IMD-0354 optimization generates potent colistin adjuvants with in vivo activity and reduced eukaryotic toxicity

The development of novel therapeutic approaches to combat infections that stem from extensively drug-resistant (XDR) gram-negative bacteria remains an area of significant unmet need. One such approach is the use of antibiotic adjuvants. Currently, colistin (polymyxin E) is used as the antibiotic of last resort for the treatment of XDR gram-negative bacterial infections. However, resistance to this antibiotic is on the rise. We previously reported that IMD-0354 (an IκB kinase-β inhibitor) and related salicylanilide adjuvants overcome colistin resistance in several gram-negative pathogens. However, this scaffold exhibits unfavorable eukaryotic toxicity, thought to arise from the salicyl moiety. Herein, we investigate the structure-activity relationship (SAR) of second-generation m-hydroxybenzanilide analogs of IMD-0354, to uncover compounds with reduced eukaryotic toxicity and IκB kinase-β inhibitory properties, while maintaining colistin adjuvant activity. We have identified new leads that lower the colistin minimum inhibitory concentration (MIC) upwards of 2048-fold against highly colistin-resistant Acinetobacter baumannii and Klebsiella pneumoniae. In particular, NDM-622 exhibits reduced HepG2 toxicity compared to IMD-0354, with an IC₅₀ of 125 ± 8.0 μM (59.4 ± 3.8 μg/mL) and a therapeutic index of ≥50. In a murine peritonitis model using a highly a colistin-resistant K. pneumoniae strain, NDM-622 and colistin together effect a decrease in colony forming units (CFUs) compared to treatment with colistin alone or vehicle controls. Preliminary mechanism-of-action (MoA) studies suggest that m-hydroxybenzanilides, including NDM-622, likely act via a mechanism distinct from IMD-0354.