Targeting Stat3 to Improve Immunotherapy

摘要 肿瘤微环境严重限制了各种免疫疗法的疗效。 对于过继T细胞治疗，需要选择/工程和扩增具有靶向性的T细胞 在保持其效应器功能和归巢能力的同时保持抗原特异性 带来了额外的挑战。我们和其他人已经证明了STAT3，一个关键 癌基因转录因子在不同的癌细胞中被成分激活 对肿瘤细胞存活很重要，在肿瘤间质免疫细胞中也被激活，并 强大的免疫抑制作用。我们的初步研究表明，靶向STAT3基因 髓系细胞消融导致肿瘤DC激活，肿瘤Treg细胞减少和 CD8+T细胞在肿瘤中的大量渗透，导致有效的抗肿瘤免疫 回应。此外，在有STAT3/-髓系隔室的小鼠中，过继 转移的CD8+T细胞增殖良好，在肿瘤引流过程中高度活化 淋巴结。我们还能够通过多光子共焦捕捉到实时图像 转移的STAT3-/-CD8+T细胞有效地渗入肿瘤的显微镜观察 增殖，导致肿瘤抗原特异性免疫反应的激活。我们有 开始了解允许STAT3在两者中的细胞和分子机制 肿瘤髓系细胞和T细胞阻碍T细胞介导的抗肿瘤免疫。在这 应用，我们建议进一步定义这些细胞和分子机制，并 从而确定其他靶点，以优化STAT3靶向和T细胞治疗。至 为了促进未来潜在的临床翻译，我们还将测试两种试剂，这两种试剂都 对肿瘤免疫环境显示出积极的作用，因为它们有可能 改进T细胞疗法。一种是新的Toll样受体(TLR)激动剂-siRNA 共轭，这是我们最近开发的一个技术平台，能够免疫 激活和靶向基因沉默所需的免疫检查点，如STAT3 在髓系和B细胞中。另一种是FDA批准的酪氨酸激酶抑制剂孙尼替尼， 我们的初步数据表明，它抑制了肿瘤细胞中的STAT3，肿瘤的侵袭 树突状细胞和髓系来源的抑制细胞。此外，我们还将测试抗肿瘤药物 慢病毒siRNAs在T细胞中沉默STAT3的效果建议进行的研究 可能导致在体内扩增和激活转移的T细胞的新策略，从而 克服T细胞治疗面临的几个主要障碍，并显著改善其 治疗癌症的疗效。

Abstract The tumor microenvironment severely limits the efficacy of various immunotherapies. For adoptive T cell therapy, the need to select/engineer and expand T cells with targeted antigen specificity while still preserving their effector function and homing capacities poses additional challenges. We and others have demonstrated that Stat3, a key oncogenic transcription factor constitutively activated in diverse cancer cells and important for tumor cell survival, is also activated in tumor stromal immune cells and potently immunosuppressive. Our preliminary studies show that targeted Stat3 gene ablation in myeloid cells results in tumor DC activation, tumor Treg cell reduction and heavy infiltration of CD8+ T cells in tumors, leading to effective antitumor immune responses. Furthermore, in mice with a Stat3-/- myeloid compartment, adoptively transferred CD8+ T cells proliferate well and are highly activated in the tumor draining lymph nodes. We were also able to capture live images by multi-photon confocal microscopy of transferred Stat3-/- CD8+ T cells efficiently infiltrating tumors, where they proliferate, leading to activation of tumor antigen-specific immune responses. We have begun to understand the cellular and molecular mechanisms that allow Stat3 in both tumor myeloid cells and T cells to impede T-cell mediated antitumor immunity. In this application, we propose to further define these cellular and molecular mechanisms and thereby identifying additional targets to optimize Stat3 targeting and T cell therapy. To facilitate potential future clinical translation, we will also test two reagents, both of which have shown positive effects on the tumor immunologic environment, for their potential to improve T cell therapies. One is a novel Toll-like receptor (TLR) agonist-siRNA conjugate, a technology platform we have recently developed, and is capable of immune activation and targeted gene silencing of desired immunologic checkpoints such as Stat3 in myeloid and B cells. The other is an FDA-approved tyrosine kinase inhibitor, sunitinib, as our preliminary data suggested it inhibited Stat3 in tumor cells, tumor-infiltrating dendritic cells and myeloid-derived suppressor cells. Additionally, we will test antitumor efficacy by silencing Stat3 in T cells ex vivo with lentiviral-siRNAs. The proposed studies may lead to new strategies to expand and activate transferred T cells in vivo, thereby overcoming several major hurdles facing T cell therapy, and significantly improve its efficacy for cancer treatment.