Preclinical Evaluation of the Antifolate QN254, 5-Chloro-N′6′-(2,5-Dimethoxy-Benzyl)-Quinazoline-2,4,6-Triamine, as an Antimalarial Drug Candidate

Abstract

Drug resistance against dihydrofolate reductase (DHFR) inhibitors-such as pyrimethamine (PM)-hasnow spread to almost all regions where malaria is endemic, rendering antifolate-based malaria treatments highly ineffective. We have previously shown that the di-amino quinazoline QN254 [5-chloro-N6-(2,5-dimethoxy-benzyl)-quinazoline-2,4,6-triamine] is active against the highly PM-resistant Plasmodium falciparum V1S strain, suggesting that QN254 could be used to treat malaria in regions with a high prevalence of antifolate resistance. Here, we further demonstrate that QN254 is highly active against Plasmodium falciparum clinical isolates, displaying various levels of antifolate drug resistance, and we provide biochemical and structural evidence that QN254 binds and inhibits the function of both the wild-type and the quadruple-mutant (V1S) forms of the DHFR enzyme. In addition, we have assessed QN254 oral bioavailability, efficacy, and safety in vivo. The compound displays favorable pharmacokinetic properties after oral administration in rodents. The drug was remarkably efficacious against Plasmodium berghei and could fully cure infected mice with three daily oral doses of 30 mg/kg. In the course of these efficacy studies, we have uncovered some dose limiting toxicity at higher doses that was confirmed in rats. Thus, despite its relative in vitro selectivity toward the Plasmodium DHFR enzyme, QN254 does not show the adequate therapeutic index to justify its further development as a single agent.

Lead Optimization of Aryl and Aralkyl Amine-Based Triazolopyrimidine Inhibitors of Plasmodium falciparum Dihydroorotate Dehydrogenase with Antimalarial Activity in Mice

“…These values are higher than reported for standard antimalarials (e.g. ED 50 's for chloroquine 1.8 mg/kg; mefloquine 3.8 mg/kg and artesunate 5.9 mg/kg) 20, 22 possibly explaining why 21 and 32 were not able to fully clear the infection. Regardless, 21 and 32 compare favorably in the Peter's test of P. berghei infection to the most potent of the N-alkyl-5-(1H-benzimidazol-1-yl)thiophene-2-carboxamides Pf DHODH inhibitors described by Booker et al , which also were reported to have an ED 50 of 10 - 15 mg/kg.…”

From crystal to compound: structure-based antimalarial drug discovery

“…Widespread use of PYR ( Figure 2) and CG, both of which target Pf DHFR-TS, has led to sitespecific mutations in the active site of the enzyme, rendering the drugs largely inactive. Current SBDD programmes now target the mutated versions of the enzyme, and use information of the restructured binding sites to design inhibitors with novel binding modes [17,108]. Conformationally constrained ligands cannot adapt to changes in the geometry of the binding site, and are therefore susceptible to the development of resistance.…”

Antifolate agents: a patent review (2006 – 2010)

“…However, additional structures of the wild-type or quadruple mutant enzyme bound to hits from an in silico screen, RJF01302 and RJF670 (PDB IDs: 3DG8, 3DGA) reported in 2009 show that the small, lead-like nature of the RJF compounds avoid the steric interactions with the S108N mutant 30 . A structure of the quadruple P. falciparum DHFR mutant, resistant to pyrimethamine, was reported in 2010 bound to QN254 (PDB ID: 3JSU), an antimalarial preclinical drug candidate that showed promising activity against P. berghei but was abandoned because of dose limiting toxicity 31 . QN254, a quinazoline inhibitor, shows an increased number of interactions relative to WR99210, a dihydrotriazine inhibitor: a water-mediated hydrogen bond with Asn 108, additional van der Waals and π-π interactions with Leu 46, Met 55 and Phe 58.…”