Small Molecule Activators of Pro-apoptotic BAX for Cancer Therapy

摘要 对细胞凋亡的抵抗是癌症的标志之一。癌细胞通过 线粒体凋亡途径，以确保肿瘤的生长、维持和对当前治疗的耐药性。 线粒体凋亡途径受bcl2家族的表达水平和相互作用的影响 蛋白质，包括促凋亡效应蛋白Bax和BAK，抗凋亡蛋白，如bcl. 2、BCL-XL、MCL-1和促凋亡蛋白BH3-Only。癌细胞最常阻断线粒体 通过上调抗凋亡的bcl2蛋白来中和BH3-only蛋白并激活bax实现细胞凋亡 还有BAK。因此，人们致力于开发抗细胞凋亡的bcl-2选择性抑制剂。 重新激活细胞凋亡的蛋白质。FDA批准的VENTOTCLAX等抑制剂依赖于该功能和 激活Bax和BAK的仅BH3蛋白的可用性。然而，在许多肿瘤中，只有BH3蛋白可以 下调、抑制或缺失，使这些肿瘤对这些抑制物不敏感，并限制其 临床应用范围更广。我们假设小分子直接激活促凋亡的bax是通过 BAX触发点是一种替代的和可能互补的药理策略，以促进细胞凋亡。 癌细胞。这种方法可以促进Bax的激活，而不依赖于BH3-Only蛋白，因此 应该有可能克服耐药肿瘤中的细胞凋亡障碍。我们实验室最近用了 独特的结构和分子洞察力和药物化学，以开发有效和选择性的化合物， 称为BAX触发位点激活剂1(BTSA1)，促进BAX激活并诱导线粒体 功能障碍和细胞凋亡。使用BTSA1，我们提供了直接激活BAX作为治疗的概念验证 靶点在急性髓系白血病，并证明直接激活Bax在体内具有良好的耐受性。在这里，我们 假设生成的BTSA1类似物具有更好的效力、口服生物利用度和药代动力学。 我们的目标是评估它们在不同癌症模型中的活性和作用机制，作为单一药物或 联合治疗，并探讨其对Bax激活和耐药的敏感和耐药机制 细胞凋亡。此外，我们的目标是确定一个临床候选的Bax激活剂，具有良好的细胞， 药理和安全性能。因此，我们提出了以下具体目标：1)表征 BTSA1的效力、选择性、促凋亡活性、体外ADME/Tox和药代动力学特性 类似物，2)确定第二代BTSA的细胞效力和作用机制，包括BTSA1.2， 使用各种白血病和实体瘤细胞单独和联合治疗并研究决定因素 敏感性和耐药性，3)确定选择的BTSA的安全性和治疗潜力 联合治疗，并研究新的生物标志物和细胞凋亡调控机制。这项建议 将推进创新的治疗策略和疗法，并提供最适合的背景 靶向癌症中Bax的激活。

ABSTRACT Resistance to apoptosis is one of the hallmarks of cancer. Cancer cells circumvent cell death through the mitochondrial apoptosis pathway to ensure tumor growth, maintenance and resistance to current treatments. The mitochondrial apoptosis pathway is governed by the expression levels and interactions of the BCL-2 family proteins, which comprise the pro-apoptotic effector proteins BAX and BAK, the anti-apoptotic proteins e.g. BCL- 2, BCL-XL, MCL-1 and the pro-apoptotic BH3-only proteins. Cancer cells most commonly block mitochondrial apoptosis by upregulating the anti-apoptotic BCL-2 proteins that neutralize BH3-only proteins and activated BAX and BAK. Therefore, efforts have focused on the development of selective inhibitors of anti-apoptotic BCL-2 proteins to re-activate apoptosis. Inhibitors such as the FDA-approved Venetoclax rely on the function and availability of BH3-only proteins to activate BAX and BAK. However, in many tumors, BH3-only proteins can be downregulated, suppressed or deleted, making these tumors insensitive to these inhibitors and limiting their broader clinical application. We hypothesized that small-molecule direct activation of pro-apoptotic BAX via the BAX trigger site is an alternative and possibly complementary pharmacological strategy to promote apoptosis in cancer cells. This approach can promote BAX activation independently of BH3-only proteins and therefore should have the potential to overcome apoptosis blockade in resistant tumors. Our laboratory recently used unique structural and molecular insights and medicinal chemistry to develop a potent and selective compound, termed BAX Trigger Site Activator 1(BTSA1) that promotes BAX activation and induces mitochondrial dysfunction and apoptosis. Using BTSA1, we provided proof-of-concept for direct BAX activation as a therapeutic target in Acute Myeloid Leukemia and demonstrated that direct BAX activation is well tolerated in vivo. Here, we hypothesized to generate BTSA1 analogues with improved potency, oral bioavailability and pharmacokinetics. Our goal is to evaluate their activity and mechanism of action in diverse cancer models as single agents or combination treatments and investigate mechanisms of sensitivity and resistance to BAX activation and apoptosis. Moreover, we aim to identify a clinical candidate BAX activator with favorable cellular, pharmacological and safety properties. Therefore, we propose the following specific aims: 1) characterize potency, selectivity, pro-apoptotic activity, in vitro ADME/Tox and pharmacokinetic properties of BTSA1 analogues, 2) determine cellular efficacy and mechanism of action of 2nd generation BTSAs, including BTSA1.2, alone and in combination treatments using various leukemia and solid tumor cells and investigate determinants of sensitivity and resistance, 3) determine safety and therapeutic potential of select BTSAs alone or in combination therapy and investigate novel biomarkers and mechanisms of apoptosis regulation. This proposal will advance an innovative therapeutic strategy and therapeutics and inform the most suitable context for targeting BAX activation in cancer.