Synergistic mode of action of a ceftazidime-hexa-arginine conjugate with resistance-overcoming properties.

We previously described that a non-toxic ceftazidime-hexa-arginine conjugate (CTZ-R6) surpassed the antibacterial potency of ceftazidime (CTZ) against multi-resistant bacteria by up to 1,000-fold and effectively reduced the bacterial load in murine bacteremia. However, the mechanistic understanding of its resistance-overcoming activity remained elusive. Here, we report on the mode of action of CTZ-R6 in gram-positive and gram-negative bacteria. Against a panel of multi-resistant Enterobacteriaceae and non-fermenters, CTZ-R6 outperformed ceftazidime. The conjugate synergized with avibactam in the same manner as CTZ, but its MICs were much lower than those of CTZ. CTZ-R6 crossed the outer membrane of IMPORTANCE: Given the global scale of antibacterial resistance, it becomes clear that numerous infections may eventually become untreatable as former, well-established treatment regimens fail. To prevent the emergence of a post-antibiotic era, strategies are being explored to extend the usefulness of clinically important antibiotics, including the synthesis of conjugates that improve the potency of established drugs. We recently demonstrated that attaching polycationic peptides to all four β-lactam subclasses improved MICs against multi-resistant gram-positive and gram-negative pathogens. The ceftazidime-hexa-arginine conjugate showed the best overall profile and improved the potency of ceftazidime against multi-resistant bacteria by up to 1,000-fold. However, the molecular mechanism of its resistance-overcoming activity remained elusive. Here, we describe which characteristics and capabilities of the peptide and the β-lactam allow them to synergize for increased potency. Mechanistic understanding is essential for further development of such conjugation approaches.