Human DNA Cross-Link Repair 1A (DCLRE1A. SNM1A); A Target Enabling Package

Cancer cells experience genomic instability, probably through a combination of excessive replicative activity and the loss of function of checkpoint and DNA repair pathways that may have contributed to the oncogenic transformation. Chemotherapy by DNA-damaging agents such as cisplatin and nitrogen mustards create DNA interstrand crosslinks (ICL), which can lead to double-strand breaks and cell death when the cells replicate their DNA. Genotoxic drugs are counteracted by the cell’s DNA damage response. Hence, it is expected that inhibiting DNA repair proteins would sensitise cells to chemotherapy.
Here we address an enzyme that participates in the repair of ICLs, DCLRE1A. The TEP includes expression clones and methods for producing the catalytic domain and high-throughput activity assays. Furthermore, we provide a crystallization system that generates thousands of reproducible crystals that allow soaking of small-molecule ligands. We provide crystal structures of several small molecule fragments and inhibitors, opening the way to development of more potent and selective inhibitors.

癌细胞会发生基因组不稳定性（genomic instability），该现象可能由过度的复制活性与检查点、DNA修复通路功能丧失共同介导，而这些因素或可促成致癌转化。采用顺铂（cisplatin）、氮芥（nitrogen mustards）等DNA损伤剂开展的化疗，可诱导DNA链间交联（DNA interstrand crosslinks, ICL）；当细胞进行DNA复制时，这类交联会引发双链断裂并最终导致细胞死亡。基因毒性药物的细胞毒性会被细胞的DNA损伤应答（DNA damage response）通路所拮抗。因此，抑制DNA修复蛋白有望使肿瘤细胞对化疗药物增敏。本研究针对参与DNA链间交联修复的酶DCLRE1A展开探讨。TEP包含表达克隆、催化结构域的制备方法以及高通量活性检测实验方案。此外，我们开发了一套结晶体系，可制备数千颗可重复的晶体，支持小分子配体浸泡实验。本研究还提供了多种小分子片段与抑制剂的晶体结构，为开发更强效、更具选择性的靶向抑制剂开辟了道路。