TOXICITY AND MITOCHONDRIAL DELIVERY OF FLAVONOL-BASED CO-RELEASING MOLECULES
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CONFERENCE: Research Week
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lecture / presentation description
Our research group previously developed a new class of photo-sensitive, flavonol-based carbon monoxide releasing molecules (CORMs) that are structurally tunable, exhibit low toxicity and exhibit both oxygen-dependent and independent CO release activity. This next-generation photo-sensitive CORM, called Flav-1, could be therapeutically useful if CO release can be targeted to mitochondria. We hypothesized that modification of the Flav-1 structure by addition of triphenylphosphonium (TPP) tail moieties of varying lengths (2 and 8 carbons) would facilitate targeting of this CORM, and thus CO release, to the mitochondria. Cytotoxicity of TPP-modified Flav-1 was assessed in human umbilical vein endothelial cells (HUVEC) using standard cell viability assays. Localization of TPP-modified Flav-1, which fluoresces when exposed to visible light, was determined using confocal microscopy; HUVECs were co-stained with Hoechst, CellMask Deep Red plasma membrane stain and MitoTracker Red for visualization of the nucleus, plasma membrane, and mitochondria, respectively. Modification of Flav-1 with either C2 or C8 TPP moieties did not increase cytotoxicity compared to the parent Flav-1 compound, though the C8 modification appeared slightly more cytotoxic than C2, perhaps reflecting greater bioavailability to the cell. Confocal microscopy of HUVEC cells treated with 100 uM Flav-1:C8 for 4 hr revealed co-localization of the CORM to the mitochondria. Exposure to laser light at 405 nm, 488 nm or both wavelengths together quenched the fluorescence of Flav-1:C2, indicating light-induced CO-release in A549 cells. Ongoing work aims to evaluate the impact of CO release from photo-triggered TPP-modified Flav-1 on the generation of reactive oxygen species via uncoupling mitochondrial respiration and subsequent effects on CO-triggered cell signaling pathways.