Fig. 8

HDAC2 inhibitors enhance the sensitivity of ccRCC to sunitinib. A and B, 786-O and A498 cells were treated with DMSO or CAY10683 (5 µM) for 24 h. Cells were collected for western blot analysis (A) and RT-qPCR assay (B). Data presents as mean ± SD with three replicates. ***, P < 0.001. C and D, 786-O and A498 cells were transfected with indicated plasmids for 24 h. Then, these cells were treated with DMSO or CAY10683 (5 µM) for another 24 h. Cells were collected for western blot analysis (C) and RT-qPCR assay (D). Data presents as mean ± SD with three replicates. Ns, not significant; **, P < 0.01; ***, P < 0.001. E and F, 786-O and A498 cells were transfected with indicated siRNAs for 24 h. Then, these cells were treated with DMSO or CAY10683 (5 µM) for another 24 h. Cells were collected for western blot analysis (C) and RT-qPCR assay (D). Data presents as mean ± SD with three replicates. Ns, not significant; **, P < 0.01; ***, P < 0.001. G, A498 and 786-O cells were transfected with indicated shRNAs for 48 h. Then, these cells were treated with vehicle (DMSO) or CAY10683 (5 µM) for another 24 h. These cells were harvested and treated with a serial dose of sunitinib. These cells were subjected to CCK-8 assay. H, 786-O cells were transfected with indicated shRNAs for 48 h. Then, these cells were treated with vehicle (DMSO) or CAY10683 (5 µM) for another 24 h. These cells were subjected to CCK-8 assay. Data presents as mean ± SD with three replicates. Ns, not significant; **, P < 0.01; ***, P < 0.001. I and J, 786-O cells were transfected with indicated shRNAs for 72 h. After puromycin selection, cells were collected and subcutaneously injected into the nude mice. The tumor mass was shown in panel I, and tumor growth curve was shown in panel J. Data presents as mean ± SD with six replicates. Ns, not significant; *, P < 0.05; **, P < 0.01; ***, P < 0.001. K, a model depicting that the HDAC2/YY1 complex transcriptionally represses the expression of YTHDC1, which prevents the downregulation of ANXA1 induced by YTHDC1 and activates the MAPK pathway to decrease the sensitivity of ccRCC to sunitinib. HDAC2 inhibitors treatment blocks the HDAC2/YY1/YTHDC1/ANXA1/MAPK axis to enhance the anti-tumor effect of sunitinib