MHC Class II Antigen Presentation In Melanoma: Impact on Immune Recognition

调节肿瘤特异性CD4T细胞反应对改进癌症免疫治疗大有可为 免疫检查点抑制、癌症疫苗和过继T细胞治疗。有效应对 免疫治疗需要T细胞识别来自癌细胞的突变多肽表位，称为新抗原。 对免疫治疗的有效反应需要能够刺激CD4和CD8T细胞的新抗原。 肿瘤特异性CD4T细胞识别由MHC II类呈递的新抗原。然而，实验或 识别具有免疫原性(诱导T细胞)的MHC II类新抗原的计算方法 回应)还没有得到很好的证实。虽然肿瘤特异性CD4T细胞表位是产生 有效的抗肿瘤T细胞对免疫治疗的反应，在体内发育的CD4调节T(Treg)细胞 肿瘤抑制其他T细胞的增殖和功能。摧毁CD4 Treg细胞是一种很有希望的方法 免疫治疗方法。然而，缺乏一种特定靶向和破坏肿瘤特异性的方法 在癌症临床试验中，CD4 Treg细胞可防止Treg细胞的耗竭进一步进展 免疫疗法。多肽-MHC四聚体可用于识别肿瘤特异性T细胞。然而，多肽-MHC 许多已知可引起T细胞反应的多肽抗原不能产生II型四聚体 与MHCⅡ类分子相互作用的低亲和力或稳定性。这一建议将推动 通过解决知识中的三个空白来解决癌症免疫学：1)无法使用四聚体来识别大多数肿瘤- 特异性CD4T细胞，2)不能针对肿瘤特异性Treg细胞进行破坏，以及3)缺乏实验 以及优先处理免疫原性MHC II类新抗原的计算工具。这项提议将利用一种新的， 仿生5模块嵌合抗原受体(5MCAR)专利技术。5MCAR由 多肽-MHC-II类胞外区与T细胞跨膜区和胞浆区的融合 受体，它便于与CD3信号复合体组装，以及由以下组成的代理共受体 CD80与Src激酶Lck融合。5MCAR在细胞毒性T细胞上表达。通过5MCAR发送信号 允许识别和杀伤表达T细胞受体的CD4T细胞，而不是识别多肽-MHC 5MCAR的II级。中心假设是5MCAR有助于免疫原性MHC的测定 II类黑色素瘤新抗原和通过破坏肿瘤特异性CD4有效治疗黑色素瘤 树突状细胞。这项建议的目的是鉴定免疫原性MHC II类新抗原并测试其疗效。 在治疗黑色素瘤中消耗新抗原特异性的CD4细胞，使用临床相关的免疫原性 小鼠黑色素瘤模型。具有5MCAR的基于细胞的分析将用于高通量筛选 用于鉴定诱导CD4常规T细胞和调节性T细胞的MHC-II类新抗原的肽库 黑色素瘤肿瘤的细胞反应。肿瘤特异性5MCAR T细胞对已确诊患者的治疗作用 单独去除新抗原特异性Treg细胞及联合免疫治疗黑色素瘤 将评估抗PD-1对检查点的抑制作用。使用生成的全面、无偏见的数据集 通过测试所有错义突变的免疫原性，一种预测免疫原性的计算模型 将创建和验证MHC II类新抗原。这项提议的影响是：1)建立一个 与西语裔服务机构亚利桑那大学的库恩斯博士合作，2)纳入了一种新的 技术创新使高通量筛选能够鉴定免疫原性MHC II类 新抗原，肿瘤特异性CD4 Treg细胞的靶向破坏，以及改进的计算方法 优先处理免疫原性MHC II类新抗原，3)通过以下方式增加奖励的影响 解决当前知识中的空白，这些空白阻碍了癌症免疫疗法的进步，最终 改善对退伍军人的癌症护理，以及4)协助未被充分代表的调查人员从事退伍军人研究。

Regulation of tumor-specific CD4 T cell responses holds great promise for improving cancer immunotherapy with immune checkpoint inhibition, cancer vaccines, and adoptive T cell therapy. Effective response to immunotherapy requires T cell recognition of mutated peptide epitopes from cancer cells, called neoantigens. Effective response to immunotherapy requires neoantigens capable of stimulating both CD4 and CD8 T cells. Tumor-specific CD4 T cells recognize neoantigens presented by MHC class II. However, experimental or computational methods for identification of MHC class II neoantigens that are immunogenic (elicit a T cell response) have not been well-established. While tumor-specific CD4 T cell epitopes are necessary to generate effective anti-tumor T cell responses to immunotherapy, CD4 regulatory T (Treg) cells that develop within tumors suppress the proliferation and function of other T cells. Destroying CD4 Treg cells is a promising immunotherapeutic approach. However, the lack of a method to specifically target and destroy tumor-specific CD4 Treg cells prevents depletion of Treg cells from advancing further in clinical trials for cancer immunotherapy. Peptide-MHC tetramers can be used to identify tumor-specific T cells. However, peptide-MHC class II tetramers cannot be generated for many peptide antigens that are known to elicit T cell responses due to low affinity or stability of the peptide interaction with MHC class II. This proposal will advance the field of cancer immunology by addressing three gaps in knowledge: 1) inability to use tetramers to identify most tumor- specific CD4 T cells, 2) inability to target tumor-specific Treg cells for destruction, and 3) lack of experimental and computational tools to prioritize immunogenic MHC class II neoantigens. This proposal will utilize a new, patented technology of a biomimetic 5-module chimeric antigen receptor (5MCAR). The 5MCAR consists of the ectodomains of peptide-MHC class II fused to the transmembrane and cytoplasmic domains of the T cell receptor, which facilitates assembly with the CD3 signaling complex, and a surrogate coreceptor composed of CD80 fused to the Src kinase Lck. The 5MCAR is expressed on a cytotoxic T cell. Signaling through the 5MCAR allows identification and killing of CD4 T cells expressing a T cell receptor than recognizes the peptide-MHC class II of the 5MCAR. The central hypothesis is that 5MCAR will facilitate determination of immunogenic MHC class II melanoma neoantigens and effectively treat melanoma tumors by destruction of tumor-specific CD4 Treg cells. The aim of this proposal is to identify immunogenic MHC class II neoantigens and test the efficacy of depleting neoantigen-specific CD4 cells in treating melanoma, using a clinically relevant, immunogenic mouse melanoma model. A cell based assay with 5MCAR will be used in high throughput screening with a peptide library to identify the MHC class II neoantigens which elicit CD4 conventional T cell and regulatory T cell responses in melanoma tumors. The efficacy of tumor-specific 5MCAR T cells in treating established melanoma tumors through depletion of neoantigen-specific Treg cells alone and in combination with immune checkpoint inhibition with anti-PD-1 will be assessed. Using the comprehensive, unbiased dataset generated from testing the immunogenicity of all missense mutations, a computational model to predict immunogenic MHC class II neoantigens will be created and validated. The impact of this proposal is to 1) establish a collaboration with Dr. Kuhns at the University of Arizona, a Hispanic Serving Institution, 2) incorporate a new technological innovation to allow high throughput screening for identification of immunogenic MHC class II neoantigens, targeted destruction of tumor-specific CD4 Treg cells, and improved computational methods for prioritization of immunogenic MHC class II neoantigens, 3) increase the impact of the Merit award by addressing gaps in the current knowledge that prevent advancement of cancer immunotherapy and ultimately improve cancer care for Veterans, and 4) assist underrepresented investigators engage in VA Research.