Synthesis and synergistic antibacterial activity of rafoxanide derivative as a novel drug for potentiating colistin activity

Antimicrobial resistance poses a significant global public health challenge in the 21st century, with multidrug resistance in gram-negative bacteria being particularly severe. Colistin (COL) is considered the "last-resort" for treating such infections. Still, its efficacy is significantly reduced due to the emergence and prevalence of mobile colistin resistance (MCR)-mediated acquired drug resistance. Based on the synergistic antibacterial mechanism between the anthelmintic rafoxanide (RAF) and COL, we synthesized a novel RAF derivative (named as dikalisuan) through a carboxymethylation method, and its structure was confirmed using ¹H NMR, ¹³C NMR, and HRMS. <i>In vitro</i> evaluation indicated that the derivatized product had no antibacterial activity (minimum inhibitory concentration [MIC] > 512 µg/mL). However, when used in combination with COL, it showed a significantly synergistic effect (fractional inhibitory concentration index [FICI] < 0.5) on <i>mcr-1-</i>positive and -negative resistant strains, including <i>Escherichia coli</i>, <i>Salmonella</i>, <i>Klebsiella pneumoniae</i>, and <i>Acinetobacter baumannii</i>. Notably, the synergistic bactericidal effect was optimal at a moderate concentration of 8 μg/mL. Furthermore, within the concentration range of 2 to 128 μg/mL, dikalisuan alone or in combination with COL (2 μg/mL) showed no significant hemolysis of red blood cells or cytotoxicity. This derivative provides a new strategy for developing adjuvants against multidrug-resistant bacteria and lays the foundation for the structural modification of anthelmintic salicylanilides.IMPORTANCEAntimicrobial resistance in gram-negative bacteria is a critical global health threat, with colistin (COL) being a "last-resort" treatment. However, rising resistance has blunted its effectiveness. This study developed a new derivative of rafoxanide (RAF). While this new compound itself is not antibacterial, it effectively potentiates COL's power against resistant bacteria when used in combination. This strategy offers a promising approach to extend the lifespan of our current antibiotics and provides a new weapon in the fight against difficult-to-treat drug-resistant bacteria.