Bactericidal Activity,Absence of Serum Effect,and Time-Kill Kinetics of Ceftazidime-Avibactam against β-Lactamase-Producing Enterobacteriaceae and Pseudomonas aeruginosa

Avibactam, a non-β-lactam β-lactamase inhibitor with activity against extended-spectrum β-lactamases (ESBLs), KPC, AmpC, and some OXA enzymes, extends the antibacterial activity of ceftazidime against most ceftazidime-resistant organisms producing these enzymes. In this study, the bactericidal activity of ceftazidime-avibactam against 18 Pseudomonas aeruginosa isolates and 15 Enterobacteriaceae isolates, including wild-type isolates and ESBL, KPC, and/or AmpC producers, was evaluated. Ceftazidime-avibactam MICs (0.016 to 32 μg/ml) were lower than those for ceftazidime alone (0.06 to ≥256 μg/ml) against all isolates except for 2 P. aeruginosa isolates (1 bla_VIM-positive isolate and 1 bla_OXA-23-positive isolate). The minimum bactericidal concentration/MIC ratios of ceftazidime-avibactam were ≤4 for all isolates, indicating bactericidal activity. Human serum and human serum albumin had a minimal effect on ceftazidime-avibactam MICs. Ceftazidime-avibactam time-kill kinetics were evaluated at low MIC multiples and showed time-dependent reductions in the number of CFU/ml from 0 to 6 h for all strains tested. A ≥3-log₁₀ decrease in the number of CFU/ml was observed at 6 h for all Enterobacteriaceae, and a 2-log₁₀ reduction in the number of CFU/ml was observed at 6 h for 3 of the 6 P. aeruginosa isolates. Regrowth was noted at 24 h for some of the isolates tested in time-kill assays. These data demonstrate the potent bactericidal activity of ceftazidime-avibactam and support the continued clinical development of ceftazidime-avibactam as a new treatment option for infections caused by Enterobacteriaceae and P. aeruginosa, including isolates resistant to ceftazidime by mechanisms dependent on avibactam-sensitive β-lactamases.     

Efficacies of Ceftazidime-Avibactam and Ceftazidime against Pseudomonas aeruginosa in a Murine Lung Infection Model

This study aimed to determine the efficacy of human-simulated plasma exposures of 2 g ceftazidime plus 0.5 g avibactam every 8 h administered as a 2-h infusion or a ceftazidime regimen that produced a specific epithelial lining fluid (ELF) percentage of the dosing interval in which serum free drug concentrations remain above the MIC (fT>MIC) against 28 Pseudomonas aeruginosa isolates within a neutropenic murine pneumonia model and to assess the impact of host infection on pulmonary pharmacokinetics. The fT>MIC was calculated as the mean and upper end of the 95% confidence limit. Against the 28 P. aeruginosa strains used, the ceftazidime-avibactam MICs were 4 to 64 μg/ml, and those of ceftazidime were 8 to >128 μg/ml. The change in log₁₀ CFU after 24 h of treatment was analyzed relative to that of 0-h controls. Pharmacokinetic studies in serum and ELF were conducted using ceftazidime-avibactam in infected and uninfected mice. Humanized ceftazidime-avibactam doses resulted in significant exposures in the lung, producing reductions of >1 log₁₀ CFU against P. aeruginosa with ceftazidime-avibactam MICs of ≤32 μg/ml (ELF upper 95% confidence limit for fT>MIC [ELF fT>MIC] of ≥19%), except for one isolate with a ceftazidime-avibactam MIC of 16 μg/ml. No efficacy was observed against the isolate with a ceftazidime-avibactam MIC of 64 μg/ml (ELF fT>MIC of 0%). Bacterial reductions were observed with ceftazidime against isolates with ceftazidime MICs of 32 μg/ml (ELF fT>MIC of ≥12%), variable efficacy at ceftazidime MICs of 64 μg/ml (ELF fT>MIC of ≥0%), and no activity at a ceftazidime MIC of 128 μg/ml, where the ELF fT>MIC was 0%. ELF fT>MICs were similar between infected and uninfected mice. Ceftazidime-avibactam was effective against P. aeruginosa, with MICs of up to 32 μg/ml with an ELF fT>MIC of ≥19%. The data suggest the potential utility of ceftazidime-avibactam for treatment of lung infections caused by P. aeruginosa.     

In Vitro Antibacterial Activity of the Ceftazidime-Avibactam Combination against Enterobacteriaceae,Including Strains with Well-Characterized β-Lactamases

The novel β-lactamase inhibitor avibactam is a potent inhibitor of class A, class C, and some class D enzymes. The in vitro antibacterial activity of the ceftazidime-avibactam combination was determined for a collection of Enterobacteriaceae clinical isolates; this collection was enriched for resistant strains, including strains with characterized serine β-lactamases. The inhibitor was added either at fixed weight ratios to ceftazidime or at fixed concentrations, with the latter type of combination consistently resulting in greater potentiation of antibacterial activity. In the presence of 4 μg/ml of avibactam, the ceftazidime MIC₅₀ and MIC₉₀ (0.25 and 2 μg/ml, respectively) were both below the CLSI breakpoint for ceftazidime. Further comparisons with reference antimicrobial agents were performed using this fixed inhibitor concentration. Against most ceftazidime-susceptible and -nonsusceptible isolates, the addition of avibactam resulted in a significant increase in ceftazidime activity, with MICs generally reduced 256-fold for extended-spectrum β-lactamase (ESBL) producers, 8- to 32-fold for CTX-M producers, and >128-fold for KPC producers. Overall, MICs of a ceftazidime-avibactam combination were significantly lower than those of the comparators piperacillin-tazobactam, cefotaxime, ceftriaxone, and cefepime and similar or superior to those of imipenem.     

Effects of Klebsiella pneumoniae Carbapenemase Subtypes,Extended-Spectrum β-Lactamases,and Porin Mutations on the In Vitro Activity of Ceftazidime-Avibactam against Carbapenem-Resistant K. pneumoniae

Avibactam is a novel β-lactamase inhibitor with affinity for Klebsiella pneumoniae carbapenemases (KPCs). In combination with ceftazidime, the agent demonstrates activity against KPC-producing K. pneumoniae (KPC-Kp). KPC-Kp strains are genetically diverse and harbor multiple resistance determinants, including defects in outer membrane proteins and extended-spectrum β-lactamases (ESBLs). Mutations in porin gene ompK36 confer high-level carbapenem resistance to KPC-Kp strains. Whether specific mechanisms of antimicrobial resistance also influence the activity of ceftazidime-avibactam is unknown. We defined the effects of ceftazidime-avibactam against 72 KPC-Kp strains with diverse mechanisms of resistance, including various combinations of KPC subtypes and ESBL and ompK36 mutations. Ceftazidime MICs ranged from 64 to 4,096 μg/ml and were lowered by a median of 512-fold with the addition of avibactam. All strains exhibited ceftazidime-avibactam MICs at or below the CLSI breakpoint for ceftazidime (≤4 μg/ml; range, 0.25 to 4). However, the MICs were within two 2-fold dilutions of the CLSI breakpoint against 24% of the strains, and those strains would be classified as nonsusceptible to ceftazidime by EUCAST criteria (MIC > 1 μg/ml). Median ceftazidime-avibactam MICs were higher against KPC-3 than KPC-2 variants (P = 0.02). Among KPC-2-Kp strains, the presence of both ESBL and porin mutations was associated with higher drug MICs compared to those seen with either factor alone (P = 0.003 and P = 0.02, respectively). In conclusion, ceftazidime-avibactam displays activity against genetically diverse KPC-Kp strains. Strains with higher-level drug MICs provide a reason for caution. Judicious use of ceftazidime-avibactam alone or in combination with other agents will be important to prevent the emergence of resistance.     

Ceftazidime-Avibactam Activity Tested against Enterobacteriaceae Isolates from U.S. Hospitals (2011 to 2013) and Characterization of β-Lactamase-Producing Strains

Ceftazidime-avibactam (MIC_50/90, 0.12/0.25 μg/ml) inhibited 99.9% (20,698/20,709) of Enterobacteriaceae isolates at ≤8 μg/ml. This compound was active against resistant subsets, including ceftazidime-nonsusceptible Enterobacter cloacae (MIC_50/90, 0.25/0.5 μg/ml) and extended-spectrum β-lactamase (ESBL) phenotype isolates. An ESBL phenotype was noted among 12.4% (1,696/13,692 isolates from targeted species) of the isolates, including 776 Escherichia coli (12.0% for this species; MIC_50/90, 0.12/0.25 μg/ml), 721 Klebsiella pneumoniae (16.3%; MIC_50/90, 0.12/0.25 μg/ml), 119 Klebsiella oxytoca (10.3%; MIC_50/90, 0.06/0.25 μg/ml), and 80 Proteus mirabilis (4.9%; MIC_50/90, 0.06/0.12 μg/ml) isolates. The most common enzymes detected among ESBL phenotype isolates from 2013 (n = 743) screened using a microarray-based assay were CTX-M-15-like (n = 307), KPC (n = 120), SHV ESBLs (n = 118), and CTX-M-14-like (n = 110). KPC producers were highly resistant to comparators, and ceftazidime-avibactam (MIC_50/90, 0.5/2 μg/ml) and tigecycline (MIC_50/90, 0.5/1 μg/ml; 98.3% susceptible) were the most active agents against these strains. Meropenem (MIC_50/90, ≤0.06/≤0.06 μg/ml) and ceftazidime-avibactam (MIC_50/90, 0.12/0.25 μg/ml) were active against CTX-M-producing isolates. Other enzymes were also observed, and ceftazidime-avibactam displayed good activity against the isolates producing less common enzymes. Among 11 isolates displaying ceftazidime-avibactam MIC values of >8 μg/ml, three were K. pneumoniae strains producing metallo-β-lactamases (all ceftazidime-avibactam MICs, >32 μg/ml), with two NDM-1 producers and one K. pneumoniae strain carrying the bla_KPC-2 and bla_VIM-4 genes. Therapeutic options for isolates producing β-lactamases may be limited, and ceftazidime-avibactam, which displayed good activity against strains, including those producing KPC enzymes, merits further study in infections where such organisms occur.     

Antimicrobial Activity of Ceftazidime-Avibactam against Gram-Negative Organisms Collected from U.S. Medical Centers in 2012

The activities of the novel β-lactam–β-lactamase inhibitor combination ceftazidime-avibactam and comparator agents were evaluated against a contemporary collection of clinically significant Gram-negative bacilli. Avibactam is a novel non-β-lactam β-lactamase inhibitor that inhibits Ambler class A, C, and some D enzymes. A total of 10,928 Gram-negative bacilli—8,640 Enterobacteriaceae, 1,967 Pseudomonas aeruginosa, and 321 Acinetobacter sp. isolates—were collected from 73 U.S. hospitals and tested for susceptibility by reference broth microdilution methods in a central monitoring laboratory (JMI Laboratories, North Liberty, IA, USA). Ceftazidime was combined with avibactam at a fixed concentration of 4 μg/ml. Overall, 99.8% of Enterobacteriaceae strains were inhibited at a ceftazidime-avibactam MIC of ≤4 μg/ml. Ceftazidime-avibactam was active against extended-spectrum β-lactamase (ESBL)-phenotype Escherichia coli and Klebsiella pneumoniae, meropenem-nonsusceptible (MIC ≥ 2 μg/ml) K. pneumoniae, and ceftazidime-nonsusceptible Enterobacter cloacae. Among ESBL-phenotype K. pneumoniae strains, 61.1% were meropenem susceptible and 99.3% were inhibited at a ceftazidime-avibactam MIC of ≤4 μg/ml. Among P. aeruginosa strains, 96.9% were inhibited at a ceftazidime-avibactam MIC of ≤8 μg/ml, and susceptibility rates for meropenem, ceftazidime, and piperacillin-tazobactam were 82.0, 83.2, and 78.3%, respectively. Ceftazidime-avibactam was the most active compound tested against meropenem-nonsusceptible P. aeruginosa (MIC₅₀/MIC₉₀, 4/16 μg/ml; 87.3% inhibited at ≤8 μg/ml). Acinetobacter spp. (ceftazidime-avibactam MIC₅₀/MIC₉₀, 16/>32 μg/ml) showed high rates of resistance to most tested agents. In summary, ceftazidime-avibactam demonstrated potent activity against a large collection of contemporary Gram-negative bacilli isolated from patients in U.S. hospitals in 2012, including organisms that are resistant to most currently available agents, such as K. pneumoniae carbapenemase (KPC)-producing Enterobacteriaceae and meropenem-nonsusceptible P. aeruginosa.     

Molecular Characterization of Carbapenem-Nonsusceptible Enterobacterial Isolates Collected during a Prospective Interregional Survey in France and Susceptibility to the Novel Ceftazidime-Avibactam and Aztreonam-Avibactam Combinations

An interregional surveillance program was conducted in the northwestern part of France to determine the prevalence of carbapenem-nonsusceptible Enterobacteriaceae (CNSE) isolates and their susceptibility to ceftazidime-avibactam and aztreonam-avibactam combinations. Nonduplicate CNSE clinical isolates were prospectively collected from six hospitals between June 2012 and November 2013. MICs of ceftazidime and aztreonam, alone or combined with a fixed concentration of avibactam (4 μg/ml), and those of carbapenems (comparator agents) were determined. MICs of ertapenem in combination with phenylalanine arginine-naphthylamide dihydrochloride (PAβN) were also determined to assess active efflux. Genes encoding carbapenemases, plasmid-mediated AmpC enzymes, extended-spectrum β-lactamases (ESBLs), and major outer membrane proteins (OMPs) were amplified and sequenced. OMPs were also extracted for SDS-PAGE analysis. Among the 139 CNSE isolates, mainly Enterobacter spp. and Klebsiella pneumoniae, 123 (88.4%) were ertapenem nonsusceptible, 12 (8.6%) exhibited reduced susceptibility to all carbapenems, and 4 Proteeae isolates (2.9%) were resistant to imipenem. Carbapenemase production was detected in only two isolates (producing OXA-48 and IMI-3). In contrast, OMP deficiency, in association with AmpCs and/or ESBLs (mainly CTX-M-9, SHV-12, and CTX-M-15), was largely identified among CNSE isolates. The ceftazidime-avibactam and aztreonam-avibactam combinations exhibited potent activity against CNSE isolates (MIC₅₀/MIC₉₀, 1/1 μg/ml and 0.5/0.5 μg/ml, respectively) compared to that of ceftazidime and aztreonam alone (MIC₅₀/MIC₉₀, 512/512 μg/ml and 128/512 μg/ml, respectively). This study reveals the in vitro activity of ceftazidime-avibactam and aztreonam-avibactam combinations against a large collection of porin-deficient enterobacterial isolates that are representative of the CNSE recovered in the northern part of France.     

Activity of Ceftazidime-Avibactam against Extended-Spectrum- and AmpC β-Lactamase-Producing Enterobacteriaceae Collected in the INFORM Global Surveillance Study from 2012 to 2014

The in vitro activity of ceftazidime-avibactam was evaluated against 34,062 isolates of Enterobacteriaceae from patients with intra-abdominal, urinary tract, skin and soft-tissue, lower respiratory tract, and blood infections collected in the INFORM (International Network For Optimal Resistance Monitoring) global surveillance study (176 medical center laboratories in 39 countries) in 2012 to 2014. Overall, 99.5% of Enterobacteriaceae isolates were susceptible to ceftazidime-avibactam using FDA approved breakpoints (susceptible MIC of ≤8 μg/ml; resistant MIC of ≥16 μg/ml). For individual species of the Enterobacteriaceae, the ceftazidime-avibactam MIC inhibiting ≥90% of isolates (MIC₉₀) ranged from 0.06 μg/ml for Proteus species to 1 μg/ml for Enterobacter spp. and Klebsiella pneumoniae. Carbapenem-susceptible isolates of Escherichia coli, K. pneumoniae, Klebsiella oxytoca, and Proteus mirabilis with a confirmed extended-spectrum β-lactamase (ESBL) phenotype, or a ceftazidime MIC of ≥16 μg/ml if the ESBL phenotype was not confirmed by clavulanic acid inhibition, were characterized further to identify the presence of specific ESBL- and plasmid-mediated AmpC β-lactamase genes using a microarray-based assay and additional PCR assays. Ceftazidime-avibactam demonstrated potent activity against molecularly confirmed ESBL-producing (n = 5,354; MIC₉₀, 0.5 μg/ml; 99.9% susceptible), plasmid-mediated AmpC-producing (n = 246; MIC₉₀, 0.5 μg/ml; 100% susceptible), and ESBL- and AmpC-producing (n = 152; MIC₉₀, 1 μg/ml; 100% susceptible) isolates of E. coli, K. pneumoniae, K. oxytoca, and P. mirabilis. We conclude that ceftazidime-avibactam demonstrates potent in vitro activity against globally collected clinical isolates of Enterobacteriaceae, including isolates producing ESBLs and AmpC β-lactamases.     

Reclaiming the Efficacy of β-Lactam–β-Lactamase Inhibitor Combinations: Avibactam Restores the Susceptibility of CMY-2-Producing Escherichia coli to Ceftazidime

CMY-2 is a plasmid-encoded Ambler class C cephalosporinase that is widely disseminated in Enterobacteriaceae and is responsible for expanded-spectrum cephalosporin resistance. As a result of resistance to both ceftazidime and β-lactamase inhibitors in strains carrying bla_CMY, novel β-lactam–β-lactamase inhibitor combinations are sought to combat this significant threat to β-lactam therapy. Avibactam is a bridged diazabicyclo [3.2.1]octanone non-β-lactam β-lactamase inhibitor in clinical development that reversibly inactivates serine β-lactamases. To define the spectrum of activity of ceftazidime-avibactam, we tested the susceptibilities of Escherichia coli clinical isolates that carry bla_CMY-2 or bla_CMY-69 and investigated the inactivation kinetics of CMY-2. Our analysis showed that CMY-2-containing clinical isolates of E. coli were highly susceptible to ceftazidime-avibactam (MIC₉₀, ≤0.5 mg/liter); in comparison, ceftazidime had a MIC₉₀ of >128 mg/liter. More importantly, avibactam was an extremely potent inhibitor of CMY-2 β-lactamase, as demonstrated by a second-order onset of acylation rate constant (k₂/K) of (4.9 ± 0.5) × 10⁴ M⁻¹ s⁻¹ and the off-rate constant (k_off) of (3.7 ± 0.4) ×10⁻⁴ s⁻¹. Analysis of the reaction of avibactam with CMY-2 using mass spectrometry to capture reaction intermediates revealed that the CMY-2–avibactam acyl-enzyme complex was stable for as long as 24 h. Molecular modeling studies raise the hypothesis that a series of successive hydrogen-bonding interactions occur as avibactam proceeds through the reaction coordinate with CMY-2 (e.g., T316, G317, S318, T319, S343, N346, and R349). Our findings support the microbiological and biochemical efficacy of ceftazidime-avibactam against E. coli containing plasmid-borne CMY-2 and CMY-69.     

In Vitro Susceptibility of Characterized β-Lactamase-Producing Strains Tested with Avibactam Combinations

Avibactam, a broad-spectrum β-lactamase inhibitor, was tested with ceftazidime, ceftaroline, or aztreonam against 57 well-characterized Gram-negative strains producing β-lactamases from all molecular classes. Most strains were nonsusceptible to the β-lactams alone. Against AmpC-, extended-spectrum β-lactamase (ESBL)-, and KPC-producing Enterobacteriaceae or Pseudomonas aeruginosa, avibactam lowered ceftazidime, ceftaroline, or aztreonam MICs up to 2,048-fold, to ≤4 μg/ml. Aztreonam-avibactam MICs against a VIM-1 metallo-β-lactamase-producing Enterobacter cloacae and a VIM-1/KPC-3-producing Escherichia coli isolate were 0.12 and 8 μg/ml, respectively.     

In Vitro Activity of Ceftaroline-Avibactam against Gram-Negative and Gram-Positive Pathogens Isolated from Patients in Canadian Hospitals from 2010 to 2012: Results from the CANWARD Surveillance Study

The in vitro activities of ceftaroline-avibactam, ceftaroline, and comparative agents were determined for a collection of bacterial pathogens frequently isolated from patients seeking care at 15 Canadian hospitals from January 2010 to December 2012. In total, 9,758 isolates were tested by using the Clinical and Laboratory Standards Institute (CLSI) broth microdilution method (document M07-A9, 2012), with MICs interpreted by using CLSI breakpoints (document M100-S23, 2013). Ceftaroline-avibactam demonstrated potent activity (MIC₉₀, ≤0.5 μg/ml) against Escherichia coli, Klebsiella pneumoniae, Klebsiella oxytoca, Proteus mirabilis, Enterobacter cloacae, Enterobacter aerogenes, Serratia marcescens, Morganella morganii, Citrobacter freundii, and Haemophilus influenzae; >99% of isolates of E. coli, K. pneumoniae, K. oxytoca, P. mirabilis, M. morganii, C. freundii, and H. influenzae were susceptible to ceftaroline-avibactam according to CLSI MIC interpretative criteria for ceftaroline. Ceftaroline was less active than ceftaroline-avibactam against all species of Enterobacteriaceae tested, with rates of susceptibility ranging from 93.9% (P. mirabilis) to 54.0% (S. marcescens). All isolates of methicillin-susceptible Staphylococcus aureus (MIC₉₀, 0.25 μg/ml) and 99.6% of methicillin-resistant S. aureus isolates (MIC₉₀, 1 μg/ml) were susceptible to ceftaroline; the addition of avibactam to ceftaroline did not alter its activity against staphylococci or streptococci. All isolates of Streptococcus pneumoniae (MIC₉₀, 0.03 μg/ml), Streptococcus pyogenes (MIC₉₀, ≤0.03 μg/ml), and Streptococcus agalactiae (MIC₉₀, 0.015 μg/ml) tested were susceptible to ceftaroline. We conclude that combining avibactam with ceftaroline expanded its spectrum of activity to include most isolates of Enterobacteriaceae resistant to third-generation cephalosporins, including extended-spectrum β-lactamase (ESBL)- and AmpC-producing E. coli and ESBL-producing K. pneumoniae, while maintaining potent activity against staphylococci and streptococci.     

Evaluation of the In Vitro Activity of Ceftazidime-Avibactam and Ceftolozane-Tazobactam against Meropenem-Resistant Pseudomonas aeruginosa Isolates

We compared ceftazidime-avibactam, ceftolozane-tazobactam, ceftazidime, cefepime, and piperacillin-tazobactam MICs for 38 meropenem-resistant Pseudomonas aeruginosa isolates. No isolates harbored carbapenemases; 74% were oprD mutants. Ceftazidime-avibactam and ceftolozane-tazobactam were active against 92% of the isolates, including 80% that were resistant to all three β-lactams. Forty-three percent of ceftazidime-avibactam-susceptible isolates and 6% of ceftolozane-tazobactam-susceptible isolates exhibited MICs at the respective breakpoints. Ceftolozane-tazobactam and ceftazidime-avibactam are therapeutic options for meropenem-resistant P. aeruginosa infections that should be used judiciously to preserve activity.     

Activity of Ceftazidime-Avibactam against Fluoroquinolone-Resistant Enterobacteriaceae and Pseudomonas aeruginosa

Ceftazidime-avibactam and comparator antibiotics were tested by the broth microdilution method against 200 Enterobacteriaceae and 25 Pseudomonas aeruginosa strains resistant to fluoroquinolones (including strains with the extended-spectrum β-lactamase [ESBL] phenotype and ceftazidime-resistant strains) collected from our institution. The MICs and mechanisms of resistance to fluoroquinolone were also studied. Ninety-nine percent of fluoroquinolone-resistant Enterobacteriaceae strains were inhibited at a ceftazidime-avibactam MIC of ≤4 mg/liter (using the susceptible CLSI breakpoint for ceftazidime alone as a reference). Ceftazidime-avibactam was very active against ESBL Escherichia coli (MIC₉₀ of 0.25 mg/liter), ESBL Klebsiella pneumoniae (MIC₉₀ of 0.5 mg/liter), ceftazidime-resistant AmpC-producing species (MIC₉₀ of 1 mg/liter), non-ESBL E. coli (MIC₉₀ of ≤0.125 mg/liter), non-ESBL K. pneumoniae (MIC₉₀ of 0.25 mg/liter), and ceftazidime-nonresistant AmpC-producing species (MIC₉₀ of ≤0.5 mg/liter). Ninety-six percent of fluoroquinolone-resistant P. aeruginosa strains were inhibited at a ceftazidime-avibactam MIC of ≤8 mg/liter (using the susceptible CLSI breakpoint for ceftazidime alone as a reference), with a MIC₉₀ of 8 mg/liter. Additionally, fluoroquinolone-resistant mutants from each species tested were obtained in vitro from two strains, one susceptible to ceftazidime and the other a β-lactamase producer with a high MIC against ceftazidime but susceptible to ceftazidime-avibactam. Thereby, the impact of fluoroquinolone resistance on the activity of ceftazidime-avibactam could be assessed. The MIC₉₀ values of ceftazidime-avibactam for the fluoroquinolone-resistant mutant strains of Enterobacteriaceae and P. aeruginosa were ≤4 mg/liter and ≤8 mg/liter, respectively. We conclude that the presence of fluoroquinolone resistance does not affect Enterobacteriaceae and P. aeruginosa susceptibility to ceftazidime-avibactam; that is, there is no cross-resistance.     

Comparative In Vitro and In Vivo Efficacies of Human Simulated Doses of Ceftazidime and Ceftazidime-Avibactam against Pseudomonas aeruginosa

The combination of ceftazidime and avibactam possesses potent activity against resistant Gram-negative pathogens, including Pseudomonas aeruginosa. We compared the efficacies of human simulated doses of ceftazidime and ceftazidime-avibactam using a hollow-fiber system and neutropenic and immunocompetent murine thigh infection models. Twenty-seven clinical P. aeruginosa isolates with ceftazidime MICs of 8 to 128 mg/liter and ceftazidime-avibactam MICs of 4 to 32 mg/liter were utilized in neutropenic mouse studies; 15 of the isolates were also evaluated in immunocompetent mice. Six isolates were studied in both the hollow-fiber system and the neutropenic mouse. In both systems, the free drug concentration-time profile seen in humans given 2 g of ceftazidime every 8 h (2-h infusion), with or without avibactam at 500 mg every 8 h (2-h infusion), was evaluated. In vivo activity was pharmacodynamically predictable based on the MIC. Ceftazidime decreased bacterial densities by ≥0.5 log unit for 10/27 isolates, while ceftazidime-avibactam did so for 22/27 isolates. In immunocompetent animals, enhancements in activity were seen for both drugs, with ceftazidime achieving reductions of ≥0.3 log unit for 10/15 isolates, whereas ceftazidime-avibactam did so against all 15 isolates. In vitro, ceftazidime resulted in regrowth by 24 h against all isolates, while ceftazidime-avibactam achieved stasis or better against 4/7 isolates. Mutants with elevated ceftazidime-avibactam MICs appeared after 24 h from 3/7 isolates studied in vitro; however, no resistant mutants were detected in vivo. Against this highly ceftazidime-nonsusceptible population of P. aeruginosa, treatment with human simulated doses of ceftazidime-avibactam resulted in pharmacodynamically predictable activity, particularly in vivo, against isolates with MICs of ≤16 mg/liter, and this represents a potential new option to combat these difficult-to-treat pathogens.     

Pharmacodynamic Target Attainment for Various Ceftazidime Dosing Schemes in High-Flux Hemodialysis

Ceftazidime is a broad-spectrum cephalosporin with high-level activity against a variety of Gram-negative pathogens, including Pseudomonas aeruginosa. Improved outcomes are associated with cumulative percentages of a 24-h period that the drug concentration exceeds the MIC under steady-state pharmacokinetic conditions (%T_MIC) of >45 to 70% of the dosing interval. Optimal dosing to achieve a 90% probability of target attainment (PTA) in patients receiving high-flux hemodialysis (HFHD) is unknown. We used existing data from six anephric adults receiving hemodialysis to construct a population model with the Pmetrics package for R. From the final model''s joint probability density, we simulated the PTA for various ceftazidime dosing regimens, HFHD schedules, and organism MICs. For HFHD every 48 h and 1 g of ceftazidime given posthemodialysis, the PTA exceeds 90% for all isolates with MICs of ≤8 μg/ml, assuming a goal of 70%T_MIC. For 72-h dialysis intervals, postdialysis dosing of 1 g is adequate for achievement of the 70%T_MIC goal only for organisms with MICs of ≤4 μg/ml, while 2 g is adequate for organisms with MICs of ≤8 μg/ml. A dose of 500 mg once daily, regardless of HFHD schedule, has a 90% PTA for organisms with MICs of ≤16 μg/ml, while 1 g once daily may achieve 100% PTA even for resistant organisms with a MIC of 32 μg/ml. Therefore, to ensure maximal ceftazidime activity, once-daily dosing of 500 mg to 1 g ceftazidime in patients receiving HFHD may be preferable for critically ill patients when MIC data are unavailable and for more resistant organisms with ceftazidime MICs of 16 to 32 μg/ml.     

First Report of Ceftazidime-Avibactam Resistance in a KPC-3-Expressing Klebsiella pneumoniae Isolate

Ceftazidime-avibactam is the first antimicrobial approved by the U.S. FDA for the treatment of carbapenem-resistant Enterobacteriaceae. Avibactam, a non-β-lactam β-lactamase inhibitor, inactivates class A serine carbapenemases, including Klebsiella pneumoniae carbapenemase (KPC). We report a KPC-producing K. pneumoniae isolate resistant to ceftazidime-avibactam (MIC, 32/4 μg/ml) from a patient with no prior treatment with ceftazidime-avibactam.     

β-Lactamase Characterization of Gram-Negative Pathogens Recovered from Patients Enrolled in the Phase 2 Trials for Ceftazidime-Avibactam: Clinical Efficacies Analyzed against Subsets of Molecularly Characterized Isolates

The correlation of the clinical efficacies of ceftazidime-avibactam and comparators (carbapenems) was evaluated against baseline Gram-negative isolates having characterized β-lactam resistance mechanisms from complicated urinary tract infection (cUTI) and complicated intra-abdominal infection (cIAI) phase 2 trials. Enterobacteriaceae displaying ceftriaxone and/or ceftazidime MICs of ≥2 μg/ml (69 isolates) and nonfermentative Gram-negative bacilli (NF-GNB [three isolates]) with ceftazidime MICs of ≥16 μg/ml were characterized for their narrow- and extended-spectrum β-lactamase (ESBL) content. Enterobacteriaceae (one isolate) and NF-GNB (three isolates) with imipenem/meropenem MICs of ≥2 and ≥16 μg/ml, respectively, were tested for carbapenemases. All cUTI E. coli had the lineage background investigated (ST131-like versus non-ST131-like). The primary efficacy endpoint was microbiological response (eradication) at test of cure (TOC) for cUTI and clinical response (inferred microbiological eradication) at TOC for cIAI. A total of 34.1% of baseline cUTI (36.4%) and cIAI (33.1%) pathogens met the MIC-based screening criteria (screen positive). All screen-positive cUTI pathogens were CTX-M-producing E. coli, except for one E. cloacae isolate with AmpC overexpression. The majority (66.7%) of screen-positive cIAI isolates produced CTX-M-type coupled with a diverse array of other β-lactamases. Similar favorable responses were observed with ceftazidime-avibactam (93.3%) and carbapenems (90.9%), when a non-ESBL Enterobacteriaceae isolate was recovered at the baseline visit. When an ESBL Enterobacteriaceae isolate was present, the favorable responses were 85.7% and 80.0% with ceftazidime-avibactam and carbapenems, respectively. Higher favorable responses were observed with ceftazidime-avibactam (75.0%) than with carbapenems (66.7%) when an ST131-like E. coli isolate was recovered at baseline, as when a non-ST131-like isolate was present (93.8% versus 86.7%, respectively). The efficacy of ceftazidime-avibactam was similar to that of carbapenems for treatment of cUTI and cIAI caused by ESBL organisms.     

Activities of Ceftazidime and Avibactam against β-Lactamase-Producing Enterobacteriaceae in a Hollow-Fiber Pharmacodynamic Model

Avibactam is a novel non-β-lactam β-lactamase inhibitor that is currently undergoing phase 3 clinical trials in combination with ceftazidime. Ceftazidime is hydrolyzed by a broad range of β-lactamases, but avibactam is able to inhibit the majority of these enzymes. The studies described here attempt to provide insight into the amount of avibactam required to suppress bacterial growth in an environment where the concentrations of both agents are varying as they would when administered to humans. Following the simulation of a single intravenous dose of the drug, ceftazidime alone had no effect on any test organism, but a ceftazidime-avibactam combination resulted in rapid killing of all of the strains, with growth suppressed for the 8 h of the study. For seven of eight strains, this was achieved with a 1-g–250-mg profile, but a 2-g–500-mg profile was necessary to completely suppress a high-level-AmpC-producing isolate. When ceftazidime was infused continuously for 24 h with a single bolus dose of avibactam, rapid killing of all of the strains was again observed, with growth suppressed for 10 to >24 h. Regrowth appeared to commence once the avibactam concentration dropped below a critical concentration of approximately 0.3 μg/ml. In a third series of studies, ceftazidime was administered every 8 h for 24 h with avibactam administered at fixed concentrations for short periods during each ceftazidime dose profile. Simulating a 1-g dose of ceftazidime, an avibactam pulse of >0.25 and <0.5 μg/ml was required to suppress growth for 24 h.     

In Vitro Activity of Biapenem plus RPX7009, a Carbapenem Combined with a Serine β-Lactamase Inhibitor,against Anaerobic Bacteria

Biapenem is a carbapenem being developed in combination with RPX7009, a new inhibitor of serine β-lactamases. Biapenem was tested alone and in combination with fixed concentrations of RPX7009 by agar dilution against 377 recent isolates of anaerobes. A separate panel of 27 isolates of Bacteroides spp. with decreased susceptibility or resistance to imipenem was also tested. Comparator drugs included meropenem, piperacillin-tazobactam, ampicillin-sulbactam, cefoxitin, ceftazidime, metronidazole, clindamycin, and tigecycline plus imipenem, doripenem, and ertapenem for the 27 selected strains. For recent consecutive strains of Bacteroides species, the MIC₉₀ for biapenem-RPX7009 was 1 μg/ml, with a MIC₉₀ of 4 μg/ml for meropenem. Other Bacteroides fragilis group species showed a MIC₉₀ of 0.5 μg/ml for both agents. The MIC₉₀s for biapenem-RPX7009 were 0.25 μg/ml for Prevotella spp., 0.125 μg/ml for Fusobacterium nucleatum and Fusobacterium necrophorum, 2 μg/ml for Fusobacterium mortiferum, 0.5 μg/ml for Fusobacterium varium, ≤0.5 μg/ml for Gram-positive cocci and rods, and 0.03 to 8 μg/ml for clostridia. Against 5 B. fragilis strains harboring a known metallo-beta-lactamase, biapenem-RPX7009 MICs were comparable to those of other carbapenems (≥32 μg/ml). Against Bacteroides strains with an imipenem MIC of 2 μg/ml, biapenem-RPX7009 had MICs of 0.5 to 2 μg/ml, with MICs of 0.5 to 32 μg/ml for meropenem, doripenem, and ertapenem. For strains with an imipenem MIC of 4 μg/ml, the MICs for biapenem-RPX7009 were 4 to 16 μg/ml, with MICs of 8 to >32 μg/ml for meropenem, doripenem, and ertapenem. The inhibitor RPX7009 had no antimicrobial activity when tested alone, and it showed little or no potentiation of biapenem versus anaerobes. Biapenem-RPX7009 showed activity comparable to that of imipenem and was superior to meropenem, doripenem, and ertapenem against imipenem-nonsusceptible Bacteroides spp.     

Role of the Outer Membrane and Porins in Susceptibility of β-Lactamase-Producing Enterobacteriaceae to Ceftazidime-Avibactam

This study examined the activity of the novel antimicrobial combination ceftazidime-avibactam against Enterobacteriaceae exhibiting different outer membrane permeability profiles, specifically with or without porins and with or without expression of the main efflux pump (AcrAB-TolC). The addition of the outer membrane permeabilizer polymyxin B nonapeptide increased the antibacterial activities of avibactam alone, ceftazidime alone, and ceftazidime-avibactam against the characterized clinical isolates of Escherichia coli, Enterobacter aerogenes, and Klebsiella pneumoniae. This enhancement of activities was mainly due to increased passive penetration of compounds since inhibition of efflux by the addition of phenylalanine-arginine β-naphthylamide affected the MICs minimally. OmpF (OmpK35) or OmpC (OmpK36) pores were not the major route by which avibactam crossed the outer membranes of E. coli and K. pneumoniae. In contrast, Omp35 and Omp36 allowed diffusion of avibactam across the outer membrane of E. aerogenes, although other diffusion channels for avibactam were also present in that species. It was clear that outer membrane permeability and outer membrane pore-forming proteins play a key role in the activity of ceftazidime-avibactam. Nevertheless, the MICs of ceftazidime-avibactam (with 4 mg/liter avibactam) against the ceftazidime-resistant clinical isolates of the three species of Enterobacteriaceae studied were ≤8 mg/liter, regardless of outer membrane permeability changes resulting from an absence of defined porin proteins or upregulation of efflux.