Targeting the Transcriptional Repressor LexA: Drug Repurposing, Synthesis and in vitro evaluation of Benzoxaboroles modulating SOS response in bacteria.

Antibiotic resistance (AMR) represents one of the most critical global health challenges, largely driven by the misuse and overuse of antibiotics, which promotes the emergence of resistant strains and undermines therapeutic efficacy. Pathogenic bacteria, in fact, can develop, through several genetic mechanisms, resistance against available drugs, thereby drastically reducing their effectiveness. Among bacteria resistance mechanism, the SOS response plays a crucial role by enabling DNA repair and promoting mutagenesis, ultimately facilitating resistance development. The key proteins in this mechanism are RecA, the sensor protein that under DNA damage activates the bacteria response through the allosteric modulation of , LexA, the effector protein, a transcriptional regulator of more than 60 genes involved in several mechanism of resistance to antimicrobial as the key regulator, LexA represents a strategic bacterial target whose inhibition could limit the onset of resistance. In this context, my thesis focused on the drug repurposing of boron containing approved drugs as inhibitors of the LexA protein. Boronic acids are excellent inhibitors of serine proteases and, by structural analogy, of LexA, as already adequately demonstrated. Therefore, the benzoxabole scaffold, was identified among 21 approved boron containing drugs for their ability to inhibit LexA in vitro and to interfere with bacterial filamentation and biofilm formation in E. coli, two processes associated with SOS response activation. First of all, docking studies were conducted to rationalize the binding orientation of the selected benzoxaborole within LexA binding site and to direct hit-to-lead optimization. Subsequentely synthetic strategies were designed and optimized to obtain a library of functionalized benzoxaborole derivatives, variously decorated. The experimental work involved the development and optimization of several reaction steps, evaluating the yield, selectivity, and stability of both intermediate and final compounds. The obtained compounds have been now directed to in vitro and in biological assays to evaluate their affinity vs LexA and their ability to modulate the SOS system. In this context, the molecules developed in this synthetic project could serve as a starting point for future investigations, potentially opening new avenues towards the development of agents able to prevent the activation of mechanisms of resistance thus prolonging the activity of available antibiotics.