IMMUNOSUPPRESSION - PRE-CLINICAL AGENTS

T. Oura¹, K. Yamashita¹, T. Suzuki¹, D. Fukumori¹, M. Watanabe¹, H. Gentaro¹, K. Wakayama¹, T. Miura², K. Okimura², K. Maeta², H. Haga³, K. Kubota³, A. Shimizu⁴, M. Taniguchi¹, T. Shimamura¹, F. Sakai⁵, H. Furukawa¹, S. Todo^1
1First Department Of General Surgery, Hokkaido University, Sapporo/JAPAN, ², Kyowa Hakko Kirin Co., Shizuoka/JAPAN, ³Department Of Pathology, Hokkaido University, Sapporo/JAPAN, ⁴, Nippon medical school, Tokyo/JAPAN, ⁵, Astellas Pharma Inc., Tsukuba/JAPAN

Body: Introduction: Previously, we have reported our preliminary data that 2-week induction treatment with ASKP1240 (4D11), a fully human anti-CD40 mAb, prolonged hepatic allograft survival in cynomolgus monkeys, however, they finally failed due to chronic rejection. On the other hand, 6-month maintenance 4D11 treatment markedly prolonged allograft survival without a sign of rejection, and one graft is still surviving for over 800 days. To investigate the underlying mechanism of 4D11 mediated graft acceptance, immunological features of this long-term surviving liver recipient were assessed. Methods: The cynomolgus monkey was transplanted liver allograft from AB blood-type identical donor (MLR-SI: 10.6). 4D11 at 10 mg/kg was given i.v. on days 0, 4, 7, 11 and 14. The treatment was maintained weekly at 5 mg/kg up-to 6 months, and was discontinued thereafter. No other immunosuppressant was given. Cellular and humoral allo-immune responses, peripheral T cell phenotypes and biopsy obtained graft histolopathology were monitored (monthly up-to 16 months, and every 3-6 month thereafter) and assessed. Results: The animal, surviving for more than 620 days after cessation of 4D11, experienced a severe ischemia/reperfusion injury (IRI) shortly after liver transplantation (LTx) and repetitive cholangitis later on (Fig. 1a). Histopathology of graft biopsy specimens basically reflected clinical events (IRI and cholangitis), but importantly, no sign of acute or chronic rejection was evident during the treatment course or even after 4D11 cessation. Cellular allo-immune responses as assessed by IFN-gamma ELISPOT assay (both direct and indirect pathways) were completely abrogated throughout the post-operative course. When examined on day 389, frequency of peripheral donor-reactive IFN-gamma producing cells was still minimal, although response was noted against 3^rd-party antigens and PHA stimulation (Fig. 1b). Peripheral donor-specific CD4⁺CD154⁺ T cells were suppressed (Pre LTx: 3.3% and Post LTx: 0.49±0.5%), and ImmuKnow® ATP values were not affected (Pre LTx: 70 ng/ml and Post LTx: 69±51 ng/ml) throughout the course. Anti-donor antibody formation (IgM and IgG) was completely abolished even after 4D11 cessation, and anti-4D11 antibody was not detected during the treatment. Effector/memory subset in the periphery, CD4⁺ (Pre LTx: 100 cells/ml and Post LTx: 99±48 cells/ml) and CD8⁺ (Pre LTx: 1715 cells/ml and Post LTx: 1141±696 cells/ml), did not show an increase. Peripheral CD4⁺Foxp3⁺ T cells was not augmented (Pre LTx: 2.4% and Post LTx: 3.1±3%). Further, intragraft CD4⁺Foxp3⁺ expression did not differ from that of acutely rejected allografts. Conclusion: The long-term liver allograft acceptance by 4D11 was associated with a strong suppression of both cellular and humoral allo-immune responses, while contribution of immuno-regulatory mechanism was not evident.

Disclosure: All authors have declared no conflicts of interest.