CETSA proteomics to investigate drug binding in BJ Ras Sv40 cells treated with MTH1 inhibitors.

Previously, we showed cancer cells rely on the MTH1 protein to prevent incorporation of otherwise deadly oxidative nucleotides into DNA and we developed MTH1 inhibitors which selectively kill cancer cells. Recently, several new and potent inhibitors of MTH1 were demonstrated to be non-toxic to cancer cells, challenging the utility of MTH1 inhibition as a target for cancer treatment. Here, we demonstrate that these recently described MTH1 inhibitors, which fail to kill cancer cells, also fail to introduce the toxic oxidized nucleotides into DNA. We further describe a new MTH1 inhibitor TH1579, an analogue of TH588, which is a potent, selective MTH1 inhibitor with good oral availability and demonstrates excellent pharmacokinetic and anti-cancer properties in vivo. Furthermore, we describe TH1579 as a best-in-class MTH1 inhibitor, which we expect to be useful in order to test the MTH1 inhibitor concept. Cellular thermal shift assay (CETSA) combined with basic reversed phase (bRP) separation and LC-MS was used to study drug binding in BJ Ras Sv40 cells treated with the MTH1 inhibitor TH1579.

既往研究表明，癌细胞依赖MTH1蛋白（MTH1 protein）以避免具有致死性的氧化核苷酸掺入DNA中，我们此前也开发了可选择性杀伤癌细胞的MTH1抑制剂。近期有研究显示，多款新型强效MTH1抑制剂对癌细胞无毒性，这对以MTH1抑制作为癌症治疗靶点的应用价值提出了挑战。本研究证实，上述新近报道的无法杀伤癌细胞的MTH1抑制剂，同样无法将有毒性的氧化核苷酸引入DNA中。本研究还报道了一种新型MTH1抑制剂TH1579——它是TH588的类似物，属于强效且高选择性的MTH1抑制剂，具备良好的口服生物利用度，并在体内展现出优异的药代动力学与抗肿瘤特性。此外，本研究将TH1579定义为同类最优MTH1抑制剂，我们认为其可用于验证MTH1抑制剂这一治疗概念的可行性。本研究采用细胞热位移测定法（Cellular Thermal Shift Assay, CETSA）结合碱性反相（basic reversed phase, bRP）分离与液相色谱-质谱联用（LC-MS）技术，针对经MTH1抑制剂TH1579处理的BJ Ras Sv40细胞开展药物结合研究。