Antimicrobial resistance is a human health issue that demands development of new antibiotics with unique mechanisms of action to combat antibiotic failure in the clinic. Methicillin-resistant Staphylococcus aureus (MRSA) has emerged as a particular resistant pathogen associated with high levels of incidence and mortality. Isolation and structural elucidation of the antibiotic natural products armeniaspirols A-C was first reported in 2012 by Sanofi. Armeniaspirol possess an unprecedented scaffold and novel multiple mechanisms of action. For these reasons the armeniaspirols were an ideal scaffold to investigate for the development of antibiotics effective against current resistant pathogens.
A series of focused derivatives were synthesized and evaluated for antibiotic activity against clinically relevant pathogens including methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococcus. Replacement of the N-methyl with N-hexyl and various N-benzyl substituents lead to a substantial increase in antibiotic activity and potency for inhibition of both ClpYQ and ClpXP, the intracellular targets of armeniaspirol.
Armeniaspirol is also known to disrupt the proton motive force (PMF) in bacteria, though initial work by the Brönstrup lab suggested this was via shuttling of protons across the membrane. With a library of analogues in hand from our previous study, we sought to characterize their disruption of the PMF. Using a voltage sensitive dye-based assay and checkerboard synergy-based assay we demonstrated that armeniaspirols disrupt the proton motive force by dissipating the electrical potential (ΔΨ) of the PMF, correcting the previous literature, which suggested they disrupted the transmembrane proton gradient (ΔpH).
Lastly, our efforts toward the total synthesis of armeniaspirol A using an oxidative chlorination transformation led to a constitutional isomer of armeniaspirol by an unexpected Lewis acid mediated rearrangement in the penultimate step. We characterized the scope of carbonyl derivatives that could undergo successful oxidation generating the key α,β-dichloro-α,β-unsaturated lactam of the armeniaspirol scaffold. Our work led to a mechanistic study demonstrating how simple ketones undergo decomposition via acylium ion formation whereas esters and amides are effectively oxidized to the desired product.
Overall, the research presented here lays the foundation for the future work to confirm safety, efficacy, and toxicity of the armeniaspirols and to synthesize new analogues with improved drug like characteristics. In addition, this thesis previews an evolution-based link between armeniaspirol and its biosynthetic precursor that suggests mid- to late-stage biosynthetic intermediates are likely to possess biological activity but via a different mechanism of action as compared to the parent natural products. Furthermore, this analysis suggests that the intermediate is likely to synergize with the natural product, increasing the fitness of the producing organism as the pathway evolves.
| Identifer | oai:union.ndltd.org:uottawa.ca/oai:ruor.uottawa.ca:10393/44943 |
| Date | 16 May 2023 |
| Creators | Darnowski, Michael |
| Contributors | Boddy, Christopher |
| Publisher | Université d'Ottawa / University of Ottawa |
| Source Sets | Université d’Ottawa |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | application/pdf |
Page generated in 0.0028 seconds