Meropenem Combined with Ciprofloxacin Combats Hypermutable Pseudomonas aeruginosa from Respiratory Infections of Cystic Fibrosis Patients

Abstract

Hypermutable Pseudomonas aeruginosa organisms are prevalent in chronic respiratory infections and have been associated with reduced lung function in cystic fibrosis (CF); these isolates can become resistant to all antibiotics in monotherapy. This study aimed to evaluate the time course of bacterial killing and resistance of meropenem and ciprofloxacin in combination against hypermutable and nonhypermutable P. aeruginosa Static concentration time-kill experiments over 72 h assessed meropenem and ciprofloxacin in mono- and combination therapies against PAO1 (nonhypermutable), PAOΔmutS (hypermutable), and hypermutable isolates CW8, CW35, and CW44 obtained from CF patients with chronic respiratory infections. Meropenem (1 or 2 g every 8 h [q8h] as 3-h infusions and 3 g/day as a continuous infusion) and ciprofloxacin (400 mg q8h as 1-h infusions) in monotherapies and combinations were further evaluated in an 8-day hollow-fiber infection model study (HFIM) against CW44. Concentration-time profiles in lung epithelial lining fluid reflecting the pharmacokinetics in CF patients were simulated and counts of total and resistant bacteria determined. All data were analyzed by mechanism-based modeling (MBM). In the HFIM, all monotherapies resulted in rapid regrowth with resistance at 48 h. The maximum daily doses of 6 g meropenem (T_>MIC of 80% to 88%) and 1.2 g ciprofloxacin (area under the concentration-time curve over 24 h in the steady state divided by the MIC [AUC/MIC], 176), both given intermittently, in monotherapy failed to suppress regrowth and resulted in substantial emergence of resistance (≥7.6 log₁₀ CFU/ml resistant populations). The combination of these regimens achieved synergistic killing and suppressed resistance. MBM with subpopulation and mechanistic synergy yielded unbiased and precise curve fits. Thus, the combination of 6 g/day meropenem plus ciprofloxacin holds promise for future clinical evaluation against infections by susceptible hypermutable P. aeruginosa.

Keywords: antibiotic resistance; carbapenem; combination therapy; epithelial lining fluid; fluoroquinolone; hypermutation; mechanism-based modeling; pharmacodynamics; pharmacokinetics.

FIG 1

Viable count profiles over 72 h for ciprofloxacin monotherapy (left column) and meropenem with ciprofloxacin combinations (middle and right columns) against nonhypermutable P. aeruginosa PAO1 (A) and hypermutable P. aeruginosa PAOΔmutS (B), CW8 (C), CW35 (D), and CW44 (E). Data are from static concentration time-kill studies for the total bacterial population. The resistant bacterial populations were quantified at 72 h on agar plates containing 10 mg/liter meropenem (M) or 1.25 mg/liter ciprofloxacin (C). For isolate CW44, the viable count profiles are shown as observed viable counts (symbols) and individual fitted profiles (lines in corresponding colors) from mechanism-based modeling. Symbols and lines below the limit of counting were plotted at the limit of counting (equivalent to one colony per agar plate); 1.0 and 0.7 log₁₀ CFU/ml for the total and resistant populations, respectively.

FIG 2

Antibacterial effect of meropenem and ciprofloxacin regimens in mono- and combination therapies against hypermutable P. aeruginosa CW44 in the dynamic hollow-fiber infection model over 8 days. The meropenem regimens were 1 g three times daily as 3-h infusions, 3 g daily as a continuous infusion, 2 g three times daily as 3-h infusions, all representative of 30% ELF penetration (left and middle columns), and 2 g three times daily as 3-h infusions representative of 60% ELF penetration (right column). The ciprofloxacin regimen was 400 mg three times daily as 1-h infusions. The top row shows the total bacterial population (CFU_all; observed viable counts, symbols, and population predicted profiles of the mechanism-based modeling [MBM] lines in corresponding colors). The bottom row shows the resistant bacterial populations (CFU_R) of meropenem and ciprofloxacin in mono- and combination therapies against CW44 quantified on 10 mg/liter meropenem (R_MEM, open symbols) or 1.25 mg/liter ciprofloxacin (R_CIP, solid symbols) containing agar plates. Symbols and lines below the limit of counting were plotted at the limit of counting (equivalent to one colony per agar plate); 1.0 and 0.7 log₁₀ CFU/ml for the total and resistant populations, respectively.

FIG 3

The mechanism-based model for bacterial growth and killing by meropenem and ciprofloxacin in mono- and combination therapies. The first population is double susceptible, i.e., it was susceptible to both meropenem and ciprofloxacin (MEM^s/CIP^s). The other two populations not shown were meropenem resistant and ciprofloxacin intermediate (MEM^r/CIPⁱ) or meropenem intermediate and ciprofloxacin resistant (MEMⁱ/CIP^r). A life cycle growth model describes the underlying biology of bacterial replication in two states for each population. The maximum killing rate constants (K_max) and related antibiotic concentrations causing 50% of K_max (KC₅₀), along with all parameter estimates, are described in Table S1 in the supplemental material (SCTK data) and Table 2 (HFIM data). Mechanistic synergy (i.e., ciprofloxacin enhancing the effect of meropenem) was included for all populations.