A simple in vitro PK/PD model system to determine time-kill curves of drugs against Mycobacteria

In vivo tuberculosis is exposed to continually changing drug concentrations for which static minimum inhibitory concentration (MIC) testing may be a poor surrogate. While in vitro approaches to determine time-kill curves for antibiotics have been widely applied in assessing antimicrobial activity against fast growing microorganisms, their availability and application for slow-growing microorganisms including Mycobacterium tuberculosis has so far been scarce. Thus, we developed a novel simple in vitro pharmacokinetic/pharmacodynamic (PK/PD) model for establishing time-kill curves and applied it for evaluating the antimicrobial activity of different dosing regimens of isoniazid (INH) against Mycobacterium bovis BCG as a surrogate for virulent M. tuberculosis. In the in vitro model M. bovis BCG was exposed to INH concentration-time profiles as usually encountered during multiple dose therapy with 25, 100 and 300mg/day in humans who are fast or slow INH metabolizers. Bacterial killing was followed over time by determining viable counts and the resulting time-kill data was analyzed using a semi-mechanistic PK/PD model with an adaptive IC(50) function to describe the emergence of insensitive populations of bacteria over the course of treatment. In agreement with previous studies, the time-kill data suggest that AUC(0-24)/MIC is the PK/PD index that is the most explanatory of the antimicrobial effect of INH. The presented in vitro PK/PD model and associated modeling approach were able to characterize the time-kill kinetics of INH in M. bovis BCG, and may in general serve as a potentially valuable, low cost tool for the assessment of antibacterial activity in slow-growing organisms in drug development and applied pharmacotherapy.