RNA Polymerase 1 inhibitors against African trypanosomes <i>in vitro</i> and in mice

Therapy of human African trypanosomiasis, fatal if not treated, has been greatly improved by orally available and safer fexinidazole, but reliable cure of late-stage central nervous system (CNS) infection remains a challenge. African trypanosomes are unique among eukaryotes in having an RNA Polymerase 1 that transcribes not only rRNA but also mRNA that encodes the abundant variable surface glycoprotein (VSG) that coats the cell membrane of bloodstream-form parasites and allows them to evade host immune defenses. RNA Pol 1 inhibitor BMH-21, an intercalator not associated with DNA damage and a lead for new cancer therapy, has reported activity against the synthesis of rRNA and VSG mRNA in <i>Trypanosoma brucei brucei</i>. We evaluated a library of BMH-21 analogs against bloodstream-form <i>T. b. brucei in vitro</i> and found limited tolerance for structural modification. Striking and time-dependent bimodal dose-response curves indicate these compounds have a complex mechanism of action. Most potent against <i>T. b. brucei</i> was benzopyridoquinazoline Compound <b>2</b>, with an EC₅₀ of 84 nM. Analysis of <i>in vitro</i> pharmacokinetic-pharmacodynamic (PK-PD) relationships by a hollow fiber model system revealed that in <i>T. b. brucei</i>, the kinetic driver of Compound <b>2</b> is drug concentration, a finding that was confirmed by dose fractionation in infected mice and that highlights the utility of the <i>in vitro</i> system for assessing antitrypanosomal PK-PD. Although Compound <b>2</b> accumulates 20-fold in mouse brain tissue vs plasma, efficacy against CNS <i>T. b. brucei</i> in mice was limited. RNA Pol 1 remains an attractive target for developing new and more selectively toxic antitrypanosomal agents.