DataSheet1_Targeting the Essential Transcription Factor HP1043 of Helicobacter pylori: A Drug Repositioning Study.pdf

Antibiotic-resistant bacterial pathogens are a very challenging problem nowadays. Helicobacter pylori is one of the most widespread and successful human pathogens since it colonizes half of the world population causing chronic and atrophic gastritis, peptic ulcer, mucosa-associated lymphoid tissue-lymphoma, and even gastric adenocarcinoma. Moreover, it displays resistance to numerous antibiotics. One of the H. pylori pivotal transcription factors, HP1043, plays a fundamental role in regulating essential cellular processes. Like other bacterial transcription factors, HP1043 does not display a eukaryote homolog. These characteristics make HP1043 a promising candidate to develop novel antibacterial strategies. Drug repositioning is a relatively recent strategy employed in drug development; testing approved drugs on new targets considerably reduces the time and cost of this process. The combined computational and in vitro approach further reduces the number of compounds to be tested in vivo. Our aim was to identify a subset of known drugs able to prevent HP1043 binding to DNA promoters. This result was reached through evaluation by molecular docking the binding capacity of about 14,350 molecules on the HP1043 dimer in both conformations, bound and unbound to the DNA. Employing an ad hoc pipeline including MMGBSA molecular dynamics, a selection of seven drugs was obtained. These were tested in vitro by electrophoretic mobility shift assay to evaluate the HP1043–DNA interaction. Among these, three returned promising results showing an appreciable reduction of the DNA-binding activity of HP1043. Overall, we applied a computational methodology coupled with experimental validation of the results to screen a large number of known drugs on one of the H. pylori essential transcription factors. This methodology allowed a rapid reduction of the number of drugs to be tested, and the drug repositioning approach considerably reduced the drug design costs. Identified drugs do not belong to the same pharmaceutical category and, by computational analysis, bound different cavities, but all display a reduction of HP1043 binding activity on the DNA.

当下，耐药细菌性病原菌是极具挑战性的公共卫生难题。幽门螺杆菌（Helicobacter pylori）是全球分布最广、适应性最强的人类病原菌之一，其定植于全球半数人口体内，可引发慢性萎缩性胃炎、消化性溃疡、黏膜相关淋巴组织淋巴瘤，甚至胃腺癌；此外，该菌对多种抗生素均表现出耐药性。HP1043作为幽门螺杆菌的关键转录因子之一，在调控细胞核心生理过程中发挥着基础性作用，与其他细菌转录因子类似，HP1043不存在真核生物同源物，上述特性使得HP1043成为开发新型抗菌策略的极具潜力的靶点。药物重定位（drug repositioning）是药物研发领域近年来兴起的创新策略：将已获批药物应用于新靶点，可大幅缩减该研发流程的时间与成本，而结合计算与体外实验的研究方法，可进一步减少需开展体内实验的化合物数量。本研究旨在筛选出可阻断HP1043与DNA启动子结合的已知药物子集，为此，我们通过分子对接（molecular docking）技术，评估了约14350个化合物在HP1043二聚体（分别处于结合DNA与未结合DNA两种构象下）的结合能力；通过搭建包含MMGBSA分子动力学分析的定制化分析流程，最终筛选得到7种候选药物，随后通过电泳迁移率变动实验（electrophoretic mobility shift assay，EMSA）对这7种药物开展体外实验，以验证其对HP1043-DNA相互作用的影响，其中3种药物表现出优异的实验结果，可显著降低HP1043的DNA结合活性。综上，本研究将计算方法与实验验证相结合，针对幽门螺杆菌的核心转录因子之一，完成了大规模已知药物的筛选工作，该方法可快速缩减需开展实验的药物数量，而药物重定位策略则大幅降低了药物研发的成本；筛选得到的药物分属不同的药学类别，通过计算分析可知它们结合HP1043的不同空腔，但均可降低HP1043的DNA结合活性。