Abstract
Objectives
We evaluated the in vivo efficacy of human-simulated regimens (HSRs) of cefiderocol, ceftazidime/avibactam, meropenem and ceftazidime/avibactam/meropenem combination against Guiana-extended spectrum (GES)-producing Pseudomonas aeruginosa isolates.
Methods
Eighteen P. aeruginosa isolates producing GES-1 (n = 5), GES-5 (n = 5) or miscellaneous GESs (combinations of GES-19, GES-20 and/or GES-26; n = 8) were evaluated. In vitro MIC testing was determined using broth microdilution. In a validated murine thigh infection model, HSRs of cefiderocol 2 g q8h as a 3 h IV infusion, ceftazidime/avibactam 2.5 g q8h as a 2 h IV infusion, meropenem 2 g q8h as a 3 h IV infusion or ceftazidime/avibactam/meropenem were administered. Change in bacterial burden relative to baseline and the proportion of isolates in each genotypic group meeting 1-log10 kill endpoint were assessed.
Results
Modal MICs (mg/L) ranged from 0.125 to 1 for cefiderocol, 4 to >64 for ceftazidime/avibactam and 2 to >64 for meropenem. Cefiderocol produced >1-log10 of kill against all 18 tested isolates. Meropenem was active against all GES-1 isolates whereas activity against GES-5 and miscellaneous GESs was lacking, consistent with the MICs. Ceftazidime/avibactam was active against all GES-1 and GES-5 isolates and retained activity against 62.5% of miscellaneous GESs including isolates with elevated MICs. For isolates where ceftazidime/avibactam failed, the addition of meropenem restored the in vivo efficacy.
Conclusions
As monotherapy, cefiderocol was active in vivo against all tested isolates. The activities of meropenem or ceftazidime/avibactam alone were variable; however, a combination of both was active against all isolates. Cefiderocol and ceftazidime/avibactam/meropenem could be valuable therapeutic options for GES-producing P. aeruginosa infections. Clinical confirmatory data are warranted.
Introduction
Infection with Pseudomonas aeruginosa can be difficult to treat due to its propensity for multiple resistance mechanisms.1 P. aeruginosa producing class A Guiana-extended-spectrum (GES) β-lactamases have been increasingly described in the USA and globally and are associated with MDR.2–4 The first GES subtypes were considered ESBLs (GES-1); however, expanded hydrolytic spectrum to carbapenems via single or double amino acid substitutions (e.g. GES-2, GES-5, GES-20) are possible.4 Additionally, the presence of multiple GES genotypes in the same isolates can result in expanded resistance, such as the tandem presence of the ESBLs GES-19 and GES-26, which was associated with carbapenem resistance.3 In addition to carbapenem resistance, GES-producing P. aeruginosa are associated with ceftolozane/tazobactam resistance, leaving few viable treatments.3 In the limited available data, GES-producing P. aeruginosa infections are also associated with increased mortality relative to infections with GES-negative strains.5
Challenges in diagnostics delay the identification of GESs. Often identification is based on the phenotypic profile of ceftazidime/avibactam-susceptible and ceftolozane/tazobactam-resistant.6,7 Due to limited commercially available detection capabilities, clinical outcomes data associated with available antibiotics are sparse. With sparse clinical data, murine models can aid in identification of best available therapy.
We characterized the in vivo efficacy of human-simulated regimens (HSRs) of cefiderocol, ceftazidime/avibactam, meropenem, and ceftazidime/avibactam/meropenem in a thigh infection model against isolates producing GES-1, GES-5 or miscellaneous (GES-19, GES-20, GES-26) β-lactamases.
Materials and methods
Ethics
This study was approved by the Institutional Animal Care and Use Committee of Hartford Hospital. All animal experiments were conducted in concordance with the standards set by the National Research Council of the National Academy of Sciences.
Antimicrobial agents
Commercially available cefiderocol, meropenem and ceftazidime were used for in vivo experiments. Analytical-grade avibactam was used for in vitro and in vivo experiments.
Isolates and MIC testing
Eighteen P. aeruginosa isolates were assessed and MICs were conducted in triplicate using broth microdilution.8,9
In vivo efficacy in the neutropenic murine thigh infection model
Specific-pathogen-free, female, CD-1 mice (20–22 g) were utilized in the model. All animals acclimatized for 48 h prior to study procedures. Groups of six animals were housed in high efficiency particulate air (HEPA)-filtered cages at controlled room temperature. Nourishment and enrichment were provided as previously described.10
Animals were pretreated with cyclophosphamide and uranyl nitrate prior to inoculation with a bacterial suspension of ∼1 × 107 cfu/mL into one thigh per animal.10,11 Cefiderocol, ceftazidime/avibactam and meropenem were administered to simulate the human plasma exposure of 2 g IV q8h as a 3 h infusion (inf), 2.5 g IV q8h as a 2 h inf, and 2 g IV q8h as a 3 h inf, respectively.10–12
Post-inoculation, groups of six mice were sacrificed at 2 h (0 h control) and the inoculated thigh was harvested to determine the baseline bacterial burden.10 Groups of six mice received sham control (24 h control), cefiderocol, ceftazidime/avibactam, meropenem, or ceftazidime/avibactam/meropenem HSRs subcutaneously for 24 h. The inoculated thigh was harvested for bacterial enumeration. Outliers were excluded if the log10 cfu/thigh was outside of Tukey’s hinges.13 Efficacy was determined as achievement of 1-log10 change of cfu/thigh relative to baseline because the 1-log10 kill endpoint in this translational model is associated with clinical success in humans.14 The proportion of isolates in each genotypic group meeting this endpoint was assessed.
Results
In vitro potency
In vitro MICs are reported in Table 1. The GES isolates were obtained from different geographical locations and produced GES-1, GES-5 or miscellaneous GESs (GES-19, GES-20, GES-26). Additional resistance genes in each isolate are presented in Table S1 (available as Supplementary data at JAC Online). Per CLSI breakpoints,8 100%, 6% and 39% of isolates were susceptible to cefiderocol, meropenem and ceftazidime/avibactam, respectively.
Table 1.
Genotypic and phenotypic information on the clinical isolates included in the in vivo model
| Isolate ID | Source | Genotypic resistance determinants | Agent MIC, mg/L (murine %fT > MIC) | ||
|---|---|---|---|---|---|
| Cefiderocol | Ceftazidime/avibactama,b | Meropenem | |||
| INT-6-19 | Middle East | GES-1 | 0.25 (100%) | 8 (88%) | 8 (75%) |
| INT-6-44 | Middle East | GES-1 | 0.25 (100%) | 16 (62%) | 16 (55%) |
| INT-12-16 | Europe | GES-1 | 0.25 (100%) | 16 (62%) | 8 (75%) |
| INT-6-8 | Middle East | GES-1 | 0.25 (100%) | 8 (88%) | 8 (75%) |
| INT-6-55 | Middle East | GES-1 | 0.5 (100%) | 8 (88%) | 2 (100%) |
| INT-5-8 | Europe | GES-5 | 1 (100%) | 64 (0%) | 16 (55%) |
| INT-3-29 | Europe | GES-5 | 0.125 (100%) | 8 (88%) | >64 (0%) |
| INT-12-34 | Europe | GES-5 | 0.125 (100%) | 4 (99%) | >64 (0%) |
| INT-10-10 | Middle East | GES-5 | 0.125 (100%) | 4 (99%) | >64 (0%) |
| INT-6-42 | Middle East | GES-5 | 0.125 (100%) | 4 (99%) | >64 (0%) |
| PSA 1866 | AR Bank #0767 | GES-20 | 1 (100%) | 64 (0%) | >64 (0%) |
| PSA 1871 | AR Bank #0772 | GES-20, GES-26 | 0.5 (100%) | >64 (0%) | 16 (55%) |
| PSA 1869 | AR Bank #0770 | GES-19, GES-26 | 0.5 (100%) | 16 (62%) | >64 (0%) |
| PSA 1867 | AR Bank #0768 | GES-19, GES-20 | 1 (100%) | 32 (34%) | >64 (0%) |
| PSA 1868 | AR Bank #0769 | GES-19, GES-26 | 1 (100%) | 64 (0%) | >64 (0%) |
| PSA 1864 | AR Bank #0765 | GES-19, GES-20 | 1 (100%) | 64 (0%) | >64 (0%) |
| PSA 1870 | AR Bank #0771 | GES-19, GES-20 | 0.5 (100%) | 64 (0%) | >64 (0%) |
| PSA 1863 | AR Bank #0764 | GES-19, GES-20 | 0.5 (100%) | 64 (0%) | >64 (0%) |
AR Bank, Antibiotic Resistance Isolate Bank; GES, Guiana extended spectrum; INT, International; PSA, P. aeruginosa.
MICs determined at fixed avibactam concentration of 4 mg/L.
%fT > MIC expressed as the ceftazidime exposure.
In vivo efficacy
The infection was established in vivo with a baseline burden of 5.25 ± 0.4 log10 cfu/thigh. All isolates displayed a robust growth (+2.7 to +4.2 change in log10 cfu/thigh). Isolates producing GES-1 were killed by ≥1-log10 when treated with any HSR (i.e. cefiderocol, meropenem, ceftazidime/avibactam or ceftazidime/avibactam/meropenem), with the highest magnitude of kill observed with cefiderocol (mean of −3.1 ± 0.7 log10 kill relative to 0 h control) and ceftazidime/avibactam/meropenem (−3.2 ± 0.6 log10 kill; Figure 1a).
Figure 1.
In vivo change in log10 cfu/thigh in each isolate treated with sham control, cefiderocol 2 g q8h 3 h infusion HSR, ceftazidime/avibactam 2.5 g q8h 2 h infusion HSR, meropenem 2 g q8h 3 h infusion HSR, or a combination of meropenem and ceftazidime/avibactam HSR against P. aeruginosa with GES β-lactamases. (a) GES-1, (b) GES-5 and (c) miscellaneous GESs such as GES-19, GES-20 and GES-26. PSA, P. aeruginosa.
Against isolates producing GES-5 (Figure 1b), meropenem failed in vivo, consistent with its phenotypic profile. Ceftazidime/avibactam produced kill against all GES-5 isolates irrespective of their in vitro susceptibility, with a mean of −2.7 ± 1.2 log10 kill. Cefiderocol and ceftazidime/avibactam/meropenem produced the highest magnitude of kill against GES-5 isolates, with −3.0 ± 0.5 and −3.0 ± 0.6 log10 kill, respectively.
Isolates producing miscellaneous GESs (Figure 1c) were consistently killed by cefiderocol and ceftazidime/avibactam/meropenem, with −2.2 ± 1.2 and −1.8 ± 1.2 log10 of kill, respectively. Meropenem in vivo activity was poor for all isolates, except for PSA 1871 with an MIC of 16 mg/L, where the pharmacodynamically optimized high-dose regimen is expected to provide sufficient exposure. Although the majority of the isolates were killed by ceftazidime/avibactam humanized exposures despite the elevated MICs, isolates PSA 1871 (MIC >64 mg/L), PSA 1867 (MIC 32 mg/L) and PSA 1863 (MIC 64 mg/L) failed to achieve the prerequisite 1-log of kill predictive of clinical efficacy in patients. Upon combination with meropenem, in vivo efficacy was restored.
Cefiderocol and ceftazidime/avibactam/meropenem produced ≥1-log10 of kill against all tested isolates. Meropenem was active against 100% of GES-1 isolates, 8% of GES-5 and 0% of miscellaneous GESs, consistent with the MICs. Ceftazidime/avibactam was active against all GES-1 and all GES-5 isolates irrespective of the in vitro susceptibility. Additionally, ceftazidime/avibactam remained active in 62.5% of miscellaneous GESs despite the elevated MICs in some isolates.
Discussion
Due to the scarcity of clinical data associated with treatment outcomes of different antibiotics against GES-producing P. aeruginosa, murine models using humanized exposures provide translational data to devise rational therapeutic strategies. Our data demonstrated that cefiderocol provided significant activity with ≥1-log10 of kill across all tested isolates expressing various GES β-lactamases. This novel cephalosporin with its siderophore-mediated cell entry retains activity against a variety of β-lactamases, and thus represents a therapeutic option for challenging pathogens.15 The in vivo kill in our data was concordant with cefiderocol’s in vitro activity and supports its use for treating infections caused by GES-producing P. aeruginosa where therapeutic options are limited.
Meropenem activity was predicted by its phenotypic profile and existing pharmacokinetics/pharmacodynamics targets. β-Lactams display time-dependent bactericidal activity, and the established benchmark for meropenem activity against P. aeruginosa is 40% fT > MIC.16 Meropenem 2 g q8h 3 h inf provides sufficient exposure up to an MIC of 16 mg/L, and thus resulted in ≥1-log10 kill against such isolates. Isolates expressing GES-1 demonstrated relatively low-level resistance (MICs ≤16 mg/L), and therefore the pharmacodynamically optimized meropenem retained activity. Conversely, meropenem activity against GES-5 and miscellaneous GESs was lacking, consistent with the elevated MICs. One exception was isolate PSA 1871 producing GES-20 and GES-26, which had a meropenem MIC of 16 mg/L, resulting in in vivo kill.
Our group has previously published murine in vivo efficacy of ceftazidime, ceftazidime/avibactam and meropenem against five GES P. aeruginosa.13 Meropenem activity was consistent with the phenotypic profile similar to the current analysis. Ceftazidime was active in vitro in 2/5 isolates, and the addition of avibactam improved the activity against ceftazidime-resistant isolates in vitro and in vivo. Due to the poor in vitro potency of ceftazidime alone in surveillance cohorts of GES-producing P. aeruginosa, it was not included in the current study.2
In the present study, isolates assessed spanned a range of in vitro MICs to ceftazidime/avibactam similar to previous findings.3 Indeed, ceftazidime/avibactam was active in vivo against GES-producing isolates that were susceptible in vitro and thus is a therapeutic option when phenotypic activity is confirmed. Despite the wide range of MICs assessed, ceftazidime/avibactam activity was generally high because it produced ≥1-log10 kill in 15/18 isolates including isolates with MIC >64 mg/L and %f T > MIC of 0. The in vivo/in vitro discordance of β-lactam/β-lactamase inhibitor combinations was previously reported with piperacillin/tazobactam and imipenem/relebactam, where in vivo efficacy was observed despite MICs elevated above the predicted pharmacodynamic breakpoints.17,18 A hypothesis for the discordance is that the higher concentration of the inhibitor in the humanized regimen relative to the fixed concentration in MIC assays may be sufficient to protect the β-lactam agent at clinically administered dosages. Confirmatory studies using varying concentrations of the β-lactamase inhibitors in MIC testing are needed.
We previously observed that GES isolates resistant to ceftazidime/avibactam were resensitized to meropenem in vivo.13 This finding of resensitization to carbapenems amongst serine β-lactamase-producing isolates resistant to ceftazidime/avibactam was previously seen with AmpC and KPC isolates.19,20 Thus a combination of meropenem plus ceftazidime/avibactam may in theory result in killing against GES-producing isolates that develop resistance to ceftazidime/avibactam by killing both susceptible and resistant subpopulations. Indeed, ceftazidime/avibactam/meropenem was efficacious against all tested isolates assessed in the model. The in vivo magnitude of kill was similar to that observed with cefiderocol and each may represent a viable therapeutic strategy. However, unlike cefiderocol, there are no approved susceptibility breakpoints or testing methodology to predict the efficacy of the combination.
In conclusion, due to the lack of clinically available molecular diagnostics the detection of GES-producing P. aeruginosa is difficult. Clinicians must rely on available phenotypic profiles to make therapeutic decisions for these difficult-to-treat pathogens. Our in vivo data suggest that when an infection with GES-producing P. aeruginosa is suspected or confirmed, the phenotypic susceptibility profiles of cefiderocol, ceftazidime/avibactam and meropenem are predictive of clinical success. As a single agent, cefiderocol produced consistent in vivo activity against the spectrum of GES subtypes studied and would, therefore, seem to be the prevailing therapeutic choice when available. In the absence of cefiderocol, a combination of ceftazidime/avibactam plus meropenem could be a valuable therapeutic option as observed in our current translational model. Clinical data assessing the outcomes of patients treated with cefiderocol or ceftazidime/avibactam/meropenem are warranted.
Supplementary Material
Acknowledgements
We would like to thank the staff from the Center for Anti-Infective Research and Development for their vital assistance in the conduct of this study.
Contributor Information
Yasmeen Abouelhassan, Center for Anti-Infective Research and Development, Hartford Hospital, 80 Seymour Street, Hartford, CT 06102, USA.
Christian M Gill, Center for Anti-Infective Research and Development, Hartford Hospital, 80 Seymour Street, Hartford, CT 06102, USA.
David P Nicolau, Center for Anti-Infective Research and Development, Hartford Hospital, 80 Seymour Street, Hartford, CT 06102, USA; Division of Infectious Diseases, Hartford Hospital, Hartford, CT, USA.
Funding
This project was internally funded by the Center for Anti-Infective Research and Development.
Transparency declarations
C.M.G. has received research grants from Cepheid, Everest Medicines, Shionogi and Entasis. D.P.N. is a consultant, speaker bureau member and has received other research grants from Abbvie, Cepheid, Merck, Paratek, Pfizer, Wockhardt, Shionogi and Tetraphase. Y.A. has no conflicts to declare.
Supplementary data
Table S1 is available as Supplementary data at JAC Online.
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