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Dose-range studies for polymyxin B (PMB) regimens of 0.75 to 12 mg/kg given every 12h (Q12h) were evaluated for bacterial killing and resistance prevention against an AmpC-overexpressing Pseudomonas aeruginosa (PA) and a blaKPC-3-harboring Klebsiella pneumoniae (KP) in 10-day in-vitro hollow fiber models. An exposure-response was observed. But all regimens failed due to regrowth. Lower-dose regimens amplified isolates that expressed transient, lower-level adaptive resistance to PMB (MICs: ≤ 4 mg/L). Higher PMB dosages amplified isolates that expressed this resistance mechanism, a higher-MIC "moderately stable" adaptive resistance, and a higher-MIC stable resistance to PMB. Failure of the highest dose regimens was solely due to subpopulations that expressed the two higher-level resistances. Total and bioactive PMB concentrations in broth declined below targeted PK profiles within hours of treatment initiation and prior to bacterial regrowth. With treatment failure, the total PMB measured in bacteria were substantially higher than in broth. But the bioactive PMB in broth and bacteria were low to non-detectable. Together these findings suggest a sequence of events for treatment failure of the clinical regimen. First, PMB concentrations in broth are diluted as PMB binds to bacteria, resulting in total and bioactive PMB in broth that are lower than targeted. Bacterial regrowth and treatment failure follow, with emergence of subpopulations that express transient lower-level adaptive resistance to PMB and possibly higher-level adaptive and stable resistances. Higher-dose PMB regimens can prevent the emergence of transient lower-level adaptive resistance but they do not prevent treatment failure due to isolates that express higher-level resistance mechanisms.

The Mycobacterium tuberculosis drug discovery effort has generated a substantial number of new/repurposed drugs for therapy for this pathogen. The arrival of these drugs is welcome, but another layer of difficulty has emerged. Single agent therapy is insufficient for patients with late-stage tuberculosis because of resistance emergence. To achieve our therapeutic ends, it is requisite to identify optimal combination regimens. These regimens go through a lengthy and expensive evaluative process. If we have a modest group of 6 to 8 new or repurposed agents, this translates into 15 to 28 possible 2-drug combinations. There is neither time nor resources to give an extensive evaluation for all combinations. We sought a screening procedure that would identify combinations that had a high likelihood of achieving good bacterial burden decline. We examined pretomanid, moxifloxacin, linezolid, and bedaquiline in log-phase growth, acid-phase growth, and nonreplicative persister (NRP) phase in the Greco interaction model. We employed the interaction term α and the calculated bacterial burden decline as metrics to rank different regimens in different metabolic states. No relationship was found between α and bacterial kill. We chose bacterial kill as the prime metric. The combination of pretomanid plus moxifloxacin emerged as the clear frontrunner, as the largest bacterial declines were seen in log phase and acid phase with this regimen and it was second best in NRP phase. Bedaquiline also produced good kill. This screening process may identify optimal combinations that can be further evaluated in both the hollow-fiber infection model and in animal models of Mycobacterium tuberculosis infection.