Activity Of Carbapenems Research Articles

Redirecting pantoprazole as a metallo-beta-lactamase inhibitor in carbapenem-resistant Klebsiella pneumoniae.

The development of resistance to carbapenems in Klebsiella pneumoniae due to the production of metallo-β-lactamases (MBLs) is a critical public health problem because carbapenems are the last-resort drugs used for treating severe infections of extended-spectrum β-lactamases (ESBLs) producing K. pneumoniae. Restoring the activity of carbapenems by the inhibition of metallo-β-lactamases is a valuable approach to combat carbapenem resistance. In this study, two well-characterized clinical multidrug and carbapenem-resistant K. pneumoniae isolates were used. The sub-inhibitory concentrations of pantoprazole and the well-reported metallo-β-lactamase inhibitor captopril inhibited the hydrolytic activities of metallo-β-lactamases, with pantoprazole having more inhibiting activities. Both drugs, when used in combination with meropenem, exhibited synergistic activities. Pantoprazole could also downregulate the expression of the metallo-β-lactamase genes bla NDM and bla VIM. A docking study revealed that pantoprazole could bind to and chelate zinc ions of New Delhi and Verona integron-encoded MBL (VIM) enzymes with higher affinity than the control drug captopril and with comparable affinity to the natural ligand meropenem, indicating the significant inhibitory activity of pantoprazole against metallo-β-lactamases. In conclusion, pantoprazole can be used in combination with meropenem as a new strategy for treating serious infections caused by metallo-β-lactamases producing K. pneumoniae.

Approaches to enhance the antimicrobial activity of carbapenems within bacterial biofilms

This review highlights current strategies to improve the anti-biofilm activity of clinically approved carbapenem-based antibiotics.

Resensitizing multidrug-resistant Gram-negative bacteria to carbapenems and colistin using disulfiram

The increasing incidence of bacterial infections caused by multidrug-resistant (MDR) Gram-negative bacteria has deepened the need for new effective treatments. Antibiotic adjuvant strategy is a more effective and economical approach to expand the lifespan of currently used antibiotics. Herein, we uncover that alcohol-abuse drug disulfiram (DSF) and derivatives thereof are potent antibiotic adjuvants, which dramatically potentiate the antibacterial activity of carbapenems and colistin against New Delhi metallo-β-lactamase (NDM)- and mobilized colistin resistance (MCR)-expressing Gram-negative pathogens, respectively. Mechanistic studies indicate that DSF improves meropenem efficacy by specifically inhibiting NDM activity. Moreover, the robust potentiation of DSF to colistin is due to its ability to exacerbate the membrane-damaging effects of colistin and disrupt bacterial metabolism. Notably, the passage and conjugation assays reveal that DSF minimizes the evolution and spread of meropenem and colistin resistance in clinical pathogens. Finally, their synergistic efficacy in animal models was evaluated and DSF-colistin/meropenem combination could effectively treat MDR bacterial infections in vivo. Taken together, our works demonstrate that DSF and its derivatives are versatile and potent colistin and carbapenems adjuvants, opening a new horizon for the treatment of difficult-to-treat infections.

Interplay between OXA-10 β-Lactamase Production and Low Outer-Membrane Permeability in Carbapenem Resistance in Enterobacterales.

The OXA-10 class D β-lactamase has been reported to contribute to carbapenem resistance in non-fermenting Gram-negative bacilli; however, its contribution to carbapenem resistance in Enterobacterales is unknown. In this work, minimum inhibitory concentrations (MICs), whole genome sequencing (WGS), cloning experiments, kinetic assays, molecular modelling studies, and biochemical assays for carbapenemase detection were performed to determine the impact of OXA-10 production on carbapenem resistance in two XDR clinical isolates of Escherichia coli with the carbapenem resistance phenotype (ertapenem resistance). WGS identified the two clinical isolates as belonging to ST57 in close genomic proximity to each other. Additionally, the presence of the blaOXA-10 gene was identified in both isolates, as well as relevant mutations in the genes coding for the OmpC and OmpF porins. Cloning of blaOXA-10 in an E. coli HB4 (OmpC and OmpF-deficient) demonstrated the important contribution of OXA-10 to increased carbapenem MICs when associated with porin deficiency. Kinetic analysis showed that OXA-10 has low carbapenem-hydrolysing activity, but molecular models revealed interactions of this β-lactamase with the carbapenems. OXA-10 was not detected with biochemical tests used in clinical laboratories. In conclusion, the β-lactamase OXA-10 limits the activity of carbapenems in Enterobacterales when combined with low permeability and should be monitored in the future.

ВЛИЯНИЕ МЕРОПЕНЕМА И ИМИЦЕНЕМА НА МИКРОБИОЦЕНОЗ И МОРФОЛОГИЮ ТОНКОГО КИШЕЧНИКА КРЫС

Background. Gut microbiota plays an important role in the human body, antimicrobial drugs intake being one of the main factors affecting microbiota composition. Antibiotics suppress normal intestinal microflora allowing for the growth and development of pathogenic flora. Objective. To study the effect of meropenem and imicenem on the microbiocenosis and morphology of the small intestine in rats. Material and methods. In an experiment on rats, we analysed the microbiocenosis of the large intestine after intraperitoneal administration of meropenem, imicinem and 0.9% NaCl, as well as examined samples of the small intestine. Results. In a comparative analysis of the control group with the meropenem and imicinem groups, there has been noted a decrease in the number of anaerobes in the meropenem group (8.6±0.14, p<0.05) as compared with the control group (9.4±0.22, p<0 .05) and the imicinem group (9.4±0.40, p<0.05); the number of bifidobacteria decreased in the meropenem group (8.1±0.16, p<0.05) in contrast to the control group (9.2±0.25, p<0.05) and the imicinem group (9.2±0.41, p<0.05). When compared with meropenem (3.8±0.44, p<0.05), imicinem (4.9±0.20, p<0.05) led to a greater growth of spore-forming anaerobes. In animals of the imicinem group, the histostructure of the jejunal mucosa was similar to the control group, while in the meropenem group, inflammatory changes in the stroma of the villi of the jejunal mucosa were observed. Conclusions. In a comparative analysis of carbapenem activity, it has been determined that these antibiotics both have a significant effect on the microbiocenosis of the large intestine and affect the mucous wall of the small intestine. These should be taken into account by clinicians in order to improve the effectiveness and safety of antibiotic therapy.

Rifampicin Enhanced Carbapenem Activity with Improved Antibacterial Effects and Eradicates Established Acinetobacter baumannii Biofilms.

Biofilm-mediated infections are critical to public health and a leading cause of resistance among pathogens, amounting to a prolonged hospital stay and increased mortality rate in the intensive care unit. In this study, the antibacterial and antibiofilm activities of rifampicin or carbapenem monotherapies were compared with rifampicin and carbapenem combination therapies against rifampicin-resistant and carbapenem-resistant Acinetobacter baumannii isolates. Among 29 CRAB isolates, 24/29 (83%) were resistant to rifampicin, with MIC values between 2-256 µg/mL. Checkerboard assays disclosed that combination therapies at FICIs between 1/8 and 1/4 improved the activity of carbapenems at subinhibitory concentrations. Time-kill kinetics indicated a 2- to 4-log reduction at 1/2 MIC rifampicin + 1/4 MIC carbapenem and 1/4 MIC rifampicin + 1/4 MIC carbapenem against the isolates, with the MIC values ranging from 2-8 µg/mL. The MTT assay revealed a dose-dependent decrease of the cell viability of established bacterial biofilm at 4 MIC rifampicin + 2 MIC carbapenems, with a percentage reduction of 44-75%, compared with monotherapies at 16 MIC. Scanning electron microscopy further confirmed bacterial cell membrane disruption, suggesting a synergism between carbapenem and rifampicin against a representative isolate. The findings demonstrated that the combination of rifampicin with carbapenems could improve antibacterial activities and eradicate established Acinetobacter baumannii biofilm.

The In vitro activity of carbapenems alone and in combination with β-lactamase inhibitors against difficult-to-treat mycobacteria; Mycobacterium tuberculosis, Mycobacterium abscessus, and Mycobacterium avium complex: A systematic review.

Difficult-to-treat mycobacterial infections are increasing globally. There is an urgent need of new treatment alternatives for multidrug-resistant Mycobacterium tuberculosis (MTB), as well as nontuberculous mycobacteria such as the Mycobacterium abscessus complex (MABC) and Mycobacterium avium complex (MAC). Recently, new carbapenems and combinations of carbapenems with β-lactamase inhibitors have become available, but activity data in vitro against mycobacteria are so far scarce. Therefore, we performed a systematic review collating the minimum inhibitory concentrations (MICs) of carbapenems, with or without a β-lactamase inhibitors for MTB, MABC, and MAC. The databases PubMed and Web of Science were searched for the relevant articles in English up until September 21, 2022. Screening of studies was performed by two independent reviewers. MIC data by recommended methods with at least five individual MICs were included. Data were reported as MIC range, MIC50, modal MIC, and/or histograms when individual MICs were available. The study protocol was registered at PROSPERO (CRD42021258537). After screening, a total of 75 studies with MIC data for carbapenems with or without β-lactamase inhibitors were included in the review. For MTB, the oral carbapenem tebipenem combined with the β-lactamase inhibitor clavulanic acid resulted in the most significant reduction of MICs. For MABC, the addition of avibactam to tebipenem resulted in a 64-fold reduction of modal MIC. Data were insufficient for the analysis of MAC. Carbapenems, and in particular the novel oral compound tebipenem, in combination with clavulanic acid for MTB and avibactam for MABC may be an untapped potential for difficult-to-treat mycobacterial infections.

The Role Of OprD Gene In The Carbapenems Resistance In Pseudomonas Aeruginosa Isolated From Burns Infection In Al-Najaf Province

Carbapenems-resistant Pseudomonas aeruginosa mutants are disseminating around the globe. 90%-100% of P. aeruginosa isolates became Carbapenems-resistant mutants in some countries like Thailand and Brazil respectively. Finding alternative therapeutic options instead of carbapenems is the global duty of researchers and institutions. The OprD gene mutation/s has the main role in carbapenems resistance appearance in P. aeruginosa due to the decrease in the outer membrane permeability for carbapenems. Aim: This study aims to assess the antimicrobial activity of carbapenems (imipenem, and meropenem) against Pseudomonas aeruginosa isolated from hospitalized burn patients in Al-Najaf/Iraq, and the role of OprD gene mutations in the resistance. Methods and materials: 76 samples were collected from hospitalized burn patients suffering from burn wounds infections, and the identity of each cultivated microbe was confirmed by VITEK compact system 2 and further confirmation for P. aeruginosa isolates via performing molecular identification by OprD gene detection, 10/12 carbapenems resistant isolates, 1/12 sensitive, and 1/12 intermediate resistant were submitted to OprD gene sequencing by Sanger method

Criteria Restricting Inappropriate Meropenem Empiricism (CRIME): a quasi-experimental carbapenem restriction pilot at a large academic medical centre

The broad-spectrum activity of carbapenems makes them appealing for empirical use; however, they are associated with development of Clostridioides difficile infection (CDI) and multidrug resistance. Selective carbapenem use is vital in maintaining their effectiveness. We examined the impact of meropenem restriction criteria on utilisation and patient outcomes. This quasi-experimental study was conducted at a single academic medical centre after medication use evaluation found frequent inappropriate meropenem utilisation. Antimicrobial stewardship-led restriction criteria were developed and implemented in February 2022. Investigators aimed to determine how restriction criteria affected meropenem utilisation across 8 weeks in the pre- (February-April 2020) versus post-implementation period (February-April 2022). The primary outcome was inappropriateness of meropenem utilisation. Secondary outcomes included days of therapy per 1000 patient-days (DOT/1000 PD), hospital length of stay (LOS), CDI Standardized Infection Ratio (SIR), and acquisition cost. Across the 8-week timeframes, reductions in inappropriate meropenem use (64.5% vs. 12.8%; P < 0.001), duration of therapy [5.8 (3.2-7.3) vs. 2.4 (1.0-5.5) days; P < 0.001] and utilisation (30.5 vs. 8.3 DOT/1000 PD; P < 0.001) pre- versus post-implementation were observed. Total meropenem orders decreased by 65% (P < 0.001). Median hospital LOS also decreased between periods [11.9 (7.8-20.4) vs. 9.2 (5.4-15.2) days], although not statistically significant (P=0.051). There was no difference in CDI SIR (0.1 vs. 0.1; P=0.99). Projected annual cost savings were ∼US$57 300. Implementation of antimicrobial stewardship-initiated restriction criteria can reduce inappropriate meropenem utilisation, overall number of orders, and total duration of therapy.

A Variant Carbapenem Inactivation Method (CIM) for Acinetobacter baumannii Group with Shortened Time-to-Result: rCIM-A.

Carbapenem-resistant Acinetobacter baumannii group organisms (CRAB) are challenging because the choice between targeted, new antibiotic drug options and hygiene measures should be guided by a timely identification of resistance mechanisms. In CRAB, acquired class-D carbapenemases (CHDLs) are active against meropenem and imipenem. If PCR methods are not the first choice, phenotypic methods have to be implemented. While promising, the carbapenemase inactivation method (CIM) using meropenem-hydrolysis is, however, hampered by poor performance or overly long time-to-result. We developed a rapid CIM (rCIM-A) with good performance using ertapenem, imipenem, and meropenem disks, 2-h permeabilization and incubation with the test strain in trypticase soy broth, and a read-out of residual carbapenem activity after 6 h, and optionally after 16–18 h. Using clinical isolates and type-strains of Acinetobacter (n = 67) not harboring carbapenemases (n = 28) or harboring acquired carbapenemases (n = 39), the sensitivity of detection was 97.4% with the imipenem disk after 6 h at a specificity of 92.9%. If the inhibition zone around the ertapenem disk at 6 h was 6 or ≤26 mm at 16–18 h, or ≤25.5 mm for meropenem, the specificity was 100%. Because of the high negative predictive value, the rCIM-A seems particularly appropriate in areas of lower CRAB-frequency.

Imitation of β-lactam binding enables broad-spectrum metallo-β-lactamase inhibitors.

Carbapenems are vital antibiotics, but their efficacy is increasingly compromised by metallo-β-lactamases (MBLs). Here we report the discovery and optimization of potent broad-spectrum MBL inhibitors. A high-throughput screen for NDM-1 inhibitors identified indole-2-carboxylates (InCs) as potential β-lactamase stable β-lactam mimics. Subsequent structure-activity relationship studies revealed InCs as a new class of potent MBL inhibitor, active against all MBL classes of major clinical relevance. Crystallographic studies revealed a binding mode of the InCs to MBLs that, in some regards, mimics that predicted for intact carbapenems, including with respect to maintenance of the Zn(II)-bound hydroxyl, and in other regards mimics binding observed in MBL-carbapenem product complexes. InCs restore carbapenem activity against multiple drug-resistant Gram-negative bacteria and have a low frequency of resistance. InCs also have a good in vivo safety profile, and when combined with meropenem show a strong in vivo efficacy in peritonitis and thigh mouse infection models.

Genetyping of Carbapenem-resistant Organisms Isolated from Clinical Isolates Received from Tertiary Care Hospitals of Ahmedabad, Gujarat

The world is seeing a continuous rise in the levels of antibiotic resistance1. Organisms develop new resistance mechanisms, emerge, and spread the resistance worldwide, making it challenging to treat common infectious diseases. In the current study, clinical isolates received between the years 2017 to 2020 were cultured and the isolated organisms were screened for antibiotic resistance; isolates with multiple drug resistance were further subjected to confirmatory screening through Combined Disc Test (CDT) and Modified Hodge Test (M.H.T.), and molecular characterization to be finally tested for gene expression analysis. Molecular characterization involved screening of genes blaVIM-2, blaKPC-3, blaNDM-1, and blaIMP-11 responsible for imparting carbapenem drug resistance2. From the laboratories of tertiary care hospitals, a total of 1452 clinical isolates were collected and identified. The organisms were subjected to antibiotic susceptibility screening and carbapenem resistance screening. The isolates found positive in the screenings were subjected to molecular characterization for genes, blaVIM-2, blaKPC-3, blaNDM-1, and blaIMP-11, responsible for imparting carbapenem drug resistance. Most of the isolates were resistant variably to aminoglycosides but were found to be resistant to fluoroquinolones and β-lactams group of antibiotics. Carbapenem activity was detected in twelve percent of total isolates and 27 percent among multidrug-resistant isolates. blaNDM-1 gene was found present in 77% isolates, and five organisms among the total number of organisms showed pan drug resistance.

Activity of β-lactam plus β-lactam-enhancer combination cefepime/zidebactam against Klebsiella pneumoniae harbouring defective OmpK35/36 porins and carbapenemases

Cefepime/zidebactam is a β-lactam/β-lactam-enhancer based novel antibiotic which is in clinical development for treating infections caused by multidrug-resistant Gram-negative bacteria. Here, in vitro activity of cefepime/zidebactam was determined against multicentre Klebsiella pneumoniae clinical isolates co-expressing serine and/or metallo-carbapenemases and defective OmpK35 and OmpK36 porins. The MICs were determined using the reference broth microdilution method. Outer membrane protein expression was assessed using SDS-PAGE and mutations in the genes encoding OmpK35 and OmpK36 were identified by DNA sequencing. Among 34 isolates studied, carbapenemase genes, blaKPC and blaOXA-48-like, were present in 18 and 11 isolates, respectively; 5 isolates harboured both blaOXA-48-like and blaNDM. Point mutations, insertions, and duplications in OmpK35 and OmpK36, which are known to impact the activity of carbapenems, were detected. Against these isolates, cefepime/zidebactam (1:1) showed a consistent activity (MICs ≤4 mg/L). In conclusion, cefepime/zidebactam overcomes enzymatic, and non–enzymatic carbapenem-impacting resistance mechanisms concurrently expressed in K. pneumoniae.

In Vitro and In Vivo Characterization of Tebipenem (TBP), an Orally Active Carbapenem, against Biothreat Pathogens.

Bacillus anthracis and Yersinia pestis, causative pathogens for anthrax and plague, respectively, along with Burkholderia mallei and B. pseudomallei are potential bioterrorism threats. Tebipenem pivoxil hydrobromide (TBP HBr, formerly SPR994), is an orally available prodrug of tebipenem, a carbapenem with activity versus multidrug-resistant (MDR) gram-negative pathogens, including quinolone-resistant and extended-spectrum-β-lactamase-producing Enterobacterales. We evaluated the in vitro activity and in vivo efficacy of tebipenem against biothreat pathogens. Tebipenem was active in vitro against 30-strain diversity sets of B. anthracis, Y. pestis, B. mallei, and B. pseudomallei with minimum inhibitory concentration (MIC) values of 0.001 - 0.008 μg/ml for B. anthracis, ≤0.0005 - 0.03 μg/ml for Y. pestis, 0.25 - 1 μg/ml for B. mallei, and 1 - 4 μg/ml for B. pseudomallei In a B. anthracis murine model, all control animals died within 52 h post challenge. The survival rates in the groups treated with tebipenem were 75% and 73% when dosed at 12 h and 24 h post challenge, respectively. The survival rates in the positive control groups treated with ciprofloxacin were 75% and when dosed 12 h and 25% when dosed 24 h post challenge, respectively. Survival rates were significantly (p=0.0009) greater in tebipenem groups treated at 12 h and 24 h post challenge and in the ciprofloxacin group 12 h post-challenge vs. the vehicle-control group. For Y. pestis, survival rates for all animals in the tebipenem and ciprofloxacin groups were significantly (p<0.0001) greater than the vehicle-control group. These results support further development of tebipenem for treating biothreat pathogens.

Synergistic Activity of Equol and Meropenem against Carbapenem-Resistant Escherichia coli.

The emergence of carbapenem-resistant Enterobacterales (CRE) seriously limits treatment options for bacterial infections. Combined drugs are an effective strategy to treat these resistant strains. This study aimed to evaluate the synergistic effect of equol and meropenem against carbapenem-resistant Escherichia coli. First, this study investigated the antibacterial activity of carbapenems on clinically isolated E. coli strains by analyzing the minimum inhibitory concentrations (MICs). The E. coli strains were all resistant to carbapenem antibiotics. Therefore, we confirmed the cause of carbapenem resistance by detecting blaKPC and blaOXA-48 among the carbapenemase genes using polymerase chain reaction (PCR) analysis. Checkerboard and time-kill analyses confirmed that equol restored the susceptibility of carbapenem-resistant E. coli to meropenem. Also, the transcription levels of specific carbapenemase genes in E. coli were significantly suppressed by equol. The study also evaluated the anti-virulence effects of equol on bacterial biofilm and motility through phenotypic and genotypic analyses. In conclusion, our results revealed that equol had a synergistic effect with meropenem on carbapenem-resistant E. coli. Therefore, this study suggests that equol is a promising antibiotic adjuvant that prevents the expression of carbapenemases and virulence factors in carbapenem-resistant E. coli.

Мониторинг инфицирования и резистентности Achromobacter xylosoxidans у взрослых пациентов с муковисцидозом

Aim: to evaluate the infection rate and resistance of isolated Achromobacter xylosoxidans to carbapenems in adult cystic fibrosis patients (CF). Patients and Methods: a retrospective analysis of the results of culture test and time-of-flight mass spectrometry MALDI-TOF was conducted: 685 sputum samples of 58 adult CF patients for a period of 5 years (2016–2020). To assess the sensitivity to imipenem and meropenem, the agar gradient diffusion and disk diffusion method were used. Results: the incidence of infection with A. xylosoxidans in adult CF patients for the period from 2016 to 2020 when monitoring a single sample of patients (n=24) to evaluate the occurrence of this pathogen with increasing age ranged from 16.6% in 2016–2017, increasing to 37.5% in 2018–2019, and with a further reduction to 20.8% (associated with disease fatal outcome in 3 of the 9 infected patients). There was no statistically significant dependence of the fatal outcome on infection with A. xylosoxidans. When analyzing the entire pool of patients (n=58) from 2016 to 2020, the release frequency of Pseudomonas aeruginosa remains approximately the same, varying from 63.3% to 46.5% and maintaining a numerical advantage in all follow-up periods, while the A. xylosoxidans infection ranges from 13.7% to 39.3%. In 2016–2018, 50% of isolates were sensitive to carbapenems, in 2018 — 53.8% of isolates, in 2019–2020 — the activity of obtained isolates decreased to 37.5% and 30.7%, respectively. Conclusion: despite the dynamics of indicators and the sample size, the dynamics over 5 years maintained a group-wide proportion of microbiome species dominated primarily by Pseudomonas infection and A. xylosoxidans. In our follow-up, the activity of carbapenems in relation to A. xylosoxidans has almost halved. KEYWORDS: cystic fibrosis, Burkholderia cepacia complex, Achromobacter xylosoxidans, Pseudomonas aeruginosa, lethality, antibiotic resistance, carbapenems. FOR CITATION: Makhmutova V.R., Gembitskaya T.E., Chermensky A.G. et al. Achromobacter xylosoxidans infection and resistance monitoring in adult cystic fibrosis patients. Russian Medical Inquiry. 2021;5(7):462–467 (in Russ.). DOI: 10.32364/2587-6821-2021-5-7-462-467.

Discovery of ANT3310, a Novel Broad-Spectrum Serine β-Lactamase Inhibitor of the Diazabicyclooctane Class, Which Strongly Potentiates Meropenem Activity against Carbapenem-Resistant Enterobacterales and Acinetobacter baumannii

The diazabicyclooctanes (DBOs) are a class of serine β-lactamase (SBL) inhibitors that use a strained urea moiety as the warhead to react with the active serine residue in the active site of SBLs. The first in-class drug, avibactam, as well as several other recently approved DBOs (e.g., relebactam) or those in clinical development (e.g., nacubactam and zidebactam) potentiate activity of β-lactam antibiotics, to various extents, against carbapenem-resistant Enterobacterales (CRE) carrying class A, C, and D SBLs; however, none of these are able to rescue the activity of β-lactam antibiotics against carbapenem-resistant Acinetobacter baumannii (CRAB), a WHO "critical priority pathogen" producing class D OXA-type SBLs. Herein, we describe the chemical optimization and resulting structure-activity relationship, leading to the discovery of a novel DBO, ANT3310, which uniquely has a fluorine atom replacing the carboxamide and stands apart from the current DBOs in restoring carbapenem activity against OXA-CRAB as well as SBL-carrying CRE pathogens.

The paradoxical in vivo activity of β-lactams against metallo-β-lactamase-producing Enterobacterales is not restricted to carbapenems.

Using murine models of infection, we previously reported the potent in vivo activity of carbapenems against MBL-producing Enterobacterales despite the observed resistance in vitro. In the current study, we examined the in vivo activity of a cefepime human-simulated regimen against MBL-producing Enterobacterales in a murine thigh infection model. A population of clinical isolates and isogenic engineered MBL-producing Enterobacterales transformants expressing MBLs but no detectable cefepime-hydrolysing serine β-lactamases were utilized. KPC-producing isolates were included as positive controls. Cefepime, piperacillin/tazobactam and meropenem MICs were determined using broth microdilution in conventional CAMHB and EDTA-supplemented (zinc-limited) broth. In vivo efficacy of a cefepime human-simulated regimen (2 g q8h as a 2 h infusion) was determined in the neutropenic murine thigh infection model against the test strains. Efficacy was measured as the change in log10 cfu/thigh at 24 h compared with 0 h controls. MBL-producing Enterobacterales strains were found to be cefepime, piperacillin/tazobactam and meropenem non-susceptible in conventional broth. Supplementation with EDTA at a concentration of 300 mg/L resulted in multi-fold reduction in the MICs and restoration of susceptibility. In accordance with the MICs generated in zinc-limited broth, administration of a cefepime human-simulated regimen was associated with substantial bacterial reductions among mice infected with MBL-producing Enterobacterales. Absence of MIC reduction in zinc-limited broth and lack of efficacy among mice infected with KPC-producing isolates were observed. For MBL-producing Enterobacterales, susceptibility testing with Mueller-Hinton broth, a zinc-rich testing medium, is flawed since it does not recapitulate the host environment, in which zinc concentrations are low.

Restoring carbapenem efficacy: a novel carbapenem companion targeting metallo-β-lactamases in carbapenem-resistant Enterobacterales.

The dissemination of MBLs compromises effective use of many β-lactams in the treatment of patients with life-threatening bacterial infections. Predicted global increases in the prevalence of MBL-producing carbapenem-resistant Enterobacterales (CRE) are being realized, yielding infections that are untreatable with existing therapies including newly approved β-lactam/β-lactamase inhibitor combinations. Developing MBL inhibitors (MBLIs) now is essential to address the growing threat that MBL-producing CRE pose to patients. A novel MBLI series was assessed by susceptibility testing and time-kill assays. Target activity and selectivity was evaluated using bacterial NDM, VIM and IMP enzyme assays and human matrix metallopeptidase enzyme assays, respectively, and cytotoxicity was assessed in HepG2 cells. In vivo efficacy of meropenem/MBLI combinations was evaluated in a mouse thigh infection model using an NDM-1-producing Escherichia coli strain. Combination of MBLIs with carbapenems reduced MICs for NDM/IMP/VIM-producing Enterobacterales by up to 128-fold compared with the carbapenems alone. Supplementation of meropenem with the promising compound 272 reduced the MIC90 from 128 to 0.25 mg/L in a panel of MBL-producing CRE clinical isolates (n = 115). Compound 272 restored the bactericidal activity of meropenem and was non-cytotoxic, potentiating the antimicrobial action of meropenem through specific inhibition of NDM, IMP and VIM. In vivo efficacy was achieved in a mouse thigh infection model with meropenem/272 dosed subcutaneously. We have developed a series of rationally designed MBLIs that restore activity of carbapenems against NDM/IMP/VIM-producing Enterobacterales. This series warrants further development towards a novel combination therapy that combats antibiotic-resistant organisms, which pose a critical threat to human health.

Dimercaptosuccinic acid in combination with carbapenems against isogenic strains of Escherichia coli producing or not producing a metallo-β-lactamase in vitro and in murine peritonitis.

Carbapenemase-producing Enterobacterales represent a major therapeutic challenge. MBLs, requiring zinc at their catalytic site, could be inhibited by meso-dimercaptosuccinic acid (DMSA), a heavy metal chelator already widely used for treating lead intoxication. To evaluate the activity of carbapenems alone or combined with DMSA against MBL-producing Escherichia coli in a severe murine peritonitis model. Isogenic strains of wild-type E. coli CFT073 producing the MBLs NDM-1, VIM-2 and IMP-1, and the control serine carbapenemases OXA-48 and KPC-3 were constructed. MIC determinations and time-kill assays were performed for imipenem, meropenem and ertapenem alone or in combination with DMSA. Infected mice were treated intraperitoneally for 24 h with imipenem, DMSA or their combination. Bacterial counts in peritoneal fluid and spleen were assessed at 24 h. DMSA in combination with each carbapenem caused a significant decrease in the MICs for all MBL-producing strains, in a concentration-dependent manner, but did not provide benefit against non-MBL strains. In mice infected with the NDM-1-producing strain, the combination of imipenem and DMSA significantly reduced bacterial counts in peritoneal fluid (P = 0.0006) and spleen (P < 0.0001), as compared with imipenem alone, with no benefit against the KPC-3-producing and CFT073 strains. DMSA concentrations in plasma of mice were comparable to those obtained in humans with a standard oral dose. DMSA restores the activity of carbapenems against MBL-producing strains, and its combination with carbapenems appears to be a promising strategy for the treatment of NDM-producing E. coli infections.