BLR&D Merit Review Research Career Scientist (RCS) Award (IK6)

摘要： 目前，使用免疫检查点抑制剂(ICI)，如α-CTLA4和/或α-PD1的治疗是 提高存活率(50%的四年存活率)，特别是在高度突变的肿瘤中。此外，结合ICIS 针对肿瘤癌基因的治疗在一些肿瘤中非常有效。 我们的假设是，免疫“冷”肿瘤对ICI治疗的反应可以通过联合ICIS来增强。 针对致癌驱动因素的治疗也诱导募集CD8+T细胞的关键趋化因子的表达 和DC向TME转化，使肿瘤相关巨噬细胞向抗肿瘤表型转化，并减少了侵袭 通过髓系来源的抑制细胞。为了验证这一假设，我们将利用免疫能力强的小鼠模型， 携带患者来源的异种移植(PDX)的人源化小鼠模型，以及人肿瘤的器官型联合培养. 与患者免疫细胞培养以确定这些特定的精确药物是否接近靶向治疗 增强对ICIS的反应，提高小鼠的存活率。 我们建议在四个方面推进黑色素瘤和乳腺癌的治疗策略。有两个 我们将为黑色素瘤研究开发的方法：1)使用免疫活性BRAFV600E/PTEN-/-， NRASQ61K、R/CDKN2A-/-或三重野生型小鼠模型、人源化PDX模型和人体3D器官 黑色素瘤与从患者体内分离的免疫细胞共培养，我们将确定是否用 Rigosertib(NRAS通路抑制剂)、CDK4/6抑制剂+MDM2拮抗剂联合ICI治疗诱导 肿瘤消退。抗肿瘤免疫反应改变的机制将被确定。2)。使用CyTOF 和sc-ngs分析，我们将对产生耐药性的黑色素瘤患者的活检组织和PBL进行特征分析。 对于免疫检查点抑制物，评估其他检查点蛋白的表达以及新的激活 驱动突变，并利用这些数据开发新的策略，用于治疗其他检查点抑制剂和 使用我们的器官培养模型进行靶向治疗。 对于我们的乳腺癌研究：1)我们将使用AKT抑制剂加紫杉醇联合ICI治疗肿瘤 治疗或在ICI治疗之前使用免疫活性小鼠模型、人源化PDX模型和器官分型 患者肿瘤培养与患者免疫细胞共培养；2)我们将检测小鼠肿瘤的生长情况 靶向缺失肿瘤组织或特定免疫细胞群中的CXCR2或CXCR4 治疗和免疫疗法。在适当的情况下，CXCR2和/或CXCR4拮抗剂可以与 靶向治疗，以提高对ICI治疗的反应。 总之，这些研究将为黑色素瘤和乳腺癌的新临床试验的设计提供关键信息。 与我们的临床合作者合作，并告知正在进行的治疗的耐药机制。黑色素瘤与乳房 癌症可能是我们退伍军人的主要问题，以及改善治疗贡献的发展，以增强 总体生存至关重要。

Abstract: Currently, therapeutic treatment with immune checkpoint inhibitors(ICI’s), such as α-CTLA4 and/or α-PD1, are increasing survival rates (>50% four-year survival), particularly in highly mutated tumors. Moreover, combining ICIs with therapies that target tumor oncogenes has been highly effective in some tumors. Our hypothesis is that response to ICI therapy in immunologically “cold” tumors can be enhanced by combining ICIs with therapies targeted to oncogenic drivers that also induce expression of key chemokines that recruit CD8+T cells and DCs to the TME and shift tumor associated macrophages to an anti-tumor phenotype, and reduce the infiltration by myeloid-derived suppressor cells. To test this hypothesis, we will utilize immune competent mouse models, humanized mouse models bearing patient-derived xenograft (PDX), and organotypic cultures of human tumor co- cultured with patient immune cells to determine if these specific precision medicine approaches to targeted therapies enhance response to ICIs, and improve survival in mice. We propose to advance therapeutic strategies for melanoma and breast cancer in four areas. There are two approaches we will develop for melanoma studies: 1) Using immune competent BRAFV600E/PTEN-/-, NRASQ61K,R/CDKN2a-/- or triple wild type mouse models, humanized PDX models and 3D organoids of human melanoma tumors co-cultured with immune cells isolated from the patient, we will determine whether treatment with either rigosertib (NRAS pathway inhibitor), CDK4/6 inhibitor +MDM2 antagonist combined with ICI therapy induces tumor regression. Mechanisms for alterations in anti-tumor immune response will be determined. 2). Using CyTOF and sc-NGS analysis we will characterize biopsy tissue and PBLs from melanoma patients who develop resistance to immune checkpoint inhibitors, evaluate the expression of other checkpoint proteins as well as activation of new driver mutations, and use these data to develop new strategies for treatment with other checkpoint inhibitors and targeted therapies using our organoid culture model. For our breast cancer studies: 1) We will treat tumors with AKT inhibitors plus paclitaxel in combination with ICI therapies or prior to ICI treatment using immune competent mouse models, humanized PDX models, and organotypic cultures of patient tumors co-cultured with patient immune cells; 2) We will examine the tumor growth in mice with targeted deletion of CXCR2 or CXCR4 in tumor tissue or in specific immune cell populations with or without targeted therapies and immune therapy. Where appropriate, CXCR2 and/or CXCR4 antagonists may be combined with the targeted therapies to enhance response to ICI therapy. Altogether these studies will provide key information for the design of new clinical trials I melanoma and breast cancer with our clinical collaborators and inform mechanisms of resistance to ongoing therapies. Melanoma and breast cancer can be major issues for our Veterans and development of improved therapeutic contributions that enhance overall survival is of key importance.