The widespread dissemination of blaKPC-possessing Klebsiella pneumoniae isolates, which are resistant to multiple antibiotics, poses a serious threat to hospitalized patients, necessitating new therapeutic strategies. Penem-1 is a novel serine-reactive β-lactamase inhibitor with potent activity against class A, C, and D enzymes and is a potent inhibitor of KPC-2 carbapenemase, but data on its in vitro activity in combination with β-lactams against clinical isolates of blaKPC-possessing K. pneumoniae are lacking. This study tested the in vitro activity of penem-1 (at a constant concentration of 4 μg/ml) combined with four β-lactams (piperacillin, aztreonam, cefotaxime, and cefepime) against 42 previously characterized blaKPC-possessing K. pneumoniae clinical isolates producing multiple β-lactamases using the agar dilution method according to CLSI criteria. Results showed that piperacillin and aztreonam had very high MIC50s (512 μg/ml) when tested alone, while cefotaxime and cefepime had lower MIC50s (64 and 32 μg/ml, respectively). Penem-1 alone had no activity against these strains (MICs > 128 μg/ml). Combinations of penem-1 with piperacillin, aztreonam, or cefotaxime remained mostly in the resistant range, but cefepime combined with penem-1 showed significantly better results, with 88.1% of strains susceptible. In conclusion, penem-1 can lower the MICs of β-lactams (usually by at least three to four 2-fold dilutions) against blaKPC-possessing K. pneumoniae clinical isolates. The combination of cefepime with penem-1 represents a possible therapeutic option for infections caused by these isolates, and further in vitro studies are needed to establish the application spectrum of β-lactam-penem-1 combinations against different multidrug-resistant Gram-negative organisms.