Establishing translational performance standards for TB therapy using rifampicin-based regimens in a male and female high-burden murine model.

BACKGROUND: Establishing efficacy benchmarks in preclinical tuberculosis (TB) models is essential for optimizing and prioritizing therapeutic regimens. However, standardized classification methods for comparing high-performing regimens are currently lacking. This study defines a quantitative framework utilizing rifampicin-based regimens in a high-burden aerosol BALB/c mouse model, incorporating both male and female mice to assess potential sex-specific treatment responses. METHODS: Mice were infected with Mycobacterium tuberculosis Erdman strain and treated for 4 or 8 weeks with rifampicin (R), rifampicin plus pyrazinamide (RZ), or rifampicin, isoniazid, and pyrazinamide (RHZ). Treatments were administered orally five days a week. The bacterial burden in the lungs and spleens was quantified by CFU enumeration. Pharmacokinetic analysis confirmed drug exposures. To establish classification benchmarks, treatment efficacy was evaluated using quartile performance thresholds and Cohen's d effect size analysis. RESULTS: All regimens reduced lung CFUs compared to controls. RHZ demonstrated a high benchmark, achieving mean reductions of 3 ± 0.5 Log(10) CFUs at 4 weeks and 4 ± 0.4 Log(10) CFUs at 8 weeks, with clearance below detection limits in most mice. The R and RZ regimens achieved intermediate reductions. No statistically significant sex differences in bacterial clearance were observed. Pharmacokinetic analysis confirmed equivalent drug exposures across sexes. Quartile ranking (> 75th percentile) and Cohen's d calculations (Cohen's d > 15) consistently classified RHZ as the benchmark high-performing regimen at both time points, showing exceptional efficacy. CONCLUSION: This study establishes a quantitative framework for evaluating TB treatments in a preclinical high-burden BALB/c mouse model. The dual-metric classification framework provides sex-inclusive, quantitative performance criteria that enhance the translational relevance of preclinical efficacy studies. This approach sets relative benchmarks that support the comparative evaluation of novel regimens and helps to align preclinical performance with clinical expectations.