Overcoming drug resistance using small molecule activators of protein phosphatase 2A

由细胞周期蛋白的异常活性引起的不受控制的细胞增殖是癌症的标志。 有丝分裂原传感器细胞周期蛋白D1的过表达是肿瘤中最常见的异常之一，增强了细胞周期蛋白D1的表达。 细胞周期蛋白依赖性激酶4和6（CDK 4/6）驱动G1→S期进程并促进细胞增殖活性 存活和耐药性。D型细胞周期蛋白表达的增加不仅是肿瘤发生所必需的， 也用于肿瘤的维持和发展。因此，异常的细胞周期蛋白D-CDK 4/6活性代表了一种可操作的 癌症治疗的目标和D型细胞周期蛋白功能是癌症管理的首要治疗目标之一。 CDK 4/6活性抑制剂已在临床上显示出前景，palbociclib、abemaciclib和ribociclib是 FDA批准用于患者。目前正在进行数百项临床试验，以评估抗肿瘤药物 这些药物在广泛的癌症类型中的作用。然而，CDK 4/6的治疗前景 抑制剂被不可避免出现的耐药性所抑制。最近的开创性研究发现了一种新的 自噬和Beclin 1调节因子1缺陷介导的对这些药物的抗性机制 （AMBRA 1），E3连接酶接头和细胞周期蛋白D1、D2和D3蛋白稳定性的主调节剂。丧失或突变 在人类癌症的重要亚组中观察到AMBRA 1的缺失，与患者存活率低相关。AMBRA 1 缺乏促进D型细胞周期蛋白的积累，过度增殖表型，和肿瘤发生，而 降低肿瘤细胞对所有三种FDA批准的CDK 4/6抑制剂的敏感性。有证据表明 D型细胞周期蛋白和非典型细胞周期蛋白D-CDK 2和p27-细胞周期蛋白D-CDK 4复合物的形成 对这些药物的耐药性构成了这一提议的基础。提出了探索战略， PP 2A小分子活化剂（SMAPs）的机制驱动应用，以克服对 CDK 4/6抑制剂在AMBRA 1缺陷的背景下。SMAPs是一类新的抗肿瘤药物， 选择性激活PP 2A全酶的一个子集，用于在多种癌症类型中有效抑制肿瘤。 该项目建立在我们发现SMAPs在所有细胞类型中有效下调细胞周期蛋白D1，D2和D3的基础上 测试.重要的是，SMAPs作为AMBRA 1非依赖性D型细胞周期蛋白“降解剂”，促进快速蛋白水解 这些分子通过蛋白酶体依赖性机制，在AMBRA 1丢失后仍然发挥作用。 基于这些发现，我们假设CDK 4/6抑制剂与SMAP的D型细胞周期蛋白联合治疗， 降解物“将增强抗肿瘤活性并逆转对由AMBRA 1驱动的CDK 4/6抑制剂的耐药性 缺陷概念验证研究将在两个具体目标中进行：（1）探索结合 CDK 4/6抑制剂和SMAPS在AMBRA 1缺陷背景下的作用，以及（2）评估SMAP-1的作用。 体内肿瘤模型中的CDK 4/6抑制剂组合。重要的是，除了解决 AMBRA 1缺陷，我们的概念验证研究结果预计将广泛适用于肿瘤窝藏 D型细胞周期蛋白水平升高和其他肿瘤相关改变导致的CDK活性异常。

Uncontrolled cell proliferation resulting from aberrant activity of cell cycle proteins is a hallmark of cancer. Overexpression of the mitogen sensor cyclin D1 is among the most frequent abnormalities in tumors, enhancing the activity of cyclin dependent kinases 4 and 6 (CDK4/6) to drive G1→S phase progression and promote cell survival and chemoresistance. Increased expression of D-type cyclins is required not only for tumorigenesis but also for tumor maintenance and progression. Thus, aberrant cyclin D-CDK4/6 activity represents an actionable target for cancer therapy and D-type cyclin function is among the top therapeutic targets for cancer management. Inhibitors of CDK4/6 activity have shown promise in the clinic and palbociclib, abemaciclib, and ribociclib are FDA-approved for use in patients. Several hundred clinical trials are currently ongoing to evaluate the antitumor effects of these agents in a broad spectrum of cancer types. However, the therapeutic promise of CDK4/6 inhibitors is dampened by inevitable emergence of resistance. Recent seminal studies have identified a novel mechanism of resistance to these agents mediated by deficiency of autophagy and beclin 1 regulator 1 (AMBRA1), an E3 ligase adaptor and master regulator of cyclin D1, D2, and D3 protein stability. Loss or mutation of AMBRA1 is seen in a significant subset of human cancers, in association with poor patient survival. AMBRA1 deficiency promotes the accumulation of D-type cyclins, a hyperproliferative phenotype, and tumorigenesis, while reducing the sensitivity of tumor cells to all three FDA-approved CDK4/6 inhibitors. Evidence that upregulation of D-type cyclins and the formation of non-canonical cyclin D-CDK2 and p27-cyclin D-CDK4 complexes underpins resistance to these agents forms the basis of this proposal. Strategies are proposed to explore the mechanism-driven application of Small Molecule Activators of PP2A (SMAPs) for overcoming resistance to CDK4/6 inhibitors in the context of AMBRA1 deficiency. SMAPs are a novel class of antitumor agents that selectively activate a subset of PP2A holoenzymes for potent tumor suppression in a variety of cancer types. This project builds on our discovery that SMAPs potently downregulate cyclins D1, D2 and D3 in all cell types tested. Importantly, SMAPs act as AMBRA1-independent D-type cyclin ‘degraders,’ promoting rapid proteolysis of these molecules via a proteasome-dependent mechanism that remains functional following loss of AMBRA1. Based on these findings, we hypothesize that combining CDK4/6 inhibitor treatment with a SMAP ‘D-type cyclin degrader’ will enhance antitumor activity and reverse resistance to CDK4/6 inhibitors driven by AMBRA1 deficiency. Proof-of-concept studies will be performed in two Specific Aims: (1) Explore the effects of combining CDK4/6 inhibitors and SMAPS in the context of AMBRA1-deficiency, and (2) Evaluate the effects of SMAP- CDK4/6 inhibitor combinations in tumor models in vivo. Importantly, in addition to addressing consequences of AMBRA1-deficiency, our proof-of-concept findings are anticipated to be broadly applicable to tumors harboring increased levels of D-type cyclins and aberrant CDK activity resulting from other tumor-associated alterations.