Roberts Allison M, Miyamoto David K, Huffman Tucker R, Bateman Leslie A, Ives Ashley N, Akopian David, Heslin Martin J, Contreras Carlo M, Rape Michael, Skibola Christine F, Nomura Daniel K
Departments of Chemistry, Molecular and Cell Biology, and Nutritional Sciences and Toxicology, 127 Morgan Hall, University of California, Berkeley , Berkeley, California 94720, United States.
Department of Molecular and Cell Biology, University of California, Berkeley , Berkeley, California 94720, United States.
ACS Chem Biol. 2017 Apr 21;12(4):899-904. doi: 10.1021/acschembio.7b00020. Epub 2017 Feb 15.
Chemical genetic screening of small-molecule libraries has been a promising strategy for discovering unique and novel therapeutic compounds. However, identifying the targets of lead molecules that arise from these screens has remained a major bottleneck in understanding the mechanism of action of these compounds. Here, we have coupled the screening of a cysteine-reactive fragment-based covalent ligand library with an isotopic tandem orthogonal proteolysis-enabled activity-based protein profiling (isoTOP-ABPP) chemoproteomic platform to rapidly couple the discovery of lead small molecules that impair pancreatic cancer pathogenicity with the identification of druggable hotspots for potential cancer therapy. Through this coupled approach, we have discovered a covalent ligand DKM 2-93 that impairs pancreatic cancer cell survival and in vivo tumor growth through covalently modifying the catalytic cysteine of the ubiquitin-like modifier activating enzyme 5 (UBA5), thereby inhibiting its activity as a protein that activates the ubiquitin-like protein UFM1 to UFMylate proteins. We show that UBA5 is a novel pancreatic cancer therapeutic target and show DKM 2-93 as a relatively selective lead inhibitor of UBA5. Our results underscore the utility of coupling the screening of covalent ligand libraries with isoTOP-ABPP platforms for mining the proteome for druggable hotspots for cancer therapy.
小分子文库的化学遗传筛选一直是发现独特新型治疗化合物的一种有前景的策略。然而，确定这些筛选中产生的先导分子的靶点仍然是理解这些化合物作用机制的一个主要瓶颈。在此，我们将基于半胱氨酸反应性片段的共价配体文库筛选与基于同位素串联正交蛋白酶解的活性蛋白谱分析（isoTOP-ABPP）化学蛋白质组学平台相结合，以快速将损害胰腺癌致病性的先导小分子的发现与潜在癌症治疗的可成药热点的鉴定联系起来。通过这种联合方法，我们发现了一种共价配体DKM 2-93，它通过共价修饰泛素样修饰激活酶5（UBA5）的催化半胱氨酸来损害胰腺癌细胞存活和体内肿瘤生长，从而抑制其作为一种将泛素样蛋白UFM1激活以对蛋白质进行UFMylation修饰的蛋白质的活性。我们表明UBA5是一种新型胰腺癌治疗靶点，并展示了DKM 2-93作为UBA5的一种相对选择性先导抑制剂。我们的数据强调了将共价配体文库筛选与isoTOP-ABPP平台相结合用于在蛋白质组中挖掘癌症治疗可成药热点的实用性。
