Design, Synthesis, and Biological Evaluation of Pyrimido[4,5- b ]indole Derivatives with Potent Activity against Multidrug-Resistant Gram-Negative Bacteria

The escalating crisis of antimicrobial resistance (AMR), particularly multidrug-resistant (MDR) Gram-negative bacterial infections, presents a grave threat to global public health. This urgent unmet medical need underscores the demand for novel antibiotics capable of overcoming existing resistance mechanisms. The ATP-binding sites of bacterial type II topoisomerases represent promising therapeutic targets against MDR Gram-negative pathogens. Pyrimido[4,5- b ]indole-based compounds (GP-1) are the first reported ATPase inhibitors of DNA gyrase with potent and broad-spectrum antibacterial activity against both MDR Gram-positive and Gram-negative pathogens; however, they are associated with hERG inhibition and suboptimal pharmacokinetic profiles. To overcome these limitations, we designed a new series of pyrimido[4,5- b ]indole-based derivatives using structure-based scaffold modification and CADD-guided optimization. This campaign culminated in the discovery of compound 65, which exhibited potent broad-spectrum antibacterial activity against MDR Gram-negative bacteria without detectable hERG liability. Moreover, 65 demonstrated improved pharmacokinetic properties, leading to enhanced in vivo efficacy compared to levofloxacin in a murine infection model.