Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, Maryland 21205, United States.
Department of Chemistry, Johns Hopkins University, 3400 North Charles Street, Baltimore, Maryland 21218, United States.
ACS Chem Biol. 2023 Oct 20;18(10):2200-2210. doi: 10.1021/acschembio.3c00118. Epub 2023 May 26.
Sterile alpha motif histidine-aspartate domain protein 1 (SAMHD1) is a deoxynucleotide triphosphohydrolase that exists in monomeric, dimeric, and tetrameric forms. It is activated by GTP binding to an A1 allosteric site on each monomer subunit, which induces dimerization, a prerequisite for dNTP-induced tetramerization. SAMHD1 is a validated drug target stemming from its inactivation of many anticancer nucleoside drugs leading to drug resistance. The enzyme also possesses a single-strand nucleic acid binding function that promotes RNA and DNA homeostasis by several mechanisms. To discover small molecule inhibitors of SAMHD1, we screened a custom ∼69 000-compound library for dNTPase inhibitors. Surprisingly, this effort yielded no viable hits and indicated that exceptional barriers for discovery of small molecule inhibitors existed. We then took a rational fragment-based inhibitor design approach using a deoxyguanosine (dG) A1 site targeting fragment. A targeted chemical library was synthesized by coupling a 5'-phosphoryl propylamine dG fragment (dGpCNH) to 376 carboxylic acids (RCOOH). Direct screening of the products (dGpCNHCO-R) yielded nine initial hits, one of which (R = 3-(3'-bromo-[1,1'-biphenyl]), ) was investigated extensively. Amide is a competitive inhibitor against GTP binding to the A1 site and induces inactive dimers that are deficient in tetramerization. Surprisingly, also prevented ssDNA and ssRNA binding, demonstrating that the dNTPase and nucleic acid binding functions of SAMHD1 can be disrupted by a single small molecule. A structure of the SAMHD1- complex indicates that the biphenyl fragment impedes a conformational change in the C-terminal lobe that is required for tetramerization.
sterile alpha motif histidine-aspartate domain protein 1 (SAMHD1) 是一种脱氧核苷酸三磷酸水解酶，以单体、二聚体和四聚体形式存在。它通过每个单体亚基上的 A1 变构位点与 GTP 结合而被激活，这诱导二聚体化，这是 dNTP 诱导四聚体化的先决条件。SAMHD1 是一个经过验证的药物靶点，源于其使许多抗癌核苷药物失活，导致药物耐药。该酶还具有单链核酸结合功能，通过多种机制促进 RNA 和 DNA 稳态。为了发现 SAMHD1 的小分子抑制剂，我们筛选了一个定制的 ∼69000 种化合物文库，以寻找 dNTPase 抑制剂。令人惊讶的是，这项工作没有产生可行的命中结果，并表明发现小分子抑制剂存在特殊的障碍。然后，我们采用基于理性的片段抑制剂设计方法，使用脱氧鸟苷 (dG) A1 位点靶向片段。通过将 5'-磷酸丙胺 dG 片段 (dGpCNH) 偶联到 376 种羧酸 (RCOOH) 来合成靶向化学文库。直接筛选产物 (dGpCNHCO-R) 得到了九个初始命中结果，其中一个 (R = 3-(3'-溴-[1,1'-联苯])，) 进行了广泛研究。酰胺是 A1 位点与 GTP 结合的竞争性抑制剂，并诱导缺乏四聚体化的无活性二聚体。令人惊讶的是，还阻止了 ssDNA 和 ssRNA 结合，表明 SAMHD1 的 dNTPase 和核酸结合功能可以被单个小分子破坏。SAMHD1-复合物的结构表明，联苯片段阻碍了 C 末端结构域发生构象变化，这是四聚体化所必需的。
