Hedgehog signaling induces PD-L1 expression and tumor cell proliferation in gastric cancer

// Jayati Chakrabarti 1 , Loryn Holokai 2 , LiJyun Syu 3 , Nina G. Steele 4 , Julie Chang 5 , Jiang Wang 6 , Syed Ahmed 7 , Andrzej Dlugosz 3, 8 and Yana Zavros 1 1 Department of Pharmacology and Systems Physiology, University of Cincinnati, Cincinnati, OH, USA 2 Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati, OH, USA 3 Department of Dermatology, University of Michigan, Ann Arbor, MI, USA 4 Division of Developmental Biology, University of Michigan, Ann Arbor, MI, USA 5 Department of Biomedical Engineering, University of Cincinnati, Cincinnati, OH, USA 6 Department of Pathology and Laboratory Medicine, University of Cincinnati College of Medicine, Cincinnati, OH, USA 7 Department of Surgery, University of Cincinnati Cancer Institute, Cincinnati, OH, USA 8 Department of Cell and Developmental Biology, University of Michigan, Ann Arbor, MI, USA Correspondence to: Yana Zavros, email: yana.zavros@uc.edu Keywords: gastric cancer organoids, PD-1; cytotoxic T lymphocytes; dendritic cells Received: August 29, 2018      Accepted: November 26, 2018      Published: December 21, 2018 ABSTRACT Tumor cells expressing programmed cell death ligand 1 (PD-L1) interact with PD-1 on CD8+ cytotoxic T lymphocytes (CTLs) to inhibit CTL effector function. In gastric cancer, the mechanism regulating PD-L1 is unclear. The Hedgehog (Hh) signaling pathway is reactivated in various cancers including gastric. Here we tested the hypothesis that Hh-induced PD-L1 inactivates effector T cell function and allows gastric cancer cell proliferation. Mouse organoids were generated from tumors of a triple-transgenic mouse model engineered to express an activated GLI2 allele, GLI2A, in Lgr5-expressing stem cells, (mTGOs) or normal mouse stomachs (mGOs). Bone marrow-derived dendritic cells (DCs) were pulsed with conditioned media collected from normal (mGO CM ) or cancer (mTGO CM ) organoids. Pulsed DCs and CTLs were then co-cultured with either mGOs or mTGOs in the presence of PD-L1 neutralizing antibody (PD-L1Ab). Human-derived gastric cancer organoids (huTGOs) were used in drug and xenograft assays. Hh/Gli inhibitor, GANT-61 significantly reduced the expression of PD-L1 and tumor cell proliferation both in vivo and in vitro . PD-L1Ab treatment induced tumor cell apoptosis in mTGO/immune cell co-cultures. GANT-61 treatment sensitized huTGOs to standard-of-care chemotherapeutic drugs both in vivo and in vitro . Thus, Hh signaling mediates PD-L1 expression in gastric cancer cells and subsequently promotes tumor proliferation.