In vitro metabolism, disposition, preclinical pharmacokinetics and prediction of human pharmacokinetics of DNDI-VL-2098, a potential oral treatment for Visceral Leishmaniasis.

The in vitro metabolism and in vivo pharmacokinetic (PK) properties of DNDI-VL-2098, a potential oral agent for Visceral Leishmaniasis (VL) were studied and used to predict its human pharmacokinetics. DNDI-VL-2098 showed a low solubility (10µM) and was highly permeable (>200nm/s) in the Caco-2 model. It was stable in vitro in liver microsomes and hepatocytes and no metabolite was detectable in circulating plasma from dosed animals suggesting very slow, if any, metabolism of the compound. DNDI-VL-2098 was moderate to highly bound to plasma proteins across the species tested (94-98%). DNDI-VL-2098 showed satisfactory PK properties in mouse, hamster, rat and dog with a low blood clearance (<15% of hepatic blood flow except hamster), a volume of distribution of about 3 times total body water, acceptable half-life (1-6h across the species) and good oral bioavailability (37-100%). Allometric scaling of the preclinical PK data to human gave a blood half-life of approximately 20h suggesting that the compound could be a once-a-day drug. Based on the above assumptions, the minimum efficacious dose predicted for a 50kg human was 150mg and 300mg, using efficacy results in the mouse and hamster, respectively.

Validation of 96-well equilibrium dialysis with non-radiolabeled drug for definitive measurement of protein binding and application to clinical development of highly-bound drugs.

Definitive plasma protein binding (PB) studies in drug development are routinely conducted with radiolabeled material, where the radiochemical purity limits quantitative PB measurement. Recent and emerging regulatory guidances increasingly expect quantitative determination of the fraction unbound (Fu) for key decision making. In the present study, PB of 11 structurally- and therapeutically-diverse drugs spanning the full range of plasma binding was determined by equilibrium dialysis of non-radiolabeled compound and was validated against the respective definitive values obtained by accepted radiolabeled protocols. The extent of plasma binding was in agreement with the radiolabeled studies; however, the methodology reported herein enables reliable quantification of Fu values for highly-bound drugs and is not limited by the radiochemical purity. In order to meet the rigor of a development study, equilibrium dialysis of unlabeled drug must be supported by an appropriately validated bioanalytical method along with studies to determine compound solubility and stability in matrix and dialysis buffer, nonspecific binding to the dialysis device, and ability to achieve equilibrium in the absence of protein. The presented methodology establishes an experimental protocol for definitive PB measurement, which enables quantitative determination of low Fu values, necessary for navigation of new regulatory guidances in clinical drug development.