Bacterial type II topoisomerases, which include DNA gyrase and its paralogous form topoisomerase IV (topoIV), are well-established and extensively studied targets in antibacterial chemotherapy. This doctoral dissertation focuses on the structural optimization of bacterial topoisomerases type II inhibitors (Novel Bacterial Topoisomerase Inhibitors; NBTI). The investigative approach includes in silico methodologies, synthetic organic chemistry for compound synthesis and a diverse array of in vitro and in vivo assays for comprehensive biological evaluations. The biological evaluation included assessment of inhibitory potency against both bacterial and human topoisomerases, antibacterial activity against a broad spectrum of bacteria and inhibition of hERG (human Ether-à-go-go-Related Gene) potassium channels. In addition, the study was extended to the determination of cardiotoxicity in zebrafish embryos (lat. Danio rerio) and in vivo efficacy in laboratory mice infected with the MRSA (Methicillin-Resistant Staphylococcus aureus) strain. Through an optimization process, a novel series of inhibitors characterized by increased selective toxicity was developed.
In the first phase, advanced predictive QSAR (Quantitative Structure-Activity Relationship) models were constructed using literature data on known NBTI inhibitors with experimentally determined in vitro inhibitory activity on recombinant enzyme. Subsequently, these models were used to predict the binding properties of newly designed virtual compounds to DNA gyrase. Selected virtual hits were synthesized and biologically evaluated and optimized compounds with inhibitory activities in the medium and low nanomolar range were found. Moreover, these compounds showed potent antibacterial activity against a broad spectrum of Gram-positive and Gram-negative bacteria, including challenging ESKAPE bacteria. An assessment of the inhibition of the hERG potassium channel also showed that some compounds exhibited weak hERG inhibition (IC50 hERG >30). Notably, certain compounds exhibited little or no toxicity in zebrafish embryos, alongside efficacy in vivo in laboratory mice infected with the MRSA strain.
The newly optimized NBTI analogs, which represent the highlight of this research, are an important contribution to the field of bacterial type II topoisomerase inhibitors. This represents progress towards the development of novel antibacterial agents. In addition, this work serves as a starting point for subsequent studies aimed at further refining and expanding the repertoire of antibacterial agents.
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