[
Antimicrob Agents Chemother,
2010]
Hybrid antimicrobials containing an antibacterial linked to a multidrug resistance (MDR) pump inhibitor make up a promising new class of agents for countering efflux-mediated bacterial drug resistance. This study explores the effects of varying the relative orientation of the antibacterial and efflux pump inhibitor components in three isomeric hybrids (SS14, SS14-M, and SS14-P) which link the antibacterial alkaloid and known substrate for the NorA MDR pump berberine to different positions on INF55 (5-nitro-2-phenylindole), an inhibitor of NorA. The MICs for all three hybrids against wild-type, NorA-knockout, and NorA-overexpressing Staphylococcus aureus cells were found to be similar (9.4 to 40.2 microM), indicating that these compounds are not effectively effluxed by NorA. The three hybrids were also found to have similar curing effects in a Caenorhabditis elegans live infection model. Each hybrid was shown to accumulate in S. aureus cells to a greater extent than either berberine or berberine in the presence of INF55, and the uptake kinetics of SS14 were found to differ from those of SS14-M and SS14-P. The effects on the uptake and efflux of the NorA substrate ethidium bromide into S. aureus cells in the presence or absence of the hybrids were used to confirm MDR inhibition by the hybrids. MDR-inhibitory activity was confirmed for SS14-M and SS14-P but not for SS14. Molecular dynamics simulations revealed that SS14 prefers to adopt a conformation that is not prevalent in either SS14-M or SS14-P, which may explain why some properties of SS14 diverge from those of its two isomers. In summary, subtle repositioning of the pump-blocking INF55 moiety in berberine-INF55 hybrids was found to have a minimal effect on their antibacterial activities but to significantly alter their effects on MDR pumps.
Lyang, Nora, Kelly, Jeffery, Xu, Jin, Hansen, Malene, Tan, Ee Phie, Nieto-Torres, Jose, Botham, Rachel, Yoon, Leonard, Zaretski, Svaitlana, Johnson, Kristen
[
International Worm Meeting,
2021]
Autophagy is an evolutionarily conserved cellular recycling process with tight links to longevity and healthspan. In particular, autophagy function declines during aging, and is dysregulated in many age-related disorders such as in neurodegenerative diseases. Therefore, identifying interventions that can boost autophagy to prevent such chronic illnesses progression is crucial to improving organismal health. Of note, autophagy is increasingly appreciated as a selective process by which specific types of cytosolic cargo, such as organelles, lipids and protein aggregates, are sequestered into double-membrane structures called autophagosomes that subsequently fuse with hydrolase-containing lysosomes to enable cargo degradation. Interestingly, accumulating evidence suggests that disruptions in selective autophagy can contribute to the development of age-related diseases. For example, chronic inhibition of lipophagy (selective lipid turnover) leads to increased accumulation of lipids, leading to obesity and diabetes. However, treatments that may target and improve selective autophagy to help relieve such illnesses remain underdeveloped. To identify novel chemical compounds that may act as selective autophagy activators, we performed a high-throughput imaging screen in human adenocarcinoma cells to uncover small molecules that activate autophagy and increase lipid clearance. Given the previous links between autophagy and aging, we tested several autophagy activator hit compounds from the screen for autophagy- and lifespan assays in C. elegans. While we found that these compounds all increased autophagosome numbers, only animals fed with small molecule A20 exhibited life- and healthspan extension, along with reduced lipid levels, as observed in human cells. Importantly, this A20-mediated lipid reduction and health benefits were not observed in autophagy mutants. Furthermore, we found that A20 could reduce PolyQ aggregate cargo load in multiple tissues, and we are currently investigating if A20 is affecting additional cytosolic cargos. Notably, inhibition of the nutrient sensor mTORC1 activates autophagy. However, A20 seemed to function independently of mTORC1, and we are currently performing studies to determine how A20 could mediate autophagy. In conclusion, we have identified a new compound A20, which may potentially be applied in future strategies to improve organismal health and alleviate age-related diseases by boosting autophagy.