免疫抑制受体IGSF9抑制T细胞增殖促进小细胞肺癌免疫逃避机制的研究

The immune therapy mediated by antibody blocking immune checkpoint has been successful in treating melanoma, non-small cell lung cancer, but has not shown clinical benefits for treating small cell lung cancer, which indicates that small cell lung cancer exploits unique escape mechanisms. Immune inhibitory receptor IGSF9 mainly expresses on central nervous system during embryonic development, however, our previous data showed that IGSF9 highly expressed in small cell lung cancer tissue samples. The role of retro-rendered IGSF9 in small cell lung cancer has not been reported. In this study, specific IGSF9 blocking antibody will be screened by IGSF9 reporter cell, and this antibody will be injected into subcutaneous xenograft, migration and PDX mice models, which proves that antibody blocking IGSF9 signal pathway can rescue T cell proliferation to inhibit the growth and migration of small cell lung cancer. Next, the paracrine pathway of IGSF9 inhibiting T cell proliferation will be investigated by RNA-seq, Co-IP, and so on. The mechanism by which IGSF9 promotes invasion and metastasis will be explored by constructing different deletions of “Ig” and “Fn” domain. In brief, this study is intended to reveal the role of IGSF9 in immune evasion and migration, and to provide a new target for immunotherapy of small cell lung cancer.

抗体封闭免疫检查点介导的免疫治疗在黑色素瘤、非小细胞肺癌中疗效显著，但对小细胞肺癌未见疗效，表明小细胞肺癌有其独特的免疫逃避机制。免疫抑制受体IGSF9表达在胚胎时期中枢神经组织，而本课题前期结果表明：小细胞肺癌组织中IGSF9蛋白表达较高。“复古”表达的IGSF9蛋白在小细胞肺癌发生、发展中的作用未见报道。本课题拟利用IGSF9报告细胞，筛选IGSF9特异阻断性抗体；应用该抗体处理小鼠皮下成瘤、侵袭转移以及PDX模型，求证抗体阻断IGSF9信号通路能够恢复T细胞增殖，进而抑制小细胞肺癌生长和侵袭转移；拟应用免疫共沉淀、RNA-seq等方法，阐明IGSF9通过旁分泌途径抑制T细胞增殖的信号通路；拟构建IGSF9蛋白胞外Ig和Fn结构域缺失体细胞，探讨IGSF9促进小细胞肺癌侵袭转移的分子机制。本课题拟揭示IGSF9蛋白在小细胞肺癌免疫逃避及侵袭转移中的作用，为小细胞肺癌免疫治疗提供新靶点。

以PD-1/PD-L1单克隆抗体为代表的肿瘤免疫药物，让肿瘤患者看到了治愈的希望。临床数据显示大约20%的肿瘤患者对PD-1/PD-L1单克隆抗体有应答，表明80%的肿瘤患者有未知的免疫逃避机制。免疫抑制受体超家族成员9（IGSF9）广泛表达在各肿瘤细胞表面，但是其在肿瘤发生、发展以及免疫逃避中的作用，未见报道。本项目阐明了IGSF9抑制T细胞增殖，促进肿瘤免疫逃避的机制，主要研究内容和结果如下：（1）免疫组化和流式细胞仪结果表明，IGSF9广泛表达在各种肿瘤细胞、肿瘤浸润T细胞、B细胞以及髓系细胞；IGSF9表达水平越高，非小细胞肺癌、肝癌、结肠癌和肾癌的恶性程度越严重；（2）与野生型小鼠相比，igsf9基因敲除鼠显著抑制肿瘤生长，而且肿瘤浸润CD3+T细胞水平显著升高；在肿瘤细胞中过表达igsf9，显著抑制肿瘤生长，而且肿瘤浸润CD3+T细胞水平显著降低；（3）制备IGSF9-ECD-Fc和IGSF9-ECD-His蛋白，与T细胞共培养，能够显著抑制T细胞增殖；在MC38细胞中过表达IGSF9-ECD，显著促进肿瘤生长，而且肿瘤浸润T细胞水平显著降低；消除小鼠体内T细胞，IGSF9或者IGSF9-ECD不再促进肿瘤生长；（4）IGSF9-ECD-Fc和IGSF9-ECD-His蛋白能够和T细胞相结合，而且该结合能够显著抑制T细胞的增殖和活性；（5）IGSF9促进肿瘤侵袭转移；（6）应用IGSF9-ECD-His蛋白免疫Balb/c小鼠，制备靶向IGSF9的单克隆抗体，该抗体能够抑制肿瘤生长，与PD-1抗体联用，协同效应显著；（7）消除抗体介导的ADCC和ADCP效应之后，anti-IGSF9仍然能够抑制肿瘤的生长和转移。.综上所述，IGSF9是一个肿瘤特异免疫检查点分子，抑制T细胞增殖，促进肿瘤细胞免疫逃避；靶向IGSF9的单克隆抗体，有望为肿瘤患者带来新的疗法。