Fragment‐Merging Strategies with Known Pyrimidine Scaffolds Targeting Dihydrofolate Reductase from Mycobacterium tuberculosis

Dihydrofolate reductase (DHFR) is a key enzyme involved in the folate pathway that has been heavily targeted for the development of therapeutics against cancer and bacterial and protozoa infections amongst others. Despite being an essential enzyme for Mycobacterium tuberculosis (Mtb) viability, DHFR remains an underexploited target for tuberculosis (TB) treatment. Herein, we report the preparation and evaluation of a series of compounds against Mtb DHFR (MtbDHFR). The compounds have been designed using a merging strategy of traditional pyrimidine‐based antifolates with a previously discovered unique fragment hit against MtbDHFR. In this series, four compounds displayed a high affinity against MtbDHFR, with sub‐micromolar affinities. Additionally, we determined the binding mode of six of the best compounds using protein crystallography, which revealed occupation of an underutilised region of the active site. Tuberculosis, caused by Mycobacterium tuberculosis (Mtb), remains a major global health problem, with rising resistance increasing the need for more therapeutic options. Through the use of fragment‐based drug discovery, a compound found from screening was merged with known inhibitors, creating compounds with sub‐micromolar affinity against Mtb dihydrofolate reductase, an essential enzyme for Mtb.