RESUMO
Blocking the CD40-CD154 interaction is reported to be effective for transplantation management and autoimmune disease models in rodents and nonhuman primates. However, clinical trials with anti-CD154 mAbs were halted because of high incidence of thromboembolic complications. Thus, we generated and characterized a fully human anti-CD40 mAb ASKP1240, as an alternative to anti-CD154 mAb. In vitro ASKP1240 concentration-dependently inhibited human peripheral blood mononuclear cell proliferation induced by soluble CD154. In addition, ASKP1240 did not destabilize platelet thrombi under physiological high shear conditions while mouse anti-human CD154 mAb (mu5C8) did. And ASKP1240 itself did not activate platelet and endothelial cells. In vivo administration of ASKP1240 (1 or 10 mg/kg, intravenously) to cynomolgus monkeys, weekly for 3 weeks, significantly attenuated both delayed-type hypersensitivity and specific antibody formation evoked by tetanus toxoid. The immunosuppressive effect was well correlated with the CD40 receptor saturation. Thus, these results suggest that ASKP1240 is immunosuppressive but not prothromboembolic, and as such appears to be a promising therapeutic candidate for the management of solid organ transplant rejection and autoimmune diseases therapy.
Assuntos
Anticorpos Monoclonais/imunologia , Antígenos CD40/imunologia , Imunossupressores/farmacologia , Animais , Anticorpos Monoclonais Humanizados , Citotoxicidade Celular Dependente de Anticorpos/efeitos dos fármacos , Reações Cruzadas , Citometria de Fluxo , Humanos , Hipersensibilidade Tardia/tratamento farmacológico , Hipersensibilidade Tardia/imunologia , Imunossupressores/imunologia , Macaca fascicularis , CamundongosRESUMO
A strategy for inhibiting CD40 has been considered as an alternative approach for immunosuppression because of undesirable effects of anti-CD154 monoclonal antibodies (mAbs). Previously, we demonstrated that ASKP1240, which is a fully human anti-CD40 mAb, significantly prolonged kidney and liver allograft survival in cynomolgus monkeys without causing thromboembolic complications. Herein, we evaluated the effect of ASKP1240 on pancreatic islet transplantation (PITx) in cynomolgus monkeys. Diabetes was induced by total pancreatectomy, and islet allografts were transplanted into the liver. Following PITx (8201-12 438 IEQ/kg), blood glucose levels normalized promptly in all animals. Control islet allografts were rejected within 9 days (n = 3), whereas ASKP1240 (10 mg/kg) given on postoperative days 0, 4, 7, 11 and 14 (induction treatment, n = 5) significantly prolonged graft survival time (GST) to >15, >23, 210, 250 and >608 days, respectively. When ASKP1240 (5 mg/kg) was administered weekly thereafter up to post-PITx 6 months (maintenance treatment, n = 4), GST was markedly prolonged to >96, >115, 523 and >607 days. During the ASKP1240 treatment period, both anti-donor cellular responses and development of anti-donor antibodies were abolished, and no serious adverse events were noted. ASKP1240 appears to be a promising candidate for immunosuppression in clinical PITx.
Assuntos
Anticorpos Monoclonais/uso terapêutico , Ligante de CD40/imunologia , Diabetes Mellitus Tipo 1/terapia , Sobrevivência de Enxerto , Transplante das Ilhotas Pancreáticas , Ilhotas Pancreáticas/imunologia , Animais , Anticorpos Monoclonais/farmacocinética , Ligante de CD40/antagonistas & inibidores , Ligante de CD40/metabolismo , Diabetes Mellitus Tipo 1/etiologia , Diabetes Mellitus Tipo 1/metabolismo , Humanos , Tolerância Imunológica , Ilhotas Pancreáticas/metabolismo , Macaca fascicularis , Masculino , Pancreatectomia/efeitos adversos , Distribuição Tecidual , Transplante HomólogoRESUMO
Blockade of the CD40-CD154 costimulatory signal is an attractive strategy for immunosuppression and tolerance induction in organ transplantation. Treatment with anti-CD154 monoclonal antibodies (mAbs) results in potent immunosuppression in nonhuman primates (NHPs). Despite plans for future clinical use, further development of these treatments was halted by complications. As an alternative approach, we have been focusing on the inhibition of the counter receptor, CD40 and have shown that a novel human anti-CD40 mAb, ASKP1240, markedly prolongs renal allograft survival in NHPs, although allografts eventually underwent chronic allograft nephropathy. On the basis of our previous findings that a CD40-CD154 costimulation blockade induces tolerance to hepatic, but not cardiac, allografts in rodents, we tested here our hypothesis that a blockade of CD40 by ASKP1240 allows acceptance of hepatic allografts in NHPs. A 2-week ASKP1240 induction treatment prolonged liver allograft survival in NHPs; however, the graft function deteriorated due to chronic rejection. In contrast, a 6-month ASKP1240 maintenance monotherapy efficiently suppressed both cellular and humoral alloimmune responses and prevented rejection on the hepatic allograft. No serious side effects, including thromboembolic complications, were noted in the ASKP1240-treated monkeys. We conclude that CD40 blockade by ASKP1240 would be a desirable immunosuppressant for clinical liver transplantation.
Assuntos
Anticorpos Monoclonais/imunologia , Antígenos CD40/imunologia , Transplante de Fígado , Animais , Anticorpos Monoclonais Humanizados , Humanos , Memória Imunológica , Teste de Cultura Mista de Linfócitos , Macaca fascicularis , Masculino , Transplante HomólogoRESUMO
Yap1p, a crucial transcription factor in the oxidative stress response of Saccharomyces cerevisiae, is transported in and out of the nucleus under nonstress conditions. The nuclear export step is specifically inhibited by H(2)O(2) or the thiol oxidant diamide, resulting in Yap1p nuclear accumulation and induction of transcription of its target genes. Here we provide evidence for sensing of H(2)O(2) and diamide mediated by disulfide bond formation in the C-terminal cysteine-rich region (c-CRD), which contains 3 conserved cysteines and the nuclear export signal (NES). The H(2)O(2) or diamide-induced oxidation of the c-CRD in vivo correlates with induced Yap1p nuclear localization. Both were initiated within 1 min of application of oxidative stress, before the intracellular redox status of thioredoxin and glutathione was affected. The cysteine residues in the middle region of Yap1p (n-CRD) are required for prolonged nuclear localization of Yap1p in response to H(2)O(2) and are thus also required for maximum transcriptional activity. Using mass spectrometry analysis, the H(2)O(2)-induced oxidation of the c-CRD in vitro was detected as an intramolecular disulfide linkage between the first (Cys(598)) and second (Cys(620)) cysteine residues; this linkage could be reduced by thioredoxin. In contrast, diamide induced each pair of disulfide linkage in the c-CRD, but in this case the cysteine residues in the n-CRD appeared to be dispensable for the response. Our data provide evidence for molecular mechanisms of redox signal sensing through the thiol-disulfide redox cycle coupled with the thioredoxin system in the Yap1p NES.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/metabolismo , Fatores de Transcrição/metabolismo , Núcleo Celular/metabolismo , Proteínas de Ligação a DNA/química , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Glutationa/metabolismo , Oxirredução , Transdução de Sinais , Tiorredoxinas/metabolismo , Fatores de Transcrição/químicaRESUMO
We have previously reported the clinical effect of NF therapy (NCS 5,000 units and 5-FU 500 mg intravenously; twice a week) on patients with advanced carcinoma of the digestive system. In the present study, NFO therapy (NCS 2,000 units and 5-FU 500 mg intravenously, Picibanil 1-2K. E. intramuscularly; twice a week) was applied for those patients to take advantage over NF modality. Treated were 62 patients with NFO and 48 were evaluated for its clinical effects. In comparison of NFO and NF, the antitumor effects were noted in 9 of 48 patients (18.8%) for NFO, and 2 of 27 (7.4%) for NF therapy judged by Koyama & Saito's criterion. If the Karnofsky's criterion was applied, I-A category or more were obtained in 10 of 48 patients (20.8%) for NFO and in 2 of 27 (7.4%) for NF therapy. In particular, NFO therapy resulted in the advantageous clinical effects on patients with hepatic and pancreatic carcinomas irrespective of primary or metastatic. The adverse effects of NFO were not more frequent than those of NF therapy. From these results, a new combination regimen, NFO, is thought to bring about a considerable benefit in the treatment of advanced carcinoma of the digestive organs.