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Anita S. Chong - One of the best experts on this subject based on the ideXlab platform.
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inhibition of herpes simplex virus type 1 by the experimental immunosuppressive agent Leflunomide
Transplantation, 2001Co-Authors: Deborah A Knight, James W Williams, Anita S. Chong, Ashley Quintin Hejmanowski, Julie E Dierksheide, James W WaldmanAbstract:Background. Despite advances in antiviral chemotherapy, herpes simplex virus type 1 (HSV-1), continues to complicate the clinical course of many allograft recipients. We have previously demonstrated that the experimental immunosuppressive agent Leflunomide inhibits production of cytomegalovirus by interference with virion assembly. We test the hypothesis that this agent exerts similar antiviral activity against HSV-1 Methods and Results. Plaque assay of virus yield from endothelial or Vero cells after inoculation with each of four clinical HSV-1 isolates demonstrated a dose-dependent reduction of virus production in the presence of pharmacologic concentrations of A77 1726, the active metabolite of Leflunomide. DNA dot blot and biochemical assay of viral DNA polymerase activity indicated that A77 does not inhibit viral DNA synthesis. Rather, as visualized by transmission electron microscopic method, this agent seems to disrupt virion assembly by preventing nucleocapsid tegumentation. Conclusions. These findings, in demonstrating that Leflunomide exerts antiviral activity against HSV-1 by mechanisms similar to those we have previously shown with cytomegalovirus, imply that this agent may possess broad spectrum activity against other herpesviruses.
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in vitro and in vivo antitumor activity of a novel immunomodulatory drug Leflunomide mechanisms of action
Biochemical Pharmacology, 1999Co-Authors: Jikun Shen, James W Williams, Leonard Blinder, Julian W Mall, Jonathan Myers, Wanyun Huang, Theodore J Saclarides, Anita S. ChongAbstract:Leflunomide, a novel immunomodulatory drug, has two biochemical activities: inhibition of tyrosine phosphorylation and inhibition of pyrimidine nucleotide synthesis. In the present study, we first showed that A77 1726 [N-(4-trifluoromethylphenyl-2-cyano-3-hydroxycrotoamide)], the active metabolite of Leflunomide, was more effective at inhibiting the tyrosine kinase activity of platelet-derived growth factor (PDGF) receptor than that of epidermal growth factor (EGF) receptor, and had no effect on the tyrosine kinase activity of the fibroblast growth factor receptor. In the presence of exogenous uridine, A77 1726 was more effective at inhibiting the PDGF-stimulated proliferation of PDGF receptor-overexpressing C6 glioma than the EGF-stimulated proliferation of EGF receptor-overexpressing A431 cells. In vivo studies demonstrated that Leflunomide treatment strongly inhibited the growth of the C6 glioma but had only a modest effect on the growth of the A431 tumor. Uridine co-administered with Leflunomide did not reverse the antitumor activity of Leflunomide on C6 and A431 tumors significantly. Quantitation of nucleotide levels in the tumor tissue revealed that Leflunomide treatment significantly reduced pyrimidine nucleotide levels in the fast-growing C6 glioma but had no effect on the relatively slow-growing A431 tumor. Whereas uridine co-administration normalized pyrimidine nucleotide levels, it had minimal effects on the antitumor activity of Leflunomide in both tumor models. Immunohistochemical analysis revealed that Leflunomide treatment significantly reduced the number of proliferating cell nuclear antigen-positive cells in C6 glioma, and that uridine only partially reversed this inhibition. These results collectively suggest that the in vivo antitumor effect of Leflunomide is largely independent of its inhibitory effect on pyrimidine nucleotide synthesis. The possibility that Leflunomide exerts its antitumor activity by inhibition of tyrosine phosphorylation or by a yet unidentified mode of action is discussed.
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novel mechanism of inhibition of cytomegalovirus by the experimental immunosuppressive agent Leflunomide
Transplantation, 1999Co-Authors: W J Waldman, James W Williams, Deborah A Knight, Nell S Lurain, Daniel M Miller, Daniel D Sedmak, Anita S. ChongAbstract:Background. Despite progress in antiviral chemotherapy, cytomegalovirus (CMV) remains a major cause of morbidity and mortality among pharmacologically immunosuppressed organ transplant recipients, frequently engaging the clinician in a struggle to balance graft preservation with control of CMV disease. Leflunomide, an inhibitor of protein kinase activity and pyrimidine synthesis, is an experimental immunosuppressive agent effective against acute and chronic allograft rejection in animal models. Because a number of CMV proteins are known to be phosphorylated, we tested the hypothesis that this agent might exert inhibitory activity against CMV. Methods and results. Plaque assays demonstrated dramatic dose-dependent attenuation of production of multiple clinical CMV isolates in Leflunomide-treated human fibroblasts and endothelial cells, common targets for CMV infection in vivo. As shown by Northern blot analysis and immunohistochemical staining, Leflunomide neither interferes with transcription of immediate early or late viral genes, nor with expression of corresponding proteins. CMV-specific DNA dot blots and biochemical enzyme assays indicated that, in contrast to currently approved anti-CMV drugs, Leflunomide exerts no inhibitory effect on the accumulation of viral DNA in infected cells, or on viral DNA polymerase activity. Rather, as visualized by transmission electron microscopy, this agent appears to act at a late stage in virion assembly by preventing tegument acquisition by viral nucleocapsids. Finally we have demonstrated equivalent inhibitory activity of Leflunomide against multi-drug-resistant CMV isolates. Conclusions. These findings imply that Leflunomide, an effective immunosuppressive agent, shows potential to concurrently attenuate a major complication of immunosuppression, CMV disease, by a novel mechanism of antiviral activity.
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inhibition of cytomegalovirus in vitro and in vivo by the experimental immunosuppressive agent Leflunomide
Intervirology, 1999Co-Authors: James W Waldman, James W Williams, Deborah A Knight, Leonard Blinder, Jikun Shen, Nell S Lurain, Daniel M Miller, Daniel D Sedmak, Anita S. ChongAbstract:Despite progress in antiviral chemotherapy, cytomegalovirus (CMV) remains a major cause of morbidity and mortality among pharmacologically immunosuppressed transplant recipients, frequently engaging the clinician in a struggle to balance graft preservation with control of CMV disease. Leflunomide, an inhibitor of protein kinase activity and pyrimidine synthesis, is an experimental immunosuppressive agent effective against acute and chronic rejection in animal models. Herein we summarize our recent studies demonstrating that Leflunomide inhibits the production of multiple clinical CMV isolates (including multi-drug-resistant virus) in both human fibroblasts and endothelial cells. In contrast to all other anti-CMV drugs currently in use, Leflunomide does not inhibit viral DNA synthesis, but rather appears to interfere with virion assembly. Finally, preliminary studies in a rat model suggest that this agent reduces viral load in vivo. These findings imply that Leflunomide, an effective immunosuppressive agent, shows potential to concurrently attenuate a major complication of immunosuppression, CMV disease, by a novel mechanism of antiviral activity.
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inhibition of jak3 and stat6 tyrosine phosphorylation by the immunosuppressive drug Leflunomide leads to a block in igg1 production
Journal of Immunology, 1998Co-Authors: Karyn F Siemasko, Hansmarti Jack, Haihua Gong, James W Williams, Anita S. Chong, Aliso FinnegaAbstract:Leflunomide is an immunosuppressive drug capable of inhibiting T and B cell responses in vivo. A number of studies demonstrate that Leflunomide functions both as a pyrimidine synthesis inhibitor and as a tyrosine kinase inhibitor. We previously reported that Leflunomide inhibits LPS-stimulated B cell proliferation, cell cycle progression, and IgM secretion. This inhibition can be reversed by the addition of exogenous uridine, suggesting that Leflunomide functions as a pyrimidine synthesis inhibitor in B cells. We report here that while the addition of uridine restored proliferation and IgM secretion to Leflunomide-treated LPS-stimulated B cells, as determined by metabolic labeling and immunoprecipitation, it did not completely restore secretion of IgG Ab. We hypothesized that Leflunomide inhibits LPS-induced IgG secretion by inhibiting tyrosine kinase activity required for isotype switch. We tested this hypothesis in a well-defined model of isotype switch, LPS plus IL-4 induction of IgG1. Leflunomide inhibited IgG1 secretion in this model in a dose-dependent manner. The signal transduction pathway utilized by IL-4 to induce IgG1 involves tyrosine phosphorylation of the IL-4 receptor, JAK1, JAK3, and STAT6 proteins induced by IL-4 binding to the IL-4R. Leflunomide diminished the tyrosine phosphorylation of JAK3 and STAT6 in the absence or presence of uridine. In gel mobility shift studies, STAT6 binding to the STAT6 DNA binding site in the IgG1 promoter decreased in the presence of Leflunomide or Leflunomide plus uridine. Taken together, these data suggest that Leflunomide acts as a tyrosine kinase inhibitor to block IgG1 production.
James W Williams - One of the best experts on this subject based on the ideXlab platform.
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Leflunomide for polyomavirus type bk nephropathy
The New England Journal of Medicine, 2005Co-Authors: James W Williams, Preston Foster, Daniel L Gillen, Basit Javaid, Robert C Harland, Marc R Garfinkel, Walter J Atwood, Predeep V Kadambi, Richard J Thistlewaite, Michael J MillisAbstract:To the Editor: Polyomavirus type BK nephropathy is an aggressively destructive disease occurring in up to 8 percent of patients with renal allografts,1,2 with rates of graft loss within one year of 30 to 65 percent.3,4 There is no therapy with proven efficacy. Leflunomide (Arava), approved for the treatment of rheumatoid arthritis, is an immunosuppressive drug, yet its active metabolite, A77 1726, has substantial antiviral activity in vitro and in animals.5 From July 2001 to April 2004, we used Leflunomide as the initial antiviral therapy in 17 patients with biopsy-proven BK nephropathy. All but one of these patients . . .
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inhibition of herpes simplex virus type 1 by the experimental immunosuppressive agent Leflunomide
Transplantation, 2001Co-Authors: Deborah A Knight, James W Williams, Anita S. Chong, Ashley Quintin Hejmanowski, Julie E Dierksheide, James W WaldmanAbstract:Background. Despite advances in antiviral chemotherapy, herpes simplex virus type 1 (HSV-1), continues to complicate the clinical course of many allograft recipients. We have previously demonstrated that the experimental immunosuppressive agent Leflunomide inhibits production of cytomegalovirus by interference with virion assembly. We test the hypothesis that this agent exerts similar antiviral activity against HSV-1 Methods and Results. Plaque assay of virus yield from endothelial or Vero cells after inoculation with each of four clinical HSV-1 isolates demonstrated a dose-dependent reduction of virus production in the presence of pharmacologic concentrations of A77 1726, the active metabolite of Leflunomide. DNA dot blot and biochemical assay of viral DNA polymerase activity indicated that A77 does not inhibit viral DNA synthesis. Rather, as visualized by transmission electron microscopic method, this agent seems to disrupt virion assembly by preventing nucleocapsid tegumentation. Conclusions. These findings, in demonstrating that Leflunomide exerts antiviral activity against HSV-1 by mechanisms similar to those we have previously shown with cytomegalovirus, imply that this agent may possess broad spectrum activity against other herpesviruses.
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in vitro and in vivo antitumor activity of a novel immunomodulatory drug Leflunomide mechanisms of action
Biochemical Pharmacology, 1999Co-Authors: Jikun Shen, James W Williams, Leonard Blinder, Julian W Mall, Jonathan Myers, Wanyun Huang, Theodore J Saclarides, Anita S. ChongAbstract:Leflunomide, a novel immunomodulatory drug, has two biochemical activities: inhibition of tyrosine phosphorylation and inhibition of pyrimidine nucleotide synthesis. In the present study, we first showed that A77 1726 [N-(4-trifluoromethylphenyl-2-cyano-3-hydroxycrotoamide)], the active metabolite of Leflunomide, was more effective at inhibiting the tyrosine kinase activity of platelet-derived growth factor (PDGF) receptor than that of epidermal growth factor (EGF) receptor, and had no effect on the tyrosine kinase activity of the fibroblast growth factor receptor. In the presence of exogenous uridine, A77 1726 was more effective at inhibiting the PDGF-stimulated proliferation of PDGF receptor-overexpressing C6 glioma than the EGF-stimulated proliferation of EGF receptor-overexpressing A431 cells. In vivo studies demonstrated that Leflunomide treatment strongly inhibited the growth of the C6 glioma but had only a modest effect on the growth of the A431 tumor. Uridine co-administered with Leflunomide did not reverse the antitumor activity of Leflunomide on C6 and A431 tumors significantly. Quantitation of nucleotide levels in the tumor tissue revealed that Leflunomide treatment significantly reduced pyrimidine nucleotide levels in the fast-growing C6 glioma but had no effect on the relatively slow-growing A431 tumor. Whereas uridine co-administration normalized pyrimidine nucleotide levels, it had minimal effects on the antitumor activity of Leflunomide in both tumor models. Immunohistochemical analysis revealed that Leflunomide treatment significantly reduced the number of proliferating cell nuclear antigen-positive cells in C6 glioma, and that uridine only partially reversed this inhibition. These results collectively suggest that the in vivo antitumor effect of Leflunomide is largely independent of its inhibitory effect on pyrimidine nucleotide synthesis. The possibility that Leflunomide exerts its antitumor activity by inhibition of tyrosine phosphorylation or by a yet unidentified mode of action is discussed.
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novel mechanism of inhibition of cytomegalovirus by the experimental immunosuppressive agent Leflunomide
Transplantation, 1999Co-Authors: W J Waldman, James W Williams, Deborah A Knight, Nell S Lurain, Daniel M Miller, Daniel D Sedmak, Anita S. ChongAbstract:Background. Despite progress in antiviral chemotherapy, cytomegalovirus (CMV) remains a major cause of morbidity and mortality among pharmacologically immunosuppressed organ transplant recipients, frequently engaging the clinician in a struggle to balance graft preservation with control of CMV disease. Leflunomide, an inhibitor of protein kinase activity and pyrimidine synthesis, is an experimental immunosuppressive agent effective against acute and chronic allograft rejection in animal models. Because a number of CMV proteins are known to be phosphorylated, we tested the hypothesis that this agent might exert inhibitory activity against CMV. Methods and results. Plaque assays demonstrated dramatic dose-dependent attenuation of production of multiple clinical CMV isolates in Leflunomide-treated human fibroblasts and endothelial cells, common targets for CMV infection in vivo. As shown by Northern blot analysis and immunohistochemical staining, Leflunomide neither interferes with transcription of immediate early or late viral genes, nor with expression of corresponding proteins. CMV-specific DNA dot blots and biochemical enzyme assays indicated that, in contrast to currently approved anti-CMV drugs, Leflunomide exerts no inhibitory effect on the accumulation of viral DNA in infected cells, or on viral DNA polymerase activity. Rather, as visualized by transmission electron microscopy, this agent appears to act at a late stage in virion assembly by preventing tegument acquisition by viral nucleocapsids. Finally we have demonstrated equivalent inhibitory activity of Leflunomide against multi-drug-resistant CMV isolates. Conclusions. These findings imply that Leflunomide, an effective immunosuppressive agent, shows potential to concurrently attenuate a major complication of immunosuppression, CMV disease, by a novel mechanism of antiviral activity.
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inhibition of cytomegalovirus in vitro and in vivo by the experimental immunosuppressive agent Leflunomide
Intervirology, 1999Co-Authors: James W Waldman, James W Williams, Deborah A Knight, Leonard Blinder, Jikun Shen, Nell S Lurain, Daniel M Miller, Daniel D Sedmak, Anita S. ChongAbstract:Despite progress in antiviral chemotherapy, cytomegalovirus (CMV) remains a major cause of morbidity and mortality among pharmacologically immunosuppressed transplant recipients, frequently engaging the clinician in a struggle to balance graft preservation with control of CMV disease. Leflunomide, an inhibitor of protein kinase activity and pyrimidine synthesis, is an experimental immunosuppressive agent effective against acute and chronic rejection in animal models. Herein we summarize our recent studies demonstrating that Leflunomide inhibits the production of multiple clinical CMV isolates (including multi-drug-resistant virus) in both human fibroblasts and endothelial cells. In contrast to all other anti-CMV drugs currently in use, Leflunomide does not inhibit viral DNA synthesis, but rather appears to interfere with virion assembly. Finally, preliminary studies in a rat model suggest that this agent reduces viral load in vivo. These findings imply that Leflunomide, an effective immunosuppressive agent, shows potential to concurrently attenuate a major complication of immunosuppression, CMV disease, by a novel mechanism of antiviral activity.
J T Ransom - One of the best experts on this subject based on the ideXlab platform.
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the immunosuppressant Leflunomide inhibits lymphocyte proliferation by inhibiting pyrimidine biosynthesis
Journal of Pharmacology and Experimental Therapeutics, 1995Co-Authors: H M Cherwinski, G Nakano, J M Young, R G Cohn, P Cheung, D J Webster, J P Caulfield, Y Z Xu, J T RansomAbstract:Leflunomide is a novel immunosuppressive compound that is effective in the treatment of animal models of autoimmune disease and human rheumatoid arthritis. The mechanism of action is unknown. Here we show that Leflunomide blocked 1) increases in nucleolar size and number, 2) upregulation of the nuclear protein antigens (PCNA and Ki-67), 3) increases in uridine incorporation and total RNA and DNA content, 4) cell cycle progression and 5) proliferation in mitogen-stimulated rat spleen mononuclear cells and human peripheral blood mononuclear cells (HPBMC). Exogenous uridine reversed the Leflunomide-dependent inhibition of the normal increase in total RNA and DNA content in mitogen-stimulated HPBMC and rat spleen cells. Uridine reversed the Leflunomide-dependent inhibition of cell cycle progression in stimulated rat cell cultures. Either uridine or cytidine, which can be converted to uridine by cytidine deaminase, reversed the antiproliferative effect of Leflunomide in HPBMC. Dihydroorotate accumulated in Leflunomide-treated human T-lymphoblastoid cells, suggesting that the compound inhibited the fourth enzyme in the pyrimidine biosynthetic pathway, dihydroorotate dehydrogenase. The results support the hypothesis that the in vitro effects of Leflunomide on T-lymphocytes are due to inhibition of de novo pyrimidine synthesis.
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the immunosuppressant Leflunomide inhibits lymphocyte proliferation by inhibiting pyrimidine biosynthesis
Journal of Pharmacology and Experimental Therapeutics, 1995Co-Authors: H M Cherwinski, G Nakano, J M Young, R G Cohn, P Cheung, D J Webster, J P Caulfield, J T RansomAbstract:Leflunomide is a novel immunosuppressive compound that is effective in the treatment of animal models of autoimmune disease and human rheumatoid arthritis. The mechanism of action is unknown. Here we show that Leflunomide blocked 1) increases in nucleolar size and number, 2) upregulation of the nuclear protein antigens (PCNA and Ki-67), 3) increases in uridine incorporation and total RNA and DNA content, 4) cell cycle progression and 5) proliferation in mitogen-stimulated rat spleen mononuclear cells and human peripheral blood mononuclear cells (HPBMC). Exogenous uridine reversed the Leflunomide-dependent inhibition of the normal increase in total RNA and DNA content in mitogen-stimulated HPBMC and rat spleen cells. Uridine reversed the Leflunomide-dependent inhibition of cell cycle progression in stimulated rat cell cultures. Either uridine or cytidine, which can be converted to uridine by cytidine deaminase, reversed the antiproliferative effect of Leflunomide in HPBMC. Dihydroorotate accumulated in Leflunomide-treated human T-lymphoblastoid cells, suggesting that the compound inhibited the fourth enzyme in the pyrimidine biosynthetic pathway, dihydroorotate dehydrogenase. The results support the hypothesis that the in vitro effects of Leflunomide on T-lymphocytes are due to inhibition of de novo pyrimidine synthesis.
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Leflunomide interferes with pyrimidine nucleotide biosynthesis
Inflammation Research, 1995Co-Authors: H M Cherwinski, N Byars, S J Ballaron, G Nakano, J M Young, J T RansomAbstract:Leflunomide is an anti-inflammatory and immunosuppressive agent which blocks proliferation of transformed cells and mitogen stimulated normal lymphocytes but does not block T cell signalling mechanisms at antiproliferative concentrations. These properties are consistent with a mechanism involving interference with nucleotide metabolism. Leflunomide had anti-proliferative activity against all cells tested here. The anti-proliferative activities could be reversed by addition of uridine or cytidine to the cultures although some species and cellular differences were observed. Purine nucleosides had no effect. Measurements of nucleotide pools in a human T cell line and mitogen stimulated rat spleen cells treated with Leflunomide showed that Leflunomide preferentially reduces pyrimidine nucleotide levels. These results indicate that inhibition of pyrimidine biosynthesis is responsible for the anti-proliferative effects of Leflunomide.
James W Waldman - One of the best experts on this subject based on the ideXlab platform.
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inhibition of respiratory syncytial virus in vitro and in vivo by the immunosuppressive agent Leflunomide
Antiviral Therapy, 2011Co-Authors: Melinda Dunn, Deborah A Knight, James W WaldmanAbstract:Background: Respiratory syncytial virus (RSV) is the primary cause of bronchiolitis and pneumonia in infants and young children worldwide and is often the cause of infections in bone marrow, solid organ transplant, cystic fibrosis and congenital heart disease patients, as well as respiratory tract disease in elderly adults. Treatment options are limited to ribavirin, which is only marginally effective, and passive immunoprophylaxis, which is very expensive. The immunosuppressive agent Leflunomide has been shown to exert potent antiviral activity against several herpesviruses and polyomavirus BK. In the current study we have tested the hypothesis that Leflunomide exerts antiviral activity against RSV. Methods: Human Hep-2 or small airway epithelial cells were inoculated with RSV and treated with A77 1726, the active metabolite of Leflunomide. Syncytia formation was assessed by immunohistochemical staining, and virus yield was measured by plaque assay. Cotton rats were intranasally inoculated with RSV, treated with Leflunomide by gavage, and pulmonary viral loads were measured by plaque assay of lung homogenates. Results: Phase contrast microscopy and immunohistochemical staining demonstrated profound attenuation of RSV-induced syncytia formation in infected cultures treated with A77 1726, the active metabolite of Leflunomide. Plaque assays of virus yield in RSV-inoculated cell cultures demonstrated potent, dose-dependent A77-mediated antiviral activity. Likewise, pulmonary viral loads in RSV-inoculated cotton rats were reduced by >3 log by Leflunomide compared with vehicle-treated controls, even when Leflunomide treatment was delayed until day 3 post-inoculation. Conclusions: These findings suggest promise for lefluno― mide as a convenient, orally administered addition to the growing arsenal of antiviral therapeutics. While specific antiviral mechanisms remain to be elucidated, Leflunomide shows unique bifunctional potential to both reduce viral load and, by virtue of its well-documented anti-iniflammatory activity, attenuate the destructive inflammation associated with RSV disease.
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inhibition of angiogenesis related endothelial activity by the experimental immunosuppressive agent Leflunomide
Transplantation, 2001Co-Authors: James W Waldman, Deborah A Knight, Alice A Bickerstaff, Gayle M Gordillo, Kathleen Orosz, Charles G OroszAbstract:Background. Leflunomide, an inhibitor of protein kinase activity and pyrimidine synthesis, is an experimental immunosuppressive agent effective in the prevention/control of acute and chronic rejection in animal models and currently in phase I clinical trials in human transplant recipients. This agent is also effective in the control of graft-versus-host disease, autoimmune reactions, and the growth of certain tumors. The importance of the endothelium in these disease processes led us to hypothesize that Leflunomide might act directly upon the endothelial cell (EC). Methods and Results. Assay of human EC colony formation demonstrated dose-dependent, Leflunomide-mediated inhibition of EC proliferation. In addition, the organization of EC into capillary-like networks, which occurs during 18 hr of incubation on Matrigel, was progressively disrupted with increasing concentrations of Leflunomide. Finally, fibrin-embedded transverse sections of murine aorta, which sprout numerous microvessels during an 11-day incubation, were inhibited from doing so in the presence of this agent. All drug concentrations used in these experiments were nontoxic and pharmacologically relevant, and none of these effects were reversible by exogenous uridine, implying that inhibition of these processes was not due to intracellular pyrimidine depletion. Furthermore, neither cyclosporine nor tacrolimus exerted inhibitory activity in any of the experiments described above. Conclusions. Data generated by these studies distinguish Leflunomide among immunosuppressants as uniquely capable of inhibiting angiogenesis-related endothelial functions and suggest additional mechanisms by which this agent might intervene in the diverse array of disease processes against which it has shown therapeutic potential.
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inhibition of herpes simplex virus type 1 by the experimental immunosuppressive agent Leflunomide
Transplantation, 2001Co-Authors: Deborah A Knight, James W Williams, Anita S. Chong, Ashley Quintin Hejmanowski, Julie E Dierksheide, James W WaldmanAbstract:Background. Despite advances in antiviral chemotherapy, herpes simplex virus type 1 (HSV-1), continues to complicate the clinical course of many allograft recipients. We have previously demonstrated that the experimental immunosuppressive agent Leflunomide inhibits production of cytomegalovirus by interference with virion assembly. We test the hypothesis that this agent exerts similar antiviral activity against HSV-1 Methods and Results. Plaque assay of virus yield from endothelial or Vero cells after inoculation with each of four clinical HSV-1 isolates demonstrated a dose-dependent reduction of virus production in the presence of pharmacologic concentrations of A77 1726, the active metabolite of Leflunomide. DNA dot blot and biochemical assay of viral DNA polymerase activity indicated that A77 does not inhibit viral DNA synthesis. Rather, as visualized by transmission electron microscopic method, this agent seems to disrupt virion assembly by preventing nucleocapsid tegumentation. Conclusions. These findings, in demonstrating that Leflunomide exerts antiviral activity against HSV-1 by mechanisms similar to those we have previously shown with cytomegalovirus, imply that this agent may possess broad spectrum activity against other herpesviruses.
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inhibition of cytomegalovirus in vitro and in vivo by the experimental immunosuppressive agent Leflunomide
Intervirology, 1999Co-Authors: James W Waldman, James W Williams, Deborah A Knight, Leonard Blinder, Jikun Shen, Nell S Lurain, Daniel M Miller, Daniel D Sedmak, Anita S. ChongAbstract:Despite progress in antiviral chemotherapy, cytomegalovirus (CMV) remains a major cause of morbidity and mortality among pharmacologically immunosuppressed transplant recipients, frequently engaging the clinician in a struggle to balance graft preservation with control of CMV disease. Leflunomide, an inhibitor of protein kinase activity and pyrimidine synthesis, is an experimental immunosuppressive agent effective against acute and chronic rejection in animal models. Herein we summarize our recent studies demonstrating that Leflunomide inhibits the production of multiple clinical CMV isolates (including multi-drug-resistant virus) in both human fibroblasts and endothelial cells. In contrast to all other anti-CMV drugs currently in use, Leflunomide does not inhibit viral DNA synthesis, but rather appears to interfere with virion assembly. Finally, preliminary studies in a rat model suggest that this agent reduces viral load in vivo. These findings imply that Leflunomide, an effective immunosuppressive agent, shows potential to concurrently attenuate a major complication of immunosuppression, CMV disease, by a novel mechanism of antiviral activity.
H M Cherwinski - One of the best experts on this subject based on the ideXlab platform.
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the immunosuppressant Leflunomide inhibits lymphocyte proliferation by inhibiting pyrimidine biosynthesis
Journal of Pharmacology and Experimental Therapeutics, 1995Co-Authors: H M Cherwinski, G Nakano, J M Young, R G Cohn, P Cheung, D J Webster, J P Caulfield, Y Z Xu, J T RansomAbstract:Leflunomide is a novel immunosuppressive compound that is effective in the treatment of animal models of autoimmune disease and human rheumatoid arthritis. The mechanism of action is unknown. Here we show that Leflunomide blocked 1) increases in nucleolar size and number, 2) upregulation of the nuclear protein antigens (PCNA and Ki-67), 3) increases in uridine incorporation and total RNA and DNA content, 4) cell cycle progression and 5) proliferation in mitogen-stimulated rat spleen mononuclear cells and human peripheral blood mononuclear cells (HPBMC). Exogenous uridine reversed the Leflunomide-dependent inhibition of the normal increase in total RNA and DNA content in mitogen-stimulated HPBMC and rat spleen cells. Uridine reversed the Leflunomide-dependent inhibition of cell cycle progression in stimulated rat cell cultures. Either uridine or cytidine, which can be converted to uridine by cytidine deaminase, reversed the antiproliferative effect of Leflunomide in HPBMC. Dihydroorotate accumulated in Leflunomide-treated human T-lymphoblastoid cells, suggesting that the compound inhibited the fourth enzyme in the pyrimidine biosynthetic pathway, dihydroorotate dehydrogenase. The results support the hypothesis that the in vitro effects of Leflunomide on T-lymphocytes are due to inhibition of de novo pyrimidine synthesis.
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the immunosuppressant Leflunomide inhibits lymphocyte proliferation by inhibiting pyrimidine biosynthesis
Journal of Pharmacology and Experimental Therapeutics, 1995Co-Authors: H M Cherwinski, G Nakano, J M Young, R G Cohn, P Cheung, D J Webster, J P Caulfield, J T RansomAbstract:Leflunomide is a novel immunosuppressive compound that is effective in the treatment of animal models of autoimmune disease and human rheumatoid arthritis. The mechanism of action is unknown. Here we show that Leflunomide blocked 1) increases in nucleolar size and number, 2) upregulation of the nuclear protein antigens (PCNA and Ki-67), 3) increases in uridine incorporation and total RNA and DNA content, 4) cell cycle progression and 5) proliferation in mitogen-stimulated rat spleen mononuclear cells and human peripheral blood mononuclear cells (HPBMC). Exogenous uridine reversed the Leflunomide-dependent inhibition of the normal increase in total RNA and DNA content in mitogen-stimulated HPBMC and rat spleen cells. Uridine reversed the Leflunomide-dependent inhibition of cell cycle progression in stimulated rat cell cultures. Either uridine or cytidine, which can be converted to uridine by cytidine deaminase, reversed the antiproliferative effect of Leflunomide in HPBMC. Dihydroorotate accumulated in Leflunomide-treated human T-lymphoblastoid cells, suggesting that the compound inhibited the fourth enzyme in the pyrimidine biosynthetic pathway, dihydroorotate dehydrogenase. The results support the hypothesis that the in vitro effects of Leflunomide on T-lymphocytes are due to inhibition of de novo pyrimidine synthesis.
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Leflunomide interferes with pyrimidine nucleotide biosynthesis
Inflammation Research, 1995Co-Authors: H M Cherwinski, N Byars, S J Ballaron, G Nakano, J M Young, J T RansomAbstract:Leflunomide is an anti-inflammatory and immunosuppressive agent which blocks proliferation of transformed cells and mitogen stimulated normal lymphocytes but does not block T cell signalling mechanisms at antiproliferative concentrations. These properties are consistent with a mechanism involving interference with nucleotide metabolism. Leflunomide had anti-proliferative activity against all cells tested here. The anti-proliferative activities could be reversed by addition of uridine or cytidine to the cultures although some species and cellular differences were observed. Purine nucleosides had no effect. Measurements of nucleotide pools in a human T cell line and mitogen stimulated rat spleen cells treated with Leflunomide showed that Leflunomide preferentially reduces pyrimidine nucleotide levels. These results indicate that inhibition of pyrimidine biosynthesis is responsible for the anti-proliferative effects of Leflunomide.