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

Bc1-2是一种抗细胞凋亡蛋白，在血液系统恶性肿瘤中经常表达失调。VENTOCLAX是FDA批准的一种选择性bcl2抑制剂，已显示出对急性髓细胞白血病(AML)和淋巴瘤的非凡疗效，尽管治疗耐药性仍然是一个挑战。靶向蛋白降解(TPD)方式，如分子胶(MG)和蛋白水解靶向嵌合体(PROTACs)，都是通过劫持E3泛素连接酶来操作的，构成了药物研究的新兴课题。靶蛋白的高效和选择性失活可以绕过几个抑制剂的限制。然而，目前还没有针对bcl-2的选择性PROTAC。在我们的前期工作中，我们设计了各种由ventoclax衍生的PROTAC，它们劫持了三个不同的E3泛素连接酶，其中一些可以降解bclxl，但不能降解bcl2。我们推测，bcl2中的赖氨酸残基不能与三元复合体中的E3连接酶结合，这就是bcl2阻碍我们典型的PROTACs的原因。为了证明这一假设，我们将系统地筛选bcl2和bclxl中赖氨酸残基和蛋白质区域，它们是触发泛素化所必需的。我们的目标是通过招募多功能的E3连接酶和改变连接子和bcl2结合部分来合理地开发bcl2选择性降解物。这类化合物可以克服与bclxl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分析进行广泛筛选，所得到的数据将用于进一步的化学优化。生化研究，包括定量蛋白质组学，将被用来评估有希望的候选者的选择性。在AML细胞系和原代细胞中，将评估其抗癌活性和对线粒体凋亡途径的干扰。BCL-2选择性PROTAC将成为深入了解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，我们的研究可能会揭示新的化学方法来克服对蛋白质降解的抵抗。

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.