No abstract

Introduction: Wnt/β-catenin pathway regulates many biological processes such as cell proliferation, stem cell renewal and tissue differentiation. Activation of Wnt/β-catenin pathway has been reported in many cancers. Tankyrase, one of the positive regulators of Wnt/β-catenin pathway, induces degradation of Axin, a negative regulator of Wnt/β-catenin pathway. Therefore, tankyrase inhibition stabilizes Axin protein and inhibits Wnt/β-catenin pathway, leading to exert antitumor effect. Recently, the role of Wnt/β-catenin pathway in immunotherapy has also gained attention; aberrant activation of Wnt/β-catenin pathway causes inactivation of dendritic cells and suppresses chemokine production resulting in paucity of CD8 T+ cells in tumor tissue. Although immune checkpoint inhibitors (ICIs), such as anti-CTLA-4 and anti-PD-1/PD-L1 antibodies have been successfully approved in some cancer therapies, some patients are still resistant to ICIs. One of the main causes is considered to be immune-desert tumor microenvironment, characterized by the absence of infiltrating lymphocytes including CD8+ T cells, cytotoxic effector cells for immunotherapy. Thus, Wnt/β-catenin pathway inhibition may overcome the resistance of anti-PD-1/PD-L1 therapy by attracting CD8+ T cells into tumor. K-476 is a highly potent and selective tankyrase inhibitor synthesized in house. In this study, we investigated whether K-476 enhanced the antitumor effect of anti-PD-L1 antibody. Furthermore, since tankyrase inhibitors are reported to have a potential gastrointestinal toxicity, we also evaluated the toxicity in mice administrated with K-476. Methods and results: Antitumor effect of K-476 combined with anti-PD-L1 antibody was evaluated in B16-derived melanoma bearing mice. Although K-476 monotherapy did not show antitumor effect, a combination with anti-PD-L1 antibody demonstrated a significant antitumor effect at doses from 50 to 200 mg/kg compared to anti-PD-L1 antibody alone, and the magnitude of the effect was comparable among the doses tested. Axin stabilization was observed in tumors after administration of K-476, suggesting that the effect was exerted through inhibition of Wnt/β-catenin pathway. The expression of chemokines, Ccl3 and Ccl4 which attract CD8+ T cells, was also upregulated. In addition, significant increase of CD8+ T cells was observed in the tumor. Gastrointestinal toxicity was not observed up to 200 mg/kg of K-476 in C57BL/6J mice. These results suggested that K-476 enhanced antitumor effect of anti-PD-L1 antibody through upregulation of Ccl3 and Ccl4 production and induction of CD8+ T cells without obvious toxicity. Conclusion: K-476 could be an attractive therapeutic agent that enhances the efficacy of anti-PD-L1 antibody. Citation Format: Haruka Kinosada, Kana Kunieda, Ryoko Okada, Minami Suzuki-Imaizumi, Motoya Mie, Toshihiko Ishii, Ryuichiro Nakai. Dual pocket binding novel tankyrase inhibitor, K-476, enhances the efficacy of immune checkpoint inhibitor by attracting CD8+ T cells into tumor [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2200.

Abstract. Lafutidine, a histamine H 2 receptor antagonist, exerts gastroprotective effects in addition to gastric antisecretory activity. The gastrointestinal protective effects of lafutidine are mediated by capsaicin-sensitive neurons, where capsaicin excites neurons by opening a member of the transient receptor potential channel family (TRPV1 ] i in PC12 cells was inhibited by capsaicin and SKF96365, an inhibitor of store-operated Ca 2+ entry, and the lafutidine response was inhibited by capsaicin but not by SKF96365. In endothelial cells, lafutidine induced an increase in [Ca 2+ ] i in a SKF96365-insensitive manner. These results suggest that lafutidine stimulates Ca 2+ entry via the capsaicin-sensitive pathway but not the SKF96365-sensitive pathway. The possible role of store-operated Ca 2+ entry induced by lafutidine on gastrointestinal function is also discussed.

Kidney fibrosis is closely associated with the progression of chronic kidney disease (CKD). Furthermore, copper‐containing secretory amine oxidases, such as lysyl oxidase (LOX) and LOX‐like 1–4 (LOXL1–4), play pivotal roles in the regulation of extracellular components and facilitate fibrosis. In this study, we investigated the regulation of LOX enzymes in human tubular epithelial HK2 cells to help clarify the role of LOX enzymes in kidney fibrosis. Among 5 LOX enzymes, LOXL2 expression is abundantly expressed in HK2 cells. LOX enzymes inhibitor, β‐aminopropionitrile, suppressed transforming growth factor‐β1 (TGF‐β1)‐promoted epithelial‐to‐mesenchymal transition processes in HK2 cells, indicating that LOX enzymes are involved in TGF‐β1‐mediated fibrotic processes. Recent studies suggest that LOX enzymes are secreted into the extracellular spaces by exosomes and promote fibrotic processes. Similar to the previous reports, we observed that exosomes secreted from HK2 cells carry LOXL2 into the extracellular spaces. Furthermore, we determined that N‐glycosylation on the asparagine residues plays a key role in LOXL2 secretion. Amino acid mutations in three asparagine residues, which can be glycosylated, suppressed the secretion of mutated LOXL2. Moreover, N‐acetylglucosaminyltransferase 5, an enzyme used for the biosynthesis of β1,6N‐acetylglucosamine‐branched N‐glycans, participated in LOXL2 secretion, and the N‐glycosylation inhibitor, glucosamine hydrochloride (GS), inhibited TGF‐β1‐mediated LOXL2 secretion and fibrotic processes. Overall, TGF‐β1 promotes LOXL2 secretion and may participate in kidney fibrosis. Our results provide novel insight into the antifibrotic properties of GS that contribute to the inhibition of CKD progression.