Discovery and development of novel anti-infective agents targeting DnaN and LasB: A dual approach against multidrug-resistant pathogens

Multidrug-resistant bacterial infections involving P. aeruginosa, and M. tuberculosis require innovative therapeutics. This thesis targets two bacterial proteins: DnaN (DNA polymerase β subunit) and LasB (P. aeruginosa virulence factor) combining direct inhibition and anti-virulence strategies. Dynamic combinatorial chemistry identified M. tuberculosis DnaN binder from acylhydrazone. Structure-activity studies delivered chemically stable heterocyclic analogues using 1,3,4-oxadiazole, 1,3,4-thiadiazole, and 1,2,4-triazole replacements. Compound AEL227 showed antimycobacterial activity (MIC = 4.74 – 9.48 μg/mL), while AEL263 exhibited no cytotoxicity at 100 μM. Benzimidazole-based phosphonate inhibitors targeted LasB. N-H donor functionality enhanced potency, with 8108 achieving sub-nanomolar activity (IC₅₀ = 0.49 nM). Co-crystallization confirmed active site hydrogen-bonding. Lead compounds demonstrated metabolic stability, minimal cytotoxicity, selectivity over human metalloproteinases, and favorable lung exposure. Cellular assays showed epithelial protection; Galleria mellonella studies demonstrated survival improvement. Additionally, fluorescence polarization and surface plasmon resonance assays for DnaN screening were established, plus an open-source plasma protein binding prediction tool was developed (R² = 0.83). This work establishes complementary antimicrobial resistance strategies: DnaN inhibitors as novel antimycobacterials and LasB inhibitors for anti-virulence therapy.