Ontogeny and Function of Tumor-Resident Innate Lymphocytes and Innate-Like T Cells

项目摘要 癌症免疫监视归因于免疫系统在抑制肿瘤发展中的作用。癌 免疫治疗方法，如检查点阻断，恢复这种功能的疲惫的T细胞， 彻底改变了癌症患者的护理尽管如此，许多患者对这种癌症模式没有反应 治疗，要求调查更广泛的肿瘤引起的免疫反应。我们有 最近表明，肿瘤生长诱导组织驻留细胞毒性先天淋巴细胞的扩增， 与NK细胞和衰竭T细胞共享不同基因表达程序的先天样T细胞。 这些细胞以高表达转录因子Hobit和溶细胞颗粒酶为特征， 本文称为杀伤先天淋巴样细胞（ILCk）和杀伤先天样T细胞（ILTCk）。值得注意的是， ILCk和ILTCk的结合导致加速的肿瘤生长。此外，肿瘤细胞表达高水平的IL-15， 并且失去E-钙粘蛋白极性，并且IL-15或E-钙粘蛋白缺乏耗尽ILCk和ILTCk， 加速肿瘤生长。基于这些发现，我们假设ILCk和ILTCk是新的谱系， 细胞毒性淋巴细胞，并且它们通过感应肿瘤细胞来源的IL-15作为细胞转化的哨兵发挥作用 和E-cadherin。为了验证这一假设，我们将首先定义ILCk和ILTCk的发育途径。通过 进行细胞转移和细胞命运作图实验以及使用谱系特异性缺陷的小鼠， 转录因子，我们将评估ILCk是否沿先天淋巴样细胞谱系分化沿着。在 此外，我们将产生T细胞受体（TCR）逆转录小鼠，并进行TCR“交换”实验， 确定不同的胸腺选择是否促进ILTCk分化。共生与诱导 还将进行造血干细胞靶向细胞命运作图实验以确定是否 ILCk和ILTCk在整个肿瘤进展过程中持续产生。其次，我们将定义 ILCk和ILTCk的功能和调节，通过初步评估Hobit表达是否标志着 功能规范，以及Hobit是否控制ILCk和ILTCk所必需的基因表达程序， 介导的癌症监测。此外，我们将利用Il 15和Il 2 rb的条件性无效等位基因和获得性- 编码转录因子Stat 5 b活性形式的功能等位基因，以确定肿瘤IL-15 作为ILCk和ILTCk的报警蛋白起作用，以及IL-15信号传导是否构成ILCk和ILTCk的限速步骤。 ILCk和ILTCk响应。最后，我们将研究肿瘤细胞与ILCk和ILTCk之间的相互作用 通过活体成像，并评估TGF-β诱导的整合素CD 103是否感知E-钙粘蛋白， 患者CDH 1热点错义突变是否促进ILCk和ILTCk介导的肿瘤逃避 癌症监测这个项目的成功完成不仅会产生机械的见解， 谱系承诺和调节肿瘤居民ILCk和ILTCk，而且还指导这种新的靶向 癌症免疫监视途径，用于治疗多种恶性肿瘤。

Project Summary Cancer immunosurveillance ascribes a role of the immune system in repressing tumor development. Cancer immunotherapy approaches such as checkpoint blockade that revives this function of exhausted T cells have revolutionized cancer patient care. Nonetheless, many patients do not respond to this modality of cancer treatment, calling for investigation of a broader spectrum of tumor-elicited immune responses. We have recently shown that tumor growth induces expansion of tissue-resident cytotoxic innate lymphocytes and innate-like T cells that share a gene expression program distinct from that of NK cells and exhausted T cells. Characterized by high expression of the transcription factor Hobit and cytolytic granzymes, these cells are herein named killer innate lymphoid cells (ILCk) and killer innate-like T cells (ILTCk). Notably, genetic depletion of ILCk and ILTCk results in accelerated tumor growth. Furthermore, tumor cells express high levels of IL-15 and lose E-cadherin polarity, and IL-15 or E-cadherin deficiency depletes ILCk and ILTCk resulting in accelerated tumor growth. Based on these findings, we hypothesize that ILCk and ILTCk are novel lineages of cytotoxic lymphocytes, and they function as sentinels of cell transformation by sensing tumor cell-derived IL-15 and E-cadherin. To test this hypothesis, we will first define the developmental pathways of ILCk and ILTCk. By performing cell transfer and cell fate-mapping experiments as well as using mice deficient in lineage-specifying transcription factors, we will assess whether ILCk are differentiated along the innate lymphoid cell lineage. In addition, we will generate T cell receptor (TCR) retrogenic mice and perform TCR “swapping” experiments to determine whether distinct thymic selection promotes ILTCk differentiation. Parabiosis and inducible hematopoietic stem cell-targeted cell fate-mapping experiments will also be performed to determine whether ILCk and ILTCk are continuously generated throughout tumor progression. Secondly, we will define the function and regulation of ILCk and ILTCk by initially assessing whether Hobit expression marks a stage of functional specification, and whether Hobit controls a gene expression program essential for ILCk and ILTCk- mediated cancer surveillance. In addition, we will utilize conditional null alleles of Il15 and Il2rb and a gain-of- function allele encoding an active form of the transcription factor Stat5b to determine whether tumor IL-15 functions as an alarmin for ILCk and ILTCk, and whether IL-15 signaling constitutes a rate-limiting step of the ILCk and ILTCk response. Finally, we will investigate the interactions between tumor cells and ILCk and ILTCk by intravital imaging, and assess whether E-cadherin is sensed by the TGF-b-induced integrin CD103, and whether patient CDH1 hot-spot missense mutations promote tumor evasion from ILCk and ILTCk-mediated cancer surveillance. Successful completion of this project will not only generate mechanistic insights into the lineage commitment and regulation of tumor-resident ILCk and ILTCk, but also guide the targeting of this novel cancer immunosurveillance pathway for therapy of a wide range of malignancies.