Application Of Omics In Antitumor Mechanistic Studies Of Acridone Derivative 8a

A new acridone derivative,2-aminoacetamido-10-(3,5-dimethoxy)-benzyl-9(10H)-acridone hydrochloride(named 8a)synthesized in our lab shows potent antitumor activity on CCRF-CEM leukemia cells,but the mechanism of action remains unclear.Herein,we report the use of an ‘omic’ approaches to study the effects of a series of acridone derivatives on cells and to shed new light on the probable antitumor mechanism of 8a.Firstly,metabolomics results showed that 23 distinct metabolites involved in five metabolic pathways were identified in 8a-treated cells compared to control.In glutathione(GSH)metabolism,GSH level and the reduced/oxidized glutathione(GSH/GSSG)ratio were significantly decreased in 8a-treated cells,while L-cysteinyl-glycine(L-Cys-Gly)and glutamate were greatly increased.In glycerophospholipid metabolism,cell membrane components phosphatidylcholines(PCs)were decreased in 8a-treated cells,while the oxidative products lysophosphatidylcholines(LPCs)were significantly increased.We further found that in 8a-treated cells,the reactive oxygen species(ROS)and lipid peroxidation product malondialdehyde(MDA)were notably increased,accompanied with decrease of mitochondrial transmembrane potential,release of cytochrome C and activation of caspase-3.Secondly,proteomics results showed that multiple cellular pathways were affected by 8a,including chromatin organization,energy metabolism,DNA repair,oxidative-stress,and apoptosis.The changes in protein expression were further verified for PKM2.Moreover,8a lowered down the expression of HEX and PFK-1.Lactate production was decreased in 8a-treated cells,indicating suppression of glycolysis.The elevated XRCC6 and decreased histone expression levels suggested increased DNA damage in 8a-treated cells,which was confirmed by the increased γ-H2 AX foci.Molecular docking of 8a with DNA demonstrated direct interactions of 8a with DNA through three hydrogen bonds and four π–π interactions,potentially explaining the mode of action that 8a damaged to DNA.Thirdly,WCL glycomics yielded 135 N-linked compounds with 83 distinct glycan compositions,among which 7 high mannose type N-glycans showed significant decrease after treatment with 8a.In comparison,CM yielded 360 compounds with 203 distinct compositions.Of these,42 N-glycans,including 12 high mannoses,4 undecorated complex/hybrid,9 fucosylated complex/hybrid,and 9 sialylated complex/hybrid,showed statistically significant changes in 8a-treated cells compared to control.These suggested that CM might be a more sensitive indicator of changes in glycosylation after treatment with acridone derivatives.All these glycosylation changes were demonstrated to be associated with the decreased expression level of DDOST in 8a-treated cells.Taken together our results suggest that the acridone derivative 8a might trigger cell death via multiple mechanisms,including oxidative stress-mediated apoptosis,energy metabolism dysfunction,DNA damage et al.The integrated omic approaches does not only shed novel insights into the pharmacological mechanistic studies of acridone derivative 8a,but also facilitate the further optimization of acridone derivatives 8a.