Navigation into new medicinal application of vanadium-based agents as antagonist against senescence

Abstract

Vanadium plays critical roles in biological systems, and vanadium-based agents have been widely evaluated for the potential treatment of certain diseases, such as diabetes in the past decades. However, the wealth of biological chemistry of vanadium endows it with more potentials in medication other than acting as anti-diabetic agents. In this thesis, the roles of vanadyl-based agents as antagonist against senescence are investigated. The in vitro primary screening among several metallo-based compounds demonstrated that the vanadyl compound, VO-Ohpic (3-hydroxypicolinate vanadium(IV)), is able to postpone accelerated cellular senescence in Lamin AG609G/G609G mouse embryonic fibroblasts (MEFs) and to repress irradiation-induced cellular senescence in wildtype MEFs. VO-Ohpic treatment is found to modify the population doubling level and cell morphology, and to improve misshaped nuclear morphology, which might be associated to its capacity of stabilizing vimentin and maintaining its subcellular perinuclear distribution, downregulate several key biomarkers of cellular senescence including p53, p21, and p16 and the DNA damage marker of γ-H2AX, and alleviate premature aging in Lamin AG609G/G609G MEFs. VO-Ohpic is found to have a huge influence on diabetic cardiomyopathy, thermogenesis and multiple pathways of neurodegeneration diseases. VO-Ohpic treatment in Lamin AG609G/G609G mice ameliorates age-dependent weight loss and significantly extent their healthspan and lifespan, increasing the median healthspan by 12.81% and the median lifespan by 13.74%. The maximum healthspan and lifespan were increased by 25.21% and 21.56%, respectively. A new strategy was developed by integration of fluorescent imaging with high throughput proteomic analysis to mine endogenous VO2+-binding proteins at proteome-wide scale in live wildtype and Lamin AG609G/G609G MEFs using the vanadyl-chelated fluorescent probe, VO-TRACER. Intense blue fluorescence is observed within MEFs achieving vanadyl-associated proteins labeling. Seven putative protein targets of vanadyl ions in living Lamin AG609G/G609G MEFs and eleven protein targets in living wildtype MEFs are subsequently identified. These proteins are predominantly located at endoplasmic reticulum (ER). Binding of vanadyl ions to selected identified proteins is further confirmed by cellular thermal shift assay. The identified proteins positively correlate to ER stress, confirming that vanadyl might exert its effect at ER. VO-Ohpic is found to downregulate the GRP78, PDIA1 and CHOP protein levels with decrease in the cell ratio of apoptosis, an increase ER Ca2+ levels, alleviation of the overloaded mitochondrial calcium and the decreased mitochondrial membrane potential during senescence. VO-Ohpic increases the G2/M and S cell ratios whereas decreases the G1 cell ratios at the late passage of MEFs. Moreover, the effect of VO-Ohpic on delaying senescence via modulating ER function in Lamin AG609G/G609G MEFs is reconfirmed by azoramide. (415 words)