Docking protein-1 in macrophage immunotherapy of gastric cancer

胃癌（GC）患者通常出现在晚期，无法进行有效治疗，导致临床预后较差。尽管胃癌在中国和世界范围内的发病率和流行率正在下降，但随着人口老龄化，胃癌的死亡率仍然很高，这是一个具有挑战性的健康问题。新的治疗方法被设计用来促进宿主对肿瘤攻击模式的免疫反应，防止免疫逃避和宿主能量。然而，在GC中，保证免疫疗法临床疗效的潜在机制，例如针对免疫检查点（PD1/PD-L1）的抗体，尚不清楚。对接蛋白-1 （DOK1）是一种细胞内适配器，可抑制致癌Ras信号传导并控制免疫受体活性。DOK1由胰岛素增敏剂（格列酮）诱导，并通过与过氧化物酶体增殖物激活受体- γ (ppar)相互作用来稳定，导致体外和体内GC生长减弱。我们最近的研究表明，DOK1抑制PD-L1的表达，促进人巨噬细胞的极化和效应功能，促进肿瘤攻击，促炎M1表型。DOK1和M1（如toll样受体）基因的改变导致胃癌患者预后不良，并且DOK1 mRNA/蛋白在肿瘤和免疫细胞区室的肿瘤进展过程中受到差异调节。因此，本研究将探索DOK1的可药物性是否可以与临床使用的免疫治疗抗体（针对PD1/PD-L1）或新型吞噬刺激剂（针对CD47）协同作用，以加强肿瘤攻击巨噬细胞反应。我们的翻译方法包括三个特定目标：1)使用生物素捕获法鉴定与DOK1结合的巨噬细胞特异性免疫受体；2)利用功能获得和CRISPR/ cas9敲除在人巨噬细胞系和原代细胞中研究DOK1免疫调节作用的细胞和分子机制；3)研究DOK1表达和巨噬细胞（髓系）浸润在GC小鼠模型和治疗患者组织中的（前）临床意义。研究结果有望深入了解DOK1在胃癌巨噬细胞中的功能机制，增强我们对癌症免疫治疗的认识，并为胃癌患者提供未来有效的治疗策略。

Gastric cancer (GC) patients usually present at an advanced stage, where effective treatment is impossible, resulting in poor clinical outcome. Although the incidence and prevalence of GC in China and world-wide is declining, the mortality from GC in an ageing population is still high and a challenging health problem. Novel therapies were designed to boost the host immune response towards a tumor-attacking mode, preventing immune evasion and host anergy. However, the underlying mechanisms that warrant clinical efficacy of immunotherapies, e.g. such as antibodies against immune checkpoints (PD1/PD-L1), in GC are unknown. Docking protein-1 (DOK1) is an intracellular adapter which inhibits oncogenic Ras signaling and controls immune receptor activities. DOK1 is induced by insulin sensitizers (glitazones) and stabilized by interaction with peroxisome proliferator-activated receptor-gamma (PPARg), resulting in attenuated GC growth in vitro and in vivo. We have recently shown that DOK1 inhibits PD-L1 expression and promotes polarization and effector functions of human macrophages towards the tumor-attacking, pro-inflammatory M1 phenotype. Alterations in DOK1 and M1 (e.g. toll-like receptors) genes conferred poor prognosis in GC patients, and DOK1 mRNA/protein was differentially regulated during tumor progression in tumor and immune cell compartments. The present proposal study shall therefore explore whether the drugability of DOK1 can be exploited to synergize with clinically-in-use immunotherapeutic antibodies (against PD1/PD-L1) or novel phagocytosis stimulating agents (against CD47) to enforce a tumor-attacking macrophage response.Our translational approach comprises three specific aims: 1) to identify macrophage-specific immune receptor(s) bound by DOK1 using biotin capture assay; 2) to investigate the cellular and molecular mechanisms underlying the immune-regulatory role of DOK1 using gain-of-function and CRISPR/Cas9-knock-out in human macrophage lines and primary cells; 3) to study the (pre)clinical significance of DOK1 expression and macrophage (myeloid lineage) infiltration in GC mouse models and patients’ tissues under therapy. Results are expected to deliver mechanistic insight into the function of DOK1 in macrophages of GC, strengthen our knowledge of cancer immunotherapy and facilitate future effective therapeutic strategies for GC patients.