Combining Immune Therapy with Targeted Therapies to Improve Melanoma Survival

转移性黑色素瘤是美国生长最快的肿瘤类型之一，我们的退伍军人患上 发展中的黑色素瘤。免疫检查点抑制剂(ICIS)免疫治疗研究进展 彻底改变了晚期/转移性黑色素瘤的治疗方法。针对PD-1/PD-L1的抗体(nivolumab， Pembrolizumab，atezolizumab)和CTLA-4(Ipilimumab)引起强大的抗黑色素瘤免疫反应1，2，3。 对于转移性疾病，已证实ipilimumab联合nivolumab或nivolumab单独治疗。 5年总存活率(OS)分别为52%和44%，许多患者经历了非凡的 回应。尽管ICIS的生存结果有所改善，但大多数患者要么没有反应，要么 对ICIS产生抵抗力。需要新的治疗策略来改善黑色素瘤患者的预后 用ICI治疗，并解决获得性耐药性。新的治疗方法的发展 对ICI治疗表现出原始或获得性抵抗的黑色素瘤患者是我们建议的重点。我们的 来自临床前模型的初步数据表明，将RAS模拟物rigosertib(RGS)与ICI相结合 显著减少黑色素瘤的生长，即使在对ICIS反应不佳的肿瘤中也是如此。RGS 治疗结果是肿瘤免疫微环境(TIME)富含CD8+T细胞、树突状细胞(DC) 和自然杀伤细胞，但不减少肿瘤内髓系来源的抑制细胞(MDSCs)。RGS 也增加了黑色素瘤细胞上CD40的表达，以促进免疫原性肿瘤细胞的死亡。在……里面 不表达CD40的肿瘤细胞，或不表达CD8+T细胞的时间，RGS的抗肿瘤活性 已经迷失了。我们假设RGS诱导的黑色素瘤细胞CD40的表达是 RGS增强的抗肿瘤免疫和在RGS+ICI中添加CD40激动剂将增强CD40- 介导的免疫原性细胞死亡。我们还建议在RGS+ICI中增加一种CXCR1/2拮抗剂 治疗将阻断MDSC向肿瘤的募集，从而延长CD8+T细胞的抗肿瘤活性。以下任一项 这些途径将进一步激活抗肿瘤免疫反应，诱导肿瘤消退。 我们将追求三个具体目标来解决这一假设。目标1：确定 RGS诱导肿瘤细胞表面CD40的表达及肿瘤细胞CD40在修复中的作用 免疫活性黑色素瘤小鼠模型对ICI的反应。目的2.评估是否增加一个 CD40激动剂抗体增强黑色素瘤对RGS+ICI的敏感性及其机制 使用小鼠模型和人类器官共培养系统进行反应的研究。肿瘤、免疫性和 转录组的变化将通过多色流式细胞术和单细胞(Sc)rnseq来评估。 分析，以提供对这种联合疗法如何调节治疗反应的机械性见解。目标3： 为了确定RGS联合CXCR1/2拮抗剂是否能使黑色素瘤对ICI重新敏感 使用与AIM类似的方法进行治疗2.意义：这些研究将导致新的治疗方法 通过提高对ICI的反应来改善我们退伍军人转移性黑色素瘤的治疗 治疗或使ICI耐药肿瘤对ICI治疗重新敏感。

Metastatic melanoma is one of the fastest growing tumor types in the US and our Veterans are at higher risk for developing melanoma. Recent therapeutic advances in immunotherapy utilizing immune checkpoint inhibitors (ICIs) have revolutionized the treatment of advanced/metastatic melanoma. Antibodies targeting PD-1/PD-L1 (nivolumab, pembrolizumab, atezolizumab) and CTLA-4 (ipilimumab) evoke powerful anti-melanoma immune responses1,2,3. For metastatic disease, treatment with ipilimumab combined with nivolumab or nivolumab alone has demonstrated 5-year overall survival (OS) rates of 52% and 44%, respectively, with many patients experiencing exceptional response. Despite improvement in survival outcomes with ICIs, the majority of patients either fail to respond or develop resistance to ICIs. Novel therapeutic strategies are needed to improve outcomes for melanoma patients treated with ICI and to address acquired resistance. Development of new therapeutic approaches for melanoma patients who exhibit primary or acquired resistance to ICI therapy is the focus of our proposal. Our preliminary data from preclinical models indicate that combining the Ras mimetic, rigosertib (RGS) with ICI significantly reduces melanoma tumor growth, even in tumors that are poorly responsive to ICIs. RGS treatment results in a tumor immune microenvironment (TIME) enriched in CD8+T cells, dendritic cells (DCs) and natural killer cells, but does not reduce intratumoral myeloid-derived suppressor cells (MDSCs). RGS also increases CD40 expression on the melanoma tumor cells to facilitate immunogenic tumor cell death. In the absence of CD40 expression by tumor cells, or without CD8+T cells in the TIME, RGS’s anti-tumor activity is lost. We hypothesize that RGS-induced expression of CD40 in melanoma cells is a key component of RGS-enhanced anti-tumor immunity and that addition of a CD40 agonist to the RGS +ICI will enhance CD40- mediated immunogenic cell death. We also propose that addition of a CXCR1/2 antagonist to the RGS+ICI therapy will block MDSC recruitment to tumor, allowing prolonged CD8+T cell anti-tumor activity. Either of these approaches would further activate the anti-tumor immune response to induce tumor regression. We will pursue three specific aims to address this hypothesis. Aim 1: To determine the mechanism by which RGS induces expression of CD40 on tumor cells and to characterize the role of tumor cell CD40 in restoring response to ICI in immunocompetent mouse models of melanoma. Aim 2. To evaluate whether addition of a CD40 agonist antibody will enhance sensitivity of melanoma tumors to RGS + ICI and define mechanisms of response using mouse models and human organoid co-culture systems. Tumoral, immune, and transcriptome changes will be evaluated through multicolor flow cytometry and single cell(sc) RNAseq analysis to provide mechanistic insight on how this combined therapy modulates response to therapy. Aim 3: To determine whether RGS combined with a CXCR1/2 antagonist, can re-sensitize melanoma tumors to ICI therapy using similar approaches as for Aim 2. Significance: These studies will result in new therapeutic approaches to improve treatment of metastatic melanoma in our Veterans by enhancing response to ICI therapies or re-sensitizing ICI-resistant tumors to treatment with ICI.