c Carbapenem-resistant bacteria represent a significant treatment challenge due to the lack of active antimicrobials available. MK-7655 is a novel ␤-lactamase inhibitor under clinical development. We investigated the combined killing activity of imipenem and MK-7655 against four imipenem-resistant bacterial strains, using a mathematical model previously evaluated in our laboratory. Time-kill studies (TKS) were conducted with imipenem and MK-7655 against a KPC-2-producing Klebsiella pneumoniae isolate (KP6339) as well as 3 Pseudomonas aeruginosa isolates (PA24226, PA24227, and PA24228) with OprD porin deletions and overexpression of AmpC. TKS were performed using 25 clinically achievable concentration combinations in a 5-by-5 array. Bacterial burden at 24 h was determined in triplicate by quantitative culture and mathematically modeled using a three-dimensional response surface. Mathematical model assessments were evaluated experimentally using clinically relevant dosing regimens of imipenem, with or without MK-7655, in a hollow-fiber infection model (HFIM). The combination of imipenem and MK-7655 was synergistic for all strains. Interaction indices were as follows: for KP6339, 0.50 (95% confidence interval [CI], 0.42 to 0.58); for PA24226, 0.60 (95% CI, 0.58 to 0.62); for PA24227, 0.70 (95% CI, 0.66 to 0.74); and for PA24228, 0.55 (95% CI, 0.49 to 0.61). In the HFIM, imipenem plus MK-7655 considerably reduced the bacterial burden at 24 h, while failure with imipenem alone was seen against all isolates. Sustained suppression of bacterial growth at 72 h was achieved with simulated doses of 500 mg imipenem plus 500 mg MK-7655 in 2 (KP6339 and PA24227) strains, and it was achieved in an additional strain (PA24228) when the imipenem dose was increased to 1,000 mg. Additional studies are being conducted to determine the optimal dose and combinations to be used in clinical investigations.

The development of β-lactam resistance during therapy could be suppressed by an optimized dosing exposure. Validation of the proposed target in a well-designed clinical study is warranted.

Antimicrobial resistance among Acinetobacter baumannii is increasing worldwide, often necessitating combination therapy. The clinical utility of using minocycline with polymyxin B is not well established. In this study, we investigated the activity of minocycline and polymyxin B against 1 laboratory isolate and 3 clinical isolates of A. baumannii. Minocycline susceptibility testing was performed with and without an efflux pump inhibitor, phenylalanine-arginine ␤-naphthylamide (PA␤N). The intracellular minocycline concentration was determined with and without polymyxin B (0.5 g/ml). Time-kill studies were performed over 24 h using approximately 10 6 CFU/ml of each strain with clinically relevant minocycline concentrations (2 g/ml and 8 g/ml), with and without polymyxin B (0.5 g/ml). The in vivo efficacy of the combination was assessed in a neutropenic murine pneumonia model. Infected animals were administered minocycline (50 mg/kg), polymyxin B (10 mg/kg), or both to achieve clinically equivalent exposures in humans. A reduction in the minocycline MIC (>4؋) was observed in the presence of PA␤N. The intracellular concentration and in vitro bactericidal effect of minocycline were both enhanced by polymyxin B. With 2 minocyclinesusceptible strains, the bacterial burden in lung tissue at 24 h was considerably reduced by the combination compared to monotherapy with minocycline or polymyxin B. In addition, the combination prolonged survival of animals infected with a minocycline-susceptible strain. Polymyxin B increased the intracellular concentration of minocycline in bacterial cells and enhanced the bactericidal activity of minocycline, presumably due to efflux pump disruption. The clinical utility of this combination should be further investigated.A cinetobacter baumannii is a common nosocomial pathogen globally which has been implicated as an etiologic agent in ventilator-associated pneumonia, skin and skin structure infections (traumatic battlefield and other wounds), urinary tract infections, meningitis, and bacteremia (1). Over the past decades, infections due to increasingly resistant strains of Acinetobacter have emerged (2, 3). This is particularly alarming because there are very limited therapeutic options for these infections (4, 5). Multidrug-resistant strains of A. baumannii have been reported to adversely affect patient outcomes. Studies have found significantly higher rates of hospital mortality in patients infected with multidrug-resistant strains than in patients with susceptible strains (6, 7).Current treatment strategies for multidrug-resistant A. baumannii include combination therapy with tigecycline, minocycline, carbapenems, polymyxins, and even daptomycin (8-11). However, the rationale of using certain agents together in a combination is not well established. In this study, the combination of minocycline and polymyxin B was studied based on a mechanistically plausible approach. Resistance to the tetracycline class in A. baumannii is commonly mediated through the upregulation of efflux pumps loca...

The emergence of resistance presents a debilitating change in the management of infectious diseases. Currently, the temporal relationship and interplay between various mechanisms of drug resistance are not well understood. A thorough understanding of the resistance development process is needed to facilitate rational design of countermeasure strategies. Using an in vitro hollow-fiber infection model that simulates human drug treatment, we examined the appearance of efflux pump (acrAB) overexpression and target topoisomerase gene (gyrA and parC) mutations over time in the emergence of quinolone resistance in Escherichia coli. Drug-resistant isolates recovered early (24 h) had 2-to 8-fold elevation in the MIC due to acrAB overexpression, but no point mutations were noted. In contrast, high-level (>64؋ MIC) resistant isolates with target site mutations (gyrA S83L with or without parC E84K) were selected more readily after 120 h, and regression of acrAB overexpression was observed at 240 h. Using a similar dosing selection pressure, the emergence of levofloxacin resistance was delayed in a strain with acrAB deleted compared to the isogenic parent. The role of efflux pumps in bacterial resistance development may have been underappreciated. Our data revealed the interplay between two mechanisms of quinolone resistance and provided a new mechanistic framework in the development of high-level resistance. Early low-level levofloxacin resistance conferred by acrAB overexpression preceded and facilitated high-level resistance development mediated by target site mutation(s). If this interpretation is correct, then these findings represent a paradigm shift in the way quinolone resistance is thought to develop.

The increasing prevalence of multidrug-resistant Gram-negative infections has led to the resurgence of systemic polymyxin B, but little is known about its pharmacokinetics. The objective of this study was to characterize the pharmacokinetics and renal disposition of polymyxin B. Eight female Sprague-Dawley rats (weight, 225 to 250 g) were administered a single intravenous polymyxin B dose (4 mg/kg of body weight). Serial serum samples were collected and assayed for major polymyxin B components using a validated ultraperformance liquid chromatography-tandem mass spectrometry method. The best-fit pharmacokinetic parameters of each component were derived and compared using one-way analysis of variance. Cumulative urine was also collected daily for 48 h and assayed for polymyxin B. Kidney drug concentrations were measured at 6 h (n ‫؍‬ 3) and 48 h (n ‫؍‬ 3) after the same dose. Additionally, three rats were administered 2 doses of intravenous polymyxin B (4 mg/kg) 7 days apart. Serial serum samples were collected pre-and post-renal insufficiency (induced by uranyl nitrate) and assayed for polymyxin B. The pharmacokinetic parameters of the major components did not appear to be significantly different (P > 0.05). Less than 1% of the dose was recovered unchanged in urine collected over 48 h following administration. Therapeutic drug concentrations persisted in kidney tissue at 48 h. The post-renal insufficiency to pre-renal insufficiency ratio of the area under the serum concentration-time curve from time zero to infinity was 1.33 ؎ 0.04. Polymyxin B components appear to have similar pharmacokinetics. Polymyxin B preferentially persists in kidneys, which suggests a selective uptake process in renal cells. A mechanism(s) other than renal excretion could be involved in polymyxin B elimination, and dosing adjustment in renal insufficiency may not be necessary.