Targeting CDK4 and CDK6 kinases in breast cancer development

项目摘要/摘要 本翻译提案侧重于使用细胞周期蛋白依赖性激酶CDK4和CDK6的抑制剂 乳腺癌的治疗。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)。抗肿瘤药物 CDK4/6 Palbociclib、Riociclib和Abemaciclib获得了来自 FDA并已被批准用于雌激素受体阳性乳腺癌的治疗。此外，这些 化合物目前正在对其他几种癌症类型进行临床试验。大量的研究表明 用CDK4/6抑制剂治疗人类癌细胞会阻止肿瘤细胞的增殖，在某些情况下 会导致肿瘤细胞衰老。与波利亚克博士合作，我们观察到肿瘤细胞表达 高水平的细胞周期蛋白D3-CDK6复合体在CDK4/6抑制后发生细胞死亡。我们发现细胞周期蛋白D3- CDK6，而不是我们分析的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，并保护肿瘤细胞免受升高的 活性氧物种。在一项独立的研究中，我们发现在小鼠和人类的肿瘤细胞中，细胞周期蛋白D- CDK4激酶调节免疫检查点蛋白--程序性死亡配体1(PD-L1)的水平。 利用小鼠肿瘤模型和体外培养的细胞，我们发现CDK4/6抑制剂的治疗效果 上调肿瘤细胞中PD-L1蛋白水平。我们还发现，联合应用CDK4/6 具有针对程序性细胞死亡蛋白1(PD-1)与其相互作用的抗体的抑制剂 配体PD-L1增加了免疫检查点治疗的疗效。在这里提出的研究中，我们将 将这些观察结果推广到乳腺癌。在目标1中，我们将与Polyak博士和Brown博士一起测试我们的 一种假说认为，通过检测人类乳腺癌中细胞周期蛋白D3和CDK6的水平，可以确定 对CDK4/6抑制剂治疗特别敏感的乳腺癌病例，如细胞周期蛋白D3/CDK6-高 抑制CDK4/6后，肿瘤可能发生肿瘤细胞死亡和肿瘤消退。我们还将探索 诱导以细胞周期蛋白D1和CDK4为主的乳腺癌细胞凋亡的途径。我们将解决 这些问题使用了人乳腺癌细胞系和患者来源的乳腺癌异种移植。 我们还将与Polyak和Brown博士合作，解决乳腺肿瘤内和肿瘤间的异质性 癌症，这可能会影响结果。在目标2中，我们验证了我们的假设CDK4/6抑制剂治疗 将提高免疫检查点治疗乳腺癌的疗效并导致肿瘤消退 并提高了小鼠乳腺癌模型的存活率。我们将用有乳房的小鼠来研究这一点 同种异体癌症移植以及自体三阴性乳腺肿瘤。预计这一事件的总体影响 提案是，它将提供手段来识别对抗癌药物特别敏感的乳腺癌病例 CDK4/6治疗，并建立涉及CDK4/6抑制剂的新型、高效的联合治疗。

Project Summary/Abstract This translational proposal focuses on the use of inhibitors of cyclin-dependent kinases CDK4 and CDK6 for breast cancer treatment. 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). Inhibitors of CDK4/6 palbociclib, ribociclib and abemaciclib received ‘Breakthrough Therapy’ designation status from the FDA and have been approved for treatment of estrogen receptor-positive breast cancers. In addition, these compounds are now in clinical trials for several other cancer types. 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. Working together with Dr. Polyak, we observed that tumor cells 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 analyzed by us (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 an independent 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 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 extend these observations to breast cancers. In Aim 1, we will work with Drs. Polyak and Brown to test our hypothesis that measuring cyclin D3 and CDK6 levels in human breast cancers may allow one to identify breast cancer cases that are particularly sensitive to CDK4/6 inhibitor treatment, as cyclin D3/CDK6-high tumors might undergo tumor cell death and tumor regression upon CDK4/6 inhibition. We will also explore ways to trigger apoptosis of breast cancers expressing predominantly cyclin D1 and CDK4. We will address these issues using xenografts of human breast cancer cell lines and patient-derived breast cancer xenografts. We will also work with Drs. Polyak and Brown to address the intra- and inter-tumoral heterogeneity of breast cancers, which may influence the outcomes. In Aim 2, we test our hypothesis that CDK4/6 inhibitor treatment would increase the efficacy of immune checkpoint therapy for breast cancers and result in tumor regression and improved survival rates in mouse breast cancer models. We will study this using mice bearing breast cancer allografts as well as autochthonous triple-negative breast tumors. The expected overall impact of this proposal is that it will provide means to identify breast cancer cases that are particularly sensitive to anti- CDK4/6 therapy, and establish novel, highly effective combinatorial treatments involving CDK4/6 inhibitors.