given, and permanent graft survival is often induced. The immunomodulator, Linomide, previously shown to stimulate T cells and prevent apoptosis, usually reduces or abolishes both tolerance induction and the graft-protective effect of the immunosuppressive drug. By chance, we observed that Linomide alone exerted a modest but unequivocal graft-protective effect in the BN to WF strain combination. This finding was analysed by simple genetic mapping of rat strains. Untreated WF recipients kept BN grafts for a median of 8 days, whereas Linomide treatment prolonged graft survival to 12. 5 days (P = 0.0001). In control groups (DA to LEW, BN to LEW, DA to WF and WF to BN), median graft survival was 5.5-7 days irrespective of whether Linomide was given. However, the BN to F1 (LEW x WF) combination also manifested slightly longer graft survival in the presence of Linomide. F1 (BN x WF) to WF grafts survived a median of 15 days without Linomide and 46 days with Linomide treatment. Both in the presence and absence of Linomide, two of the control graft combinations [F1 (BN x DA) to WF and F1 (BN x WF) to BN] manifested 6-7-day graft survival. Taken together, our results suggest a delicate balance between unresponsiveness and rejection, while a single agent (Linomide) may either cause on its own long-term survival of allografts in one setting or rejection despite optimal immunosuppression in another setting.

Topics: Adjuvants, Immunologic; Animals; Graft Rejection; Graft Survival; Heart Transplantation; Hydroxyquinolines; Immune Tolerance; Rats; Rats, Inbred Lew; Rats, Inbred WF; Transplantation, Homologous

The aim of this study was to ascertain the potential usefulness of the antiangiogenic compound linomide for treatment of von Hippel-Lindau (VHL)-related tumors. Paraganglioma tissue fragments obtained at surgery from a VHL type 2a patient were transplanted s.c. to male BALB/c nu/nu (nude) mice: (a) 2-3-mm fragments for "prevention" experiments; and (b) 2-3-mm fragments allowed to grow to 1 cm for "intervention" studies. Both groups received either 0.5 mg/ml linomide in drinking water or acidified water and were followed until tumor diameter reached 3 cm or for 4 weeks. In both the prevention and intervention experiments, a significant diminution of tumor size and weight was observed in the drug-treated animals. In vivo nuclear magnetic resonance analysis of tumor blood flow in linomide-treated animals showed localization of blood vessels almost exclusively to the periphery of the poorly vascularized tumors with a significant reduction of both vascular functionality and vasodilation. Histological examination of tumors from linomide-treated animals revealed marked avascularity. Treated animals also displayed a 2.4-fold reduction of tumor vascular endothelial growth factor mRNA levels. Taken together, our data indicate that in VHL disease, therapy directed at inhibition of constitutively expressed VEGF induction of angiogenesis by VHL tumors may constitute an effective medical treatment.

Topics: Administration, Oral; Angiogenesis Inhibitors; Animals; Cell Division; Humans; Hydroxyquinolines; Magnetic Resonance Imaging; Male; Mice; Mice, Nude; Neovascularization, Pathologic; Paraganglioma; Transplantation, Heterologous; Tumor Cells, Cultured; von Hippel-Lindau Disease

Linomide is a potent immunomodulator that either enhances or suppresses certain immunological processes. Of particular interest is this compound's capacity to inhibit a variety of organ-specific autoimmune diseases. Here, we report on the effects of linomide on several immunological reactions elicited by endotoxin (LPS), both in vivo and in vitro. In rats and mice linomide inhibited the elicitation of endotoxin-induced uveitis (EIU), an acute inflammatory eye disease that develops within 24 h following footpad injection of LPS. Linomide also inhibited the production of TNF-alpha and IL-6 by LPS-stimulated rat and mouse macrophage monolayers. On the other hand, treatment with linomide significantly increased the levels of IL-1beta (mice and less in rats), IL-6 (rats), and TNF-alpha (mice) in serum samples collected 2 h following injection with LPS. The increased production of proinflammatory cytokines in linomide-treated mice was also indicated by the enhanced lethal effect of LPS in these mice. The finding of elevated levels of these cytokines in animals with suppressed EIU is also in line with previous observations of an inverse relationship between EIU severity and levels of TNF-alpha. Data recorded here underscore the unique capacity of linomide to both enhance and suppress the immune system.

Topics: Adjuvants, Immunologic; Animals; Aqueous Humor; Cells, Cultured; Dose-Response Relationship, Drug; Endotoxins; Female; Hydroxyquinolines; Interleukin-1; Interleukin-6; Lipopolysaccharides; Macrophages; Mice; Mice, Inbred C3H; Rats; Rats, Inbred Lew; Tumor Necrosis Factor-alpha; Uveitis

We have recently reported that the tricyclic antidepressants (TCAs) imipramine, clomipramine, and citalopram induce apoptosis in human peripheral lymphocytes. This system is well suited for studies on the pathophysiology/physiology of apoptosis regulation. Apoptosis was determined using both DNA gel electrophoresis and flow cytometric analysis. TCA-induced apoptosis in lymphocytes was monitored in the presence of the protein synthesis inhibitor cycloheximide (CHX), the RNA synthesis inhibitor actinomycin D (Act D), the antioxidant reduced glutathione (GSH), the nuclease inhibitor aurintricarboxylic acid (ATA), the cytokine interleukin-2 (IL-2), and the immunostimulator linomide. CHX and Act D failed to prevent and actually enhanced TCA-induced apoptosis in lymphocytes, indicating that protein and RNA syntheses are not required for this process. Exogenous IL-2, GSH, and ATA protected the lymphocytes from apoptosis induced by TCAs in a dose-dependent manner, whereas linomide had no effect on TCA-induced apoptosis under our in vitro conditions. Our data demonstrate that TCA-induced apoptosis in human lymphocytes shares many common features with other stimuli-induced apoptotic processes.

Topics: Antidepressive Agents, Tricyclic; Apoptosis; Aurintricarboxylic Acid; Cycloheximide; Dactinomycin; Dose-Response Relationship, Drug; Glutathione; Humans; Hydroxyquinolines; Interleukin-2; Lymphocytes

Linomide is a new synthetic immunomodulator which exerts prominent anti-autoimmune effects in various experimental models. Recently, it was tested in clinical trials to patients suffering from multiple sclerosis and showed to inhibit the activity of the disease. Therefore, due to its pharmacological importance, we attempted elucidate its structure using one-dimensional and two-dimensional nuclear magnetic resonance (NMR) techniques and study its conformational properties using a combination of two-dimensional NMR spectroscopy and molecular modeling. The conformational analysis of linomide was based on the measurement of interproton nuclear Overhauser enhancement (NOE) values obtained from a two-dimensional NMR spectrum and a number of molecular modeling techniques used to calculate the low energy conformers of this compound. This information will serve as an aid to synthetic chemists whom their research activity is focused on developing linomide analogs with better biological profiles.

Topics: Adjuvants, Immunologic; Hydroxyquinolines; Magnetic Resonance Spectroscopy; Models, Molecular; Molecular Structure

Angiogenesis and antiangiogenesis, as applied to oncology, are phenomena in which (1) tumors acquire a new blood vascular system from the host that is needed for their growth progression and metastasis; and (2) factors are produced that interfere with neovascularization, thereby inhibiting growth and metastasis of the tumor. Linomide, a chemical antiangiogenesis agent, inhibited the growth of transplanted tumors in mice and rats and inhibited the early development of metastasizing tumors induced in the prostate-seminal vesicle (P-SV) complex of genetically susceptible Lobund-Wistar (L-W) rats.. L-W rats with small induced P-SV tumors were treated with a recommended dosage of linomide (100 mg/kg BW/day) by the intraperitoneal and oral routes. The rats were monitored for the next 1-2 months, and the primary and metastatic tumors were compared with related data in drug-free tumor-bearing control rats.. P-SV tumors in linomide-treated and untreated control rats continued to grow, except that in the former (1) the tumors were marginally smaller, (2) the centers of the primary P-SV tumors had failed to grow, (3) the peripheral areas of the tumors contained actively proliferating tumor cells, and (4) metastatic P-SV tumors in the lungs were disrupted with focal areas of necrosis, but areas of intact tumor cells survived. Spread of tumor cells into the peritoneal cavity was not inhibited. Rats on orally administered linomide lived significantly longer than rats inoculated by the intraperitoneal route and untreated control rats. The dosage of linomide used showed evidence of toxicity.. Although primary and metastatic P-SV tumors were damaged in L-W rats treated with linomide, this antiangiogenic drug was of minimal therapeutic benefit to rats in which a palpable P-SV tumor had developed before onset of treatments.

Topics: Animals; Antineoplastic Agents; Body Weight; Genital Neoplasms, Male; Hydroxyquinolines; Male; Neoplasm Metastasis; Neovascularization, Pathologic; Prostatic Neoplasms; Rats; Rats, Wistar; Seminal Vesicles

Oral linomide, (quinoline-3-carboxamide), has been shown to prevent autoimmune insulitis, islet destruction, and diabetes in NOD mice treated at an early stage of the disease, but confers only partial protection in animals with advanced disease. Reg protein, the gene product of a complementary DNA isolated from a regenerating rat islet library, has been previously shown to induce expansion of beta-cell mass in pancreatectomized rats. To determine the effect of treatment combining immunomodulation and Reg protein on advanced autoimmune diabetes, we treated female NOD mice with oral linomide and i.p. Reg protein injections. In 14-week-old animals with less severe disease (glucose tolerant), treatment with each agent alone resulted in amelioration of diabetes, as did treatment with Reg alone in 5-week-old prediabetic mice. In 14-week-old animals with more severe disease (glucose intolerant), only treatment with the combination of both agents, but not that with each separately, resulted in amelioration of diabetes. Our study suggests that treatment aimed at abrogation of autoimmunity combined with expansion of beta-cell mass constitutes a potential therapeutic approach for treatment of insulin-dependent diabetes mellitus.

Topics: Adjuvants, Immunologic; Animals; Autoimmune Diseases; Blood Glucose; Calcium-Binding Proteins; Diabetes Mellitus, Type 1; Female; Glucose Tolerance Test; Hydroxyquinolines; Immunotherapy; Insulin; Islets of Langerhans; Lithostathine; Mice; Mice, Inbred NOD; Nerve Tissue Proteins; Pancreas

The blocking of angiogenesis provides a novel therapeutic target to inhibit tumour spreading. In this study, we investigated the effect of linomide on angiogenesis induced in vivo by highly angiogenic breast carcinoma cells. The rabbit cornea was used to assess neovascular growth in the absence of a tumour mass. MCF-7 cells stably transfected with the cDNA encoding for vascular endothelial growth factor 121 (VEGF121) (V12 clone) were used to elicit a potent VEGF-dependent corneal angiogenesis. After tumour cell implant, albino rabbits received 100 mg kg(-1) day(-1) linomide for 5 consecutive days. Daily observation of neovascular progression indicated that linomide blocked angiogenesis. The antiangiogenic effect of linomide was apparent within 48 h from the beginning of the treatment and was both angiosuppressive and angiostatic. The block of neovascular growth lasted over 10 days from treatment suspension, and preformed vessels, which had regressed, remained dormant, suggesting the persistence of unfavourable conditions for capillary progression. Linomide (50-200 microg ml[-1]) was not cytotoxic in vitro on resting capillary endothelial cells but blocked endothelial cell replication induced by VEGF. Our data indicate that linomide can efficiently and persistently block VEGF-dependent angiogenesis in vivo in the absence of a growing tumour mass. These data suggest that linomide could be a chemopreventive drug in breast cancer patients and a valuable tool in clinical settings in which metastatic spreading occurs in the absence of a detectable tumour mass.

Topics: Animals; Antineoplastic Agents; Breast Neoplasms; Carcinoma; Corneal Neovascularization; Endothelial Growth Factors; Endothelium, Vascular; Female; Hydroxyquinolines; Lymphokines; Neoplasm Proteins; Rabbits; Transfection; Transplantation, Heterologous; Tumor Cells, Cultured; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors

Paclitaxel, bropirimine and linomide therapy was evaluated in a murine prostate cancer model. All drugs were effective in impeding tumor growth but the mechanisms of action varied. Paclitaxel inhibited bcl-2 expression suggesting an apoptotic mechanism. Bropirimine, while inhibiting bcl-2 expression also significantly depressed tumor necrosis factor-alpha (TNF-alpha) expression. In the bropirimine treated group there was also a correlation between angiogenesis and cyclin D expression. Finally, linomide significantly decreased angiogenesis. Since the mechanism of action of these drugs differ, combining them at lower doses might maintain therapeutic efficacy while reducing toxicity.

Topics: Animals; Antineoplastic Agents; Cell Division; Cytosine; Hydroxyquinolines; Immunohistochemistry; Male; Mice; Models, Biological; Paclitaxel; Prostatic Neoplasms; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Inbred F344

Linomide (quinoline-3-carboxamide) is a synthetic immunomodulator that suppresses several experimental autoimmune diseases. Here we report the effects of Linomide on experimental autoimmune encephalomyelitis (EAE), a CD4+ T cell-mediated animal model of multiple sclerosis (MS). EAE induced in Lewis rats by inoculation with homogenized guinea pig spinal cord and Freund's complete adjuvant was strongly suppressed by Linomide administered daily subcutaneously from the day of inoculation. Linomide dose-dependently delayed the interval between immunization and onset of clinical EAE, and reduced severity of EAE symptoms. These clinical effects were associated with dose-dependent down-modulation of myelin antigens-induced T cell responses and by suppression of the proinflammatory cytokines IFN-gamma and TNF-alpha, and upregulation IL-4, IL-10 and TGF-beta as evaluated by in situ hybridization for mRNA expression in spleen mononuclear cells and spinal cord sections. These findings suggest that Linomide could be useful in certain T cell dependent autoimmune diseases.

Topics: Acute Disease; Adjuvants, Immunologic; Animals; Cytokines; Dose-Response Relationship, Drug; Encephalomyelitis, Autoimmune, Experimental; Histocompatibility Antigens Class II; Hydroxyquinolines; Immunohistochemistry; Male; Rats; Rats, Inbred Lew; RNA, Messenger; T-Lymphocytes

Proinflammatory cytokine overproduction, as well as synthesis of the inducible form of nitric oxide synthase (iNOS), are known to play a major role in HIV-1-triggered disease. AIDS patients show increased serum tumour necrosis factor (TNF)-alpha and interferon (IFN)-gamma levels, which synergize with HIV-1-produced nitric oxide (NO) to augment viral replication. Linomide has strong immunomodulatory effects in animals and humans, yielding promising clinical benefits in several pathological disorders including septic shock and autoimmune disease, processes largely mediated by overproduction of these cytokines. In peripheral T cells, linomide also prevents apoptosis triggered by a variety of stimuli, including superantigens, dexamethasone and vaccinia virus.. Linomide inhibits production of proinflammatory cytokines such as TNF-alpha, interleukin-1 beta and IFN-gamma, as well as iNOS synthesis. The SCID-hu-PBL mouse model was used to analyse the effect of linomide on HIV-1 infection. T-cell frequency was characterized in reconstituted animals, and the frequency of infected mice and viral load of infected animals were studied.. Linomide promotes an increase in human CD4+ T-cell counts in the peritoneal cavity of HIV-1-infected, linomide-treated mice. Linomide also prevents human TNF-alpha and IFN-gamma production, as well as iNOS expression and affects the viral load, promoting potent suppression of HIV-1 infectivity as detected in peritoneal cavity and spleen.. The combination of linomide's properties, namely, blockage of proinflammatory cytokine and NO production, as well as prevention of apoptosis, is of paramount interest, making linomide a potential candidate for combating HIV-1 infection or preventing some of its associated pathological manifestations.

Topics: Adjuvants, Immunologic; Animals; Anti-HIV Agents; CD3 Complex; CD4-Positive T-Lymphocytes; Cytokines; HIV Envelope Protein gp120; HIV Infections; HIV-1; Humans; Hydroxyquinolines; Interferon-gamma; Leukocytes, Mononuclear; Mice; Mice, SCID; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Peritoneal Cavity; Spleen; Viral Load; Virus Replication

Neuronal death after injury or disease could result from imbalanced cytokine expression. Linomide (LS-2616, quinoline-3-carboxamide), a synthetic immunomodulator with effects on cytokine production, suppresses autoimmune diseases of the nervous system. Here adult mice were pre-treated with 200 mg/kg/day of Linomide for 9 days, after which the sciatic nerves were crushed. After another 10 days of Linomide treatment the dorsal root ganglia were dissected out and stained for apoptosis, either immediately or after 2 days in culture, which increases cell death. Superior cervical ganglia were also cultured for 2 days. The Linomide pretreatment profoundly reduced (approximately 60-80%) the injury-induced apoptotic death of neurons and satellite cells in both systems. The results suggest that modulation of the inflammatory cytokine cascade is a promising road to nerve cell rescue.

Topics: Adjuvants, Immunologic; Animals; Apoptosis; Cell Death; Ganglia, Spinal; Hydroxyquinolines; Male; Mice; Mice, Inbred Strains; Nerve Crush; Neurons; Sciatic Nerve

Linomide (N-phenylmethyl-1,2-dihydro-4-hydroxyl-1-methyl-2-oxoquinoline-3-carboxa mide) is a novel immunomodulator with a potent anti-tumoral activity. This study was undertaken to test the effect of Linomide on basic fibroblast growth factor (bFGF) induced angiogenesis in vivo, which manifests itself in an increased number of blood vessels per unit of cell infiltrated area. Subcutaneously implanted polyvinyl alcohol sponges (PVS) in guinea pigs were used as a model system to quantitate angiogenesis in vivo. Oral treatment with Linomide was able to reduce significantly the bFGF induced blood vessel growth and proliferation within the implanted PVS, relative to untreated controls. In addition, Linomide significantly reduced the bFGF mediated augmentation of protein and collagen content in the implanted PVS, indicating an inhibition in the deposition of extracellular matrix (ECM). We conclude that the potent inhibition of bFGF induced angiogenesis by Linomide in vivo in addition to immunomodulatory effects may have potentially important clinical applications.

Topics: Administration, Oral; Animals; Fibroblast Growth Factor 2; Guinea Pigs; Hydroxyquinolines; Immunohistochemistry; Neovascularization, Physiologic

Topics: Adjuvants, Immunologic; Animals; Cytokines; Female; Gene Expression Regulation; Hydroxyquinolines; Myasthenia Gravis; Rats; Rats, Inbred Lew; RNA, Messenger; Th1 Cells; Th2 Cells; Time Factors; Transcription, Genetic

Linomide is a synthetic immunomodulator that down-regulates autoimmune response without inducing systemic immunosuppression. Linomide effectively inhibits severe experimental autoimmune diseases, like experimental allergic encephalomyelitis (EAE), a model of multiple sclerosis (MS). Here we report that Linomide suppresses nitric oxide (NO) production by microglia and astrocytes derived from newborn rats and prevented oligodendrocyte damage. Linomide strongly inhibited interleukin (IL) 1 betamRNA expression on glial cells, suggesting a potential mechanism for inhibition of NO production by Linomide. These results demonstrate that Linomide-mediated inhibition of NO production by glial cells could explain the preventive and therapeutic effects of Linomide in EAE and perhaps also MS. However, Linomide at higher dose [correction of doss] (10(-5) M) resulted in direct oligodendrocyte damage.

Topics: Adjuvants, Immunologic; Animals; Animals, Newborn; Cell Survival; Cells, Cultured; Dose-Response Relationship, Drug; Hydroxyquinolines; Interleukin-1; L-Lactate Dehydrogenase; Microglia; Nitric Oxide; Oligodendroglia; Rats; Rats, Inbred Lew; RNA, Messenger

Both Linomide (quinoline-3-carboxamide) and tolerization with self-antigens have been demonstrated to successfully ameliorate demyelinating disease in experimental autoimmune encephalomyelitis (EAE). Based on the autoimmune hypothesis of multiple sclerosis (MS), both agents have been tested in clinical trials but have been found to be toxic or not efficacious. We investigated the efficacy of these immunomodulators in an alternative experimental model of MS, a virus-induced demyelinating disease. Oral administration of Linomide to Theiler's virus-infected mice beginning either at time of infection or at day 15 post-infection (p.i.) resulted in an increased percentage of spinal cord quadrants with demyelination. Administration of Linomide beginning at day 15 p.i. increased lesion size as compared to infected control-treated mice. Treatment with 80 mg kg(-1) day(-1) of Linomide beginning at the time of infection significantly increased the number of Theiler's murine encephalomyelitis virus (TMEV)-positive cells mm(-2) of spinal cord white matter. There were no differences in the amount of remyelination between mice treated with Linomide or water. However, chronically infected mice treated with Linomide had severely reduced spontaneous vertical activity as measured using a activity wheel. Oral tolerization of mice with mouse or bovine myelin had no effect on virus-induced demyelination or virus antigen expression. The contrasting results obtained between the TMEV model and the autoimmune model of demyelination do not support recent reports suggesting that the underlying mechanism of demyelination in the Theiler's model is autoimmune.

Topics: Adjuvants, Immunologic; Administration, Oral; Animals; Cattle; Disease Models, Animal; Female; Hydroxyquinolines; Immune Tolerance; Mice; Mice, Inbred Strains; Multiple Sclerosis; Myelin Sheath; Poliomyelitis; Theilovirus; Treatment Failure

Susceptibility to autoimmunity has been associated with polarization of Th1/Th2 balance in immune system towards the Th1-type of reactivity. We report here that orally administered quinoline-3-carboxamide (Linomide) selectively downregulates Th1 response in BALB/c and SJL mice, leading to reduction of autoimmunity in the BALB/c and SJL models of experimental allergic encephalomyelitis (EAE). This was shown by prevention of EAE in Th1 responding SJL mice and partial downregulation of EAE in Th2-prone BALB/c mice. In a BALB/c model of EAE, in which infection with Semliki Forest A7 virus (SFV-A7) is used for enhancement of autoimmunity, clinical signs of EAE were reduced while mortality due to viral infection in the CNS was enhanced. Selective downregulation of the Th1 response by Linomide also rendered initially resistant SJL mice susceptible to SFV-A7 CNS infection. This was shown by immunohistochemical detection of extensive deposits of viral antigen in numerous perivascular foci within the CNS and abolished virus antigen-specific lymphocyte reactivity in Linomide-treated SJL mice. In addition, analysis of spleen cell cytokine mRNA production profile revealed decreased number of IFN-gamma producing cells in both SJL and BALB/c mice, reduced number of IL-12p40 producing cells in SJL and increased number of 12p40 producing cells in BALB/c mice along with slightly increased IL-4 production in both strains of mice. These results indicate that oral treatment with Linomide induces selective downregulation of Th1 reactivity causing reduction of autoimmunity and increased susceptibility to SFV-A7 CNS infection. Selective downregulation of Th1 response is a desired effect in the treatment of autoimmune diseases but our results suggest that the benefits have to be balanced against the possible loss in immunoprotection against pathogens.

Topics: Adjuvants, Immunologic; Alphavirus Infections; Animals; Antibody Formation; Antigens, Viral; Autoimmunity; Central Nervous System; Cytokines; Disease Susceptibility; Encephalomyelitis, Autoimmune, Experimental; Female; Hydroxyquinolines; Mice; Mice, Inbred BALB C; Mice, Inbred Strains; RNA, Messenger; Semliki forest virus; Spleen; Th1 Cells

Multiple sclerosis (MS) is characterized by high levels of circulating mononuclear cells (MNC) that respond to myelin proteins like myelin basic protein (MBP) in vitro by expressing mRNA of both pro-inflammatory cytokines, e.g. interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha) and lymphotoxin (LT) that may make MS worse, and anti-inflammatory cytokines like IL-4, IL-10 and transforming growth factor-beta (TGF-beta) that may act beneficially. Substances that down-regulate cytokines such as TNF-alpha or promote IL-10 or TGF-beta can be anticipated to affect MS beneficially.. In situ hybridization to detect and enumerate IFN-gamma, TNF-alpha, LT, IL-4, IL-10 and TGF-beta mRNA expressing blood MNC after stimulation with myelin basic protein (MBP), control antigens and without antigen in presence and absence of Linomide (roquinimex, LS-2616) was employed. In parallel, ELISPOT assay to detect MBP- and PHA-reactive IFN-gamma secreting blood MNC+/-Linomide was used.. Here we report that Linomide, a synthetic immunomodulator, at concentrations effective in vivo reduces the number of MBP-reactive TNF-alpha and increases MBP-reactive IL-10 and TGF-beta mRNA expressing MNC from MS patients' blood when analysed in vitro. Compared to dexamethasone, Linomide up-regulated levels of blood MNC expressing mRNA of TGF-beta after culture in presence of MBP.. Changes of cytokine balance towards a production of anti-inflammatory cytokines could be a desirable effect to be evaluated in future drug studies of Linomide-like substances. At present, Linomide is not evaluable in MS clinical trials due to side-effects.

Topics: Adjuvants, Immunologic; Adult; Aged; Cytokines; Female; Humans; Hydroxyquinolines; Inflammation; Male; Middle Aged; Multiple Sclerosis; RNA, Messenger

Topics: Animals; Antineoplastic Agents; Humans; Hydroxyquinolines; Interferon-gamma; Interleukin-12; Interleukin-18; Macrophages; Neoplasms, Experimental; Plasminogen Activator Inhibitor 2

Tumor-associated macrophages (TAMs) can either promote angiogenesis (i.e., the formation of new blood vessels) in tumors by secreting tumor necrosis factor-alpha (TNF-alpha) or inhibit angiogenesis by producing granulocyte-macrophage colony-stimulating factor (GM-CSF), which in turn stimulates production of the antiangiogenic protein plasminogen activator inhibitor type 2 (PAI-2). We tested, alone or in combination, the anti-prostate cancer activity of agents that perturb macrophage function.. By use of enzyme-linked immunosorbent assays, we measured the effects of Linomide (roquinimex), thalidomide, pentoxifylline, and genistein on TNF-alpha and GM-CSF production in vitro by virally transformed RAW 264.7 mouse macrophages and on PAI-2 production in vitro by human macrophages. The antitumor effects of these agents were tested in vivo on transplanted Dunning R-3327 MAT-Lu rat prostate cancers; TAM numbers and blood vessel densities in these cancers were determined by use of immunocytochemistry.. Linomide selectively inhibited mouse macrophage secretion of TNF-alpha but not of GM-CSF; however, thalidomide, pentoxifylline, and genistein inhibited the production of both cytokines. Linomide, but not thalidomide or pentoxifylline, increased production of PAI-2 by human macrophages. When administered to rats bearing MAT-Lu tumors, each of the tested agents reduced TAM numbers (Linomide, by 46%; thalidomide, by 94%; pentoxifylline, by 71%; and genistein, by 96%). However, all of the agents reduced tumor blood vessel density and tumor growth, with Linomide being the most effective (44% reduction in blood vessel density and 69% inhibition of tumor growth). None of the other agents potentiated Linomide's antitumor effect.. Linomide is unique among the antiangiogenic agents tested, in that it inhibits the stimulatory effects of TAMs on tumor angiogenesis without eliminating their antiangiogenic effects, and may thus prove to be more effective against prostate cancer.

Topics: Animals; Antineoplastic Agents; Enzyme-Linked Immunosorbent Assay; Genistein; Granulocyte-Macrophage Colony-Stimulating Factor; Humans; Hydroxyquinolines; Macrophages; Male; Mice; Neovascularization, Pathologic; Pentoxifylline; Prostatic Neoplasms; Rats; Thalidomide; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha

Experimental autoimmune neuritis (EAN) is an animal model that shares clinical, pathological and electrophysiological features with the human disease Guillain-Barre syndrome. The synthetic immunomodulatory substance linomide has been successfully used to prevent the development of several experimental autoimmune models in laboratory animals and has been proved to be beneficial in modulating the course of multiple sclerosis in humans. In the present study we demonstrate that oral administration of linomide prevents the development of clinical and histopathological signs of EAN in Lewis rats, inoculated with the P2 (60-70) synthetic peptide. The immunomodulatory effect of linomide on this experimental model of disease was associated with marked apoptosis of lymphocytes in thymus and spleen early after starting the treatment. Furthermore, a downregulation of the endothelial expression of the adhesion molecules ICAM-1 at the target site and LFA-1 on lymphocytes could also contribute to the absence of inflammatory cells in the neuraxis.

Topics: Adjuvants, Immunologic; Animals; Apoptosis; Female; Hydroxyquinolines; Immunohistochemistry; Intercellular Adhesion Molecule-1; Lymphocyte Function-Associated Antigen-1; Lymphocytes; Myelin P2 Protein; Neuritis, Autoimmune, Experimental; Rats; Rats, Inbred Lew; Sciatic Nerve

Linomide (quinoline-3-carboxamide) is a synthetic immunomodulator that suppresses several experimental autoimmune diseases. Here we report the effects of Linomide on chronic progressive and/or relapsing experimental autoimmune encephalomyelitis (PR-EAE), a CD4+ T cell mediated animal model of multiple sclerosis (MS). PR-EAE induced in DA rats by inoculation with homogenized guinea pig spinal cord and Freund's complete adjuvant, was strongly suppressed by Linomide administered daily subcutaneously from the day of inoculation. Linomide dose-dependently delayed the interval between immunization and onset of clinical PR-EAE, reduced severity and relapse of clinical PR-EAE, and shortened clinical PR-EAE. These clinical effects were associated with the down-modulation of CNS antigen-induced T cell responses and production of proinflammatory cytokines (IFN-gamma and TNF-alpha) as well as with upregulation of IL-4 (except in spleen MNC), IL-10 and TGF-beta in both spleen MNC and the spinal cord. These effects indicate that Linomide can suppress PR-EAE and may mediate its suppressive effects by regulation of cytokines.

Topics: Adjuvants, Immunologic; Animals; Body Weight; Cell Division; Cells, Cultured; Cytokines; Dose-Response Relationship, Drug; Encephalomyelitis, Autoimmune, Experimental; Guinea Pigs; Hydroxyquinolines; Leukocytes, Mononuclear; Male; Rats; Rats, Inbred Strains; RNA, Messenger; Spinal Cord; Spleen; T-Lymphocytes; Time Factors

The immunomodulatory drug Linomide (PNU-212616) is an efficient inhibitor of experimental autoimmune encephalomyelitis (EAE) and a variety of other models of autoimmunity. The mechanism of action of the drug is, however, incompletely resolved. It was recently suggested that Linomide might exert its immunomodulatory activity by stimulation of the hypothalamic-pituitary-adrenal axis. To investigate the relevance of this mechanism of action, we monitored the plasma levels of endogenous corticosterone after treatment with Linomide, and also directly compared the inhibitory activity of the drug on acute EAE induced in sham or adrenalectomized SJL/N mice. Treatment with Linomide resulted in a dose related inhibition of EAE in line with previously reported results. Upon onset of clinical signs of EAE, there was a 7-10 fold elevation of plasma corticosterone from the normal baseline level. Administration of Linomide did however not by itself result in any change in plasma corticosterone levels, neither at the pre-symptomatic phase of the disease nor during acute short term treatment. In adrenal ectomized animals immunized for EAE, paralytic disease developed rapidly and was of a more severe and fatal nature as compared to sham-operated controls. Treatment with Linomide had a profound inhibitory effect on development of paralytic disease in both the ectomized and sham-operated groups. These results strongly suggest that Linomide does not exert its immunomodulatory activity via the release of endogenous glucocorticoids.

Topics: Adjuvants, Immunologic; Adrenalectomy; Animals; Corticosterone; Encephalomyelitis, Autoimmune, Experimental; Female; Glucocorticoids; Hydroxyquinolines; Mice; Mice, Inbred C57BL

Nitric oxide is involved as a messenger molecule in a large number of physiologic and pathologic responses. Local generation of high nitric oxide output through the expression of the calcium-independent, cytokine-inducible form of nitric oxide synthase (iNOS) can result in either protective or damaging effects. The development of drugs that specifically suppress iNOS expression or inhibit its activity may therefore provide an excellent therapeutic tool for treatment of a diverse set of dysfunctions, including asthma, inflammatory processes, and autoimmune disease. We show compelling evidence that linomide, an immunomodulator known to ameliorate autoimmune diseases, prevents accumulation in the macrophages of mRNA encoding iNOS in mice injected with LPS. This effect is partially mediated by the blocking of TNF-alpha and IL-beta production by activated macrophages. Here, we also present evidence that kidneys from MRL/Mp-lpr/lpr mice express high iNOS levels when the mice develop glomerulonephritis. The administration of linomide to MRL/Mp-lpr/lpr mice significantly decreases iNOS mRNA levels and prevents the development of glomerulonephritis, extending the half-life of mice of this strain. This linomide effect is compatible with its role in preventing the development of autoimmune disease and extends its possible use to other pathologic manifestations associated with iNOS expression, such as the systemic lupus erythematosus-associated glomerulonephritis present in MRL/Mp-lpr/lpr mice.

Topics: Adjuvants, Immunologic; Animals; Enzyme Induction; Hydroxyquinolines; Kidney; Lipopolysaccharides; Lupus Erythematosus, Systemic; Macrophage Activation; Mice; Mice, Inbred MRL lpr; Nephritis; Nitric Oxide Synthase; Shock, Septic

Suppressive effects of the synthetic immunomodulatory drug Linomide have been shown in several autoimmune models, including antibody-mediated experimental autoimmune myasthenia gravis (EAMG), a model for human myasthenia gravis (MG). To define the mechanisms underlying EAMG suppression, we injected Linomide subcutaneously at different doses into Lewis rats immunized with Torpedo acetylcholine receptor (AChR) in complete Freund's adjuvant (CFA), and investigated AChR-specific T and B cell responses, and the levels of lymph node cells expressing mRNA of different cytokines after AChR stimulation in vitro. Both 160 and 16, but not 1.6, mg/kg/day of Linomide effectively suppressed clinical muscle weakness, accompanied by decreased AChR-induced T and B cell responses. Linomide also suppressed the mRNA expression of the Th1 cytokines IFN-gamma, IL-12 and TNF-alpha as well as the Th2 cytokines IL-4 and IL-10, which are important in the immunopathogenesis of EAMG by promoting antibody production. There were no differences for IL-1beta, IL-6, lymphotoxin or TGF-beta expression in Linomide-treated vs nontreated control EAMG rats. We conclude that Linomide suppresses clinical EAMG as well as B and T cell responses to AChR by counteracting the production of AChR-induced Th1 and Th2 cytokines.

Topics: Adjuvants, Immunologic; Animals; Antibodies; Cell Division; Cytokines; Female; Hydroxyquinolines; Immunoglobulin G; Lymphocytes; Monocytes; Muscle Weakness; Myasthenia Gravis; Rats; Rats, Inbred Lew; Receptors, Cholinergic; RNA, Messenger

Linomide is a p.o. active antiangiogenic agent that has been demonstrated to be effective in suppressing the in vivo growth of rat and human prostatic cancer xenografts. The present studies were conducted to determine whether the angiogenic molecules, vascular endothelial growth factor/vascular permeability factor (VEGF/VPF) and basic fibroblast growth factor (bFGF) are expressed in vitro by DU-145, PC-3, TSU-PR1, and LnCaP human prostate cancer cell lines and whether Linomide inhibits the secretion of these angiogenic molecules. Additionally, two different androgen-responsive human prostatic cancer xenograft models (i.e., PC-82 and A-2) were used to determine whether androgen ablation-induced reduction in tumor growth is associated with a reduction in tumor VEGF and/or bFGF levels. These studies demonstrated that both VEGF and bFGF proteins are expressed to different degrees in the human prostatic cancer cell lines. The secretion of VEGF but not bFGF is up-regulated by hypoxia. Linomide is unable to inhibit either basal or hypoxia-induced secretion of VEGF. Linomide also has no effect on secreted bFGF levels. Castration inhibited tumor VEGF but had no effect on bFGF levels in both the androgen-responsive PC-82 and A-2 human prostatic cancers when grown in severe combined immunodeficient mice. When given in combination, castration potentiated the inhibition of tumor growth induced by Linomide alone. This potentiation is not due to a further inhibition in tumor VEGF levels induced by castration. Although both castration and Linomide inhibit angiogenesis, the former accomplishes it by inhibiting VEGF secretion, whereas the latter has multiple effects at several steps in the angiogenic process other than VEGF secretion. Based on their different but complementary mechanisms of action, simultaneous combination of androgen ablation with Linomide enhances the anti-prostatic cancer efficacy compared to either monotherapies alone and warrants testing in humans.

Topics: Adenocarcinoma; Androgens; Animals; Cell Hypoxia; Drug Screening Assays, Antitumor; Endothelial Growth Factors; Fibroblast Growth Factor 2; Gene Expression Regulation, Neoplastic; Humans; Hydroxyquinolines; Lymphokines; Male; Mice; Mice, SCID; Neoplasm Proteins; Neoplasm Transplantation; Neoplasms, Hormone-Dependent; Neovascularization, Pathologic; Orchiectomy; Prostatic Neoplasms; Specific Pathogen-Free Organisms; Tumor Cells, Cultured; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors

Anti-proliferative drugs have been used for immunosuppression since the introduction of clinical transplantation. Most transplant centres include azathioprine (Aza) and cyclosporine (CyA) in their standard regimens, despite several controlled studies having failed to confirm the benefit of this combination. Aza is still the most commonly used anti-proliferative drug, although no major differences in immunosuppressive or toxic effects have been shown between Aza and cyclophosphamide (Cph). Cph as an adjunct to CyA has never been tested in a randomized study. Recently, mycophenolate mofetil (MMF) has been developed as the most selective inhibitor of T- and B-cell proliferation and promoted as an adjunct to CyA treatment. In the present study, the additive or synergistic effects of these three anti-proliferative agents in combined treatment with CyA have been investigated using a rat cardiac transplantation model in which the immunomodulator linomide (Lin) was included as a potentiator of rejection. As single drug treatment, CyA, Cph and MMF, but not Aza, exerted a beneficial effect on graft survival. This prolongation of graft survival was abrogated when any one drug was administered together with Lin. The addition of MMF, Aza or Cph to CyA plus Lin treatment improved the graft survival significantly, thus demonstrating each of the anti-proliferative drugs to exert additive or synergistic effects in conjunction with cyclosporine. MMF seemed to be the most effective and least toxic of the drugs tested.

Topics: Adjuvants, Immunologic; Animals; Azathioprine; Cyclophosphamide; Cyclosporine; Drug Synergism; Drug Therapy, Combination; Graft Survival; Heart Transplantation; Hydroxyquinolines; Immunosuppressive Agents; Male; Mycophenolic Acid; Rats; Rats, Inbred Lew; Rats, Inbred WKY

Linomide (quinoline-3-carboxamide, LS-2616), a synthetic immunomodulator, protects animals against a variety of experimental autoimmune diseases. In experimental autoimmune encephalomyelitis, an animal model of multiple sclerosis (MS), linomide blocks both the clinical and histological signs of the disease, without inducing generalized immunosuppression. In the first clinical trial in patients with MS, linomide was shown to inhibit the progression of the disease. In the present study we investigated several aspects of the mechanisms of action of this immunomodulator. We found that linomide can inhibit acute EAE even when given as pretreatment, prior to induction of disease (days - 10 to 0). This inhibitory effect was reversed by adoptive transfer of naive spleen cells. A short course (7 days) of linomide treatment also inhibited EAE, especially when administered immediately after disease induction. Spleen cells from linomide-treated mice failed to present myelin antigens to T-cell lines in vitro. The defective antigen presentation was normalized by anti-oxidants such as 2-mercaptoethanol. The proportion of Mac1+ cells in the spleens of linomide-treated mice was significantly reduced and macrophage growth was inhibited in long term cultures of spleen cells derived from linomide-treated animals. Our findings suggest that the effect of linomide on EAE may be attributed, at least in part, to inactivation of antigen presenting cells, possibly following a short period of over-stimulation and increased oxidant production. This mechanism may play a universal role in the regulation of autoimmune reactivity and merits further investigation.

Topics: Adjuvants, Immunologic; Animals; Antigen-Presenting Cells; Autoimmunity; Cell Adhesion; Cell Count; Cell Division; Cells, Cultured; Concanavalin A; Encephalomyelitis, Autoimmune, Experimental; Female; Hydroxyquinolines; Lipopolysaccharides; Macrophages; Macrophages, Peritoneal; Mice; Reference Values; Spleen; Time Factors

Linomide (LS-2616, quinoline-3-carboxamide) has been reported to exert a diverse range of effects on the immune system. On one hand, this drug was found to stimulate the immune system and to enhance activities such as DTH or allograft rejection. On the other hand, linomide was shown to inhibit the induction of experimental autoimmune encephalomyelitis and myasthenia gravis, as well as the development of diabetes in non-obese diabetic (NOD) mice. Here we report the effects of linomide in animals immunized with uveitogenic retinal antigens. Treatment with linomide completely inhibited the development of experimental autoimmune uveoretinitis (EAU) in mice immunized with interphotoreceptor retinoid-binding protein and markedly suppressed EAU in rats immunized with S-antigen (S-Ag). In addition, linomide-treated rats exhibited reduced antibody production and lymphocyte proliferative response to S-Ag. In contrast to these suppressive activities, linomide treatment did not affect the development of adoptively transferred EAU in rats and moderately enhanced the DTH reactions to S-Ag in immunized rats in which EAU and other immune responses to this antigen were suppressed.

Topics: Adjuvants, Immunologic; Animals; Arrestin; Autoimmune Diseases; Female; Hydroxyquinolines; Hypersensitivity, Delayed; Immunization; Lymphocyte Activation; Male; Mice; Rats; Rats, Inbred Lew; Retinitis; Uveitis

Experimental autoimmune neuritis (EAN) is a T-cell mediated autoimmune disease of the peripheral nervous system, in which macrophages and T-cells feature prominently in nerve lesions. EAN represents a counterpart to Guillain-Barré syndrome in humans. In the present study, we investigated the in vitro and in vivo effects of Linomide (LS-2616, quinoline-3-carboxamide), a synthetic immunomodulatory compound, on macrophages in relation to EAN. Linomide strongly suppressed IFN-gamma and lipopolysaccharide (LPS)-induced IL-1 beta, TNF-alpha and IL-6 mRNA expression in macrophages in vitro as demonstrated by in situ hybridisation. Linomide administered daily subcutaneously from the day of inoculation completely prevented the development of clinical symptoms of EAN. Linomide administered from day 9 post immunisation (p.i.) significantly suppressed clinical EAN. Macrophages from Linomide-treated EAN rats showed decreased IL-1 beta, TNF-alpha and IL-6 mRNA expression in response to IFN-gamma and LPS. LPS-induced nitric oxide production by macrophages was also suppressed by Linomide in vitro. Linomide, however, does not affect macrophage death and release of lactate dehydrogenase. We conclude that Linomide may exert its actions in EAN and perhaps also in other autoimmune disease models, by suppressing macrophage functions.

Topics: Adjuvants, Immunologic; Animals; Down-Regulation; Hydroxyquinolines; Interleukin-1; Interleukin-6; Lipopolysaccharides; Macrophages; Male; Neuritis, Autoimmune, Experimental; Nitric Oxide; Rats; Rats, Inbred Lew; RNA, Messenger; Tumor Necrosis Factor-alpha

Escape from "castration inhibition," be it surgical or chemically induced, is still the major problem in prostate cancer treatment. New agents that can be given as adjuvant therapy are needed. Linomide has demonstrated both anti-tumor and anti-angiogenic activity with little toxicity in the Dunning R-3327 rat prostate tumor system. Therefore it was deemed essential to study the efficacy of this drug in the adjuvant situation.. Linomide, roquinimex, was administered 3 times a week i.p. alone or in conjunction with castration to rats bearing the Dunning R-3327 PAP rat prostate tumor and its effect on tumor growth analyzed. Similar experiments, in which Linomide 25 mg./kg./day was given in the drinking water were carried out in rats with the Dunning R-3327 G tumor. The effect of treatment on blood vessel density and blood flow in the tumor was also assessed using an image analysis system.. Linomide, 2.5 & 40 mg./kg., administered from the day after castration inhibited the regrowth of the Dunning R-3327 PAP tumors In addition, Linomide 40 mg./kg. administered after tumor regrowth occurred following castration(week 10) inhibited further tumor growth. Inhibition of tumor regrowth after castration was also found in the Dunning G tumor. When Linomide treatment was stopped regrowth of the tumors occurred, either in the same animal or on transplantation to new intact hosts, demonstrating that the tumor cells were still viable. Tumor blood vessel density was decreased both after castration and Linomide treatment alone, 40 and 32% respectively. On combination of castration and Linomide a 60% decrease in blood vessel density was found. This was significantly different from either of the two treatments given alone. The enhancement on combining castration and Linomide was confirmed by a further decrease in blood flow, from 19 and 22 to 12 ml. per minute/gm. tissue respectively.. Linomide, an anti-angiogenic drug, inhibits escape from "castration inhibition".

Topics: Animals; Antineoplastic Agents; Castration; Chemotherapy, Adjuvant; Humans; Hydroxyquinolines; Male; Neovascularization, Pathologic; Prostatic Neoplasms; Rats; Rats, Inbred Strains

Topics: Animals; Antineoplastic Agents; Breast Neoplasms; Female; Humans; Hydroxyquinolines; Male; Neovascularization, Pathologic; Prostatic Neoplasms; Rats; Tamoxifen

Linomide (quinoline-3-carboxamide), a well tolerated, orally administered compound was recently shown to be effective in the prevention and treatment of several autoimmune diseases in experimental animal models. We have investigated its effect on specific humoral immune responses directed to T-cell-dependent soluble or particulate antigens and to a T cell-independent antigen in several mouse strains. Linomide administered after antigen priming did not affect primary and secondary antibody responses directed to T-cell particulate antigens (SRBC) or soluble antigens given with or without complete Freund's Adjuvant (CFA). Linomide treatment given prior to antigen priming did not affect the antibody response to a soluble antigen (TNP-KLH) given with an adjuvant. In contrast, dose-dependent down regulation of primary antibody responses was observed when T cell-dependent (BSA-dextran) or T-cell-independent (TNP-Ficoll) antigens were administered in an immunogenic form without adjuvant after starting Linomide treatment. The primary anti-SRBC antibody response was also suppressed by high dose Linomide given prior to immunization although normal secondary responses were retained. It is worth noting that no immunosuppressive effects on antibody responses were found at low dose ranges which effectively reversed T cell dependent autoimmune manifestation.

Topics: Animals; Antibody Formation; Antigen Presentation; Antigens, T-Independent; Autoantibodies; Autoantigens; Autoimmune Diseases; Dextrans; Diabetes Mellitus, Type 1; Drug Administration Schedule; Encephalomyelitis, Autoimmune, Experimental; Female; Ficoll; Freund's Adjuvant; Haptens; Hemocyanins; Hydroxyquinolines; Immunization; Immunologic Factors; Mice; Mice, Inbred BALB C; Mice, Inbred DBA; Mice, Inbred NOD; Myelin Basic Protein; Peptide Fragments; Serum Albumin, Bovine; Solubility; Trinitrobenzenes

In a recent double-blind, phase II study, conducted in our department, we showed that Linomide-treated MS patients had significantly less active lesions (in serial monthly MRI tests) and a tendency for clinical stabilization. Here we present the immunological evaluation of the patients who participated in this study and propose a novel mechanism by which Linomide downregulates autoreactivity. Peripheral blood leukocytes (PBLs), serum, and CSF samples were obtained at two to four time points over the 6 months of the trial. Flow cytometric analysis (FACS) of the CD5/CD19, CD4/CD8, CD14/CD3, CD16/CD3, CD45RA/CD4, and CD45RO/CD4 surface markers on PBLs was performed and the levels of the IL-1beta, IL-2, IL-4, IL-6, IL-10, TNF-alpha, IFN-gamma, and IL-2R were also examined. White blood counts of Linomide-treated patients were consistently elevated throughout the treatment period (P = 0.002-0.04). Cytokines levels in serum and CSF were highly fluctuating and we could not detect any clear trend as a result of Linomide treatment. FACS analysis showed that Linomide treatment significantly increased the percentage of the CD4+/CD45RA+ cells (from 35.5% at baseline to 42.3% at week 24; P = 0.02), and decreased CD4+/CD45RO+ lymphocytes (62.6% at baseline vs 53.7% at week 24, P = 0.02). Linomide also induced a transient increase in the NK-cells, the NK 1.1 cells, and the CD5 B-cells (P = 0.02). Upregulation of naive CD45RA T-lymphocytes and parallel downregulation of memory CD45RO cells seems to be one of the main mechanisms by which Linomide inhibits MS activity and may represent an alternative immunomodulating approach for the treatment of MS and autoimmunity in general.

Topics: Adjuvants, Immunologic; Adult; Blood; CD4-Positive T-Lymphocytes; Cytokines; Disease Progression; Down-Regulation; Female; Humans; Hydroxyquinolines; Leukocyte Common Antigens; Leukocyte Count; Lymphocyte Subsets; Male; Middle Aged; Multiple Sclerosis; Up-Regulation

Linomide is a synthetic compound that affects various immunological functions and inhibits experimental autoimmune encephalomyelitis (EAE). In the present study we evaluated the effect of linomide on the HPA axis functions under basal and stress-induced conditions and examined whether the effect of linomide on the HPA axis is involved in linomide-induced amelioration of EAE in rats. Linomide caused a significant increase of serum ACTH and corticosterone (CS). The adrenocortical response to various stress modalities as well as the negative feedback exerted by glucocorticoids was not affected. The marked reduction of thymus weight following linomide treatment was abrogated in adrenalectomized rats. The induction of EAE in adrenalectomized rats was completely inhibited by linomide treatment. These results suggest that the increased CS levels induced by linomide are responsible for the decrease in thymus weight but do not play a role in the therapeutic effect of this drug in EAE.

Topics: Adjuvants, Immunologic; Adrenal Cortex; Adrenalectomy; Adrenocorticotropic Hormone; Animals; Corticosterone; Dexamethasone; Encephalomyelitis, Autoimmune, Experimental; Feedback; Glucocorticoids; Hydroxyquinolines; Male; Organ Size; Rats; Rats, Inbred Strains; Reference Values; Stress, Physiological; Thymus Gland

Linomide, a quinoline-3-carboxamide, has a pleiotropic immune modulating capacity and inhibits development as well as progression of disease in animal models of autoimmunity. Linomide treatment of mice resulted in a dramatic, dose-dependent decrease of the thymic cell number shortly after the start of administration. Flow cytometric analysis revealed that the major thymocyte subset, the early immature type CD4+CD8+ thymocytes, were reduced in number by 75%, mature CD4+CD8- or CD4-CD8+ thymocytes were less sensitive to treatment. The polyclonal T cell activator Con A (Concanavalin A) was used together with IL-2 to evaluate the potential proliferative responsiveness of ex vivo thymocytes. Thymocytes from mice treated with Linomide exhibited a more vigorous proliferation than control cultures. An effect shown to not only be due to the enrichment of mature thymocytes in the cultures from Linomide treated animals, but also when purified, mature thymocytes (CD4+CD8- and CD4-CD8+) were cultured with Con A and IL-2, these cells responded with a significantly enhanced proliferation. In vivo Linomide treatment did not result in increased plasma concentrations of corticosterone and treatment of adrenalectomized mice resulted in a reduction of thymocytes which was comparable to the effect in intact mice, indicating that glucocorticoids (GC) are not major mediators of Linomide-induced thymocyte deletion. In addition to this, and supporting a glucocorticoid independent mode of action, Linomide treatment of thymocytes in vitro resulted in a significant increase in the number of apoptotic cells, specifically in the CD4+CD8+ subset, implicating apopotosis as one component in the course of thymocyte reduction. In addition to this, in vivo treatment with Linomide resulted in an identical pattern to that seen in vitro in that there was significantly increased apoptosis only in the CD4+CD8+. These data indicate that Linomide modifies thymocyte development using a glucocorticoid independent pathway and results in the increased apoptosis of the CD4+CD8+ subset.

Topics: Adjuvants, Immunologic; Adrenalectomy; Animals; Apoptosis; CD4 Antigens; CD4-Positive T-Lymphocytes; CD8 Antigens; CD8-Positive T-Lymphocytes; Cell Differentiation; Cells, Cultured; Concanavalin A; Corticosterone; Dose-Response Relationship, Drug; Female; Glucocorticoids; Hydroxyquinolines; Interleukin-2; Lymphocyte Activation; Lymphocyte Count; Mice; Mice, Inbred C57BL; T-Lymphocyte Subsets; Thymus Gland

Repeated administration of monoclonal antibodies (mAb) directed against the CD4 lymphocyte receptor may induce specific, long-lasting unresponsiveness to fully MHC-mismatched cardiac allografts in rats without additional immunosuppression. We assessed the effect of a single dose of murine anti-rat depleting anti-CD4 mAb (OX-38) on allograft survival in high- and low-responder rat strain combinations. Isogenic strains of DA (RT1av1), PVG (RT1c), AUG (RT1c), and WF (RT1u) rats were used. Recipients in antibody treated groups were given one dose of 5 mg/kg OX-38 mAb on the day of transplant, a dose which was shown to effectively deplete (or block) circulating CD4+ T cells. Other groups were treated for 10 days with cyclosporin A (CsA) and/or Linomide, a novel immunomodulator, which is the first compound able to fully eliminate the effect of CsA in the rat cardiac allograft model. The DA strain was identified as a low-responder to the allogeneic haplotype RT1c (PVG or AUG), but not to RT1u (WF), and developed true tolerance following RT1c grafting and OX-38 or low-dose CsA (5 mg/kg) induction, as verified by the response to retransplantation of a graft from the same donor strain or a third-party challenge. PVG recipients of DA grafts were characterized by high response and only modest (OX-38; median 9.5 days) or moderate (CsA; 23.5 days) prolongation of graft survival. Contrasting graft survival results were obtained in the low-responder combination, either very early rejection (at 10 days) or permanent graft survival (> 100 days). Linomide challenge affected CsA treatment in the high-responder combination but not tolerance induction in the low-responder combination, or the effect of OX-38. It was concluded that in rat heart transplantation a single-dose anti-CD4 mAb therapy may induce permanent donor-specific unresponsiveness in a low-responder strain combination, and that anti-CD4 mAb seems to be unique among immunosuppressive agents while being resistent to challenge by Linomide.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Monoclonal; CD4 Antigens; CD4-CD8 Ratio; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cyclosporine; Heart Transplantation; Hydroxyquinolines; Immunoglobulin G; Immunosuppressive Agents; Male; Mice; Rats; Rats, Inbred Strains; Species Specificity; Transplantation, Homologous

1. Linomide (N-phenylmethyl-1,2-dihydro-4-hydroxyl-1-methyl-2-oxoquinoline-3-carb oxa mide) inhibits vascular proliferation and has been proposed as an antiangiogenic drug. We have investigated the vascular effect of linomide in rabbit aortic and saphenous vein ring preparations and in rat cultured vascular smooth muscle cells (VSMCs). 2. Linomide (25-300 micrograms ml-1) did not alter the basal tone of the preparations. The drug induced a concentration-dependent relaxant effect in aortic rings with endothelium, preconstricted by noradrenaline (NA), 5-hydroxytryptamine (5-HT) and by the thromboxane mimetic U46619. 3. The degree of relaxation induced by linomide was significantly reduced by exposure to the cyclooxygenase inhibitors indomethacin (3 microM) and acetylsalicylic acid (500 microM), and was not influenced by pretreatment with the nitric oxide synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME) (100 microM) in aortic rings with endothelium, preconstricted with NA. 4. Endothelium removal significantly reduced the relaxant response to linomide in aortic ring preparations. 5. A concentration-dependent relaxant response was observed also in rabbit saphenous vein preparations deprived of endothelium and preconstricted either by NA or U46619. The degree of relaxation obtained in a high potassium solution was consistently smaller than that observed in NA-pretreated venous preparations. 6. The vasorelaxant effect of linomide was consistently blunted by the adenylate cyclase inhibitor SQ 22536 (50 microM), both in intact aortic rings and in those deprived of endothelium. 7. In rat cultured vascular smooth muscle cells, linomide (100-200 micrograms ml-1) induced a significant increase in cyclic AMP levels, which was blocked by exposure to 50 microM SQ 22536. 8. In endothelium-deprived aortic ring preparations, the linomide-induced relaxant effect was greatly reduced in high potassium medium (KCl = 25 mM). Pretreatment with the ATP potassium channel inhibitor glibenclamide (3 microM) significantly reduced the linomide-induced relaxation. 9. The results show that linomide possesses a vasorelaxant effect which is attributable to both endothelium-dependent and -independent properties. While the former component of the drug's activity is apparently due to the release of a prostanoid from endothelial cells, the endothelium-independent mechanism involved in linomide relaxation is linked to cyclic AMP accumulation and to ATP-sensitive potassium channel ac

Topics: Animals; Aorta; Cyclic AMP; Endothelium, Vascular; Hydroxyquinolines; Male; Muscle, Smooth, Vascular; Potassium Channel Blockers; Rabbits; Rats; Rats, Wistar; Saphenous Vein; Vasodilation

Donor-derived cell-mediated immunotherapy has been shown to be an effective tool for reinduction of remission in chronic myeloid leukaemia (CML) patients who have relapsed post-bone marrow transplantation (BMT). Linomide, quinoline-3-carboxamine (LS 2616), is a new immunomodulator shown to increase the number of NK precursors in mice in addition to upregulating the quantity of CD56(+), CD3(-) and CD16(+) NK cells in the peripheral blood of patients following autologous BMT (ABMT). We investigated the in vitro effects of Linomide on NK activity of normal human donors. Large granular lymphocytes (LGLs) and NK cells were incubated overnight with Linomide (0.02-4.8 mg/ml), recombinant human interleukin-2 (IL-2, 75 IU/ml), or a combination of both. Linomide, at 0.02-0.3 mg/ml, augmented IL-2-induced proliferation of LGLs and NK cells in an inversely proportional manner. In contrast, Linomide at 0.6-4.8 mg/ml inhibited IL-2-induced proliferation of LGLs and NK cells in a dose-dependent manner. Linomide was able to potentiate phytohemaglutinin-induced CD3(+) cell proliferation. In addition, supernatants derived from Linomide treated CD3(+) T cells were able to mimic the direct stimulatory effect of Linomide on activated NK cell proliferation. These supernatants were found to have low levels of tissue necrosis factor-alpha (TNF-alpha) and interleukin-1 (IL-1) and therefore Linomide stimulation of NK and T cell proliferation may be due to its inhibitory effect on the secretion of these cytokines by activated CD3(+) T cells. Linomide had no effect on cytotoxicity nor on the phenotypic expression of resting and IL-2-activated LGLs or NK cells. In view of our results, Linomide could possibly play a potential role in adoptive cell-mediated immunotherapy post-BMT.

Topics: Adjuvants, Immunologic; Cytotoxicity, Immunologic; Humans; Hydroxyquinolines; Interleukin-2; Killer Cells, Natural; Lymphocyte Activation; Phytohemagglutinins

Topics: Adjuvants, Immunologic; Antibodies, Monoclonal; Graft vs Host Disease; Humans; Hydroxyquinolines; Immunotherapy; Interleukin-2; Leukemia, Myeloid, Acute; Leukocyte Transfusion; Neoplasm, Residual

There are two distinct phases during prostatic carcinogenesis with regard to tumor blood vessel development. During the first or prevascular phase, which may persist for years, cells that have undergone some but not all of the transformation steps undergo a limited amount of net growth, producing premalignant prostatic intraepithelial neoplastic (PIN) lesions. Most of these PIN lesions do not continue net growth and do not progress to produce histologically detectable cancer. Even the PIN lesions that do progress to cancer remain of limited virulence unless they undergo conversion to the second or angiogenic phase. Once this angiogenic phase is reached, new blood vessel development is greatly enhanced within the cancer. It is this enhanced tumor angiogenesis which allows these cancers both to grow continuously and to metastasize. Thus, inhibition of angiogenesis should be an effective chemopreventive approach for prostatic carcinogenesis. Linomide is a low molecular weight, water-soluble agent with excellent p.o. absorption and bioavailability. We have previously demonstrated that daily p.o. treatment with Linomide has antiangiogenic abilities against a series of rat and human prostatic cancer xenografts growing in vivo. In the present studies, we have demonstrated using Matrigel in in vivo angiogenesis assays that daily p.o. Linomide at 25 mg/kg/day inhibits angiogenesis induced by tumor necrosis factor alpha, acidic fibroblast growth factor, basic fibroblast growth factor, and vascular endothelial growth factor. Using an N-methylnitrosourea initiation-androgen promotion model, Linomide was given p.o. at a daily dose as high as 25 mg/kg/day for at least 1 year without major toxicity while inhibiting the development of seminal vesicle/prostate cancers in male rats by >50%. Dose-response analysis demonstrated that a Linomide blood level of 50-100 microM is optimal for such chemoprevention. In addition, Linomide treatment at a dose of 25 mg/kg/day was able to inhibit by approximately 60% the incidence of N-methylnitrosourea and approximately 50% of 7,12-dimethyl-benz(a)anthracine-induced mammary carcinogenesis in female rats.

Topics: Androgens; Animals; Anticarcinogenic Agents; Carcinogens; Female; Genital Neoplasms, Male; Growth Substances; Hydroxyquinolines; Male; Mammary Neoplasms, Experimental; Neovascularization, Pathologic; Prostatic Neoplasms; Rats; Rats, Inbred Strains; Rats, Wistar; Seminal Vesicles; Sensitivity and Specificity; Tumor Necrosis Factor-alpha

Fas is an apoptosis-signaling receptor molecule expressed in vivo on thymocytes, liver, heart, and ovary. In vivo administration of the anti-Fas Jo2 antibody in mice induces severe apoptotic liver damage leading to fulminant hepatitis and death. Linomide, a quinoline 3-carboxamide, inhibits apoptosis of B and T cells induced by various stimuli including viruses, superantigens, and glucocorticoids. Mice treated with linomide survived the lethal effect of anti-Fas antibody, did not accumulate ceramide in hepatocytes, and recovered liver structure and function within 96 h of anti-Fas injection, as confirmed by histology and glutamic oxalacetic transaminase, glutamic pyruvic transaminase, and lactate dehydrogenase levels. Surviving mice showed severe depletion of cortical thymocytes, but medullar thymic cells expressing high CD3 and Fas levels also survived the treatment with anti-Fas in the presence of linomide. Heart, lung, and ovary showed no signs of apoptosis promoted by Fas ligation. These results suggest that linomide prevents cell death triggered by Fas ligation and can be useful for therapeutic intervention in fulminant hepatitis.

Topics: Animals; Antibodies; Apoptosis; Ceramides; fas Receptor; Female; Hydroxyquinolines; Liver; Male; Mice; Mice, Inbred BALB C; Thymus Gland

We have previously reported that linomide, a quinoline-3-carboxamide, has antitumor effects against prostatic cancers in vivo through its ability to inhibit tumor angiogenesis. Subsequently, we reported that linomide inhibits several steps in the process of angiogenesis, including direct effects on endothelial cell proliferation and their chemotactic migration and invasion. Besides these direct effects, linomide's antiangiogenic activity also involves indirect effects secondary to inhibition of tumor infiltration of macrophages and their ability to secrete the angiogenic factor tumor necrosis factor-alpha (TNF-alpha). The current studies were conducted to gain insight into the mechanism by which linomide inhibits macrophage TNF-alpha secretion. The virally transformed RAW 264.7 mouse macrophage cell line was used as a model system. Chronic in vitro exposure (7 days) to 81-650 microM linomide is cytostatic to RAW cells. Such chronic exposure to linomide significantly decreased (P < 0.05) RAW cells' baseline ability to secrete TNF-alpha and also their ability to up-regulate TNF-alpha secretion in response to lipopolysaccharide (LPS) challenge. Ribonuclease protection assays demonstrated that linomide's ability to inhibit baseline and LPS-challenged TNF-alpha secretion is not functioning at the mRNA level, because steady-state levels of TNF-alpha mRNA do not change in response to linomide. Linomide's ability to inhibit TNF-alpha secretion is not associated with an increase in cell-associated TNF-alpha levels. Immunoprecipitation experiments demonstrated that linomide did not inhibit the normal proteolytic processing of the initial 26 kDa plasma membrane-bound TNF-alpha to the secreted 17 kDa soluble form. These results demonstrate that linomide inhibits TNF-alpha secretion by inhibition of the synthesis of the TNF-alpha protein. Linomide's ability to inhibit TNF-alpha protein synthesis is not due to an inhibition of general protein synthesis or secretion and is not mediated via a change in cyclic adenosine monophosphate levels.

Topics: Androgen Antagonists; Animals; Cell Division; Cell Line, Transformed; Cyclic AMP; Homeostasis; Hydroxyquinolines; Intracellular Membranes; Mice; Protein Synthesis Inhibitors; RNA, Messenger; Tumor Necrosis Factor-alpha

The objective of this study was to assess the beneficial effects of an early administration of low dose linomide, a new immunomodulator, in an animal model of experimental systemic lupus erythematosus (SLE).. Experimental SLE was induced in naive BALB/c mice, by immunization with anti-DNA mAb (MIV-7). Control Mice immunized with irrelevant human IgM served as controls. The immunized mice were treated with linomide (0.1 mg/ml in the drinking water), four weeks prior to the first immunization, at an early stage of the disease induction (one month after boost injection), or at a later stage (3 months following boost immunization). The treatment duration was 3 months in all schedules. The follow-up studies continued for 8 weeks after discontinuation of the treatment. The presence in the serum of autoantibodies against ssDNA, dsDNA histones, phospholipids and an irrelevant autoantigen-pyruvate dehydrogenase, was determined by enzyme-linked immunosorbent assay (ELISA). The clinical parameters assessed included erythrocyte sedimentation rate, peripheral blood cell counts and proteinuria.. There was a 50-64% decrease in autoantibody levels in the sera of mice immunized with anti-DNA (MIV-7) mAb at the early stage of experimental SLE in mice which received linomide for a period of 3 months. No effect of linomide was noted in mice which received the drug during the later stages of experimental SLE when the disease was fully developed. Linomide had a preventive effect on the induction of experimental SLE in naive mice, when the treatment was initiated before the induction of the disease. This effect was abolished following cessation of the treatment.. Linomide proved to be effective at the early stages of induction of the experimental SLE. However, the autoantibody levels rose following discontinuation of the therapy.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Antinuclear; Antibodies, Monoclonal; Autoimmune Diseases; Disease Models, Animal; Drug Evaluation, Preclinical; Humans; Hydroxyquinolines; Immunization, Passive; Lupus Erythematosus, Systemic; Mice; Mice, Inbred BALB C

In order to assess the mechanism of action of the quinoline-3-carboxyamide linomide as an antiangiogenic drug, the effect of linomide was studied in vitro on postcapillary endothelial cells exposed to vascular endothelial growth factor (VEGF). Linomide did not block the spontaneous replication of endothelial cells, but significantly suppressed endothelial cell growth and migration elicited by VEGF. It is concluded that linomide appears to be an effective tool to inhibit VEGF-dependent angiogenesis.

Topics: Capillaries; Cell Division; Cell Line; Cell Movement; Endothelial Growth Factors; Endothelium, Vascular; Hydroxyquinolines; Neovascularization, Pathologic

Oral therapy with linomide protects prediabetic nonobese diabetic (NOD) mice from insulin-dependent diabetes mellitus. The mechanisms by which linomide exerts its protective effect are not fully understood. A decreased TCR-mediated activity of the GTP-GDP binding p21(ras) proto-oncogene is associated with prediabetes in NOD mice. However, the role of this signal transduction defect in the pathogenesis of autoimmune diabetes is not known. The TCR-mediated and protein kinase C-induced activations of p21(ras) were determined in mononuclear cells from lymph nodes of linomide-treated and untreated prediabetic NOD mice. TCR cross-linking by Con A induced an increase of 13 +/- 6.8% and a decrease of 0.8 +/- 1.8% in p21(ras) activity in the linomide-treated group and the untreated controls, respectively. Cell stimulation with PMA resulted in a 15 +/- 2% increase in p21(ras) activity in the linomide-treated mice and a 10 +/- 11.4% decrease in the untreated mice. Protein levels of p21(ras) and its regulatory elements, the GTPase-activating protein and the guanine nucleotide-releasing factor, mSOS, were comparable in both groups. We, therefore, conclude that prevention of autoimmune diabetes by linomide is associated with up-regulation of the p21(ras) T cell signal transduction defect in NOD mice.

Topics: Adjuvants, Immunologic; Animals; Diabetes Mellitus, Type 1; Female; Hydroxyquinolines; Male; Mice; Mice, Inbred NOD; Proto-Oncogene Proteins p21(ras); Receptors, Antigen, T-Cell; Up-Regulation

Linomide (LS-2616, quinoline-3-carboxamide) has strong immunomodulating effects in animal models, inhibiting toxic shock, progressive autoimmune disease and cancer. In humans, linomide strongly reduced the appearance of new lesions in multiple sclerosis yet enhanced immune responses after bone marrow transplantation. In contrast to these clear effects in vivo, attempts to show an effect of linomide in vitro have not been successful and its mode of action remains to be elucidated. Here we show that at concentrations effective in vivo, linomide is active on human peripheral blood mononuclear cells (PBMC), severely inhibiting the induction by Staphylococcus aureus enterotoxin B of mRNA of three cytokine genes expressed in Th1 cells, those for IFN-gamma, IL-2, and tumor necrosis factor-beta. Yet, cell viability was not affected by linomide. The extent of inhibition is dose-dependent on linomide. Linomide also blocked induction of IL-2 and IFN-gamma mRNA by phytohemagglutinin. The inhibitory effect is expressed immediately but can be enhanced significantly by a prolonged exposure of PBMC to linomide, reaching 10-fold. These results support the concept that linomide antagonizes the activation of Th1 cells during a cellular immune response.

Topics: Adjuvants, Immunologic; Cells, Cultured; Cytokines; Dose-Response Relationship, Immunologic; Enterotoxins; Gene Expression Regulation; Humans; Hydroxyquinolines; Interferon-gamma; Interleukin-2; Leukocytes, Mononuclear; Lymphotoxin-alpha; RNA, Messenger; Staphylococcus aureus; Superantigens; Th1 Cells

Topics: Adjuvants, Immunologic; Autoimmune Diseases; Bone Marrow Transplantation; Graft vs Host Disease; Humans; Hydroxyquinolines; Immunosuppression Therapy; Immunotherapy; Lymphocyte Depletion; T-Lymphocytes; Transplantation, Homologous

The idioptypic (Id) determinant of immunoglobulin expressed on the cell surface of malignant B cells represents a prototypical tumor-associated antigen (TAA), which has been used in a purified soluble form for active immunization in experimental tumor models and human hematological malignancies. Using a spontaneous transplantable murine model of B cell leukemia/lymphoma (BCL1), we have demonstrated the expression of the B7 costimulatory molecules in addition to the previously described Id determinant and class II major histocompatibility antigens. Intact irradiated BCL1 cells bearing these distinct determinants induced long lasting antitumor immunity in naive syngeneic mice. Induction was dose-dependent and most effective when three doses of 30 x 10(6) intact irradiated BCL1 cells were given at intervals of 7-10 days. The induced immunity protected 96% of 28 mice inoculated with a lethal dose of 10(5)-10(6) nonirradiated BCL1 cells and 85% of 27 mice given a second challenge, whereas control mice died on day 20 after inoculation with 10(6) BCL1 cells. Adoptive transfer of splenocytes derived from immune mice did not induce leukemia in syngeneic recipients. Such splenocytes, harvested more than 365 days following immunization and administered together with fresh BCL1 cells to adoptive recipients, were able to confer protection for 90 days, even following a second challenge given 104 days after the first one. BCL1 immune splenocytes transferred into BCL1-bearing mice exerted a therapeutic effect, preventing leukemia onset for at least 180 days. Our results demonstrate the ability of tumor cells to trigger effective anti-tumor immunity. These findings could ultimately be applied to the prevention of tumor relapse in treatment of hematological and other malignancies expressing TAA, class II MHC antigen and costimulatory molecules.

Topics: Animals; Antigens, Neoplasm; B7-1 Antigen; Histocompatibility Antigens Class II; Hydroxyquinolines; Immunoglobulin Idiotypes; Immunoglobulin M; Immunotherapy, Adoptive; Lymphoma, B-Cell; Mice; Mice, Inbred BALB C; Receptors, Antigen, B-Cell; Survival Analysis; Vaccines

Linomide, a quinoline-3-carboxamide, has the ability to inhibit the growth of prostatic cancer in vivo but not in vitro (T. Ichikawa et al., Cancer Res., 52: 3022-3028, 1992). The reason for this discrepancy is that linomide inhibits tumor growth not directly but indirectly in vivo via its ability to inhibit the angiogenic response induced within the growing prostatic cancer (J. Vukanovic, et al., Cancer Res., 53: 1833-1837, 1993). Tumor associated macrophages can stimulate angiogenesis via their ability to secrete various cytokines, particularly tumor necrosis factor alpha (TNF-alpha). Treatment of rats with linomide decreases significantly (P < 0.05), by more than 50%, the number of tumor associated macrophages within both locally invasive (i.e., from 20-40 to 10 macrophages/high power field) and highly metastatic primary prostatic cancers (i.e., from 60-70 to 15-37 macrophages/high power field). Monocytes/macrophages isolated from linomide treated rats had a decreased ability to secrete TNF-alpha when challenged in vitro with the bacterial endotoxin, lipopolysaccharide [i.e., 702 +/- 76 (SEM) ng of TNF-alpha/10(5) monocytes/macrophages from control versus 401 +/- 2 ng of TNF-alpha/10(5) monocytes/macrophages from linomide treated rats]. In addition, when rats were treated with linomide and than challenged with lipopolysaccharide in vivo, the resulting elevation in serum TNF-alpha was inhibited by approximately 50% (i.e., 4.56 +/- 1.8 ng/ml of TNF-alpha in control versus 2.9-2.2 ng/ml depending upon the dose of linomide). The ability of linomide to decrease monocyte/macrophage secretion of TNF-alpha is not immediate, however, since the secretion of TNF-alpha induced by lipopolysaccharide challenge of monocytes/macrophages isolated from untreated animals is not decreased by acute (i.e., < 4 h) linomide treatment in vitro. These results demonstrate that the ability of linomide to inhibit the secretion of TNF-alpha by monocytes/macrophages requires either additional time or host factors. To test if natural killer (NK) cells might be one of the additional host factors required for the in vivo abilities of linomide, prostatic cancer bearing rats were treated with appropriate antiserum to deplete NK cells and then tested for their response to linomide treatment. These studies demonstrated that the antitumor, antimetastatic, and antimacrophage effects of linomide were unaffected by NK cell depletion.(ABSTRACT TRUNCATED AT 400 WORDS)

Topics: Animals; Hydroxyquinolines; Killer Cells, Natural; Lipopolysaccharides; Lung Neoplasms; Macrophages; Male; Monocytes; Neovascularization, Pathologic; Prostatic Neoplasms; Rats; Rats, Inbred Strains; Tumor Necrosis Factor-alpha

Linomide, a quinoline-3-carboxamide, has growth-inhibitory effects against a series of Dunning R-3327 rat prostatic cancers in vivo [Ichikawa et al.: Cancer Res 52:3022-3028, 1992]. In addition, we have demonstrated that daily linomide treatment can inhibit angiogenic responses in nontumor-bearing rats and reduce tumor blood flow in tumor-bearing rats [Vukanovic et al.: Cancer Res 53:1833, 1993]. In the present study we have demonstrated that the reduced tumor blood flow is due to linomide's ability to inhibit tumor angiogenesis, as documented by decreased number of blood vessels in prostatic carcinomas growing in rats treated daily with linomide. Due to linomide's ability to inhibit tumor angiogenesis, and since tumor angiogenesis is required not only for the growth of the primary cancer but also for its ability to metastasize, the effect of linomide on metastasis was directly tested using a quantitation metastasis assay. These in vivo experiments demonstrated that daily linomide treatment decreased by 3-fold the extent of dissemination of cancer cells to the lungs. To test if this antimetastatic response is due to direct effects of linomide on the metastatic cells themselves as well as an induced effect upon inhibition of tumor angiogenesis, additional studies were performed. These studies demonstrated that linomide is not converted in vivo to metabolite(s) which are directly cytotoxic or cytostatic to the prostatic cancer cells themselves. These studies also demonstrated that linomide does not decrease the attachment, migration, or invasive abilities of metastatic cancer cells. These results suggest that the major mechanism for the antitumor and antimetastatic effects of linomide is via its inhibition of tumor angiogenesis. Additional studies have demonstrated that in vivo linomide treatment results in the apoptotic death of thymocytes. This cytotoxic effect is not required for linomide's antitumor effect, nor is it due to elevated plasma levels of glucocorticoid.

Topics: Alkylation; Animals; Antineoplastic Agents; Cell Adhesion; Cell Death; Cell Movement; Glucocorticoids; Hydroxyquinolines; Male; Neoplasm Invasiveness; Neoplasm Metastasis; Neovascularization, Pathologic; Prostatic Neoplasms; Rats; Rats, Inbred Strains; Thymus Gland; Time Factors

Human prostatic cancer cells have a remarkably low rate of proliferation even when they have metastasized to the bone and have become androgen independent (Berges et al., Clin. Cancer Res., 1:473-480, 1995). Due to this low proliferation, patients with such androgen-independent metastatic prostatic cancer cells are rarely treated successfully with the presently available chemotherapeutic agents. Therefore, new approaches are urgently needed which are not dependent on the rate of cancer cell proliferation for their effectiveness. One such approach is to inhibit the angiogenic response within localized and metastatic cancer deposits, since the resultant hypoxia-induced tumor cell death does not require cell proliferation. We have previously demonstrated that the quinoline-3-carboxamide, linomide, is an p.o. active agent which inhibits tumor angiogenesis and thus blood flow in a variety of rat prostatic cancers independent of their growth rate, androgen sensitivity, or metastatic ability. Because of its antiangiogenic effects, linomide treatment induces the hypoxic death of rat prostatic cancer cells, thus inhibiting their net growth and metastases. To determine whether human prostatic cancer cells are similarly sensitive to hypoxia-induced death caused by linomide inhibition of tumor angiogenesis, androgen-independent TSU and PC-3 human prostatic cancer cells were xenotransplanted into SCID mice that were either untreated or treated p.o. with linomide. These studies demonstrated that linomide treatment decreases microvessel density in both androgen-independent human prostatic cancers. Microvessel density was decreased from 1.8 +/- 0.4% of the total area in control tumors to 1.0 +/- 0.2% in linomide-treated TSU tumors [i.e., a 44% decrease in microvessel density (P < 0.05)]. Similarly, a 56% decrease (P < 0.05) was observed in the microvessel density of PC-3 tumors (i.e., 2.7 +/- 0.8% of the area in control tumor versus 1.2 +/- 0.2% in the linomide-treated tumors). This inhibition of angiogenesis increased cell death in both TSU and PC-3 cancer cells. This is reflected in both an increase in the area of necrosis and an increase in the apoptotic index in non-necrotic areas. In untreated TSU tumors, 40 +/- 2% of tumor volume was necrotic. Linomide treatment increased this necrotic percentage to 59 +/- 2% [i.e., 48% increase (P < 0.05)]. Linomide therapy also increased apoptotic cell death in non-necrotic tumor areas. In the untreated TSU tumors, 2.9 +/- 0.6% o

Topics: Adenocarcinoma; Animals; Apoptosis; Cell Division; Humans; Hydroxyquinolines; In Vitro Techniques; Male; Mice; Mice, SCID; Neoplasm Transplantation; Neovascularization, Pathologic; Prostatic Neoplasms; Transplantation, Heterologous; Tumor Cells, Cultured

Roquinimex is being used for posttransplantation immunotherapy of autologous bone marrow transplantation for acute and chronic myelogenous leukemia. This immunotherapeutic agent is a cytokine inducer and may induce an autologous graft-versus-host (GVH) and graft-versus-tumor reactions.. Our purpose was to examine patients undergoing this immunotherapy for clinical signs and symptoms of acute GVH reactions and to correlate these symptoms with their clinical outcome.. We studied eight patients receiving requinimex therapy.. We found autologous GVH reactions in three of eight patients (38%) treated with this immunotherapy. Their disease was manifested by localized or widespread violaceous papules that on histologic evaluation were compatible with a grade II GVH reaction. The acute cutaneous GVH reaction was associated with eccrine sweat gland necrosis, a dermatologic toxicity usually associated with chemotherapy.. Long-term studies of larger numbers of patients treated with this immunotherapy will determine whether these GVH reactions confer significant, sustained, antitumor effects.

Topics: Acute Disease; Adjuvants, Immunologic; Bone Marrow Transplantation; Female; Graft vs Host Reaction; Humans; Hydroxyquinolines; Leukemia, Myeloid; Male; Middle Aged; Necrosis; Skin; Skin Diseases; Sweat Glands; Transplantation, Autologous

Small bowel transplantation may be complicated not only by rejection but also by graft-versus-host reaction (GVHR). So far, little is known about the association between these two immunological reactions, partly because of the lack of standardized, reproducible experimental models for such studies. In this work, a rat model in which GVHR and rejection occur simultaneously was established. When transplanting small bowel grafts from BN donors to Lewis recipients and thereafter treating the grafts locally with the immunomodulating substance LS-2616, a clinically visible GVHR occurred on day 6, at the same time the first signs of rejection became visible. The GVHR was confirmed by immunohistochemical staining for MHC class II-positive cells in liver and ear skin biopsy specimens. An obvious quantitative difference in the number of positive cells in both organs was observed when local treatment was compared with oral LS-2616 treatment or with findings in organs from untreated animals. We conclude that GVHR and rejection are not mutually exclusive and thus may occur simultaneously, and that this pharmacological model might facilitate further studies of the impact of GVHR on graft rejection and recipient survival.

Topics: Adjuvants, Immunologic; Animals; Graft Rejection; Graft vs Host Reaction; Histocompatibility Antigens Class II; Hydroxyquinolines; Immunoenzyme Techniques; Intestine, Small; Liver; Male; Rats; Rats, Inbred BN; Rats, Inbred Lew; Time Factors; Transplantation, Homologous

Topics: Adjuvants, Immunologic; Administration, Oral; Animals; Autoimmunity; Encephalomyelitis, Autoimmune, Experimental; Hydroxyquinolines; Killer Cells, Lymphokine-Activated; Mice; Rats; Recurrence

Roquinimex (Roq) is an immunomodulator known to stimulate cellular immune responses. It is currently used for immunotherapy after bone marrow transplantation (BMT). One of the major features of this compound is an enhancement of natural killer (NK) cell activity and numbers. We studied the in vitro effect of Roq on human peripheral blood NK and adherent lymphokine-activated killer cell (ALAK) activities. In cultures supplemented with recombinant interleukin 2 (rIL-2) (1000 U ml-1) and Roq a significant increase in NK and LAK function was observed without a parallel increase in cell numbers. We also examined the generation of NK cells from human bone marrow (BM) immature progenitors, obtained by purging with 4-hydroperoxycyclophosphamide (4HC). NK cell numbers and activity were both increased when cultures with rIL-2 (10 U ml-1) were supplemented with Roq. These results confirm findings obtained in vivo and in vitro in the murine system and suggest that Roq is an active agent on these lymphoid populations. These properties and good tolerability make Roq an attractive tool for immunotherapy.

Topics: Adjuvants, Immunologic; Bone Marrow; Bone Marrow Cells; Bone Marrow Purging; Cell Count; Cell Division; Cells, Cultured; Cyclophosphamide; Hematopoietic Stem Cells; Humans; Hydroxyquinolines; Killer Cells, Lymphokine-Activated; Killer Cells, Natural; Leukemia, Erythroblastic, Acute; Leukocytes, Mononuclear; Lymphocyte Activation; Phenotype; Stimulation, Chemical; Tumor Cells, Cultured

Topics: Adjuvants, Immunologic; Animals; Autoimmunity; Combined Modality Therapy; Diabetes Mellitus, Type 1; Female; Hydroxyquinolines; Islets of Langerhans Transplantation; Mice; Mice, Inbred NOD; Transplantation, Isogeneic

Topics: Adjuvants, Immunologic; Animals; Cyclosporine; Drug Synergism; Drug Therapy, Combination; Graft Survival; Heart Transplantation; Hydroxyquinolines; Immunosuppressive Agents; Mycophenolic Acid; Rats; Rats, Inbred Lew; Time Factors; Transplantation, Homologous

Topics: Adjuvants, Immunologic; Animals; Antilymphocyte Serum; Cyclosporine; Drug Synergism; Female; Graft Survival; Heart Transplantation; Hydroxyquinolines; Immune Tolerance; Immunosuppressive Agents; Male; Rats; Rats, Inbred Strains; T-Lymphocytes; Time Factors; Transplantation, Homologous

Topics: Adjuvants, Immunologic; Animals; Graft Rejection; Graft vs Host Reaction; Histocompatibility Antigens Class II; Hydroxyquinolines; Intestine, Small; Rats; Rats, Inbred BN; Rats, Inbred Lew; Transplantation, Homologous

The efficacy of combined treatment with the novel vitamin D analogue MC 1288 (20-epi-1 alpha,25-dihydroxycholecalciferol) and cyclosporine A (CyA) in preventing rejection following organ transplantation was evaluated in the heterotopic cardiac allograft model. Wistar/Kyoto rats received hearts from PVG donors and were subsequently treated with MC 1288 and CyA in various dose combinations. Administration of MC 1288 0.1 microgram/kg together with CyA 5 mg/kg produced significantly prolonged graft survival times as compared with single therapy with MC 1288 0.1 microgram/kg (p < 0.01) or CyA 5 mg/kg (p < 0.001). MC 1288 and CyA were tested also in combination with the immunomodulating drug LS-2616, which, in several laboratories, is used as a model for the evaluation of new combinations of immunosuppressive drugs. LS-2616 abolishes the immunosuppressive effect of CyA and the transplanted hearts are thus rejected at the same time as grafts of untreated recipients, i.e. on day 8. The immunosuppressive effect of MC 1288 is also counteracted by LS-2616, but the effect is not absolute and the median graft survival time is 11.0 days. Combined treatment with MC 1288 and CyA in the presence of LS-2616 resulted in a median graft survival time of 14.0 days. The results obtained in our experiments indicate at least an additive effect of MC 1288 and CyA thus suggesting that in the future MC 1288, or other vitamin D analogues, might be used to control rejection, either alone, in combination with CyA or in combination with other immunosuppressive drugs with other mechanisms of action.

Topics: Adjuvants, Immunologic; Animals; Calcitriol; Calcium; Cyclosporine; Drug Therapy, Combination; Graft Survival; Heart Transplantation; Homeostasis; Humans; Hydroxyquinolines; Immunosuppressive Agents; Rats; Rats, Inbred WKY; Transplantation, Homologous; Vitamin D

Linomide (Roquinimex) has antitumor activity when given in vivo (but not when applied in vitro) that has been attributed to immune host mechanisms. Recent studies, however, suggest that Linomide may also possess antiangiogenic properties. The aim of the present study was to evaluate the antiangiogenic effect of Linomide using an intravital microscopic technique. Syngeneic pancreatic islets were isolated and implanted into the dorsal skinfold chamber of Syrian golden hamsters. This model allows detailed repeated in vivo observations and quantitative analysis of revascularization of pancreatic islet grafts. The neovascularization process of the islets is a highly reproducible phenomenon that is completed within about 2 weeks, resulting in a microvascular network very similar to that of islets in situ. The plasma concentration profile of Linomide following a single oral dose of the compound was determined. The elimination of Linomide was fast, the half-life being 2.6 +/- 0.2 h. Due to the short half-life, the hamsters were given Linomide twice a day. One group of animals (n = 9) was force-fed Linomide (100 mg/kg per day) from the day of implantation throughout the 2-week observation period, and the results were compared with those obtained in a nontreated control group (n = 7). At days 6, 10, and 14 after implantation, the neo-vasculature of the islets was examined. In the control group, 91% +/- 4% (mean +/- SEM) of the islets showed the first signs of angiogenesis at day 6, whereas in the Linomide-treated group the corresponding value was 48% +/- 12%. At days 10 and 14, the "take-rate" in the control group increased to 94% +/- 3% for day 0 and to 94% +/- 4% (n = 6) for day 14, whereas in the treated group the corresponding take-rate was 67% +/- 11% and 72% +/- 12%, respectively. The functional capillary density in the control group at days 6, 10, and 14 was 223 +/- 17,348 +/- 29, and 495 +/- 29 cm-1, respectively, and that in the Linomide treated group was 91 +/- 28, 181 +/- 43, and 229 +/- 47 cm-1, respectively. These results demonstrate that Linomide suppresses the neovascularization of the islet grafts by both delaying the onset of and reducing the percentage of islets displaying angiogenesis as well as by decreasing the rate of proliferation of capillary endothelium of the revascularized islets.

Topics: Animals; Antineoplastic Agents; Cricetinae; Hydroxyquinolines; Islets of Langerhans; Islets of Langerhans Transplantation; Mesocricetus; Neovascularization, Pathologic; Transplantation, Heterotopic

Linomide, a synthetic immunomodulator, increases natural killer (NK) activity and markedly activates several lymphocyte populations in both experimental animals and humans. It has been shown to ameliorate the autoimmune manifestations of lupus-like disease in MRL/lpr mice and the clinical and pathological signs of acute and chronic-relapsing experimental autoimmune encephalomyelitis (EAE) in SJL/J mice. We examined the effect of linomide (100 mg/kg/day; administered in drinking water) on rabbits and rats with experimental autoimmune myasthenia gravis (EAMG). Following immunization with Torpedo acetylcholine receptor (AChR), all control rabbits developed clinical signs of severe weakness and exhibited a decrement of muscle action potential upon repetitive stimulation. In contrast, mild signs of weakness appeared in only two of five linomide-treated rabbits, with EMG borderline positive in one of them. Booster immunization with Torpedo AChR induced severe relapse and death in two EAMG control rabbits, whereas the two linomide-treated animals remained free of myasthenic symptoms. The serum level of antibodies against both Torpedo and rat AChR were markedly suppressed in the linomide-treated animals. Similar inhibition of clinical signs of EAMG was observed in the EAMG rat model. Furthermore, the in vitro proliferative response of lymph node cells to Torpedo AChR and the purified protein derivative of Mycobacterium tuberculosis was significantly lower in the linomide-treated EAMG rats than in the controls. Linomide may constitute a new immunomodulating agent for the treatment of myasthenia gravis.

Topics: Adjuvants, Immunologic; Animals; Autoantibodies; Electromyography; Female; Hydroxyquinolines; Immunization; Lymphocyte Activation; Myasthenia Gravis; Rabbits; Rats; Rats, Inbred Lew; Receptors, Cholinergic

The vitamin D analogue MC 1288 (20-epi-1 alpha,25-dihydroxycholecalciferol) was tested here for its possible immunosuppressive properties in vivo using different rat transplantation models. MC 1288, in a dose of 0.1 microgram/kg daily, administered intraperitoneally for 10 days, was found to be effective in prolonging cardiac allograft survival. Untreated recipients rejected their grafts around day 8 while MC 1288 treatment delayed rejection until day 22 (P < 0.001). Addition of the immunostimulatory drug LS-2616 (Linomide) reduced the immunosuppressive effect of MC 1288 and rejection occurred around day 11. The immunosuppressive effect of MC 1288 on rejection following small bowel transplantation was determined by measuring the amounts of hyaluronan (HA) secreted into the intestinal lumen. On day 6 post-transplantation the amounts of intraluminal HA in untreated animals was 29.2 +/- 5.3 ng/min and cm, while in MC 1288-treated animals it was just 5.0 +/- 1.6 ng/min and cm (P < 0.01). We conclude that MC 1288 has immunosuppressive effects that may make it suitable for the prevention of graft rejection.

Topics: Adjuvants, Immunologic; Animals; Calcitriol; Graft Rejection; Graft Survival; Graft vs Host Reaction; Heart Transplantation; Hyaluronic Acid; Hydroxyquinolines; Immunosuppressive Agents; Injections, Intraperitoneal; Intestinal Mucosa; Intestine, Small; Male; Rats; Rats, Inbred Lew; Rats, Inbred WKY; Transplantation, Homologous

Oral administration of the synthetic immunomodulating drug quinoline-3-carboxamide (Linomide) in the drinking water to 5-week-old female non-obese diabetic (NOD) mice resulted in complete protection from insulitis and maintenance of normal glucose tolerance for over 40 weeks (impaired glucose tolerance: treated n = 2 of 18; control n = 17 of 18, p < 0.0001). Delayed administration of the drug at 16 weeks resulted in slowing of the progression to diabetes when assessed at 42 weeks (treated with diabetes n = 7 of 25; control with diabetes 25 of 43, p < 0.0234). No gross changes of immune system cell phenotype or function were observed in the Linomide-treated group. Adoptive transfer of spleen and lymph node cells from treated female NOD mice into sub-lethally irradiated male recipients failed to transfer diabetes, whereas a similar transfer of cells obtained from untreated age-matched controls resulted in diabetes in all secondary recipients (diabetes in control group n = 12 of 13; in Linomide group n = 0 of 11, p < 0.0001). Linomide pretreatment of the secondary recipients also inhibited the transfer of diabetes (diabetes in pretreated group n = 2 of 9, control group n = 12 of 13, p < 0.015), as did adoptive co-transfer of cell mixtures obtained from treated female NOD mice, free of diabetes, and from diabetic NOD female mice (diabetes in Linomide group n = 4 of 9; in control group 7 of 7, p < 0.0337).(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adjuvants, Immunologic; Administration, Oral; Animals; Death; Diabetes Mellitus, Type 1; Dose-Response Relationship, Drug; Female; Hydroxyquinolines; Immunophenotyping; Immunotherapy, Adoptive; Islets of Langerhans; Lymph Nodes; Lymphocyte Activation; Lymphocyte Transfusion; Male; Mice; Mice, Inbred NOD; Spleen; Time Factors

Topics: Adjuvants, Immunologic; Animals; Azathioprine; Drug Synergism; Graft Survival; Guanidines; Heart Transplantation; Hydroxyquinolines; Immunosuppressive Agents; Male; Rats; Rats, Inbred Strains; Rats, Inbred WKY; Time Factors; Transplantation, Homologous

Programmed cell death (PCD) is involved in the physiological regulation of lymphocyte turnover, as well in the antigen-driven selection of T and B cells. Here it is shown that the immunomodulator linomide (quinoline-3-carboxamide) inhibits the apoptotic decay of peripheral T lymphocytes in response to three different stimuli. First, linomide reduces the superantigen-mediated apoptosis and deletion of specific T lymphocytes of both the CD4+ and the CD8+ subsets without affecting other superantigen-triggered phenomena such as T cell expansion and anergy. Second, linomide abolishes the T lymphopenia and inhibits PCD of splenic CD4+ and CD8+ T cells induced by exogenous glucocorticoids. This effect is restricted to peripheral T lymphocytes and does not concern thymocytes. Finally, linomide abolishes the development of lymphopenia that follows infection with vaccinia virus, while reducing PCD of CD4+ and CD8+ peripheral T cells. The anti-apoptotic effect of linomide could account for its immunostimulatory properties and might be relevant to the treatment of immunodeficiencies associated with an increased apoptotic decay of T lymphocytes.

Topics: Adjuvants, Immunologic; Animals; Apoptosis; Clonal Anergy; Dexamethasone; Enterotoxins; Flow Cytometry; Hydroxyquinolines; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Receptors, Antigen, T-Cell, alpha-beta; Spleen; Superantigens; T-Lymphocyte Subsets; Vaccinia

The effects of the immunomodulating substance LS-2616 (linomide) on graft-versus-host reaction (GVHR) were investigated in a semi-syngeneic small bowel transplantation model. The entire bowel of Lewis donors were transplanted heterotopically into (Lewis x BN) F1 hybrids. Both untreated animals and animals treated with LS-2616, in a daily dose of 160 mg/kg, developed a lethal GVHR. The median survival time in untreated animals was 14.5 days while in LS-2616 treated animals it was just eight days (p < 0.01). LS-2616 in combination with cyclosporin A (CyA), 15 mg/kg given orally on days 0-20, did not seem to alter the survival times compared with CyA treatment alone; 56% of the animals treated with CyA survived for more than 100 days and after combined treatment with CyA/LS-2616 there were 50% permanent survivors. Also the effect of earlier sensitization of the donor on the course of GVHR was investigated. Hearts from BN rats were transplanted heterotopically to the neck vessels of Lewis rats. The hearts were rejected on about day six; five days later the bowels were harvested and transplanted into (Lewis x BN) F1 hybrids. The median survival time in this group was 12.5 days. Taken together our results, in combination with earlier findings, suggest that, at the level of effector mechanisms, GVHR is not an exact mirror image of rejection. Also, LS-2616 appears to be a useful tool for further studies of the mechanisms of action of GVHR.

Topics: Adjuvants, Immunologic; Animals; Cyclosporine; Drug Administration Schedule; Graft vs Host Disease; Graft vs Host Reaction; Heart Transplantation; Hydroxyquinolines; Interleukin-2; Intestine, Small; Male; Molecular Structure; Rats; Rats, Inbred BN; Rats, Inbred Lew; Stimulation, Chemical; T-Lymphocytes; Transplantation, Heterotopic

Topics: Adult; Carcinoma; Child, Preschool; Dermatofibrosarcoma; Drug Eruptions; Female; Graft vs Host Disease; Granulocyte Colony-Stimulating Factor; Granulocyte-Macrophage Colony-Stimulating Factor; Granuloma; Hidradenitis; Histiocytosis, Sinus; Humans; Hydroxyquinolines; Lymphatic Diseases; Male; Middle Aged; Myxedema; Nevus; Paraproteinemias; Skin Diseases; Syphilis; Xanthomatosis

Linomide (N-phenylmethyl-1,2-dihydro-4-hydroxyl-1-methyl-2-oxoquinoline-3-carboxa mide) has a reproducible in vivo antitumor effect against a series of both androgen responsive and independent Dunning R-3327 rat prostatic cancers. This antitumor effect of linomide is host mediated. One possible mechanism involving the host is that linomide has antiangiogenic activity. An indication that linomide treatment has antiangiogenic activity is the observation that prostatic cancers from linomide treated rats have more focal necrosis than sized matched tumors from untreated rats. To directly test if linomide has antiangiogenic activity, a newly developed Matrigel based quantitative in vivo angiogenic assay was used. These experiments demonstrated that linomide has dose dependent, antiangiogenic activity in vivo in the rat. Additional studies demonstrated that due to its antiangiogenic activity, linomide treatment of rats bearing prostate cancers resulted in a more than 40% decrease in tumor blood flow. Blood flow to a variety of non-tumor bearing organs was not decreased suggesting that linomide selectively inhibits angiogenesis and does not induce loss of established blood vessels. Using as a model the response of human umbilical vein endothelial cells to linomide treatment in a variety of in vitro assays, linomide was demonstrated to have cytostatic but not cytotoxic effect on human umbilical vein endothelial cells at a medium concentration of > or = 100 micrograms/ml. In addition, both endothelial cell chemotactic migration and invasion are steps in angiogenesis inhibited by linomide treatment.

Topics: Adjuvants, Immunologic; Animals; Antineoplastic Agents; Cell Adhesion; Cell Division; Cell Movement; Cells, Cultured; Endothelium, Vascular; Humans; Hydroxyquinolines; Male; Neovascularization, Pathologic; Prostatic Neoplasms; Rats; Rats, Sprague-Dawley

Topics: Adjuvants, Immunologic; Animals; Autoimmune Diseases; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Hydroxyquinolines; Mice; Multiple Sclerosis; Rats

Linomide (LS-2616, quinoline-3-carboxamide) is a synthetic immunomodulator that stimulates natural killer cell activity and activates several lymphocytic subpopulations in experimental animals and humans. In this study we determined the effect of oral treatment with linomide on the development of experimental autoimmune encephalomyelitis, an animal model for immune-mediated human demyelinating disorders. Experimental autoimmune encephalomyelitis was induced in SJL/J mice and in an outbred strain of rats (Sabra) by subcutaneous injection of spinal cord homogenate in adjuvant followed by inoculation with Bordetella pertussis. Linomide was administered in drinking water, at an estimated dose of 50 to 100 mg/kg/day. None of the linomide-treated mice (0/41) and Sabra rats (0/15) developed any clinical or pathological signs of experimental autoimmune encephalomyelitis, whereas almost all control animals (48/53 and 18/19, respectively) were severely paralyzed and 64.5% died from the disease. Lymphocytes obtained from linomide-treated animals had reduced in vitro proliferative responses to guinea pig myelin basic protein, proteolipid protein of the myelin, and tuberculin-purified protein derivative, unlike antigen-independent proliferation which was rather unaffected. Natural killer cell activity (tested by a cytotoxic assay on radiolabeled YAC-1 target cells) was significantly enhanced in mice treated with linomide. Our results indicate that modulation of the immune system with linomide leads to complete inhibition of experimental autoimmune encephalomyelitis in the absence of systemic immunosuppression. Linomide could therefore be of use in future clinical trials for the treatment of human autoimmune demyelinating disorders.

Topics: Adjuvants, Immunologic; Animals; Autoimmune Diseases; Dose-Response Relationship, Drug; Encephalomyelitis, Autoimmune, Experimental; Female; Hydroxyquinolines; Killer Cells, Natural; Lymph Nodes; Lymphocytes; Mice; Rats; Spleen

Linomide is a synthetic immunomodulator that enhances natural killer cell activity and significantly activates several lymphocytic cell subpopulations in both experimental animals and humans. In this study we examined the effect of linomide (80 mg per kg per day in drinking water) on mice with chronic-relapsing experimental autoimmune encephalomyelitis (CR-EAE), a T-cell-mediated organ-specific autoimmune disease that resembles human multiple sclerosis. None of the mice (n = 17) that were treated with linomide from day 7 after disease induction developed any clinical or histopathological signs of CR-EAE, as compared to 19 of 20 untreated controls that were severely paralyzed and had extensive demyelinating lesions in the central nervous system. Linomide-treated animals were also resistant to an induced attack by a booster injection with a murine spinal cord homogenate. When administered to mice exhibiting severe clinical signs of paralysis, linomide inhibited both spontaneous and induced relapses. Linomide treatment protected mice from passively induced CR-EAE as well, when given from the day of injection with myelin-basic-protein-specific lymphocytes. Lymphocytes obtained from linomide-treated mice had a reduced in vitro proliferative response to the myelin basic protein and to the tuberculin purified protein derivative, whereas the mitogenic response to concanavalin A was not affected. Natural killer cell and lymphokine-activated killer cell activities were enhanced. These results suggest that linomide regulates autoimmunity in the absence of systemic immunosuppression. Since linomide is very well tolerated in experimental animals and humans, it might be used in the treatment of multiple sclerosis.

Topics: Adjuvants, Immunologic; Animals; Biomarkers; Chronic Disease; Encephalomyelitis, Autoimmune, Experimental; Female; Hydroxyquinolines; Killer Cells, Lymphokine-Activated; Killer Cells, Natural; Lymphocyte Activation; Mice; Mice, Inbred Strains; Recurrence; T-Lymphocytes

Intravenous injections of 50 micrograms Staphylococcus aureus enterotoxin B (SEB) or bacterial lipopolysaccharide (LPS) are lethal, provided that mice are simultaneously sensitized with either N-galactosamine (GalN) or the anti-glucocorticoid RU-38486. Similar to the synthetic glucocorticoid (GC) receptor agonist dexamethasone, pharmacological doses of the immunomodulator linomide (quinoline-3-carboxamide) prevent death in all four models of lethal septic shock (LPS + GalN, LPS + RU-38486, SEB + GalN, and SEB + RU-38486) and inhibit the secretion of tumor necrosis factor, one of the major intermediate effector molecules of SEB and LPS toxicity. In this system, cyclosporine A (CsA), although effective in suppressing SEB toxicity, fails to counteract the lethal effect of LPS. This observation, together with the fact that linomide acts in the presence of excess amounts of GC receptor antagonist, indicates that linomide functions in a different way to that of known immunosuppressive agents like CsA and GC.

Topics: Adjuvants, Immunologic; Animals; Apoptosis; Enterotoxins; Hydroxyquinolines; Lipopolysaccharides; Male; Mice; Mice, Inbred BALB C; Shock, Septic; Tumor Necrosis Factor-alpha

The effects of the anti picornaviral drug WIN 54954 (5-(5-(2.6-dichloro-4-(4.5-dihydro-2-oxazolyl)phenoxy)pentyl)-3-me thyl- isoxazole) and the immune modulator LS 2616 (Quinoline-3-carboxamide) on plasma cachectin/TNFa and g-interferon (IFN-g) responses were investigated during the clinical course of a myocarditic coxsackievirus B3 (CB3) infection in the mouse. Virus as well as inflammatory and necrotic lesions were found in the hearts on days 4 and 7 post inoculation (p.i.), respectively. This was demonstrated using in situ virus RNA hybridization and immune histological techniques with monoclonal antibodies against lymphocyte subsets. The CB3 infection increased TNFa levels during the first three days of disease. This response was suppressed by WIN 54954 and LS 2616. IFN-g was decreased in infected mice in the late phase of the disease (day 11). Therapy, however, was protective, and WIN 54954 even tended to increase the IFN-g response at day 5, corresponding to the time when viremia peaks. These results indicate that cytokines may serve as prognostic markers in the therapy of infectious diseases and also that WIN 54954 and LS 2616 are both possible candidates for treatment of coxsackievirus infections in man. It is suggested that a combined antiviral and immune stimulatory treatment could be of future value.

Topics: Adjuvants, Immunologic; Animals; Antiviral Agents; Coxsackievirus Infections; Enterovirus B, Human; Female; Hydroxyquinolines; Interferon-gamma; Isoxazoles; Mice; Mice, Inbred BALB C; Myocarditis; Tumor Necrosis Factor-alpha

Topics: Animals; Cyclosporine; Drug Administration Schedule; Drug Therapy, Combination; Guanidines; Heart Transplantation; Hydroxyquinolines; Immunosuppressive Agents; Male; Rats; Rats, Inbred Strains; Transplantation, Homologous

Topics: Adjuvants, Immunologic; Animals; Cyclosporine; Drug Synergism; Graft Survival; Guanidines; Heart Transplantation; Hydroxyquinolines; Immunosuppressive Agents; Male; Rats; Rats, Inbred Strains; Rats, Inbred WKY; Transplantation, Heterotopic; Transplantation, Homologous

The immunostimulator LS-2616 abolishes the effect of cyclosporin A in a rat cardiac transplantation model. The present paper compares the characteristics of rejection obtained under different immunosuppressive regimens with and without additional LS-2616 application in the same model. Cyclosporin A (CyA, 10 mg/kg daily), prednisolone (15 mg/kg daily), or 15-deoxyspergualin (2, 5, or 10 mg/kg daily), all given from the day of transplantation until day 9, protected the grafts during the treatment period. The addition of LS-2616 (160 mg/kg, day -1 until stop) resulted in a total abrogation of the immunosuppressive effect of CyA and prednisolone. However, LS-2616 could only partially or not at all reverse the effect of 15-deoxyspergualine. These results show a certain drug selectivity of LS-2616 in promoting rejection of immunosuppressed allografts. Further studies with LS-2616 may be of benefit in evaluating the mode of action of different immunosuppressive compounds and, thus, contribute to finding more effective antirejection therapies.

Topics: Adjuvants, Immunologic; Animals; Cyclosporine; Graft Survival; Guanidines; Heart Transplantation; Hydroxyquinolines; Immunosuppressive Agents; Models, Animal; Prednisolone; Rats; Rats, Inbred Strains; Rats, Inbred WKY; Time Factors

Topics: Adjuvants, Immunologic; Animals; Cyclosporins; Drug Evaluation, Preclinical; Graft Rejection; Guanidines; Heart Transplantation; Hydroxyquinolines; Immunosuppressive Agents; Prednisolone; Rats; Rats, Inbred Strains; Transplantation, Homologous

Linomide (N-phenylmethyl-1,2-dihydro-4-hydroxyl-1-methyl-2-oxo-quinoline-3- carboxamide) is a quinoline 3-carboxamide which previously has been demonstrated to produce immunomodulator and antitumor effects when given in vivo. To test the possible antitumor effects of linomide against prostatic cancers, rats bearing five distinct Dunning R-3327 rat prostatic cancer sublines were treated daily with i.p. injections of linomide. These studies demonstrated that linomide has a reproducible antitumor effect against all of the prostatic cancers tested regardless of their growth rate, degree of morphologic differentiation, metastatic ability, or androgen responsiveness. This antitumor effect is observed only in vivo, not in vitro, and involves a cytotoxic response of the prostatic cancer cells. This cytotoxic response results in the retardation of the growth rate (i.e., increased tumor volume doubling time) of primary prostatic cancers and in metastatic lesions. Linomide's growth retardation is reversible, and thus continuous daily treatment with linomide is required for maximal antitumor response. Pretreatment of rats with linomide before tumor inoculation has no effect in addition to that produced by initiating linomide treatment at the time of tumor inoculation. No enhancement of either natural killer cell number or natural killer cell cytotoxic activity is induced by linomide treatment in the tumor-bearing rats. In addition, depletion of natural killer cell activity via injections of asialo-GM1 antiserum does not prevent the antitumor effects of linomide in vivo. Likewise, the antitumor effects of linomide are also produced in prostatic cancer-bearing athymic nude rats. These results suggest that the requirement for host involvement in the antitumor effects of linomide against rat prostatic cancers may involve both immune and nonimmune host mechanism(s) (e.g., antiangiogenesis).

Topics: Adjuvants, Immunologic; Animals; Antineoplastic Agents; Cell Division; Cell Line; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Hydroxyquinolines; Killer Cells, Natural; Lung Neoplasms; Male; Neoplasm Transplantation; Prostatic Neoplasms; Rats; Rats, Inbred Strains; Spleen; Tumor Cells, Cultured

The immunomodulator LS-2616 (Linomide) induces rejection of cyclosporine-protected rat cardiac allografts. The aim of this study was to characterize this rejection in the presence of CsA and to test LS-2616 in other models of permanent graft acceptance in the rat. PVG rat hearts were transplanted heterotopically to Wistar/Kyoto (Wi/Ky) rat recipients on day 0. The recipients were treated orally on days 0-9 with CsA (10-40 mg/kg) and/or with LS-2616 (2.5-160 mg/kg) starting at different times (day -7 -+5) until the day of complete rejection. The addition of LS-2616 (day -1--stop) to CsA (10 mg/kg) resulted in a dose-dependent antagonism of the immunosuppressive effect of CsA with daily doses of 2.5-160 mg/kg. Furthermore, the results were similar, irrespective of whether LS-2616 treatment (160 mg/kg) was started on day -7, -1, +1, +3, or +5. LS-2616 (160 mg/kg) pretreatment of the recipient for 7 days before transplantation was considerably less effective. CsA (20 mg/kg) for 14 days after a PVG to DA transplantation resulted in permanent graft survival. This was not abrogated by LS-2616. Neither was rejection induced in long-term surviving grafts of RT1.C incompatible Lewis recipients. Our data suggest that LS-2616 activates already stimulated and sensitized T cells that are otherwise controlled by CsA.

Topics: Animals; Cyclosporins; Dose-Response Relationship, Drug; Drug Administration Schedule; Graft Rejection; Graft Survival; Heart Transplantation; Hydroxyquinolines; Immunohistochemistry; Kidney Transplantation; Male; Myocardium; Rats; Rats, Inbred Strains; Rats, Inbred WKY; Time Factors; Transplantation, Homologous

MRL lpr/lpr mice develop a generalized autoimmune disease associated with a massive accumulation in the peripheral lymphoid organs of abnormal, phenotypically immature T cells. Both the lymphoadenopathy and the autoimmunity are thymus-dependent and likely to arise from an aberrant pathway of intrathymic differentiation. We here present the marked beneficial effects acquired in MRL lpr/lpr mice after in vivo administration of a novel immunomodulator called linomide. The highly altered pattern of thymic subpopulations in MRL lpr/lpr mice is normalized after linomide-treatment. Concomitantly, the peripheral T cell compartment, which in MRL lpr/lpr is highly deficient in producing and responding to IL-2, gains substantial functional reactivities. We propose that linomide acts by correcting the abnormal T cell development in autoimmune MRL lpr/lpr mice. This new immunomodulator may be a useful tool for providing insight into both the pathogenesis of autoimmune disorders and intrathymic differentiation.

Topics: Adjuvants, Immunologic; Animals; Autoimmune Diseases; Cell Count; Cell Differentiation; Hydroxyquinolines; Lymphocyte Activation; Mice; Mice, Mutant Strains; Phenotype; T-Lymphocyte Subsets; T-Lymphocytes

NK cells originate from progenitors in the bone marrow and maturate independently of other lymphoid organs. NK cell maturation represents an important site for regulation of the level of NK activity and constitutes a potentially interesting target for therapeutic intervention. The effect of the immunomodulator Linomide (carboxamide-3-quinoline) on the regeneration of NK cells was studied in vivo after depletion of mature NK cells. Linomide significantly, although to various extent, accelerated the maturation of NK cells after specific depletion with antibodies to asialomonoganglioside, treatment with cyclophosphamide or lethal irradiation and syngeneic bone marrow grafting. Examination of the target cell spectrum lysed by spleen effector cells during Linomide treatment as well as studies of phenotype, clearly indicated that the effector cells studied were NK cells. Treatment of mice for 4 days with Linomide increased the frequency of bone marrow NK cell progenitors from 1/11,900 to 1/6,000 as judged by limiting dilution analysis. Direct addition of Linomide in vitro had no effect on cultures of mature NK cells from spleen, but had an additive effect to IL-2 on the generation of NK cells when added to bone marrow cultures. Our study indicates that different mechanisms exist for the regulation of progenitor and mature NK cells, and that the immunomodulator Linomide represent a potentially important tool for investigating the mechanisms governing NK cell maturation.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Monoclonal; Antigens, Differentiation; Bone Marrow; Bone Marrow Cells; Cytotoxicity, Immunologic; Hematopoiesis; Hydroxyquinolines; Immunity, Innate; Killer Cells, Natural; Mice; Mice, Inbred C57BL; Spleen

Quinoline-3-carboxamide (LS 2616) is a broadly acting immunostimulator with anti-inflammatory effects in Coxsackie virus B3-induced myocarditis in female BALB/c mice. This infection caused extensive inflammatory and necrotic lesions in the myocardium 7 days after inoculation (6.8% of tissue section area). The damaged area was reduced (to 3.7% (p less than 0.05] and the lethality decreased when LS 2616 was administered over 14 days, starting 7 days before the inoculation. The response pattern of lymphocyte subsets in situ in myocardial inflammatory lesions was elucidated by a newly developed immune histochemical staining technique. LS 2616 increased the number of class II-expressing cells 3-fold (p less than 0.01) and the CTL, Ts:Th cell ratio by 55% (p less than 0.05), whereas Lyt-1+ and TIB+ cells were unaffected. After 7 days of LS 2616 treatment, spleen lymphocyte activity tended to increase (T cells by 21% (NS) and B cells by 60% (p less than 0.05), respectively). The activity of NK cells increased by 51% (p less than 0.01). LS 2616 may thus have potential in therapy of human inflammatory disorders, such as myocarditis.

Topics: Adjuvants, Immunologic; Animals; Coxsackievirus Infections; Enterovirus B, Human; Female; Hydroxyquinolines; Lymphocytes; Mice; Mice, Inbred BALB C; Myocarditis; Premedication; Spleen; Thymus Gland

The effect of the quinoline-3-carboxamide LS-2616 (Linomide), given alone or together with cyclosporine, was studied in the first set cardiac allograft transplantation model in the rat. PVG rat hearts were transplanted heterotopically to Wistar/Kyoto rat recipients on day 0. The recipients were given LS-2616 orally on day -1 to rejection and/or CsA orally on days 0-9. In untreated animals rejection occurred on days 8-9, as judged by the absence of palpable pulsations. Treatment with CsA (5 or 10 mg/kg) resulted in prolongation of graft survival to days 17-21, i.e., the rejection occurred 8-10 days after cessation of treatment. LS-2616 in a dose of 160 mg/kg did not in itself have any impact on graft survival, but when given in doses of 40 or 160 mg/kg simultaneously with CsA (10 mg/kg), the effect of CsA was totally abolished. Animals treated with LS-2616 together with CsA had slightly lower trough blood levels than those treated with CsA alone. This interaction with CsA pharmacokinetics does not explain the results, as doubling of the CsA dose to 20 mg/kg, which well compensated for the difference in blood levels, was not sufficient to reverse the effect of LS-2616. To our knowledge this is the first compound known to abolish the effect of CsA. The mechanism is unknown, but is is possible that studies on the interaction between these two drugs will shed further light on the molecular basis of their modes of action.

Topics: Adjuvants, Immunologic; Administration, Oral; Animals; Cyclosporins; Drug Combinations; Drug Therapy, Combination; Graft Rejection; Heart Transplantation; Hydroxyquinolines; Male; Rats; Rats, Inbred WKY

Topics: Adjuvants, Immunologic; Animals; Cyclosporins; Graft Rejection; Graft Survival; Heart Transplantation; Hydroxyquinolines; Male; Rats; Rats, Inbred Strains; Rats, Inbred WKY; Transplantation, Homologous

In the current study we examine parameters affecting the metastasis of ocular tumors of in vivo derived B16F10 melanoma. In C57BL/6J beige (bg/bg) mice, with low NK activity, metastasis to the lungs was increased and survival time decreased. In C57BL/6J normal (+/+) mice treatment with PK136, a highly specific monoclonal anti-NK antibody (Ab), caused a depletion of NK cytotoxic activity, as demonstrated using a standard 51Cr release assay. In animals bearing ocular tumors, treatment with PK136 Ab resulted in significantly increased pulmonary metastasis and an altered pattern of metastasis. The effect of combined treatment protocols using LS2616 (linomide) and cyclophosphamide (Cy) was examined in enucleated and unenucleated animals. Treatment with LS2616 and Cy resulted in a significant decrease in mean pulmonary metastases (MPM), a decreased frequency of metastasis to the submandibular lymph nodes and an increase in mean survival time. In enucleated mice this combined treatment protocol resulted in apparent cures, the lowest MPM and the longest survival time observed. When tumor-bearing mice were treated with either silica, carrageenan or sublethal gamma irradiation, no effect on metastasis or survival was observed. This study demonstrates the importance of the NK cell as a primary effector cell for the control of metastasis from in vivo derived ocular B16F10 melanoma.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Monoclonal; Cyclophosphamide; Eye Enucleation; Eye Neoplasms; Female; Hydroxyquinolines; Killer Cells, Natural; Lung Neoplasms; Male; Melanoma; Mice; Mice, Inbred C57BL

An autoimmune disease in MRL lpr/lpr (MRL/l) mice is associated with a plethora of T-cell abnormalities, one of them being impaired delayed-type hypersensitivity (DTH). Three immunomodulating drugs, cyclophosphamide (Cy), cyclosporin A (CsA) and LS 2616, all with beneficial effects on autoimmune diseases, have been examined with regard to their potential effects on DTH in female MRL/l mice and in healthy control mice. All three drugs, given as a single dose at the time of sensitization, increased DTH reactivity of oxazolone in healthy control mice, while only two of them, Cy and LS 2616, significantly augmented the defective DTH response in MRL/l mice.

Topics: Adjuvants, Immunologic; Animals; Autoimmune Diseases; Cyclophosphamide; Cyclosporins; Dermatitis, Contact; Female; Hydroxyquinolines; Mice; T-Lymphocytes

The quinoline-3-carboxamide, Linomide, has been shown to possess potent immunomodulatory activity. We have evaluated the effect of Linomide in the type II collagen induced arthritis in mice. Treatment with Linomide (1.25-80 mg/kg/day) from the day of immunization strongly suppressed the arthritic response. On the other hand, initiation of treatment (20-80 mg/kg/day) at the onset of arthritis resulted in an increased severity of the arthritis. These potent and contradictory effects of Linomide, depending on treatment regime, indicates that central immuno-regulatory functions are affected and that this compound may be a useful tool for the understanding of autoimmune mechanisms.

Topics: Adjuvants, Immunologic; Animals; Antibodies; Arthritis; Arthritis, Experimental; Collagen; Hydroxyquinolines; Male; Mice; Mice, Inbred DBA; Time Factors

The effects of the immunomodulatory drug Linomide (LS-2616) have been investigated on two variants of collagen-induced arthritis (CIA) in Lewis rats, i.e. arthritis induced either with heterologous (bovine) or with homologous (rat) collagen type II (CII). Treatment with Linomide from the day of immunization (prophylactic) had a mild ameliorative effect on the severity of arthritis in the heterologous CIA, while the homologous CIA was strongly augmented. In both models, Linomide treatment caused a more severe arthritis when given from onset of clinical signs of disease and onwards (therapeutic). Serum antibody levels to CII were significantly decreased by prophylactic Linomide treatment in rats immunized with heterologous CII, while elevated levels of anti-rat CII antibodies were seen in the homologous model. No effect on antibody levels was seen with the therapeutic treatment regime. The opposing effects of prophylactic treatment with Linomide in heterologous versus homologous CIA indicate that the immune response to an autoantigen may be regulated differently from that to a foreign antigen. These results further strengthen the view that heterologous and homologous CIA should be regarded as separate experimental models, and that the studies on homologous CIA may represent a novel approach for future studies of autoimmune responses and evaluation of anti-rheumatic drugs.

Topics: Adjuvants, Immunologic; Animals; Antibody Formation; Arthritis; Autoantibodies; Autoantigens; Collagen; Female; Hydroxyquinolines; Rats

We have studied the effects of immunotherapy in coxsackievirus B3-induced myocarditis in male BALB/c mice. A single i.p. injection of the synthetic interferon inducer poly I:C conferred an almost total protection from lethality when administered at 0 h or 24 h after infection. Poly I:C treatment at 48 h after infection, as well as daily i.p. injections of the quinoline-3-carboxamide LS 2616, a new stimulator of NK-cell activity, gave no protection from lethality. The inflammatory lesions and necrosis in the ventricular myocardium 7 days after virus inoculation (3.1% of section area) were reduced in the poly I:C (24 h) treated group (1.0% of tissue section area). A less pronounced reduction was seen in the LS 2616 and poly I:C (48 h) treated groups (1.7 and 1.9% of tissue section area, respectively). The response patterns of the studied lymphocyte subpopulations were different with these two compounds, TIB+ (pre-B)-cells increased with poly I:C treatment (49%), but decreased with the LS 2616 treatment (65%). The Lyt 1+ (pan T)-cells responded similarly. Poly I:C (24 h) and LS 2616 treatment tended to increase the number of class II expressing cells (1.9- and 2.9-fold, respectively). The titer of neutralizing antibodies to coxsackievirus B3 was significantly increased in the LS 2616 treated group (1:80) but not significantly so in the poly I:C treated groups (1:40) as compared to the infected and non-infected control groups (1:20 and less than 1:5, respectively).

Topics: Adjuvants, Immunologic; Animals; Antibodies; Coxsackievirus Infections; Enterovirus B, Human; Hydroxyquinolines; Immunohistochemistry; Male; Mice; Mice, Inbred BALB C; Myocarditis; Poly I-C

In experiments using cultured cells, LS2616 has been shown to decrease growth of primary tumors and pulmonary metastasis of murine melanoma. In the current study, we examine the efficacy of LS2616 for the prophylactic and therapeutic treatment of metastases from ocular and flank inoculations of the highly aggressive in vivo derived B16F10 melanoma in C57BL/6J mice. Experimental animals were treated with 160 mg/kg/day of this drug in drinking water, until they became moribund or died. When mice were pretreated for 7 days and inoculated subcutaneously (sc) or intracamerally (ic) with 10(5) in vivo derived B16F10 tumor cells, the mean number of pulmonary metastases was significantly reduced, and the incidence of pulmonary metastases decreased. In ocular experiments, when pretreatment with drug was combined with enucleation at day 7, the mean number of lung nodules was significantly reduced, the incidence of metastasis to the lung and lymph nodes decreased and survival increased. An apparent cure rate of 31% was observed. Treatment beginning on the day of enucleation (day 7) resulted in a reduction of pulmonary metastases, a decrease in metastasis to the lungs and lymph nodes and no change in survival. LS2616 did not alter tumorigenicity of either sc or ic inoculations. In an in vivo neutralization assay, spleen cells of mice treated for 7 days with LS2616 demonstrated an increase in cytostatic or cytotoxic activity when incubated with B16F10 melanoma cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Eye Enucleation; Eye Neoplasms; Female; Hydroxyquinolines; Lung Neoplasms; Male; Melanoma; Mice; Skin Neoplasms; Spleen; Tumor Cells, Cultured

LS 2616 (Linomide) is a new immunomodulator that enhances natural killer (NK)-cell activity, delayed-type hypersensitivity reaction, mitogen responsiveness of T cells, and antibody production. It suppresses tumor growth and reduces metastases in animal experiments. In a phase I clinical trial, the maximal tolerated dose will not necessarily be the maximally effective dose, so both effect and toxicity parameters have to be monitored. Because of the pleiotropic function of the drug, several biological responses have to be analyzed. Furthermore, different malignancies are associated with different immunologic disturbances both qualitatively and quantitatively, necessitating the use of normal controls and baseline assessments as well as a range of different malignancies. The pharmacokinetic features will differ from the kinetics of the biological responses, and thus both drug concentration and effect parameters will be followed over time. Repeated doses will give information needed for tailoring of optimal schedules for administration.

Topics: Adjuvants, Immunologic; Dose-Response Relationship, Drug; Drug Administration Schedule; Drug Evaluation; Humans; Hydroxyquinolines; Neoplasms

The autoimmune MRL/Mp-lpr/lpr (MRL/l) mouse spontaneously develops sialadenitis with a morphological and phenotypical pattern similar to that seen in human Sjögren's syndrome (SS). This makes the MRL/l mouse a suitable model for therapeutical studies of autoimmune sialadenitis. We have, by histological and immunohistochemical techniques, analyzed the therapeutical effect of treatment with LS2616, a recently synthesized oxokinolinamide derivative, on sialadenitis in submandibular glands of MRL/l mice. The results were compared with effects obtained after treatment with cyclophosphamide (CY) and physiologic saline. Administration of both LS2616 and CY to MRL/l mice has previously been found to result in prolongation of survival and amelioration of organ pathology. However, only CY treatment reduced sialadenitis, while LS2616 increased the semiquantitatively assessed focal inflammation of salivary glands in 6 months old mice. No differences in T-cell phenotypes of infiltrating lymphoid cells in salivary glands between different treatment regims could be noted. However, the frequency of B-cells in the sialadenitis was decreased in the CY treated group. In contrast, CY but not LS2616 treatment normalized expression of T-helper and cytotoxic T-cell phenotypes as well as reduced the B-cell portion in lymph nodes. It is concluded that CY treatment can suppress sialadenitis although both LS2616 and CY are effective in prolongation lifespan of MRL/l mice. This may implicate different immunopathogenic mechanisms for development of sialadenitis versus other organ lesions in the autoimmune disease of MRL/l mice.

Topics: Adjuvants, Immunologic; Animals; Autoimmune Diseases; Cyclophosphamide; Hydroxyquinolines; Male; Mice; Salivary Gland Diseases; Sialadenitis; Submandibular Gland Diseases

Spleen cells from mice treated with LS2616 display a highly increased response to the polyclonal T cell lectin ConA. The total number of splenic T cells, and the relative ratios between L3T4+ and Lyt-2+ T cells were not altered by LS2616 treatment. By dissecting the overall ConA response it was found that the number of ConA-inducible, IL-2 reactive T cells was unaffected, while ConA-induced IL-2 production was enhanced after LS2616 treatment. Spleen cells from LS2616 treated mice, depleted of G10 adherent macrophages (M phi) and reconstituted with M phi from untreated mice displayed normal levels of ConA responses. M phi depleted spleen cells from untreated animals, cocultured with M phi enriched populations from LS2616 treated animals resulted in an increased ConA response. Furthermore, spleen cells from treated mice were found to be excellent stimulators for alloantigen-induced T cell responses; when used as responders in MLC, however, these cells were comparable to responders from non-treated animals. Taken together the results demonstrate that LS2616 exerts an immunostimulatory effect on M phi, which indirectly facilitates polyclonal and antigen-specific T cell responses. The possible implications of this observation on various immunoregulatory events are discussed.

Topics: Animals; Concanavalin A; Hydroxyquinolines; Interleukin-2; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Macrophage Activation; Macrophages; Mice; Spleen; T-Lymphocytes

Treatment of Sprague-Dawley rats with the compound LS 2616, a quinoline-3-carboxamide, enhanced the delayed-type hypersensitivity response (DTH) to recall antigens as judged by the specific accumulation of inflammatory cells after challenge with Bordetella pertussis in the pleural space. LS 2616 was effective when given both before sensitization and at the time of secondary antigen challenge. The effect of LS 2616 on DTH was dose-dependent. LS 2616 was highly effective in enhancing DTH in rats with suppressed cell-mediated immunity after treatment with anti-thymocyte globulin or prednisolone, indicating its possible value in the immunosuppressed state.

Topics: Adjuvants, Immunologic; Animals; Antigens, Bacterial; Antilymphocyte Serum; Bordetella pertussis; Female; Hydroxyquinolines; Hypersensitivity, Delayed; Immunization; Interleukin-1; Interleukin-2; Rats; Rats, Inbred Strains; T-Lymphocytes

The carboxamide-quinoline LS 2616 is a novel immunomodulator augmenting natural killer (NK) cell activity and T-lymphocyte related effector functions. To investigate the possible usefulness of LS 2616 in immunotherapy of tumors, the effect of the substance on growth and metastasis of the B16-F10 melanoma in syngeneic C57BL/6 mice was investigated. Treatment with LS 2616 from the time of s.c. inoculation of B16-F10 cells significantly reduced tumor take. Continuous treatment of mice with LS 2616 initiated 4 days prior to i.v. injection of tumor cells reduced the number of pulmonary metastases by 85%. When treatment with LS 2616 was started 4 days after i.v. injection of tumor cells, a time when established tumor foci were readily detectable in the lungs, a significant reduction in the number of pulmonary metastases resulted. LS 2616 significantly reduced the number of spontaneous pulmonary metastases developing from a B16-F10 tumor growing in the footpad. When treatment with LS 2616 was initiated after the establishment of grossly visible spontaneous pulmonary metastases, no significant effect on the number of metastases was found after 2 weeks of treatment. However, combined treatment with a dose of cyclophosphamide which in itself was ineffective resulted in a statistically significant 70% reduction in the number of remaining pulmonary metastases. Injection of antibodies to asialomonoganglioside which strongly reduce NK cell activity in various organs was used as a probe for the involvement of NK cells in the effects of LS 2616 on the B16-F10 tumor. The therapeutic efficiency of LS 2616 on tumor take when given from the time of s.c. inoculation, on the number of i.v. induced pulmonary metastases when treatment was started before tumor cell injection, as well as the spontaneous development of pulmonary metastases during exposure to the substance was abrogated by simultaneous injection with antibodies to asialomonoganglioside. In contrast, the beneficial effects of LS 2616 on already established i.v. produced or spontaneous pulmonary metastases were unaltered in mice made NK cell deficient by injection of anti-asialomonoganglioside antibodies. In conclusion, LS 2616 has potent antitumor activities mediated by NK cells as well as non-NK cell related defense mechanisms.

Topics: Adjuvants, Immunologic; Animals; Antineoplastic Combined Chemotherapy Protocols; Cyclophosphamide; Hydroxyquinolines; Interleukin-2; Killer Cells, Natural; Lung Neoplasms; Melanoma; Mice; Mice, Inbred C57BL

Autoimmune MRL/1 mice were treated with a recently developed substance with immunomodulating properties, LS-2616. Treatment was initiated at the age of 8 weeks, before the onset of clinically apparent disease, and at 16 weeks of age, after development of established lupus disease. Beneficial therapeutic effects were obtained, even when LS-2616 was administered at the lowest dose tested (1 mg/mouse/week) to 16-week-old mice. The effects of LS-2616 on longevity, as well as on development of lymphadenopathy, splenomegaly, glomerulonephritis, and vasculitis, were pronounced and were comparable with those of cyclophosphamide. The results obtained suggest a potential role for LS-2616 in the treatment of autoimmune disease in humans.

Topics: Adjuvants, Immunologic; Animals; Autoimmune Diseases; Cyclophosphamide; Drug Evaluation, Preclinical; Female; Glomerulonephritis; Hydroxyquinolines; Immunity; Lymphatic Diseases; Mice; Mice, Inbred Strains; Splenomegaly; Vasculitis

Autoimmune (NZB X NZW) F1 female hybrid mice were treated with LS-2616, a recently developed substance with immunomodulating properties. Treatment was initiated at the age of 4 months (i.e. at the early stage of the disease) as well as at 7 months of age (i.e. after the development of established lupus-like disease). Control groups treated with cyclophosphamide and physiological saline were also studied. Beneficial therapeutic effects were obtained regardless of when the treatment was initiated and the dose of LS-2616 administered (1 and 8 mg/mouse/week). The effects of LS-2616 on longevity, splenomegaly and glomerulonephritis were pronounced and sometimes comparable to those of cyclophosphamide (1.8 mg/mouse/week). The results obtained suggest that LS-2616 may be useful in the treatment of autoimmune disease in man.

Topics: Animals; Autoimmune Diseases; Complement C3; Female; Hydroxyquinolines; Immunoglobulins; Lupus Erythematosus, Systemic; Lupus Nephritis; Mice

The quinoline-3-carboxamide LS 2616 administered to mice in drinking water increased spontaneous cytotoxicity against YAC-1 cells in a dose-dependent manner. The enhancement of spontaneous cytotoxicity was found to be mediated by NK cells, as judged by their lack of adherence to nylon wool columns, relative resistance to treatment with antibodies to Thy-1.2 and complement, and almost total abrogation after depletion of asialo-GM1+ cells. Enhancement of NK activity was evident after 2 days of treatment, was maximal after 4 days, and remained elevated during the 14-day exposure period studied. NK activity returned to control levels 4 days after cessation of treatment. NK activity was significantly increased in spleen, peripheral blood, lymph nodes, and bone marrow of LS 2616-treated mice, while activity in peritoneal exudate cells and thymus remained low. LS 2616 was able to elevate NK activity in several mouse strains studied, including mice homozygous for the beige gene. Serum interferon levels were not increased during treatment with LS 2616. Combined injection of the interferon inducer Poly I:C and LS 2616 did not increase NK activity above that of animals injected with Poly I:C alone. However, Poly I:C, in contrast to LS 2616, increased NK activity in peritoneal exudate cells. Studies at the single cell level revealed that LS 2616 increased NK activity by increasing the number of lytically active cells via recruitment of new target-binding cells and not by increasing the lytic activity of pre-existing binders.

Topics: Adjuvants, Immunologic; Animals; Binding Sites; Cell Line; Cytotoxicity, Immunologic; Hydroxyquinolines; Interferons; Killer Cells, Natural; Mice; Mice, Inbred A; Mice, Inbred AKR; Mice, Inbred BALB C; Mice, Inbred C3H; Mice, Inbred C57BL; Mice, Inbred CBA; Species Specificity; Tissue Distribution