Abstract A95: A genome-wide CRISPR screen identifies novel ligands for the Siglec family of glyco-immune checkpoint receptors

Abstract

Abstract Cancer cells are frequently characterized by extensive remodeling of their cell surface glycocalyx, the complex pattern of sugar modifications that decorate cell surface proteins. In particular, cancer cells often overexpress an unusually high density of sialic acid glycans on their plasma membrane. This modification shields tumors from immune detection through engagement of inhibitory Siglec (Sialic acid-binding immunoglobulin-type lectin) receptors, which are widely expressed in innate immune cells. Blockade of these Siglec-based immune checkpoints has shown the potential to increase immune activity against highly sialylated cancer cells. The specific glycosylated ligands that serve as counter-receptors for Siglecs, however, are not well characterized. Here we describe a strategy for better defining these immune checkpoints using a CRISPR-based genetic screening approach. A library of gRNAs was screened using FACS to identify genes whose ablation significantly reduced cell-surface binding of recombinantly expressed, fluorescently labeled Siglecs. We thereby generated a list of genes with putative roles in driving the cell-surface expression of sialic acid-containing Siglec ligands. Our screen correctly identified known genes involved in sialic acid biosynthesis, validating the specificity of this approach. Additionally, this screen revealed that knockdown of a single heavily O-glycosylated protein plasma membrane protein dramatically reduced binding of Siglec7, the Siglec receptor expressed specifically in NK cells. Follow-up work has shown that Siglec7 binds to this ligand with high affinity and that genetic blockade of this interaction potentiates NK cell killing activity in vitro. This study has thus defined a novel and specific glyco-immune interaction with potential significance in driving tumor escape from innate immune surveillance. Citation Format: Simon Wisnovsky, Carolyn Bertozzi. A genome-wide CRISPR screen identifies novel ligands for the Siglec family of glyco-immune checkpoint receptors [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2018 Nov 27-30; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(4 Suppl):Abstract nr A95.