Glyco-immune checkpoint receptors, molecules that inhibit immune cell activity following binding to glycosylated cell-surface antigens, are emerging as attractive targets for cancer immunotherapy. Defining biologically relevant ligands that bind and activate such receptors, however, has historically been a significant challenge. Here, we present a CRISPRi genomic screening strategy that allowed unbiased identification of the key genes required for cell-surface presentation of glycan ligands on leukemia cells that bind the glyco-immune checkpoint receptors Siglec-7 and Siglec-9. This approach revealed a selective interaction between Siglec-7 and the mucin-type glycoprotein CD43. Further work identified a specific N-terminal glycopeptide region of CD43 containing clusters of disialylated O-glycan tetrasaccharides that form specific Siglec-7 binding motifs. Knockout or blockade of CD43 in leukemia cells relieves Siglec-7-mediated inhibition of immune killing activity. This work identifies a potential target for immune checkpoint blockade therapy and represents a generalizable approach to dissection of glycan–receptor interactions in living cells.

糖免疫检查点受体与糖基化细胞表面抗原结合后抑制免疫细胞活性的分子正在成为癌症免疫治疗的有吸引力的靶点。然而，定义结合和激活此类受体的生物学相关配体在历史上一直是一个重大挑战。在这里，我们提出了一种 CRISPRi 基因组筛选策略，该策略允许无偏见地识别与糖免疫检查点受体 Siglec-7 和 Siglec-9 结合的白血病细胞上聚糖配体的细胞表面呈递所需的关键基因。这种方法揭示了 Siglec-7 和粘蛋白型糖蛋白 CD43 之间的选择性相互作用。进一步的工作确定了 CD43 的特定 N 端糖肽区域，其中包含形成特定 Siglec-7 结合基序的二唾液酸化 O-聚糖四糖簇。敲除或阻断白血病细胞中的 CD43 可缓解 Siglec-7 介导的免疫杀伤活性抑制。这项工作确定了免疫检查点阻断疗法的潜在目标，并代表了一种剖析活细胞中聚糖-受体相互作用的通用方法。