dinitrobenzenes and Inflammation

The author has devised a novel approach to the immunotherapy of cancer based on modification of autologous tumor cells with the hapten, dinitrophenyl (DNP). This technology is being developed by AVAX Technologies (MO, USA) as a treatment for melanoma under the brand name, M-Vax. The treatment program consists of multiple intradermal injections of DNP-modified autologous tumor cells mixed with bacille Calmette-Guerin as an immunological adjuvant. Administration of DNP vaccine to patients with metastatic melanoma induces a unique reaction--the development of inflammation in metastatic masses. Following DNP-vaccine treatment, almost all patients develop delayed-type hypersensitivity (DTH) to autologous, DNP-modified melanoma cells and about half also exhibit DTH to autologous, unmodified tumor cells. The toxicity of the vaccine is mild, consisting mainly of papules or pustules at the injection sites. Clinical trials have been conducted in two populations of melanoma patients: Stage IV with measurable metastases, and clinical Stage III patients rendered tumor-free by lymphadenectomy. There were 11 antitumor responses in 83 patients with measurable metastases: two complete, four partial and five mixed. In 214 Stage III patients the 5-year overall survival rate was 44%, which compares favorably with the reported surgical rate of 20-25%. In both populations, the induction of DTH to unmodified autologous tumor cells was associated with significantly longer survival. This is a platform technology that is adaptable to other human cancers and early trials indicate immunological activity in ovarian and renal cell carcinomas.

Topics: Adjuvants, Immunologic; Antigens, Neoplasm; Cancer Vaccines; Clinical Trials as Topic; Dinitrobenzenes; Haptens; Humans; Hypersensitivity, Delayed; Inflammation; Lung Neoplasms; Melanoma; Neoplasm Staging; T-Lymphocytes

We have devised a novel approach to active immunotherapy based on modification of autologous cancer cells with the hapten, dinitrophenyl (DNP). The treatment program consists of multiple intradermal injections of DNP-modified autologous tumor cells mixed with BCG. Administration of DNP-vaccine to patients with metastatic melanoma induces a unique reaction - the development of inflammation in metastatic masses. Histologically, this consists of infiltration of T lymphocytes, most of which are CD8+. These T cells usually produce gamma interferon in situ. Moreover, they represent expansion of T cell clones with novel T cell receptor structures. Occasionally, administration of DNP-vaccine results in partial or complete regression of measurable metastases. The most common site of regression has been small lung metastases. Administration of DNP-vaccine to patients in the post-surgical adjuvant setting produces a more striking clinical effect. We have treated 214 patients with clinically evident stage III melanoma who had undergone lymphadenectomy. With a median follow-up time of 4.4 years (1.8-10.4 years) the 5-year overall survival (OS) rate is 47% (one nodal site = 51%, two nodal sites = 33%). These results appear to be comparable to those obtained with high dose interferon. More recent studies suggest that this therapeutic approach is also applicable to ovarian cancer. There appear to be no insurmountable impediments to applying this approach to much larger numbers of patients or to developing it as a standard cancer treatment.

Topics: Antigens, Neoplasm; Autoantigens; Cancer Vaccines; Combined Modality Therapy; Dinitrobenzenes; Female; Haptens; Humans; Inflammation; Lymphocytes, Tumor-Infiltrating; Neoplasm Metastasis; Neoplasms; Ovarian Neoplasms

We have devised a novel approach to active immunotherapy based on modification of autologous cancer cells with the hapten, dinitrophenyl (DNP). The treatment program consists of multiple intradermal injections of DNP-modified autologous tumor cells mixed with BCG. Administration of DNP-vaccine to patients with metastatic melanoma induces a unique reaction- the development of inflammation in metastatic masses. Histologically, this consists of infiltration of T lymphocytes, most of which are CD8+. These T cells usually produce interferon-gamma in situ. Moreover, they represent expansion of T-cell clones with novel T-cell receptor (TCR) structures. Occasionally, administration of DNP-vaccine results in regression of measurable metastases. The most common site of regression has been small lung metastases. Administration of DNP-vaccine to patients in the postsurgical adjuvant setting produces a more striking clinical effect. Of 62 patients with clinically evident stage III melanoma who had undergone lymphadenectomy, the 5-year relapse-free survival rate was 45% and the overall survival rate was 58%. These results appear to be better than those obtained with high-dose interferon, although a randomized phase III trial is required to prove that point. A recent phase I study suggests that this therapeutic approach is also applicable to stage III ovarian cancer. There appear to be no insurmountable impediments to applying this approach to much larger numbers of patients or to developing it as a standard cancer treatment.

Topics: Animals; Cancer Vaccines; Clinical Trials as Topic; Dinitrobenzenes; Female; Haptens; Humans; Hypersensitivity, Delayed; Immunotherapy, Active; Inflammation; Lung Neoplasms; Male; Melanoma; Mycobacterium bovis; Neoplasm Metastasis; Ovarian Neoplasms; T-Lymphocytes; Tumor Cells, Cultured

Phyllostachys nigra (PN) is an herbal medicine that originates from the inner bark of Phyllostachys nigra Munro var. henosis Stapf or Phyllostachys bambusoides Siebold et Zuccarini. It has long been used to relieve fever and to treat diarrhea and inflammation. PN has been shown to possess inhibitory effects on pneumonia, intestinal inflammation, tumors, and fatigue. However, its potential efficacy in the treatment of atopic dermatitis (AD) has not been extensively studied or reported.. The objective of this research was to investigate the impact of PN on HaCaT and HMC-1 cells, as well as its potential in an experimental model of AD induced by 1-chloro-2,4-dinitrobenzene (DNCB).. We analyzed the anti-inflammatory efficacy of PN in HaCaT cells and HMC-1 cells using ELISA and PCR, and investigated invasion of inflammatory cell, change of dermis and epidermis, and the SCORAD index in AD-like mice model. We also measured the MAPK signaling pathway using the dorsal tissue of mice.. Our results show that PN reduced the expressions of TARC, GM-CSF, TNF-α, MCP-1, and IL-6 in vitro. PN also decreased the SCORAD index, thickening of epidermis and dermis, and inhibited the invasions of mast cells and eosinophils as well as CD4. These results demonstrate that PN could be an effective alternative medicine for allergic inflammatory disease.

Topics: Animals; CD8-Positive T-Lymphocytes; Chemokines; Cytokines; Dermatitis, Atopic; Dinitrobenzenes; Dinitrochlorobenzene; Inflammation; Interleukin-6; Mice; Mice, Inbred BALB C; NF-kappa B; Skin

Active immunotherapy of cancer with therapeutic vaccines has been the subject of experimental and clinical studies for at least 50 years. Our approach has employed 1) autologous, human cancer cells because of extensive evidence that tumor rejection antigens may differ between multiple tumors of the same histology; 2) the immunopotentiating drug, cyclophosphamide; and 3) haptens, particularly dinitrophenyl. Multiple clinical trials in 455 patients with melanoma and ovarian cancer have shown that administration of haptenized vaccines at the proper dosage-schedule regularly induces T cell-mediated immunity to autologous tumor cells as measured by delayed-type hypersensitivity. Moreover, the vaccine causes changes in the tumor site suggestive of an immune reaction, including inflammation and infiltration with CD8+ T lymphocytes that are activated and produce cytokines. The T cell response is oligoclonal, and dominant Vβ families differ between patients. Studies of measurable metastases show clinically important tumor regression. Commercial development of this technology is clearly feasible.

Topics: Antigens, Neoplasm; Cancer Vaccines; Dinitrobenzenes; Female; Humans; Inflammation; Melanoma

Atopic dermatitis (AD) is a chronic, relapsing skin disease accompanied by itchy and dry skin. AD is caused by complex interactions between innate and adaptive immune response. AD treatment include glucocorticoids and immunosuppressants. However, long-term treatment can have serious side effects. Thus, an effective AD treatment with fewer side effects is required. Natural materials, including herbal medicines, have potential applications.. This study evaluated the in vivo and in vitro therapeutic effects of BS012, a mixture of Asarum sieboldii, Platycodon grandiflorum, and Cinnamomum cassia extracts, on AD and investigated the underlying metabolic mechanisms.. The anti-inflammatory effects of BS012 were assessed using a mouse model of AD induced by 1‑chloro-2,4-dinitrobenzene (DNCB) and in tumor necrosis factor-alpha/interferon-gamma (TNF-α/IFN-γ) stimulated normal human epidermal keratinocytes (NHEKs). In DNCB-induced mice, total dermatitis score, histopathological analysis, and immune cell factors were assessed to evaluate the anti-atopic activity. In TNF-α/IFN-γ-stimulated NHEKs, pro-inflammatory cytokines, chemokines, and related signaling pathways were investigated. Serum and intracellular metabolomics were performed to identify the metabolic mechanism underlying the therapeutic effects of BS012 treatment.. In DNCB-induced mice, BS012 showed potent anti-atopic activity, including reducing AD-like skin lesions and inhibiting the expression of Th2 cytokines and thymic stromal lymphopoietin. In TNF-α/IFN-γ-stimulated keratinocytes, BS012 dose-dependently inhibited the expression of pro-inflammatory cytokines and chemokines by blocking nuclear factor-kappa B and signal transducer and activator of transcription signaling pathways. Serum metabolic profiles of mice revealed significant changes in lipid metabolism related to inflammation in AD. Intracellular metabolome analysis revealed that BS012 treatment affected the metabolism associated with inflammation, skin barrier function, and lipid organization of the stratum corneum.. BS012 exerts anti-atopic activity by reducing the Th2-specific inflammatory response and improving skin barrier function in AD in vivo and in vitro. These effects are mainly related to the inhibition of inflammation and recovery of metabolic imbalance in lipid organization. BS012, a novel combination with strong activity in suppressing the Th2-immune response, could be a potential alternative for AD treatment. Furthermore, the metabolic mechanism in vivo and in vitro using a metabolomics approach will provide crucial information for the development of natural products for AD treatment.

Topics: Animals; Anti-Inflammatory Agents; Asarum; Chemokines; Cinnamomum aromaticum; Cytokines; Dermatitis, Atopic; Dinitrobenzenes; Dinitrochlorobenzene; Humans; Inflammation; Interferon-gamma; Lipids; Mice; Mice, Inbred BALB C; Platycodon; Skin; Tumor Necrosis Factor-alpha

Gut microbiota are involved in the onset and development of chronic intestinal inflammation. The recently described endocannabinoidome (eCBome), a diverse and complex system of bioactive lipid mediators, has been reported to play a role in various physio-pathological processes such as inflammation, immune responses and energy metabolism. The eCBome and the gut microbiome (miBIome) are closely linked and form the eCBome - miBIome axis, which may be of special relevance to colitis.. Colitis was induced in conventionally raised (CR), antibiotic-treated (ABX) and germ-free (GF) mice with dinitrobenzene sulfonic acid (DNBS). Inflammation was assessed by Disease Activity Index (DAI) score, body weight change, colon weight-length ratio, myeloperoxidase (MPO) activity and cytokine gene expression. Colonic eCBome lipid mediator concentrations were measured by HPLC-MS /MS.. GF mice showed increased levels of anti-inflammatory eCBome lipids (LEA, OEA, DHEA and 13- HODE-EA) in the healthy state and higher MPO activity. DNBS elicited reduced inflammation in GF mice, having lower colon weight/length ratios and lower expression levels of Il1b, Il6, Tnfa and neutrophil markers compared to one or both of the other DNBS-treated groups. Il10 expression was also lower and the levels of several N-acyl ethanolamines and 13-HODE-EA levels were higher in DNBS-treated GF mice than in CR and ABX mice. The levels of these eCBome lipids negatively correlated with measures of colitis and inflammation.. These results suggest that the depletion of the gut microbiota and subsequent differential development of the gut immune system in GF mice is followed by a compensatory effect on eCBome lipid mediators, which may explain, in part, the observed lower susceptibility of GF mice to develop DNBS-induced colitis.

Topics: Animals; Colitis; Dinitrobenzenes; Inflammation; Lipids; Mice

Huang-Lian-Jie-Du Decoction is a traditional Chinese medicine formula which has long been used to treat inflammatory skin disease including AD. However, Gardeniae Fructus, a component herb of HLJDD, has noticeable toxicity in liver and kidney. We therefore replaced Gardeniae Fructus with Dictamni Cortex with a hope to derive at a modified HLJDD (MHLJDD) with better safety profile.. The present study aimed to develop MHLJDD and identify its active fraction as innovative therapeutic agents for AD using 2,4-dinitrobenzene (DNCB) and calcipotriol (MC903)-sensitized mouse models of AD.. MHLJDD and the combination of the 1-butanol-soluble-fraction and the water-soluble-fraction (MHLJDD-F) were given intragastrically to the DNCB-induced mice and MC903-induced mice for two weeks. The body weight, dorsal skin/ear thickness and severity of AD symptoms of the mice were measured throughout the study. Scratching behaviors were observed after drug treatment. The blood and dorsal skin/ear tissues of mice were harvested for histopathological examination and biochemical analyses.. The results revealed that DNCB- and MC903-induced AD symptoms, including skin thickening, dryness, erythema and excoriations, in the dorsal skin and ears were significantly alleviated in the MHLJDD and MHLJDD-F-treated mice. Ceramides content and protein expressions of filaggrin and loricrin were also up-regulated after treatment with MHLJDD and MHLJDD-F. In addition, skin inflammation induced by DNCB and MC903 were markedly suppressed in the MHLJDD and MHLJDD-F-treated mice, and the action mechanisms involve suppression of the release of inflammatory cytokines, as well as downregulation of the activation of NF-κB and MAPKs pathways. Besides, MHLJDD and MHLJDD-F could reverse the abundance of gut microbiota induced by DNCB in mice.. MHLJDD and MHLJDD-F could markedly relieve AD-like symptoms induced by DNCB and MC903 in mice through, at least in part, improving the epidermal barrier function and inhibiting skin inflammation via suppressing the activation of NF-κB and MAPKs pathways and regulation of the gut microflora dysbiosis. This study reported for the first time that MHLJDD and its active fraction could be used as innovative therapeutic agents for AD.

Topics: Animals; Coptis chinensis; Cytokines; Dermatitis, Atopic; Dinitrobenzenes; Dinitrochlorobenzene; Disease Models, Animal; Inflammation; Mice; Mice, Inbred BALB C; NF-kappa B; Skin

Huang-Lian-Jie-Du Decoction (HLJDD), is a well-known traditional Chinese herbal formula first written in the Tang dynasty. In Chinese medicine practice, HLJDD is commonly prescribed to treat various inflammatory skin diseases, such as atopic dermatitis (AD) and psoriasis.. The present study aimed at investigating the therapeutic effect of HLJDD extract (HLJDE) and to elucidate the underlying molecular mechanisms of action in the 1-chloro-2,4-dinitrobenzene (DNCB)-induced AD-like mice.. Female Balb/c mice were sensitized with DNCB for three days. After sensitization, mice were challenged with DNCB every three days and orally administrated with HLJDE (150, 300 and 600 mg/kg) daily from day 14 to day 29 for consecutive 16 days. At the end of experiment, the clinical AD scores of the mice were calculated to evaluate the therapeutic effect of HLJDE, and serum, ears and dorsal skin of the mice were collected for unravelling molecular mechanisms.. HLJDE significantly reduced the clinical symptoms in the AD-like mice by inhibiting eosinophil and mast cell infiltration, suppressing the production of Th2-associated cytokine (IL-4) and pro-inflammatory cytokines (TNF-α). In addition, HLJDE significantly suppressed the NF-κB and MAPKs pathways. Moreover, HLJDE was able to accentuate filaggrin expression in the skin lesion when compared to the sensitized mouse without treatment.. HLJDE significantly improved the AD-like symptoms on the DNCB-sensitized mice through mitigating the production of inflammatory mediators via suppressing MAPKs and NF-κB pathways. Additionally, the elevated expression of filaggrin in the skin lesion by HLJDE contributes to the recovery of dysfunctional skin barrier on the DNCB-sensitized mice.

Topics: Animals; Anti-Inflammatory Agents; Cytokines; Dermatitis, Atopic; Dinitrobenzenes; Drugs, Chinese Herbal; Eosinophils; Female; Inflammation; Mast Cells; Mice; Mice, Inbred BALB C; Mitogen-Activated Protein Kinases; NF-kappa B; Plant Extracts; Signal Transduction; Skin; Skin Diseases; Tumor Necrosis Factor-alpha

Angiotensin II, the main effector of renin angiotensin system, plays an important role in the inflammatory process and most of its effects are mediated through the AT1 receptor activation. However, the knowledge about the AT2 receptor involvement in this process is still evolving. We previously found that in an experimental model of colitis, AT2 receptor activation can contribute to the impairment of the muscle contractility in vitro in the course of inflammation. Here, we investigated the potential alleviating effects of the in vivo treatment of PD123319 (1-[[4-(Dimethylamino)-3-methylphenyl]methyl]-5-(diphenylacetyl)-4,5,6,7- tetrahydro-1H-imidazo[4,5-c]pyridine-6-carboxylic acid ditrifluoroacetate), AT2 receptor antagonist, in 2,4-dinitrobenzene sulfonic acid (DNBS)-induced rat model of colitis. The effects of i.p PD123319 (0.3, 3 and 10 mg/kg) administration to rats subjected to intra-rectal DNBS instillation were investigated. The study revealed that the colon injury and the inflammatory signs were ameliorated by PD123319 when visualized by the histopathological examination. The colon shortening, myeloperoxidase activity, and colonic expression of IL-1β, IL-6 and iNOS were downregulated in a dose-dependent manner in DNBS-induced colitis rats treated with PD123319 and the anti-oxidant defense machinery was also improved. The mechanism of these beneficial effects was found in the ability of PD123319 to inhibit NF-κB activation induced by DNBS. The colonic contractility in inflamed tissues was also improved by PD123319 treatment. In conclusion, our data have demonstrated previously that undescribed proinflammatory effects for the AT2 receptors in DNBS-induced colitis in rats in which they are mediated likely by NF-κB activation and reactive oxygen species generation. Moreover, when the inflammatory process is mitigated by the AT2 receptor antagonist treatment, the smooth muscle is able to recover its functionality.

Topics: Angiotensin II; Angiotensin II Type 2 Receptor Blockers; Animals; Colitis; Colon; Dinitrobenzenes; Imidazoles; Inflammation; Male; Oxidative Stress; Pyridines; Rats; Rats, Wistar; Receptor, Angiotensin, Type 2

In search of safer treatments for inflammatory bowel disease in subjects not responding to, or showing adverse effects to TNF-α antagonists, we tested three novel indoline carbamates in the 2,4-dinitrobenzene sulfonic acid (DNBS) model of colitis in rats. The compounds have anti-inflammatory activity in other disease models in mice.. AN827 (3-(2-(methoxy carbonyl) ethyl) indolin-4-ylethyl methyl) carbamate (0.1 or 1mg/kg), AN680 (3-(2-(methoxy carbonyl) ethyl) indolin-6-ylethyl methyl) carbamate (1.25 or 2.5mg/kg) and AN917 (3-(3-amino propyl) indolin-4-ylethyl methyl) carbamate (1 or 2mg/kg), 5-aminosalycilic acid (5-ASA) (1 or 100mg/kg) or saline (1ml/kg) were administered rectally 1h after intracolonic administration of DNBS, (35mg/kg in 30% alcohol). Disease severity was assessed four days after DNBS administration by change in body weight, colon weight, area of ulceration, myeloid peroxidase (MPO) activity, colonic TNF-α, IL-6 and IL-1β levels. Histopathological scoring was performed after staining colon sections with hematoxylin and eosin and with antibodies to CD68 and CD11b.. AN827 (0.1 and 1mg/kg), AN680 (2.5mg/kg) and AN917 (2.0mg/kg) significantly reduced all macroscopic and microscopic parameters of colitis, colonic pro-inflammatory cytokines, TNF-α, IL-1β and IL-6 and MPO activity by about 80%.. The indoline derivatives largely prevented the symptoms of colitis and were 500-50 times more potent and more effective than 5-ASA. It may be worth evaluating them in models of established colitis. Since AN827 is strongly bound by plasma proteins no adverse effects are expected if compound is absorbed into the circulation after rectal administration.

Topics: Administration, Rectal; Animals; Anti-Inflammatory Agents; Colitis; Dinitrobenzenes; Dose-Response Relationship, Drug; Indoles; Inflammation; Male; Mice; Rats; Rats, Wistar; Ulcer

Occupational exposure to chemicals is an important cause of asthma. Recent studies indicate that IgE antibodies enhance sensitization to chemicals in the skin.. We investigated whether IgE might similarly promote the development of airway inflammation following inhalation of a contact sensitizer.. A model of chemical-induced asthma is described in which introduction of the low-molecular-weight compound, trinitrobenzene sulphonic acid (TNBS), via the respiratory tract was used for both sensitization and challenge. The role of IgE antibodies in the immune response to inhaled TNBS in this model was assessed by comparing the responses of wild-type (WT) and IgE-deficient (IgE(-/-)) mice on the BALB/c background. Reconstitution of circulating IgE levels by intravenous injection of IgE antibodies into IgE(-/-) mice before sensitization was performed to confirm the role of IgE in any differences observed between the responses of WT and IgE(-/-) mice.. Intranasal challenge of TNBS-sensitized (but not sham-sensitized control mice) induced intense pulmonary inflammation. Macrophages, eosinophils and lymphocytes, including T, B, natural killer and natural killer T cells, were recruited to the airway and the animals displayed bronchial hyperresponsiveness (BHR) to methacholine. Serum levels of murine mast cell protease-1 (mMCP-1) were elevated suggesting mast cell activation. In contrast, the development of airway inflammation, recruitment of lymphocytes, induction of BHR and production of mMCP-1 were all significantly attenuated in IgE-deficient mice. Reconstitution of IgE(-/-) mice with IgE (of unrelated antigen specificity) before sensitization partially restored these features of asthma.. Our data indicate that IgE antibodies non-specifically enhance the development of airway inflammation induced by exposure to chemical antigens.

Topics: Airway Resistance; Animals; Asthma; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cell Count; Chemokine CCL2; Dinitrobenzenes; Disease Models, Animal; Eosinophils; Haptens; Immunization; Immunoglobulin E; Inflammation; Killer Cells, Natural; Lung; Metaplasia; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Knockout; Natural Killer T-Cells; Occupational Diseases; T-Lymphocytes; Trinitrobenzenesulfonic Acid

Efficient priming of adaptive immunity depends on danger signals provided by innate immune pathways. As an example, inflammasome-mediated activation of caspase-1 and IL-1beta is crucial for the development of reactive T cells targeting sensitizers like dinitrofluorobenzene (DNFB). Surprisingly, DNFB and dinitrothiocyanobenzene provide cross-reactive Ags yet drive opposing, sensitizing vs tolerizing, T cell responses. In this study, we show that, in mice, inflammasome-signaling levels can be modulated to turn dinitrothiocyanobenzene into a sensitizer and DNFB into a tolerizer, and that it correlates with the IL-6 and IL-12 secretion levels, affecting Th1, Th17, and regulatory T cell development. Hence, our data provide the first evidence that the inflammasome can define the type of adaptive immune response elicited by an Ag, and hint at new strategies to modulate T cell responses in vivo.

Topics: Animals; Antigens; Caspase 1; Cross Reactions; Dinitrobenzenes; Dinitrofluorobenzene; Humans; Immune Tolerance; Immunity, Innate; Inflammation; Interleukin-1beta; Interleukin-2; Interleukin-6; Mice; Signal Transduction; T-Lymphocytes, Regulatory; Th1 Cells

Administration of a vaccine consisting of autologous melanoma cells modified with a hapten, dinitrophenyl (DNP), induces T cell infiltration of metastatic sites. We have reported an analysis of these infiltrating T cells, indicating that certain TCR-Vbeta gene segments are greatly overexpressed. In this study, we investigate the rearrangement of the TCR-Vbeta as well as the junctional diversity in T cells infiltrating melanoma metastases following treatment with DNP vaccine. In 19 of 26 control specimens, V-D-J length analysis showed the expected polyclonal patterns. In contrast, postvaccine tumors from 9 of 10 patients showed dominant peaks of V-D-J junction size in one or more Vbeta families. Dominant peaks were seen most frequently in six Vbeta families (Vbeta7, 12, 13, 14, 16, and 23) and were never seen in seven others. Further analysis of the oligoclonal Vbeta products showed dominant peaks in the J region as well. Of particular interest was the finding that Vbeta and Jbeta peaks were similar in inflamed metastases obtained at different times or from different sites from the same patient. Although 6 of 10 patients expressed HLA-A1, there was no common pattern of TCR rearrangements among them. Finally, the amplified PCR products from seven of these specimens were cloned and sequenced and the amino acid sequence of the complementarity-determining region 3 was deduced. In six of seven specimens, the same complementarity-determining region 3 sequence was repeated in at least two clones and in five of seven in at least three clones. Our study indicates that DNP vaccine induces the expansion of particular T cell clones that may be agents of its antitumor effects.

Topics: Cancer Vaccines; Clone Cells; Cloning, Molecular; Complementarity Determining Regions; Dinitrobenzenes; Gene Rearrangement, beta-Chain T-Cell Antigen Receptor; Humans; Inflammation; Lymph Nodes; Lymphocytes, Tumor-Infiltrating; Melanoma; Receptors, Antigen, T-Cell, alpha-beta; T-Lymphocyte Subsets; Transcription, Genetic

Treatment of metastatic melanoma patients with an autologous vaccine modified by the hapten, dinitrophenyl (DNP), produces a striking immunological effect: the induction of clinically evident inflammatory responses in metastatic tumors. Histological examination shows these tumors to be infiltrated with T lymphocytes. We studied the expression of activation markers on those cells and compared them with matched peripheral blood lymphocytes (PBL) and with lymphocytes extracted from metastases before treatment with DNP-conjugated vaccine. The median fraction of cells that were T cells in post-vaccine tumors was 41%, as compared with 9% in pre-treatment tumors, and those T cells were predominantly CD8+ (mean CD8/CD4 ratio = 5.0). A high proportion of both pre- and post-treatment infiltrating T cells expressed HLA-DR (mean +/- SE = 48% +/- 4%), CD69 (56% +/- 7%), and ganglioside GD3 (68% +/- 5%). This distinguished them from matched PBL in which expression of those markers was significantly lower (HLA-DR = 10% +/- 2%; CD69 = 2% +/- 0.4%; GD3 = 49% +/- 4%). These changes were not accompanied by increased cell-surface expression of interleukin-2 (IL-2) receptors, either CD25 or p75, which were expressed by 1%-2% and 12% of tumor-infiltrating lymphocytes (TIL), respectively. The pattern of activation marker expression that we identified appears to be characteristic of tissue T cells with the memory phenotype. The low expression of IL-2 receptors could indicate functional impairment of TIL in situ, perhaps because of inhibitory molecules produced by melanoma cells.

Topics: Biomarkers; Dinitrobenzenes; Flow Cytometry; Gangliosides; Haptens; HLA-DR Antigens; Humans; Immunotherapy, Active; Inflammation; Lymphocyte Activation; Lymphocytes, Tumor-Infiltrating; Melanoma; Phenotype; T-Lymphocytes; Vaccines, Conjugate

The percutaneous administration of in vitro haptenated epidermal cells (EC) has become established as a procedure to produce contact sensitivity (CS) in experimental animals for routine use. The cells have also been found to elicit a significant delayed-type skin reaction by intradermal test in the animals sensitized by painting the skin with the hapten. The fate of 2,4-dinitrophenylated (DNP) isogeneic epidermal cell suspensions (EC) injected intradermally was investigated histologically in intact or 2,4-dinitrochlorobenzene (DNCB)-sensitized strain 13 guinea pigs to study the role of the cells in CS. DNP-EC were found to proliferate actively in the dermis and formed EC nests with central keratinization and then elicited inflammatory reaction associated with necrosis of the epidermal structures 7 days after injection in the intact animals. DNP-EC injected intradermally into the animals which had received and reacted against DNCB underwent a suppression of EC proliferation. These findings are discussed in relation to the role of the haptenated EC in CS.

Topics: Animals; Dinitrobenzenes; Dinitrochlorobenzene; Epidermis; Guinea Pigs; Haptens; Hypersensitivity, Delayed; Inflammation; Injections, Intradermal; Male; Oxazolone; Skin Diseases

Specificity of plasmacellular infiltration was studied using a guinea pig peritoneal inflammation model. Acute and chronic inflammations were induced by repeated injections of either of two non-crossreacting antigens (DNP-BSA and PPD). With an enzyme-immunohistochemical sandwich procedure allowing quantitation of DNP-BSA-specific plasma cells, specificity of plasmacellular infiltration could be demonstrated. DNP-BSA-specific antibody-forming cells were found not to enter inflammatory reactions elicited by PPD. Our data support the hypothesis that virtually all plasma cells in a chronic inflammatory exudate release antibodies specific for antigens that are locally available, and that such antigens are likely to play a central role in the perpetuation of chronicity.

Topics: Animals; Antibody Specificity; Antibody-Producing Cells; Cell Movement; Chronic Disease; Dinitrobenzenes; Female; Guinea Pigs; Inflammation; Peritoneal Cavity; Plasma Cells; Serum Albumin, Bovine; Tuberculin