Discovery of BCL-2-selective degraders for applications in hematological malignancies

BCL-2是一种抗凋亡蛋白，在血液恶性肿瘤中经常失调。维奈托克是FDA批准的选择性BCL-2抑制剂，已证明对急性髓性白血病（AML）和淋巴瘤具有非凡的疗效，尽管治疗耐药性仍然是一个挑战。靶向蛋白降解（TPD）模式，如分子胶（MG）和蛋白水解靶向嵌合体（PROTAC），两者都通过劫持E3泛素连接酶来操作，构成药物研究的新兴主题。靶蛋白的有效和选择性失活可以规避几种抑制剂的限制。然而，目前没有可用于BCL-2的选择性PROTAC。在我们的初步工作中，我们设计了各种维奈托克衍生的PROTAC，它们劫持了三种不同的E3泛素连接酶，其中一些降解BCL-XL，但不降解BCL-2。我们假设BCL-2中的赖氨酸残基对三元复合物内的E3连接酶的不可接近性解释了BCL-2对我们的示例性PROTAC的阻碍。为了证明这一假设，我们将系统地筛选BCL-2和BCL-XL中对引发泛素化至关重要的赖氨酸残基和蛋白质区域。我们的目标是合理地开发BCL-2-选择性降解剂，通过招募通用的E3连接酶和改变接头和BCL-2结合部分。这样的化合物可以克服与BCL-XL抑制剂和降解剂相关的毒性限制。BCL-2 PROTAC将使用高通量、基于荧光的蛋白质稳定性测定进行广泛筛选，所得数据将用于进一步的化学优化。生物化学研究，包括定量蛋白质组学，将被用来评估有前途的候选人的选择性。在AML细胞系和原代细胞中，将评价抗癌活性和对线粒体凋亡途径的干扰。BCL-2选择性PROTAC将是深入了解BCL-2功能的有力工具，并可能导致新的癌症治疗方法的开发。更广泛地说，我们的研究可能揭示新的化学方法来克服蛋白质降解的阻力。

BCL-2 is an antiapoptotic protein that is frequently dysregulated in hematological malignancies. Venetoclax, an FDA-approved selective BCL-2 inhibitor, has demonstrated extraordinary efficacy against acute myeloid leukemia (AML) and lymphoma, although treatment resistance remains a challenge. Targeted protein degradation (TPD) modalities such as molecular glues (MGs) and proteolysis targeting chimeras (PROTACs), both operating by hijacking E3 ubiquitin ligases, constitute emerging topics of drug research. The efficient and selective inactivation of target proteins can circumvent several inhibitor limitations. However, no selective PROTACs are currently available for BCL-2. In our preliminary work, we designed various venetoclax-derived PROTACs that hijack three distinct E3 ubiquitin ligases, some of which degrade BCL-XL but not BCL-2. We postulate that the inaccessibility of lysine residues in BCL-2 to the E3 ligase inside the ternary complex accounts for the hindrance of BCL-2 to our exemplary PROTACs. To prove this hypothesis, we will systematically screen lysine residues and protein areas in BCL-2 and BCL-XL that are essential to trigger ubiquitination. We aim to rationally develop BCL-2-selective degraders by recruiting versatile E3 ligases and altering the linker and BCL-2-binding moieties. Such compounds may overcome toxicity constraints associated with BCL-XL inhibitors and degraders. BCL-2 PROTACs will be extensively screened using a high-throughput, fluorescence-based protein stability assay, and the resulting data will be used for further chemical optimization. Biochemical research, including quantitative proteomics, will be utilized to evaluate the selectivity of promising candidates. In AML cell lines and primary cells, the anticancer activity and interference with the mitochondrial apoptotic pathway will be evaluated. BCL-2-selective PROTACs will be powerful tools for gaining a deeper understanding of BCL-2 functions and may lead to the development of novel cancer treatments. More broadly, our studies may reveal new chemical approaches to overcome resistance to protein degradation.