Fig. 7.

WQQ-345 was identified as a novel BCAT1 inhibitor with antitumor potency. a Chemical structure of WQQ-345. b Enzyme-inhibition effects of WQQ-345 on recombinant human BCAT1 protein. c Binding model of WQQ-345 with BCAT1. Left panel shows WQQ-345 (magenta sticks) bound at the active site of BCAT1. The cofactor PLP was depicted in orange. Right panel shows surface representation of BCAT1 (gray surface) with WQQ-345 (magenta sticks). d Colony formation assays were performed to examine the growth of 67R cells treated with gabapentin or WQQ-345 (n = 5). e Quantification of relative colony formation of 67R cells treated with vehicle control, ASK120067, WQQ-345 and drug combination (n = 4). f Quantification of α-KG levels in 67R cells treated with vehicle control, gabapentin or WQQ-345 for 3 days (n = 4). g Western blot analysis of the expression of BCAT1 and H3K27me3 in 67R cells upon WQQ-345 treatment for 3 days. Data are shown as representative images (left) and a quantitative graph (right) (n = 4). h ECAR analysis of 67R cells with or without WQQ-345 treatment for 3 days. i Western blot analysis of the expression of indicated glycolytic enzymes in 67R cells upon WQQ-345 treatment for 3 days. The results are shown as representative images (left) and a quantitative graph (right) (n = 4). j The in vivo antitumor activity of WQQ-345 was evaluated in a 67R xenograft tumor model. 67R tumor-bearing mice were given oral treatments of PBS control or WQQ-345 twice daily for 43 days, and tumor volume was monitored. k The expression of BCAT1, H3K27me3 and indicated glycolysis related enzymes in 67R xenograft tumors at the endpoint of drug treatment was assessed by Western blot assay and shown as representative images (left) and a quantitative graph (right) (n = 6). *p < 0.05, **p < 0.01, ***p < 0.001; ns, not significant. Data in (j) are expressed as the mean ± SEM, and other data are expressed as the mean ± SD