Comparative pharmacokinetics of seven propargyl-linked antifolate antibiotics in the mouse.

Antimicrobial resistance (AMR) to existing antibiotics poses a critical global health challenge, with significant morbidity and mortality from bacterial infections. Methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant strains (VISA, VRSA) are among the most pressing threats, particularly for vulnerable populations. To combat this crisis, the development of novel therapeutic strategies is imperative. We report the pharmacokinetic evaluation of a promising class of propargyl-linked diaminopyrimidine dihydrofolate reductase (DHFR) inhibitors with potent activity against drug-resistant bacteria, including MRSA and VISA strains. Previous studies have demonstrated minimum inhibitory concentration (MIC) values below 1 μg.mL-1 for several compounds in this series. Here, we detail the development and validation of an LC-QQQ bioanalytical method for seven propargyl-linked diaminopyrimidine analogues. Pharmacokinetic studies in a murine model across intravenous (IV), intraperitoneal (IP), and oral (PO) routes revealed substantial variability in parameters such as half-life (t₁/₂), area under the curve (AUC), and peak plasma concentration (Cmax). Compound 38C1 demonstrated favorable solubility, a higher maximum tolerated dose, and oral bioavailability of 20 %, making it a lead candidate. Pharmacokinetic-to-MIC ratio analyses showed that 38C1 maintained plasma concentrations significantly above MIC values for multiple S. aureus strains, including MRSA and VISA. These findings highlight 38C1 as a promising antifolate candidate for further development. Ongoing studies will assess its efficacy in infection models and refine delivery strategies to maximize therapeutic potential while mitigating resistance development.