Novel therapeutic approaches with CDK4/6 inhibitors for melanoma treatment

项目摘要/摘要 这项提案的总体目标是提高CDK4/6抑制剂治疗黑色素瘤的疗效，以及 寻找使用CDK4/6抑制剂治疗黑色素瘤患者的新的、高效的联合疗法。 细胞周期蛋白依赖性激酶CDK4和CDK6是被激活的核心细胞周期机制的组成部分 当与它们的调控亚单位相互作用时，D型细胞周期蛋白(细胞周期蛋白D1、D2和D3)。化学缓蚀剂 目前已用于240多项不同癌症类型的临床试验，包括 黑色素瘤。大量研究表明CDK4/6对人癌细胞的治疗作用 抑制剂会阻止肿瘤细胞的增殖，在某些情况下还会导致肿瘤细胞衰老。在我们的预赛中 研究中，我们观察到高水平表达细胞周期蛋白D3-CDK6复合体的肿瘤细胞株 CDK4/6抑制后死亡。我们发现了细胞周期蛋白D3-CDK6，但没有发现其他几种类型的细胞周期蛋白D-CDK4/6 复合体(如细胞周期蛋白D1-CDK4)，在调节肿瘤细胞代谢中起限速作用，以及 保护肿瘤细胞免受活性氧水平升高的影响。在另一项研究中，我们发现在 在小鼠和人类肿瘤细胞中，细胞周期蛋白D-CDK4激酶调节免疫检查点蛋白的水平， 程序性死亡配体1(PD-L1)。利用小鼠癌症模型和体外培养的细胞，我们发现 用CDK4/6抑制剂治疗后，肿瘤细胞中PD-L1蛋白水平显著上调。我们 还发现，联合使用CDK4/6抑制剂和针对相互作用的抗体 程序性细胞死亡蛋白1(PD-1)及其配体PD-L1增强免疫效力 检查点疗法。在这里提出的研究中，我们将测试涉及CDK4/6的新的治疗方法 黑色素瘤的体内抑制治疗。在目标1中，我们将测试我们的假设，即测量细胞周期蛋白D3 转移性黑色素瘤中CDK6的水平可以帮助人们识别出 可接受CDK4/6抑制剂治疗，因为它们将经历肿瘤细胞(和黑色素瘤)的死亡 转移细胞)对CDK4/6的抑制。我们还将探索通过以下方式触发对黑色素瘤细胞的杀伤 CDK4/6抑制剂，在黑色素瘤不表达高周期蛋白D3/CDK6的情况下。我们将解决这些问题 问题使用非常大和独特的黑色素瘤患者来源的异种移植，和短期 原代人转移性黑色素瘤细胞的培养。在目标2中，我们将检验我们的假设CDK4/6 抑制剂治疗将增加免疫检查点治疗黑色素瘤的疗效，并导致 小鼠黑色素瘤模型中肿瘤的消退和存活率的提高。我们还将研究如何治疗 CDK4/6抑制剂对荷黑色素瘤小鼠机体抗肿瘤免疫反应的影响 水平。这项提案的预期总体影响是，它将识别出以下黑色素瘤的子集 对抗CDK4/6治疗特别敏感，并寻找新的、高效的联合治疗方法 涉及CDK4/6抑制剂的黑色素瘤患者。

Project Summary/Abstract The overall goal of this proposal is to enhance the efficacy of CDK4/6 inhibitors for melanoma treatment, and to identify novel, highly effective combinatorial treatments for melanoma patients involving CDK4/6 inhibitors. Cyclin dependent kinases CDK4 and CDK6 are components of the core cell cycle machinery that are activated upon interaction with their regulatory subunits, the D-type cyclins (cyclins D1, D2 and D3). Chemical inhibitors of the CDK4/6 kinase are currently being used in over 240 clinical trials for different cancer types, including melanomas. A large number of studies demonstrated that treatment of human cancer cells with CDK4/6 inhibitors blocks tumor cell proliferation, and in some cases causes tumor cell senescence. In our preliminary studies, we observed that tumor cell lines expressing high levels of cyclin D3-CDK6 complexes undergo cell death upon CDK4/6 inhibition. We found that cyclin D3-CDK6, but not several other types of cyclin D-CDK4/6 complexes (such as cyclin D1-CDK4), plays a rate-limiting role in regulating tumor cell metabolism, and protects tumor cells against elevated levels of reactive oxygen species. In another study, we found that in mouse and human tumor cells, cyclin D-CDK4 kinase regulates the levels of an immune checkpoint protein, programmed death-ligand 1 (PD-L1). Using mouse cancer models and in vitro cultured cells, we found that treatment with CDK4/6 inhibitors results in a strong upregulation of PD-L1 protein levels in tumor cells. We also found that combined administration of CDK4/6 inhibitors with antibodies that target the interaction between programmed cell death protein 1 (PD-1) and its ligand PD-L1 increased the efficacy of immune checkpoint therapy. In the studies proposed here, we will test novel therapeutic approaches involving CDK4/6 inhibition in vivo for treatment of melanomas. In Aim 1, we will test our hypothesis that measuring cyclin D3 and CDK6 levels in metastatic melanomas may allow one to identify tumor cases that are particularly amenable to CDK4/6 inhibitor treatment, as they would undergo death of tumor cells (and of melanoma metastatic cells) upon CDK4/6 inhibition. We will also explore ways to trigger killing of melanoma cells by CDK4/6 inhibitors, in case of melanomas not expressing high cyclin D3/CDK6 levels. We will address these issues using a very large and unique collection of melanoma patient-derived xenografts, and short-term cultures of primary human metastatic melanoma cells. In Aim 2, we will test our hypothesis that CDK4/6 inhibitor treatment would increase the efficacy of immune checkpoint therapy for melanomas, and result in tumor regression and improved survival rates in mouse melanoma models. We will also study how treatment of melanoma-bearing mice with CDK4/6 inhibitors affects the anti-tumor immune response at the organismal level. The expected overall impact of this proposal is that it will identify a subset of melanomas that are particularly sensitive to anti-CDK4/6 therapy, and identify novel, highly effective combinatorial treatments for melanoma patients involving CDK4/6 inhibitors.