Fig. 7: HIFα negatively regulates FKBP10, and inhibition of FKBP10 enhances the antitumor effects of PT2385.
A HK2, Caki1, ACHN and RCC4 cells were exposed to 21% O2 or 1% O2 for 24 h. B RCC4 cells were treated with DMOG (1 mM) or MG132 (10 μM) for 24 h. C Western blot analysis of the indicated proteins in 786 O cells overexpressing VHL. D, E Western blot analysis of the indicated proteins in 786 O cells with HIF2α knockdown or HIF1α overexpression. F The mRNA levels of FKBP10 in 786 O cells with HIF2α knockdown or HIF1α overexpression were examined by qRT‒PCR. G HIFα binding sites (HBSs) were obtained from the JADPAR promoter database within the human FKBP10 promoter region. CUT&RUN assays and qRT‒PCR were performed with primers containing candidate HBS sites in the FKBP10 promoter. H Western blot analysis of the indicated proteins in 786O-NC or 786O-shFKBP10 cells treated with PT2385 (20 µM) or PBS. I Cell proliferation abilities were measured by colony formation assays. Cell invasion abilities were measured by Transwell assays. Scale bar: 10 µm. Data are presented as the means ± SD and were independently replicated three times. J Clinical information for patient #1 from the patient’s cancer tissues. K Tumors from the shControl, PT2385+shControl, shFKBP10+PBS and PT2385+shFKBP10 groups are shown. L, M Tumor growth curves and tumor weight of each group. N Representative HE and IHC staining of the indicated proteins in tumors. O Schematic diagram to illustrating that FKBP10 mediated by HIFα promotes ccRCC progression via LDHA phosphorylation. Statistical analyses were performed with a two-tailed unpaired Student’s t test (* P < 0.05, **P < 0.01, ***P < 0.001).
