Mechanistic simulations explain paradoxical saquinavir metabolism during in vitro vectorial transport study.
We were confronted by an unexpected observation: in a Transwell study of confluent, Cyp3A4 and P-gp-expressing Caco-2 cells, higher intracellular saquinavir levels, yet less first metabolite (M7) formation were observed following apical dosing, compared to basal dosing. To test two seemingly plausible hypothesized explanations, we constructed an in silico working analogue using the synthetic method. Neither mechanism alone was sufficient, but when combined and tuned within the analogue, the results generated were a semi-quantitative match to the experimental data. After 60 cycles, more of the simulated dose was present within analogue cells as parent drug after apical dosing. Furthermore, less M7 was present after apical dose. The paradox disappeared by having simulated drugs equilibrate among separate intracellular zones. Building, studying, and exploring mechanistic explanations for complex wet-lab phenomena using the new methods improved insight into the referent system, while providing a straightforward, scientific means of testing the plausibility of mechanistic hypotheses.
Lam
Tai Ning
TN
Dept. of Bioengineering and Therapeutic Sciences, 513 Parnassus Ave., S-926, University of California, San Francisco, CA 94143, USA. tai.ning,lam@ucsf.edu
Hunt
C Anthony
CA
D001693:Biological Transport, Active / Q000502:physiology; D018938:Caco-2 Cells; D003198:Computer Simulation; D017320:HIV Protease Inhibitors / Q000493:pharmacokinetics; D006801:Humans; D008657:Metabolic Clearance Rate; D008954:Models, Biological; D019258:Saquinavir / Q000493:pharmacokinetics*; D015398:Signal Transduction / Q000502:physiology*
D017320:HIV Protease Inhibitors; D019258:Saquinavir