Abstract PO-096: The synaptic protein Netrin G1 ligand (NGL-1) modulates tumorigenesis and immunosuppression in pancreatic cancer
Authors: Costa Debora Barbosa Vendramini , Francescone Ralph , Franco-Barraza Janusz , Luong Tiffany , Steele Nina , Allen Benjamin , Pasca di Magliano Marina , Ogier Charline , Astsaturov Igor , Cai Kathy Q. , Klein-Szanto Andres J. , Wang Huamin , Campbell Kerry , Cukierman Edna 
Published: November 2021

Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest types of cancer, with a 5-year survival of 10%. A major feature of PDAC is the presence of a dense fibrous stroma, due to the expansion of cancer associated fibroblasts (CAFs) and their extracellular matrix. This unique environment represents a challenge for therapies as it promotes immunosuppression, limits access to nutrients, and excludes or inactivates antitumor immune cells. Recently, we identified the ectopic expression of the neuronal protein Netrin G1 Ligand (NGL-1) in PDAC tissue, including its novel expression in immune cells and CAFs. However, the roles of NGL-1 in the tumor microenvironment (TME) of PDAC and in immune cell function are unknown and warranted further investigation. The contribution of NGL-1 to PDAC tumorigenesis was assessed by measuring the expression of NGL-1 in different models of PDAC and by orthotopically injecting PDAC cells in wild type (WT) or NGL-1 full body knockout mice (KO). Using our in vitro 3D system we evaluated if NGL-1+ CAFs, compared to NGL-1 knockdown (KD) CAFs, produced less immunosuppressive factors and were able to rescue PDAC cell survival under nutrient deprivation. For NGL-1 dependent immune cell functions we isolated naïve immune cells from WT and KO mice and performed ex-vivo functional assays. NGL-1 expression in fibroblasts correlated with disease development in different models of PDAC, and myeloid, T and NK cells from tumor bearing mice tended to overexpress NGL-1 when compared with cells from naïve mice. Accordingly, NGL-1 KO mice orthotopically injected with PDAC cells developed smaller tumors with decreased secretion of immunosuppressive factors, increased presence of CD8+ T cells and CD4+ T cells expressing less pro-tumor markers. Single cell RNA sequencing data from tumors from KO mice showed downregulation of pro-tumor genes in different cell populations, with the fibroblastic populations differing between WT and KO mice. In order to evaluate the contribution of the immune system for tumorigenesis in WT and KO mice, we performed bone marrow chimeras and depletion of specific immune cells. Functionally, CD8+ and CD4+ T cells from KO mice proliferated more when stimulated in vitro, suggesting that NGL-1 could represent a functional brake for T cells, inhibiting their anti-tumor capacity. The lack of NGL-1 in stimulated bone marrow-derived macrophages decreased pro-inflammatory cytokine secretion, further suggesting a functional role for NGL-1 in myeloid cells. Of note, NGL-1 KD CAFs did not support PDAC cell survival in vitro and produced less immunosuppressive cytokines, which was phenocopied by the treatment with a peptide targeting NGL-1. Translationally, we assessed the overall survival of 140 PDAC patients according to NGL-1 expression in the TME, where low expression of NGL-1 in CAFs and immune cells correlated with better survival of PDAC patients. Overall, this suggests NGL-1 as potential new target in PDAC, that could be manipulated in different compartments in pancreatic cancer.

Citation Format: Debora Barbosa Vendramini Costa, Ralph Francescone, Janusz Franco-Barraza, Tiffany Luong, Nina Steele, Benjamin Allen, Marina Pasca di Magliano, Charline Ogier, Igor Astsaturov, Kathy Q. Cai, Andres J. Klein-Szanto, Huamin Wang, Kerry Campbell, Edna Cukierman. The synaptic protein Netrin G1 ligand (NGL-1) modulates tumorigenesis and immunosuppression in pancreatic cancer [abstract]. In: Proceedings of the AACR Virtual Special Conference on Pancreatic Cancer; 2021 Sep 29-30. Philadelphia (PA): AACR; Cancer Res 2021;81(22 Suppl):Abstract nr PO-096.