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The widespread dissemination of blaKPC-possessing Klebsiella pneumoniae isolates represents a serious threat for hospitalized patients (4, 8). blaKPC-possessing K. pneumoniae isolates are resistant to all β-lactams, commercially available β-lactam-β-lactamase inhibitor combinations, quinolones, and frequently to aminoglycosides (5, 8). Furthermore, blaKPC-possessing K. pneumoniae isolates can manifest a phenotype of nonsusceptibility to tigecycline and colistin (6, 7). As a result, new therapeutic strategies against infections due to blaKPC-possessing K. pneumoniae isolates should be quickly developed.
Penem-1 (Wyeth Research) is a novel serine-reactive β-lactamase inhibitor with potent activity against class A, C, and D enzymes (1, 10). We recently demonstrated that this methylidene penem is a potent inhibitor of the KPC-2 carbapenemase (with a Km or Ki of 60 nM and a kcat/kinact of 250, where kinact is the rate constant of enzyme inactivation) (9). Data regarding the in vitro activity of penem-1 in combination with various commercially available β-lactams against blaKPC-possessing K. pneumoniae clinical isolates are not yet available.
In this work, we tested the in vitro activity of penem-1 in combination with four β-lactams against a collection of 42 previously characterized blaKPC-possessing K. pneumoniae clinical isolates that produce multiple β-lactamases (5). MICs of β-lactams and β-lactams combined with penem-1 (at a constant concentration of 4 μg/ml) were determined by the agar dilution method according to Clinical and Laboratory Standards Institute (CLSI) criteria (2) using cation-adjusted Mueller-Hinton agar (BBL; Becton Dickinson) and a Steers replicator that delivers 104 CFU/10-μl spot. Piperacillin, cefotaxime, aztreonam (Sigma Chemical Co.), and cefepime (Apotex Corp.) were combined with penem-1. The following ATCC control strains were used: Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853, and K. pneumoniae ATCC 700603. MIC results, including those for the combinations with penem-1, were interpreted according to the CLSI criteria established for the β-lactams when tested alone (3).
As shown in Fig. Fig.1,1, piperacillin and aztreonam exhibited very high MICs (MIC50s, ≥512 μg/ml) when tested against blaKPC-possessing K. pneumoniae isolates whereas cefotaxime and cefepime exhibited lower MICs (MIC50s, 64 and 32 μg/ml, respectively). When penem-1 was tested alone, this inhibitor did not show activity against blaKPC-possessing K. pneumoniae strains (MICs for all strains, >128 μg/ml) (data not shown). When penem-1 was combined with piperacillin, aztreonam, or cefotaxime, the MICs remained mostly in the resistant range. In contrast, cefepime in combination with penem-1 showed significantly better results, with an overall proportion of susceptible strains of 88.1% (Fig. (Fig.11).
In conclusion, we show here that penem-1 possesses good ability to lower the MICs of β-lactams (usually by at least three to four 2-fold dilutions from those of the β-lactam alone) when tested against blaKPC-possessing K. pneumoniae clinical isolates. This new β-lactamase inhibitor undergoes novel reaction chemistry when acylated in the active site and is superior to clavulanate and tazobactam (5, 9). The combination with cefepime represents a possible therapeutic option against infections due to blaKPC-possessing K. pneumoniae isolates. Further in vitro studies should be performed to establish the possible spectrum of application of β-lactam-penem-1 combinations against different types of multidrug-resistant Gram-negative organisms.
This work was supported by the Veterans Affairs Merit Review Program (R.A.B.), the National Institutes of Health (grant no. R03-AI081036 to R.A.B.), and the Geriatric Research Education and Clinical Center VISN 10 (R.A.B.). We thank Louis B. Rice, David L. Paterson, Michael R. Jacobs, Gerri S. Hall, and Stephen G. Jenkins for providing K. pneumoniae isolates. We also thank Tarek S. Mansour for the kind gift of penem-1.
Published ahead of print on 1 February 2010.