Design, Synthesis and Biological Evaluation of DNA Gyrase and Topoisomerase IV Inhibitors as Novel Antibacterials

The spread of antibiotic resistance is a great threat to medicine. We are in desperate need of new antibiotics to replace those for which resistance is widespread. The bacterial type two topoisomerases, DNA gyrase and topoisomerase IV, are well validated antimicrobial targets. The fluoroquinolone antibiotics target the DNA binding site of these enzymes, but no currently available antibiotics target the ATP binding site. It is hoped that simultaneously targeting the ATP binding sites of these two bacterial enzymes will slow the emergence of resistance.
This thesis describes the investigation of a series of pyridine 3-carboxamide DNA gyrase inhibitors, extending knowledge of the structure activity relationships of these compounds and making assessment of their inhibition of the topoisomerase IV enzyme in addition to their inhibition of DNA gyrase. Many of the inhibitors proved to be highly potent inhibitors of both DNA gyrase and topoisomerase IV with the most potent compound having IC50s of 24 nM and 87 nM against the DNA gyrase and topoisomerase IV enzymes respectively. These compounds also showed promising antibacterial activity with MICs as low as 1 μg/ml against S. aureus. Additionally, in vitro ADME profiling of key compounds from this series was carried out.
A subunit of the DNA gyrase protein was produced for use in crystallisation studies. Cocrystallisation of the protein with four inhibitor molecules was carried out and four crystal structures were solved revealing the binding poses of the pyridine 3-carboxamide compounds bound in the ATP site of GyrB.
Efforts were also made to identify a new series of DNA gyrase inhibitors. De novo design and virtual high throughput screening were utilised and a number of compounds were purchased and assessed for their ability to inhibit DNA gyrase. Two of the compounds showed activity in the enzyme assay. A scaffold hopping approach was also used to rationally design new series of potential inhibitors. Three compound series were designed and synthesis was carried out towards them. A series of quinazolinones proved to be the most synthetically tractable and a small library was produced and assessed for their biological activity. Several of the compounds were active against the DNA gyrase enzyme with IC50s as low as 1.1 μM.