Genetic and immunological dissection of coinhibitory crosstalks between human T cells and cancer cells

项目摘要/摘要 这项提议的目标是识别新的协同抑制检查点分子，这些分子正向工作 目前针对癌症患者的检查站，如PD-1。尽管取得了前所未有的成功 检查点阻断免疫疗法作为一种治疗多种癌症的方法， 仍然存在显著的个体间差异，从无反应到完全缓解 全身转移性病变。各种肿瘤内在参数，如肿瘤突变 负荷、新抗原负荷和错配修复途径缺陷，仅显示有限的预测性 取得良好临床结果的力量。因此，对这种肿瘤内在因素的补充，它是 貌似合理的假设是人类有迄今为止未知的协同抑制机制 目前已知的免疫检查点和一小部分有生殖系或体细胞的人类 这些途径的突变可能受益于由以下因素引发的异常强大的抗肿瘤免疫 检查站封锁免疫疗法。识别这样的正交通路将显著地 促进多种癌症患者的健康；例如，15种癌症用于 哪一种培溴利珠单抗已经被指出。目前的项目旨在解剖人类基因 人T细胞与癌细胞共抑制串扰的免疫学基础 两个具体目标。具体目的1是阐明PD-L1/L2介导的分子机制 抑制T细胞活化，不依赖PD-1。这一目标得到了初步观察的支持 遗传性完全性PD-1缺乏症患者的原代和永生化T细胞，AS 以及PD-1阴性的人T细胞系Jurkat和HuT78，都对珠蛋白-1的共同抑制有反应。 固定化PD-L1/L2。CD83被认为是PD-L1介导的免疫抑制的候选分子。 拟议的工作包括通过RNA测序和表面分析寻找更多的候选 通过RNA沉默和CRISPR引导的受体分析和功能验证 淘汰赛。特异性目的2是研究CD83在人T细胞中的免疫调节功能。 以前的研究表明，CD83既可以作为免疫抑制受体，也可以作为配体 人类T细胞，尽管其对抗性受体(S)，下游信号，并与抗肿瘤相关 豁免权基本上仍然难以捉摸。建议的工作包括CD83的生化特性 利用基因敲除和慢病毒对有无癌细胞共培养的人T细胞的信号传导 CD83基因的挽救。拟议的项目将有助于理清生化和免疫学基础。 对人类T细胞共抑制电路中癌细胞之间的串扰也有实质性的支持作用 申请者的科学投资组合，并加快向独立调查人员的发展 开创了人类遗传学癌症免疫学和免疫疗法的先河。

PROJECT SUMMARY / ABSTRACT The goal of this proposal is to identify novel coinhibitory checkpoint molecules operating orthogonally to currently targeted checkpoints in cancer patients, such as PD-1. Despite the unprecedented success of checkpoint blockade immunotherapy as a therapeutic approach against multiple types of cancer, significant inter-individual variability remains, ranging from no response to complete remission of systemically metastasized lesions. Various tumor-intrinsic parameters, such as tumor mutational burden, neoantigen burden, and mismatch repair pathway deficiencies, only show limited predictive power for favorable clinical outcomes. Therefore, complementary to such tumor-intrinsic factors, it is plausible to hypothesize that humans have hitherto unknown coinhibitory mechanisms orthogonal to currently known immune checkpoints and that a minor fraction of humans with germline or somatic mutations in such pathways could benefit from exceptionally potent antitumor immunity triggered by checkpoint blockade immunotherapy. Identification of such orthogonal pathways would significantly enhance the health of patients with multiple types of cancer; for instance, the 15 types of cancer for which pembrolizumab has been indicated. The current project aims at dissecting the human genetic and immunological basis of coinhibitory crosstalk between human T cells and cancer cells by tackling the two specific aims. Specific Aim 1 is to delineate the molecular mechanisms of PD-L1/L2-mediated inhibition of T cell activation independent from PD-1. This aim is supported by preliminary observations that both primary and immortalized T cells from a patient with inherited complete PD-1 deficiency, as well as PD-1-negative human T cell lines Jurkat and HuT78, all responded to co-inhibition by bead- immobilized PD-L1/L2. CD83 was identified as a candidate for PD-L1-mediated immunosuppression. The proposed work includes searching for additional candidates through RNA sequencing and surface receptor profiling and functionally validating the candidates via RNA silencing and CRISPR-guided knockout. Specific Aim 2 is to characterize the immunoregulatory function of CD83 in human T cells. Previous studies suggest that CD83 can serve as both an immunosuppressive receptor and ligand for human T cells, although its counterreceptor(s), downstream signaling, and relevance to antitumor immunity remain mostly elusive. The proposed work includes biochemical characterization of CD83 signaling in human T cells with or without cancer cell coculture utilizing genetic knockout and lentiviral rescue of CD83 gene. The proposed project will help untangle the biochemical and immunological basis of human T cell coinhibitory circuits in the crosstalk between cancer cells and also substantially bolster the applicant's scientific portfolio and expedite the growth toward an independent investigator pioneering human genetics of cancer immunology and immunotherapy.