Dihydrofolate reductase inhibitors – applications to Chagas disease and dental caries

Dihydrofolate reductase (DHFR) is a ubiquitous enzyme present in almost all eukaryote and prokaryote organisms. This enzyme participates in the folate metabolism, which is essential for the synthesis of DNA, RNA, and proteins and is thus vital to all living cells. The inhibitors of the enzymes within the folate pathway are referred to as antifolates. Antifolates have been used as therapeutic drugs against various diseases. The research presented in this dissertation focus on the discovery inhibitors of Trypanosoma cruzi DHFR (TcDHFR) as potential treatments of Chagas disease and inhibitors of Streptococcus mutans ( SmDHFR) as potential agents for the treatment and prevention of dental caries.

Taking advantage of the subtle differences in the architecture of T. cruzi and the human DHFRs, we have designed and synthesized a series of TMQ analogs as selective inhibitors of TcDHFR. While retaining the nanomolar activity of TMQ, the study resulted in the identification of inhibitors that are about 500 fold more selective. Attempts have also been made to discover new structural scaffolds of the TcDHFR inhibitors using in silico of commercial compound libraries against TcDHFR active site using the software FlexX integrated in LeadIT 2.1.0. Five new structural scaffolds of TcDHFR inhibitors with low micromolar / nanomolar activities were identified. The best compound identified from these studies displayed 33% inhibition of TcDHFR at 25 nM and about 30 fold selectivity towards inhibiting TcDHFR.

Streptococcus mutans (S. mutans) is the major etiological agent for dental caries. The formation of tenacious biofilms is the hallmark of S. mutans induced pathogenesis of dental caries. Thus, drugs inhibiting S. mutans biofilms have the potential to be used in the treatment and prevention of this disease. With the objective of identifying potential antibiofilm agents, TMQ and several of its analogs have been evaluated against S. mutans growth and biofilm. This study revealed that these compounds strongly inhibited S. mutans growth and biofilm. They were also found to be potent inhibitors of S. mutans DHFR. The least active compounds against biofilm and growth also had a markedly decreased activity against SmDHFR indicating that their antibacterial effects are caused by SmDHFR inhibition.