pyrophosphate and Neoplasms

Ribonucleotide reductase (RR) is an essential multi-subunit enzyme found in all living organisms; it catalyzes the rate-limiting step in dNTP synthesis, namely, the conversion of ribonucleoside diphosphates to deoxyribonucleoside diphosphates. As expression levels of human RR (hRR) are high during cell replication, hRR has long been considered an attractive drug target for a range of proliferative diseases, including cancer. While there are many excellent reviews regarding the structure, function, and clinical importance of hRR, recent years have seen an increase in novel approaches to inhibiting hRR that merit an updated discussion of the existing inhibitors and strategies to target this enzyme. In this review, we discuss the mechanisms and clinical applications of classic nucleoside analog inhibitors of hRRM1 (large catalytic subunit), including gemcitabine and clofarabine, as well as inhibitors of the hRRM2 (free radical housing small subunit), including triapine and hydroxyurea. Additionally, we discuss novel approaches to targeting RR and the discovery of new classes of hRR inhibitors.

Topics: Catalytic Domain; Diphosphates; Enzyme Inhibitors; Humans; Neoplasms; Ribonucleotide Reductases

DNA sequencing is critical to identifying many human genetic disorders caused by DNA mutations, including cancer. Pyrosequencing is less complex, involves fewer steps, and has a superior limit of detection compared with Sanger sequencing. The fundamental basis of pyrosequencing is that pyrophosphate is released when a deoxyribonucleotide triphosphate is added to the end of a nascent strand of DNA. Because deoxyribonucleotide triphosphates are sequentially added to the reaction and because the pyrophosphate concentration is continuously monitored, the DNA sequence can be determined.. To demonstrate the fundamental principles of pyrosequencing.. Salient features of pyrosequencing are demonstrated using the free software program Pyromaker ( http://pyromaker.pathology.jhmi.edu ), through which users can input DNA sequences and other pyrosequencing parameters to generate the expected pyrosequencing results.. We demonstrate how mutant and wild-type DNA sequences result in different pyrograms. Using pyrograms of established mutations in tumors, we explain how to analyze the pyrogram peaks generated by different dispensation sequences. Further, we demonstrate some limitations of pyrosequencing, including how some complex mutations can be indistinguishable from single base mutations. Pyrosequencing is the basis of the Roche 454 next-generation sequencer and many of the same principles also apply to the Ion Torrent hydrogen ion-based next-generation sequencers.

Topics: Base Sequence; Diphosphates; DNA Mutational Analysis; DNA, Neoplasm; Genotype; High-Throughput Nucleotide Sequencing; Humans; Mutation; Neoplasms; Polymerase Chain Reaction; Sequence Analysis, DNA; Software

Upon receiving the Nobel Prize in Physiology or Medicine in 1987, Susumu Tonegawa referred to the then recent discovery of the γδ T-cell receptor and stated that "while the function of the T cells bearing this receptor is currently unknown (…) these T cells may be involved in an entirely new aspect of immunity". [Tonegawa, S., Scand. J. Immunol. 1993. 38: 303-319]. Twenty-five years of intense research later this ambivalent view still holds true. Immunologists now appreciate that γδ T cells indeed represent a highly intriguing "new aspect of immunity" that is unique and distinct from conventional lymphocytes, yet even scientists in the field still struggle to understand the molecular basis of γδ T-cell responses, especially with respect to the enigmatic mode of antigen recognition. Here, we portray the peculiar responsiveness of human Vγ9/Vδ2 T cells to microorganisms, tumor cells and aminobisphosphonates, in an attempt to integrate the corresponding - and at times confusing - findings into a "theory of everything" that may help explain how such diverse stimuli result in similar γδ T-cell responses via the recognition of soluble low molecular weight phosphoantigens.

Topics: Animals; Diphosphates; Humans; Immunotherapy; Infections; Lymphocyte Activation; Neoplasms; Receptors, Antigen, T-Cell, gamma-delta; T-Lymphocytes

Vascular disrupting agents (VDAs) have presented a new kind of anti-cancer drug in recent years. VDAs take advantage of the weakness of established tumor endothelial cells and their supporting structures. In contrast to anti-angiogenic therapy, which inhibits the outgrowth of new blood vessels, vascular targeting treatments selectively attack the existing tumor vasculature. Here we summarized the anti-tumor activities, mechanisms and clinical applications of small molecule VDAs.

Topics: Angiogenesis Inhibitors; Animals; Antineoplastic Agents; Bibenzyls; Diphosphates; Endothelial Cells; Humans; Molecular Structure; Neoplasms; Neovascularization, Pathologic; Oligopeptides; Organophosphorus Compounds; Serine; Stilbenes; Tubulin Modulators; Xanthones

Human Vgamma2Vdelta2 T cells play important roles in mediating immunity against microbial pathogens and have potent anti-tumor activity. Vgamma2Vdelta2 T cells recognize the pyrophosphorylated isoprenoid intermediates (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMBPP), an intermediate in the foreign 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway, and isopentenyl pyrophosphate (IPP), an intermediate in the self-mevalonate pathway. Infection with bacteria and protozoa using the MEP pathway leads to the rapid expansion of Vgamma2Vdelta2 T cells to very high numbers through preferential recognition of HMBPP. Activated Vgamma2Vdelta2 T cells produce proinflammatory cytokines and chemokines, kill infected cells, secrete growth factors for epithelial cells, and present antigens to alphabeta T cells. Vgamma2Vdelta2 T cells can also recognize high levels of IPP in certain tumors and in cells treated with pharmacological agents, such as bisphosphonates and alkylamines, that block farnesyl pyrophosphate synthase. Activated Vgamma2Vdelta2 T cells are able to kill most tumor cells because of recognition by T-cell receptor and natural killer receptors. The ubiquitous nature of the antigens converts essentially all Vgamma2Vdelta2 T cells to memory cells at an early age. Thus, primary infections with HMBPP-producing bacteria are perceived by Vgamma2Vdelta2 T cells as a repeat infection. Extensive efforts are underway to harness these cells to treat a variety of cancers and to provide microbial immunity.

Topics: Antigen Presentation; Diphosphates; Humans; Immunologic Memory; Lymphocyte Activation; Neoplasms; Protein Prenylation; Receptors, Antigen, T-Cell, gamma-delta; T-Lymphocyte Subsets

Vascular targeting agents (VTAs) for the treatment of cancer are designed to cause a rapid and selective shutdown of the blood vessels of tumors. Unlike antiangiogenic drugs that inhibit the formation of new vessels, VTAs occlude the pre-existing blood vessels of tumors to cause tumor cell death from ischemia and extensive hemorrhagic necrosis. Tumor selectivity is conferred by differences in the pathophysiology of tumor versus normal tissue vessels (e.g., increased proliferation and fragility, and up-regulated proteins). VTAs can kill indirectly the tumor cells that are resistant to conventional antiproliferative cancer therapies, i.e., cells in areas distant from blood vessels where drug penetration is poor, and hypoxia can lead to radiation and drug resistance. VTAs are expected to show the greatest therapeutic benefit as part of combined modality regimens. Preclinical studies have shown VTA-induced enhancement of the effects of conventional chemotherapeutic agents, radiation, hyperthermia, radioimmunotherapy, and antiangiogenic agents. There are broadly two types of VTAs, small molecules and ligand-based, which are grouped together, because they both cause acute vascular shutdown in tumors leading to massive necrosis. The small molecules include the microtubulin destabilizing drugs, combretastatin A-4 disodium phosphate, ZD6126, AVE8062, and Oxi 4503, and the flavonoid, DMXAA. Ligand-based VTAs use antibodies, peptides, or growth factors that bind selectively to tumor versus normal vessels to target tumors with agents that occlude blood vessels. The ligand-based VTAs include fusion proteins (e.g., vascular endothelial growth factor linked to the plant toxin gelonin), immunotoxins (e.g., monoclonal antibodies to endoglin conjugated to ricin A), antibodies linked to cytokines, liposomally encapsulated drugs, and gene therapy approaches. Combretastatin A-4 disodium phosphate, ZD6126, AVE8062, and DMXAA are undergoing clinical evaluation. Phase I monotherapy studies have shown that the agents are tolerated with some demonstration of single agent efficacy. Because efficacy is expected when the agents are used with conventional chemotherapeutic drugs or radiation, the results of Phase II combination studies are eagerly awaited.

Topics: Angiogenesis Inhibitors; Antibodies, Monoclonal; Cell Division; Clinical Trials as Topic; Diphosphates; Genetic Therapy; Humans; Hypoxia; Immunotoxins; Ligands; Models, Biological; Necrosis; Neoplasms; Organophosphorus Compounds; Peptides; Radioimmunotherapy; Stilbenes; Time Factors; Up-Regulation; Xanthones

Iron deficiency is the most common nutritional deficiency in advanced cancer patients and causes anaemia. Iron deficiency anaemia treatment (i.e. intravenous or oral iron administration) has been demonstrated to be effective but is often associated with adverse reactions. Micronised microencapsulated ferric pyrophosphate (MMFP) is a recently developed formulation characterised by a higher intestinal bioavailability due to the small particle size distribution at nanometer level. The aim of this study was to evaluate the efficacy of an oral administration of 30 mg of MMFP associated with 80 mg of ascorbic acid in advanced cancer patients with hyposideraemia.. This was an observational prospective cohort study (10 months) conducted on 42 adult patients with advanced cancer and serum iron levels lower than 60 μg/dL. All patients received one capsule/day for 30 days of a supplement containing 30 mg of MMFP and 80 mg of ascorbic acid. At enrolment (T0) and at 30 days (T1) patients were subjected to blood sampling for evaluation of serum iron, ferritinaemia and blood count. In addition, any undesirable effects reported by patients were evaluated.. MMFP treatment increased sideraemia from 36.1±8.37 μg/dL to 73.22±28.60 μg/dL, haemoglobin from 10.43±1.09 g/dL to 11.52±1.90 g/dL, and ferritinaemia from 42.10±16.90 ng/mL to 123.33±55.79 ng/mL. No adverse effects were noted from the use of MMFP supplementation.. The supplementation of 30 mg/d of MMFP in combination with 80 mg/d of ascorbic acid in advanced cancer patients with hyposideraemia led to a significant increase in sideraemia and ferritinaemia. Moreover, in some of the patients whose serum iron level did not increase, an increase in haemoglobin was observed.

Topics: Administration, Intravenous; Administration, Oral; Aged; Anemia, Iron-Deficiency; Diphosphates; Female; Humans; Iron; Iron Deficiencies; Male; Middle Aged; Neoplasms; Pilot Projects; Prospective Studies

This phase I, open-label, randomized, 2-part crossover study assessed the safety, pharmacokinetics and relative bioavailability of single doses of the anticancer MET inhibitor foretinib (formerly known as GSK1363089, EXEL-2880 and XL-880) free base tablet formulation compared to a bisphosphate salt capsule formulation (Part 1), and assessed the safety, efficacy, and pharmacokinetics of the bisphosphate salt capsule administered 3 times a week in cancer patients (Part 2).. In Part 1, patients were randomized in a crossover manner to receive a single oral dose of foretinib formulated as a bisphosphate salt capsule (240 mg; 183 mg free base equivalent) followed one week later by a single dose of a free base tablet (180 mg), or vice versa where the treatment sequence was reversed. In Part 2, patients self-administered oral doses of bisphosphate salt capsules (200 mg) 3 times a week until disease progression.. Twelve patients with solid tumors were enrolled and completed Part 1, and 10 patients continued into Part 2. Most AEs were mild or moderate in severity. The most common drug-related AEs were fatigue, diarrhea, and nausea. The least-squares (LS) mean total area under the curve was 3144 and 3514 ng*h/mL for the free base tablet and bisphosphate salt capsule, respectively, with a ratio of 0.89 (90% confidence interval, CI: 0.69, 1.16). The LS mean maximal concentration (Cmax) was 81.6 and 98.5 ng/mL for the free base and bisphosphate salt, respectively, with a ratio of 0.83 (90% confidence interval, CI: 0.67, 1.02). The time to reach Cmax was ∼4 h for both formulations. The pharmacokinetics of foretinib were not clinically different between the 2 formulations. Of the 10 patients assessed for efficacy, 3 patients achieved stable disease.. Foretinib was well tolerated as single doses of both the free base and bisphosphate salt formulations. The pharmacokinetics and relative bioavailability of the 2 formulations were not clinically different. The bisphosphate salt formulation was well tolerated on a 3-times a week dosing schedule, and reached steady-state plasma concentration after 2 weeks.

Topics: Administration, Oral; Adult; Aged; Angiogenesis Inhibitors; Anilides; Area Under Curve; Biological Availability; Capsules; Chemistry, Pharmaceutical; Cross-Over Studies; Diphosphates; Drug Compounding; Female; Humans; Least-Squares Analysis; Male; Middle Aged; Neoplasms; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-met; Quinolines; Tablets; Treatment Outcome; Young Adult

Preclinical studies show that OXi4503 (combretastatin A1 diphosphate, CA1P) is more potent than other clinically evaluated vascular-disrupting agents.. Escalating doses of OXi4503 were given intravenously over 10 minutes on days 1, 8, and 15 every 28 days to patients with advanced solid tumors.. Doses were escalated in single-patient cohorts from 0.06 to 1.92 mg/m(2), then expanded cohorts to 15.4 mg/m(2) in 43 patients. Common adverse drug reactions were hypertension, tumor pain, anemia, lymphopenia, and easily controllable nausea/vomiting and fatigue. Five patients experienced different drug-related dose-limiting toxicities, atrial fibrillation, increased troponin, blurred vision, diplopia, and tumor lysis. Prophylactic amlodipine failed to prevent adverse events. Pharmacokinetics showed dose-dependent linear increases in peak plasma concentrations and area under the curve value of OXi4503. One partial response was seen in a heavily pretreated patient with ovarian cancer. Dynamic contrast-enhanced MRI confirmed a dose effect and showed significant antivascular effects in 10 of 13 patients treated at doses of 11 mg/m(2) or higher.. The maximum tolerated dose was 8.5 mg/m(2) but escalation to 14 mg/m(2) was possible with only temporary reversible cerebrovascular toxicity by excluding hypertensive patients. As a tumor response was seen at 14 mg/m(2) and maximum tumor perfusion reductions were seen at doses of 11 mg/m(2) or higher, the recommended phase II dose is from 11 to 14 mg/m(2).

Topics: Adult; Aged; Angiogenesis Inhibitors; Antineoplastic Agents; Diphosphates; Female; Humans; Magnetic Resonance Imaging; Male; Middle Aged; Neoplasms; Stilbenes; Treatment Outcome; Young Adult

OXi4503 is a tubulin-binding vascular disrupting agent that has recently completed a Cancer Research UK-sponsored phase I trial. Preclinical studies demonstrated early drug-induced apoptosis in tumour endothelial cells at 1-3 h and secondary tumour cell necrosis between 6 and 72 h.. To capture both possible outcomes of OXi4503 treatment on cell death, plasma samples for analysis by M30 and M65 ELISAs, which measure different circulating forms of cytokeratin 18 as biomarkers of apoptosis and necrosis, respectively, were collected from patients entered into the trial at early (4/6 h) and later time points (24h, day 8 and day 15).. OXi4503 induced a selective dose-dependent elevation in M30 antigen levels (apoptosis) at 4/6 h and a similar elevation in M65 antigen levels at 24 h (necrosis) consistent with its preclinical cell death profile. For the purposes of investigating potential biomarker relationships to patient characteristics, the trial population was divided into three groups based on radiological and clinical response: (a) early progression, (b) progressive disease and (c) stable disease (SD)/partial response. A significant increase in antigen concentrations was measured by M65 at 24 h in the SD group compared with the two other groups (P=0.015, mean increase 30.9%).. These results provide pharmacodynamic evidence of drug mechanism of action in cancer patients and highlight the M65 ELISA as a potentially useful biomarker assay of response to OXi4503.

Topics: Antineoplastic Agents; Apoptosis; Biomarkers, Tumor; Diphosphates; Enzyme-Linked Immunosorbent Assay; Humans; Keratin-18; Neoplasms; Neovascularization, Pathologic; Peptide Fragments; Stilbenes

Vgamma9Vdelta2 (gammadelta) T lymphocytes, a critical peripheral blood lymphocyte subset, are directly cytotoxic against many solid and hematologic tumor types. Vgamma9Vdelta2 T lymphocytes can be selectively expanded in vivo with BrHPP (IPH1101) and IL-2. The present phase I trial was conducted with the aim of determining the maximum-tolerated dose (MTD) and safety of IPH1101 combined with a low dose of IL-2 in patients with solid tumors.. A 1-h intravenous infusion of IPH11 was administered alone at cycle 1, combined with a low dose of SC IL-2 (1 MIU/M(2) d1 to d7) in the subsequent cycles (day 1 every 3 weeks). The dose of IPH1101 was escalated from 200 to 1,800 mg/m(2).. As much as 28 patients with solid tumors underwent a total of 109 treatment cycles. Pharmacodynamics data demonstrate that gammadelta T lymphocyte amplification in humans requires the co-administration of IL-2 and is dependent on IPH 1101 dose. Dose-limiting toxicity occurred in two patients at a dose of 1,800 mg/m(2): one grade 3 fever (1 patient) and one grade 3 hypotension (1 patient) suggesting cytokine release syndrome immediately following the first infusion. At lower doses the treatment was well tolerated; the most frequent adverse events were mild fever, chills and abdominal pain, without exacerbation in the IL-2 combined cycles.. IPH1101 in combination with SC low-dose IL-2 is safe, well tolerated and induces a potent gammadelta T lymphocyte expansion in patients. Its clinical activity will be evaluated in phase II clinical trials.

Topics: Adult; Aged; Antineoplastic Agents; Diphosphates; Dose-Response Relationship, Drug; Female; Humans; Interleukin-2; Male; Maximum Tolerated Dose; Middle Aged; Neoplasms; T-Lymphocyte Subsets; T-Lymphocytes

Vascular disrupting agents (VDA) cause acute shutdown of abnormal established tumor vasculature, followed by massive intratumoral hypoxia and necrosis. However, a viable rim of tumor tissue invariably remains from which tumor regrowth rapidly resumes. We have recently shown that an acute systemic mobilization and homing of bone marrow-derived circulating endothelial precursor (CEP) cells could promote tumor regrowth following treatment with either a VDA or certain chemotherapy drugs. The molecular mediators of this systemic reactive host process are unknown. Here, we show that following treatment of mice with OXi-4503, a second-generation potent prodrug derivative of combretastatin-A4 phosphate, rapid increases in circulating plasma vascular endothelial growth factor, stromal derived factor-1 (SDF-1), and granulocyte colony-stimulating factor (G-CSF) levels are detected. With the aim of determining whether G-CSF is involved in VDA-induced CEP mobilization, mutant G-CSF-R(-/-) mice were treated with OXi-4503. We found that as opposed to wild-type controls, G-CSF-R(-/-) mice failed to mobilize CEPs or show induction of SDF-1 plasma levels. Furthermore, Lewis lung carcinomas grown in such mice treated with OXi-4503 showed greater levels of necrosis compared with tumors treated in wild-type mice. Evidence for rapid elevations in circulating plasma G-CSF, vascular endothelial growth factor, and SDF-1 were also observed in patients with VDA (combretastatin-A4 phosphate)-treated cancer. These results highlight the possible effect of drug-induced G-CSF on tumor regrowth following certain cytotoxic drug therapies, in this case using a VDA, and hence G-CSF as a possible therapeutic target.

Topics: Animals; Antibodies, Monoclonal; Antibodies, Monoclonal, Humanized; Antineoplastic Combined Chemotherapy Protocols; Bevacizumab; Chemokine CXCL12; Diphosphates; Endothelial Cells; Granulocyte Colony-Stimulating Factor; Hematopoietic Stem Cell Mobilization; Humans; Melanoma; Mice; Mice, Inbred C57BL; Mice, Nude; Mice, Transgenic; Neoplasms; Prodrugs; Stem Cells; Stilbenes; Vascular Endothelial Growth Factor A; Xenograft Model Antitumor Assays

(1R,2R-diaminocyclohexane)(dihydropyrophosphato) platinum(II), also abbreviated as RRD2, belongs to a class of potent antitumor platinum cytostatics called phosphaplatins. Curiously, several published studies have suggested significant mechanistic differences between phosphaplatins and conventional platinum antitumor drugs. Controversial findings have been published regarding the role of RRD2 binding to DNA in the mechanism of its antiproliferative activity in cancer cells. This prompted us to perform detailed studies to confirm or rule out the role of RRD2 binding to DNA in its antiproliferative effect in cancer cells. Here, we show that RRD2 exhibits excellent antiproliferative activity in various cancer cell lines, with IC

Topics: Antineoplastic Agents; Cisplatin; Diphosphates; DNA; Neoplasms; Organoplatinum Compounds; Oxaliplatin; Platinum

Topics: Amyloidosis; Cardiomyopathies; Diphosphates; Humans; Neoplasms; Prealbumin; Technetium Tc 99m Pyrophosphate

The availability of specific stimuli to induce the anticancer cytotoxicity of human TCRVγ9-expressing T lymphocytes has allowed the development of γδ T cell-based cancer immunotherapies. However, the stringent dependence of such strategies on the inherently toxic IL-2 has raised safety concerns for patients, justifying a search for alternative methods for inducing γδ T cell stimulation. IL-33 is a γ-chain receptor-independent cytokine of the IL-1 superfamily that is expressed by endothelial cells from a tumor microenvironment and can sustain Th1 and Th2 immune responses. Therefore, we investigated its ability to support the stimulation of human TCRVγ9(+) γδ T cells. In this study, we report that IL-33 efficiently sustained the in vitro activation of Vγ9 T lymphocytes by synthetic phosphoantigens, zoledronate, and a BTN3A1 Ab in the absence of an exogenous supply of IL-2. IL-33 was as potent as IL-2 in allowing the proliferative amplification of Vγ9 T cells isolated from PBMC following activation by the synthetic phosphoantigen bromohydrin pyrophosphate. IL-33 also induced an identical maturation into TNF-α- and IFN-γ-producing Th1 effector memory cells, and IL-33-stimulated cells showed an equivalent cytotoxicity for various tumor cells in vitro. Finally, we found that the bioactivity of IL-33 on the Vγ9 T cell was indirectly mediated through contact with CD4 T cells and IL-2 production by CD4 T cells and Vγ9 T cells themselves. These data posit IL-33 as an alternative to IL-2 for Vγ9 T cell-based cancer immunotherapies.

Topics: Antigens, CD; Butyrophilins; Cell Proliferation; Cells, Cultured; Diphosphates; Diphosphonates; Endothelial Cells; Humans; Imidazoles; Immunotherapy; Interferon-gamma; Interleukin-2; Interleukin-33; Leukocytes, Mononuclear; Lymphocyte Activation; Neoplasms; Receptors, Antigen, T-Cell, gamma-delta; Th1 Cells; Tumor Necrosis Factor-alpha; Zoledronic Acid

The response of tissues to radiation with mild temperature hyperthermia is dependent on the interval between the two modalities. This study investigated the effect that the vascular disrupting agent OXi4503 had on this time-interval interaction.. The normal right rear foot of female CDF1 mice or foot-implanted C3H mammary carcinomas were locally irradiated (230 kV X-rays) and heated (41.5 °C for 60 min) by foot immersion in a water bath. OXi4503 (50 mg/kg) was injected intraperitoneally 1.5 h before irradiating. Irradiation was performed either in the middle of the heating period (simultaneous treatment) or at 1 or 4 h prior to starting the heating (sequential treatments). Response was the percentage of mice showing local tumour control at 90 days or skin moist desquamation between days 11-23. From the radiation dose response curves the dose producing tumour control (TCD(50)) or moist desquamation (MDD50) in 50% of mice was calculated.. The TCD(50) and MDD50 values for radiation alone were 54 Gy and 29 Gy, respectively. Simultaneously heating the tissues enhanced radiation response, the respective TCD(50) and MDD50 values being significantly (chi-square test, p < 0.05) reduced to 33 Gy and 14 Gy. A smaller enhancement was obtained with a sequential treatment in both tissues. OXi4503 enhanced the radiation response of tumour and skin. Combined with radiation and heat, the only effect was in tumours where OXi4503 prevented the decrease in sensitisation seen with the sequential treatment.. Combining OXi4503 with a sequential radiation and heat treatment resulted in a 1.4-fold therapeutic gain.

Topics: Animals; Diphosphates; Female; Hyperthermia, Induced; Mice; Mice, Inbred C3H; Neoplasms; Radiation, Ionizing; Stilbenes

Bispecific antibodies have been successfully introduced into clinical application. γδ T cells are of special interest for tumor immunotherapy, due to their recognition of pyrophosphates that are overproduced by many tumor cells resulting in HLA-nonrestricted tumor cell killing. Here we describe in detail a [(Her2)2 × Vγ9] tribody construct that targets human Vγ9 T cells to HER2-expressing tumor cells. The direct comparison with other selective Vγ9 T cell agonists including phosphoantigens and nitrogen-containing bisphosphonates revealed the superiority of the [(Her2)2 × Vγ9] tribody in triggering γδ T cell-mediated tumor cell killing with negligible induction of γδ T cell death. In contrast, phosphoantigens and bisphosphonates are potent inducers of γδ T cell proliferation but less efficient enhancers of γδ T cell-mediated tumor cell killing. Collectively, our data identify unique properties of a γδ T cell-targeting [(Her2)2 × Vγ9] tribody which make it an attractive candidate for clinical application in γδ T cell-based tumor immunotherapy.

Topics: Antibodies, Bispecific; CD8-Positive T-Lymphocytes; Cell Line, Tumor; Cell Proliferation; Cytotoxicity, Immunologic; Diphosphates; Diphosphonates; Humans; Immunotherapy; Lymphocyte Activation; Neoplasms; Receptor, ErbB-2; Receptors, Antigen, T-Cell, gamma-delta

Phosphorylation of 2-(3-hydroxy-5-methoxyphenyl)-6,7-methylenedioxy-1H-quinolin-4-one (1) afforded diphosphate 2. We found that, upon treatment with methanol under mild conditions, 2 can undergo facile and highly regioselective dephosphorylation to give the monophosphate 3, with a phosphate group remaining on the phenyl ring. The details of the dephosphorylation process were postulated and then probed by LC-MS and HPLC analyses. Furthermore, as a preliminary study, the water soluble monophosphate prodrug 4 was tested for antitumor activity against a MCF-7 xenograft nude mice model.

Topics: Animals; Antineoplastic Agents; Diphosphates; Humans; MCF-7 Cells; Mice; Neoplasms; Phosphorylation; Prodrugs; Quinolinium Compounds; Solubility; Structure-Activity Relationship; Xenograft Model Antitumor Assays

Phosphoantigens enable the access to a new anti-tumoral and anti-infectious therapeutic pathway, based on innate immunity through the selective activation of Tγ9δ2 lymphocytes. The first proof of concept of this new immunotherapy approach was demonstrated with the synthetic phosphoantigen named bromohydrin pyrophosphate (BrHPP, IPH 1101) which was administrated in racemic form to about 200 patients in six clinical trials with good safety and promising early signals of efficacy in type C viral hepatitis and follicular non-Hodgkin's lymphoma. Enantiopure samples of BrHPP in gram scale are required for further studies on structure-bioactivity relationship. Thus we developed two complementary synthetic pathways, the first using transformation of a chiral compound and the second involving asymmetric synthesis starting from a prochiral building-block. The synthesis of a second-generation phosphoantigen, N-HDMAPP, which bears a phosphoramidate moiety, was also investigated.

Topics: Diphosphates; Humans; Immunotherapy; Molecular Structure; Neoplasms; Organophosphorus Compounds; Stereoisomerism

Tumour growth promotes the expansion of CD4(+) CD25(+) FoxP3(+) regulatory T cells (Tregs) which suppress various arms of immune responses and might therefore contribute to tumour immunosurveillance. In this study, we found an inverse correlation between circulating Treg frequencies and phosphoantigen-induced gammadelta T-cell proliferation in cancer patients, which prompted us to address the role of Tregs in controlling the gammadelta T-cell arm of innate immune responses. In vitro, human Treg-peripheral blood mononuclear cell (PBMC) co-cultures strongly inhibited phosphoantigen-induced proliferation of gammadelta T cells and depletion of Tregs restored the impaired phosphoantigen-induced gammadelta T-cell proliferation of cancer patients. Tregs did not suppress other effector functions of gammadelta T cells such as cytokine production or cytotoxicity. Our experiments indicate that Tregs do not mediate their suppressive activity via a cell-cell contact-dependent mechanism, but rather secrete a soluble non-proteinaceous factor, which is independent of known soluble factors interacting with amino acid depletion (e.g. arginase-diminished arginine and indolamine 2,3-dioxygenase-diminished tryptophan) or nitric oxide (NO) production. However, the proliferative activity of alphabeta T cells was not affected by this cell-cell contact-independent suppressive activity induced by Tregs. In conclusion, these findings indicate a potential new mechanism by which Tregs can specifically suppress gammadelta T cells and highlight the strategy of combining Treg inhibition with subsequent gammadelta T-cell activation to enhance gammadelta T cell-mediated immunotherapy.

Topics: Antigens; Cell Communication; Cell Proliferation; Cells, Cultured; Cytotoxicity, Immunologic; Diphosphates; Forkhead Transcription Factors; Humans; Immune Tolerance; Immunity, Innate; Interleukin-2 Receptor alpha Subunit; Lymphocyte Culture Test, Mixed; Neoplasms; Phosphoproteins; Receptors, Antigen, T-Cell, gamma-delta; T-Lymphocyte Subsets; T-Lymphocytes, Regulatory; Tumor Cells, Cultured

The unique responsiveness of Vgamma9Vdelta2 T-cells, the major gammadelta subset of human peripheral blood, to non-peptidic prenyl pyrophosphate antigens constitutes the basis of current gammadelta T-cell-based cancer immunotherapy strategies. However, the molecular mechanisms responsible for phosphoantigen-mediated activation of human gammadelta T-cells remain unclear. In particular, previous reports have described a very slow kinetics of activation of T-cell receptor (TCR)-associated signal transduction pathways by isopentenyl pyrophosphate and bromohydrin pyrophosphate, seemingly incompatible with direct binding of these antigens to the Vgamma9Vdelta2 TCR. Here we have studied the most potent natural phosphoantigen yet identified, (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP), produced by Eubacteria and Protozoa, and examined its gammadelta T-cell activation and anti-tumor properties.. We have performed a comparative study between HMB-PP and the anti-CD3epsilon monoclonal antibody OKT3, used as a reference inducer of bona fide TCR signaling, and followed multiple cellular and molecular gammadelta T-cell activation events. We show that HMB-PP activates MEK/Erk and PI-3K/Akt pathways as rapidly as OKT3, and induces an almost identical transcriptional profile in Vgamma9(+) T-cells. Moreover, MEK/Erk and PI-3K/Akt activities are indispensable for the cellular effects of HMB-PP, including gammadelta T-cell activation, proliferation and anti-tumor cytotoxicity, which are also abolished upon antibody blockade of the Vgamma9(+) TCR Surprisingly, HMB-PP treatment does not induce down-modulation of surface TCR levels, and thereby sustains gammadelta T-cell activation upon re-stimulation. This ultimately translates in potent human gammadelta T-cell anti-tumor function both in vitro and in vivo upon transplantation of human leukemia cells into lymphopenic mice,. The development of efficient cancer immunotherapy strategies critically depends on our capacity to maximize anti-tumor effector T-cell responses. By characterizing the intracellular mechanisms of HMB-PP-mediated activation of the highly cytotoxic Vgamma9(+) T-cell subset, our data strongly support the usage of this microbial antigen in novel cancer clinical trials.

Topics: Animals; Antigens, Bacterial; CD3 Complex; Cell Line, Tumor; Cytotoxicity, Immunologic; Diphosphates; Endocytosis; Extracellular Signal-Regulated MAP Kinases; Humans; Interleukin-2; Ligands; Lymphocyte Activation; Mice; Mice, SCID; Mitogen-Activated Protein Kinase Kinases; Molecular Mimicry; Neoplasms; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Receptors, Antigen, T-Cell, gamma-delta; Signal Transduction; T-Lymphocytes; Transcription, Genetic

Human Vgamma2Vdelta2 T cells recognize nonpeptide antigens, such as isoprenoid pyrophosphomonoester intermediates, alkylamine compounds, and bisphosphonate drugs, as well as some tumor cells. Although attempts have been made to derive novel cancer immunotherapies based on the discovery of these unconventional antigens, effective therapies remain to be developed. Here, we synthesized a series of pyrophosphate-containing compounds and examined the chemical requirements for the recognition of pyrophosphomonoester antigens by gammadelta T cells. The structural analysis clearly demonstrated that a proximal methylene moiety plays a crucial role in the stimulatory activity of the antigens. For optimal gammadelta T cell proliferation, we find that the use of human serum albumin was preferred and that pyrophosphomonoesters were superior to nitrogen-containing bisphosphonate compounds. Using these techniques, we have successfully expanded gammadelta T cells from healthy donors as well as from cancer patients using one of the most active compounds, 2-methyl-3-butenyl-1-pyrophosphate (2M3B1PP). The resulting expanded gammadelta T cells exhibited potent, cytotoxic activity against a wide variety of tumor cell lines. Even gammadelta T cells from a patient with advanced liver carcinoma efficiently responded to 2M3B1PP and exhibited strong cytotoxic activity against tumor cells. The pretreatment of tumor cells with nonpeptide antigens was essential for efficient cytotoxicity via TCR-gammadelta. The present study suggests a novel strategy for cancer immunotherapy using synthetic small pyrophosphate-containing compounds and nitrogen-containing bisphosphonates.

Topics: Antibody Specificity; Antigens, Neoplasm; Antineoplastic Agents; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Proliferation; Clone Cells; Diphosphates; Diphosphonates; Flow Cytometry; Humans; Immunotherapy; Interleukin-2; Jurkat Cells; Models, Molecular; Monocytes; Neoplasms; T-Lymphocytes

Human Vgamma2/Vdelta2(+) gammadelta T cells respond to low molecular-mass nonpeptide Ags in a gammadelta TCR-dependent manner. Although requirements of Ag presentation have remained controversial, we have indicated that specific responses of the primary gammadelta T cells to pamidronate were dependent on monocytic adherent cells for Ag presentation. Here, we show that human tumor cells can efficiently present aminobisphosphonate and pyrophosphomonoester compounds to gammadelta T cells, inducing specific proliferation and IFN-gamma production. gammadelta TCR dependency of the response to Ag-pulsed tumor cells was confirmed by using a Jurkat line transfected with a Vgamma2/Vdelta2 gammadelta TCR. Furthermore, gammadelta T cells exhibited markedly enhanced cytotoxicity against the Ag-pulsed tumor cells as compared with untreated tumor cells. Survey of a number of human tumor cell lines of different origins revealed that the majority of them became susceptible for gammadelta T cell-mediated cytotoxicity following the Ag pulsing except for breast cancer lines so far examined, while normal PHA blast cells remained resistant. The results not only imply a unique mode of nonpeptide Ag recognition by human gammadelta T cells but also may provide a novel strategic clue for immunotherapy of human malignancy.

Topics: Cytotoxicity, Immunologic; Diphosphates; Diphosphonates; Humans; Immunotherapy; Interferon-gamma; Lymphocyte Activation; Neoplasms; Pamidronate; Receptors, Antigen, T-Cell, gamma-delta; T-Lymphocytes; Tumor Cells, Cultured

Topics: Aged; Calcium Pyrophosphate; Diagnosis, Differential; Diphosphates; Female; Humans; Neoplasms; Radiography; Temporomandibular Joint; Temporomandibular Joint Disorders

Data on 97 patients were analysed. Of these, 42 received combined therapy, 30 radiotherapy and 25 chemotherapy. The utmost therapeutic effect (by the selected criteria) was obtained with combined treatment (88.1%). Radionuclide signs of the efficacy of radiotherapy were noted in 70% of the patients, whereas 23.3% did not respond. The least effect was obtained with chemotherapy: the absence of improvement or deterioration were noted in 48% of the patients. The correlation of the data obtained with the clinical ones (the painful syndrome) was observed. A high sensitivity of the method does not only help reveal pathological skeletal changes but also plan adequate therapy.

Topics: Bone Neoplasms; Combined Modality Therapy; Diphosphates; Diphosphonates; Female; Humans; Male; Neoplasms; Radionuclide Imaging; Technetium; Technetium Compounds; Technetium Tc 99m Pyrophosphate

Topics: Adolescent; Child; Child, Preschool; Diagnosis, Differential; Diphosphates; Female; Humans; Infant; Male; Neoplasms; Radionuclide Imaging; Sodium Pertechnetate Tc 99m; Technetium; Technetium Tc 99m Pyrophosphate

Topics: Diphosphates; Doxorubicin; Heart; Heart Diseases; Humans; Neoplasms; Radionuclide Imaging; Technetium

Technetium-99m pyrophosphate was utilized for myocardial imaging in 15 patients on adriamycin treatment for neoplasia. We have noted abnormal accumulation of the pyrophosphate in several patients, particularly in those in whom the so-called poor-risk factors were operative, namely prior radiation, cyclophosphamide therapy, and ischemic heart disease.

Topics: Adult; Aged; Antineoplastic Agents; Diphosphates; Doxorubicin; Female; Heart Diseases; Humans; Male; Middle Aged; Neoplasms; Radionuclide Imaging; Technetium

Topics: Cardiomyopathies; Diphosphates; Doxorubicin; Humans; Myocardium; Neoplasms; Radionuclide Imaging; Technetium

99Tcm-Sn pyrophosphate bone scans of 250 patients referred for skeletal metastatic survey were analysed to determine the frequency of abnormal extra-osseous localization and the various pathological causes. Twenty-six patients demonstrated abnormal extra-osseous concentration. There were three false positives. Sixty-five per cent of the extra-osseous lesions concentrating pyrophosphate were malignant (carcinoma of lung and breast, metastatic hepatic carcinoma, chondrosarcoma) and the remainder were benign lesions, e.g. sarcoidosis, soft-tissue calcification, post-surgical and irradiation sites. An incidental finding was the unusual frequency of accumulation of pyrophosphate in various joints, especially the knees and shoulders in asymptomatic patients above 70 years of age.

Topics: Age Factors; Aged; Bone Neoplasms; Breast Neoplasms; Diphosphates; False Positive Reactions; Humans; Joints; Liver Neoplasms; Lung Neoplasms; Middle Aged; Neoplasm Metastasis; Neoplasms; Radionuclide Imaging; Sarcoidosis; Technetium; Tin

Topics: Adenine; Adenosine Triphosphate; Animals; Antimetabolites; Ascites; Carbon Isotopes; Diphosphates; Metabolism; Mice; Neoplasms; Neoplasms, Experimental; Nucleosides; Nucleotides; Phosphoribosyl Pyrophosphate; Research; Tissue Culture Techniques; Xylose

Topics: Antineoplastic Agents; Diphosphates; Humans; Neoplasms

Topics: Cervix Uteri; Diphosphates; Female; Humans; Neoplasms; Thiamine; Thiamine Pyrophosphate

Topics: Animals; Characidae; Diphosphates; Naphthoquinones; Neoplasms; Radiation-Sensitizing Agents; Radiotherapy; Sodium; Vitamin K

Topics: Animals; Characidae; Diphosphates; Drug Therapy; Neoplasms; Sodium; Vitamin K

Topics: Animals; Characidae; Diphosphates; Humans; Ions; Naphthoquinones; Neoplasms; Sodium; Tetracycline