Expanding the Landscape of Dual Action Antifolate Antibacterials through 2,4-Diamino-1,6-dihydro-1,3,5-triazines

Antibiotics that operate via multiple
mechanisms
of action are a promising strategy to combat growing resistance. Previous
studies have shown that dual action antifolates formed from a pyrroloquinazolinediamine
core can inhibit the growth of bacterial pathogens without developing
resistance. In this work, we expand the scope of dual action antifolates
by repurposing the 2,4-diamino-1,6-dihydro-1,3,5-triazine (DADHT)
cycloguanil scaffold to a variety of derivatives designed to inhibit
dihydrofolate reductase (DHFR) and disrupt bacterial membranes. Dual
mechanism DADHTs have activity against a variety of target pathogens,
including Mycobacterium tuberculosis, Mycobacterium abscessus, and Pseudomonas aeruginosa, among other ESKAPEE organisms. Through X-ray crystallography, we confirmed engagement
of the Escherichia coli DHFR target
and found that some DADHTs stabilize a previously unobserved conformation
of the enzyme but, broadly, bind in the occluded conformation. Using in vitro inhibition of purified E. coli and Staphylococcus aureus DHFR and
disruption of E. coli membranes, we
determined that alkyl substitution of dihydrotriazine at the 6-position
best optimizes the DADHT’s two mechanisms of action. By employing
both mechanisms, the DADHT spectrum of activity was extended beyond
the scope of traditional antifolates. We are optimistic that the dual
mechanism approach, particularly through the action of antifolates,
offers a unique means of combating hard-to-treat bacterial infections.