Modulation of tumor susceptibility to NK cells induced by JAK family genes

描述(由申请人提供)：NK细胞是天然免疫反应的主要效应者，通过其有效的细胞溶解活性来对抗感染性病原体和恶性转化。与T和B细胞不同，NK不识别经典的主要组织相容性复合体(MHC)中的抗原，但它们在没有特异性抗原识别的情况下溶解肿瘤细胞。尽管一些研究表明NK细胞对靶细胞的杀伤能力依赖于几种抑制性或激活性受体的表达，但参与靶细胞对NK细胞易感性的分子机制仍不清楚。我们假设，使用基因筛查策略将识别调节肿瘤细胞对免疫系统敏感性的新途径。为了验证这一假设，我们开发了一种基于细胞-细胞相互作用shRNA的筛选方法，以IM-9肿瘤细胞为靶细胞，NK细胞为效应者。在调节靶细胞对NK活性敏感性的基因中，我们发现了JAK家族的两个成员(JAK1和JAK2)。独立实验证实，虽然JAK1和JAK2的靶细胞更容易受到NK细胞活性的影响，但对JAK家族的其他两个成员(JAK3和TYK2)没有影响。初步的基因表达谱分析表明，靶细胞敲除JAK1基因显著调节了TRAIL/FASL途径中的基因表达。已知这些途径可在NK细胞参与后诱导靶细胞凋亡。在目前的研究中，我们建议：1)进行一系列实验，以更好地了解JAK1和JAK2是如何通过TRAIL/FASL途径调节肿瘤敏感性的。2)分析JAK基因如何调节一系列原发肿瘤细胞亚群对NK或T细胞活性的敏感性。3)开始定义可能的下游分子，这些分子也参与诱导肿瘤对NK细胞的敏感性。NK细胞是机体免疫监视的主要效应者之一，在巡视血液、脾、骨髓等淋巴器官清除病毒感染和肿瘤转化细胞方面发挥着重要作用。更好地了解NK细胞如何识别其靶点将有助于改善癌症患者的临床预后，并将进一步推动NK细胞治疗在预防和有效治疗癌症的临床应用中的进展。 公共卫生相关性：NK细胞是针对感染、病原体和恶性转化的先天免疫反应的主要效应者，其有效的细胞溶解活性。尽管已有研究表明，NK细胞杀伤靶细胞的能力依赖于多种抑制性或激活性受体的表达，但其调控靶细胞对NK细胞易感性的分子机制仍不十分清楚。为了确定能够诱导肿瘤细胞对天然免疫系统易感性的新基因，我们开发了一种基于细胞-细胞相互作用shRNA的筛选方法。针对1000多个基因的多个独立shRNAs以高通量的方式转导到肿瘤靶细胞系中，并检测其对NK细胞的敏感性。两个JAK家族成员的基因被发现显著调节肿瘤对NK细胞的敏感性。在本研究中，我们拟探讨JAK基因在诱导肿瘤细胞对NK细胞活性的敏感性中的作用。我们预计我们的研究将：1)更好地确定JAK基因如何调节肿瘤对NK细胞活性的敏感性的机制。2)分析JAK基因如何调节一系列原发肿瘤细胞亚群对NK或T细胞活性的敏感性。3)开始定义可能的下游分子，这些分子也参与诱导肿瘤对NK细胞的敏感性。

DESCRIPTION (provided by applicant): NK cell are the primary effectors of the innate immune response against infectious pathogens and malignant transformations through their efficient cytolytic activity. Unlike T and B cells, NK do not recognize antigens in the context of classical major histocompatibility complex (MHC) but they lyse tumor cells without specific antigen recognition. Although several studies have shown that the ability of NK cells to lyse the target is dependent on the expression of several inhibitory or activating receptors, the molecular mechanism that are involved in the target cell susceptibility to NK cells are still not well understood. We hypothesized that the use of a genetic screening strategy would identify novel pathways that modulate tumor cell susceptibility to the immune system. To test this hypothesis, we developed a cell-cell interaction shRNA based screening using IM- 9 tumor cells as target and NK cells as effectors. Among the genes that modulated target cell susceptibility to NK activity we found 2 members of the Jak family (Jak1 and Jak2). Independent experiments confirmed that while target cells knock-down for Jak1 and Jak2 were more susceptible to NK cell activity no effect was noticed with the other 2 members of the Jak family (Jak3 and TYK2). Preliminary gene expression profile analysis showed that target cells knock down for Jak1 significantly modulated the expression of genes in the TRAIL/FASL pathway. These pathways are known to induce target apoptosis after NK cells engagement. In the current study we propose: 1) a series of experiments to better understand how Jak1 and Jak2 can modulate tumor susceptibility through the TRAIL/FASL pathway. 2) Analyze how Jak genes modulate susceptibility of a series of primary tumor cell subpopulation to NK or T cell activity. 3) Start to define possible downstream molecules that are also involved in the induction of tumor sensitivity to NK cells. NK cells represent one of the main effectors of immune surveillance in the body, playing an important role in patrolling the bloodstream, spleen, bone marrow and other lymphoid organs to eliminate virus infected and tumor-transformed cells. A better understanding of how NK cells recognize their targets will help to improve clinical outcome in cancer patients and will lead to further advances in the clinical application of NK cell therapy to prevent and effectively treat cancer. PUBLIC HEALTH RELEVANCE: NK cell are the primary effectors of the innate immune response against infections pathogens and malignant transformations through their efficient cytolytic activity. Although several studies have shown that the ability of NK cells to lyse the target is dependent on the expression of several inhibitory or activating receptors, the molecular mechanism that regulate the target cell susceptibility to NK cells are still not well understood. To identify new genes that can induce tumor cell susceptibility to the innate immune system we developed a cell-cell interaction shRNA based screening. Multiple independent shRNAs, targeting more than 1,000 genes, were transduced in a tumor target cell line in an high-throughput way and tested for their susceptibility to NK cells. Two members of the Jak family genes were found to significantly modulate tumor susceptibility to NK cells. In the current study we propose to investigate the involvement of the Jak genes in the induction of tumor cell susceptibility to the NK cell activity. We anticipate that our studies will: 1) better determine the mechanisms on how Jak genes modulate tumor susceptibility to NK cell activity. 2) Analyze how Jak genes modulate susceptibility of a series of primary tumor cell subpopulation to NK or T cell activity. 3) Start to define possible downstream molecules that are also involved in the induction of tumor sensitivity to NK cells.