Fig 2: STEAP1 expression is upregulated and associated with poor survival in patients with liver cancer. Publicly accessible gene expression profiling datasets, (A) GSE14520 and (B) GSE36376, were obtained from the Gene Expression Omnibus and analyzed to evaluate STEAP1 expression in patients with liver cancer. Associations between STEAP1 expression and (C) OS and (D) RFS in GSE14520, and (E) OS in TCGA were evaluated using the Kaplan-Meier method. STEAP1, six-transmembrane epithelial antigen of the prostate 1; OS, overall survival; RFS, recurrence-free survival; TCGA, The Cancer Genome Atlas.

Fig 3: Inhibition of STEAP1 suppresses c-Myc and its downstream target genes in liver cancer cell lines. Two different liver cancer cell lines (HepG2 and Hep3B) were transfected with non-targeting control siRNA and siSTEAP1. c-Myc expression was evaluated by RT-qPCR and western blotting. RT-qPCR in (A) HepG2 and (B) Hep3B cells. Western blotting in (C) HepG2 and (D) Hep3B cells. Data of RT-qPCR experiments are shown as the mean of quadruplicate measurements ± SD. **P<0.01 vs. control. (E) Scatterplot representation of the c-Myc targeted genes validated by PCR array in HepG2 cells. siRNA, small interfering RNA; STEAP1, six-transmembrane epithelial antigen of the prostate 1; RT-qPCR, reverse transcription-quantitative PCR.

Fig 4: Knockdown of STEAP1 leads to inhibition of cell proliferation in different liver cancer cell lines. Two different liver cancer cell lines, HepG2 and Hep3B, were transfected with non-targeting control siRNA and two independent siRNAs targeting (siSTEAP1_1 and siSTEAP1_2). Efficiency of knockdown was evaluated using RT-qPCR and western blotting. RT-qPCR in (A) HepG2 and (B) Hep3B cells. Western blotting in (C) HepG2 and (D) Hep3B. Data of RT-qPCR experiments are shown as the mean of quadruplicate measurements ± SD. Cell proliferation of two different liver cancer cell lines, (E) HepG2 and (F) Hep3B, was evaluated using a WST-1 assay at 0, 24, 48 and 96 h after siRNA transfection. Data are presented as the mean of triplicate measurements ± SD. *P<0.05 and **P<0.01 vs. control. RT-qPCR, reverse transcription-quantitative PCR; siRNA, small interfering RNA; STEAP1, six-transmembrane epithelial antigen of the prostate 1.

Fig 5: STEAP1 silencing triggers G1 arrest in liver cancer cell lines. Two different liver cancer cell lines, (A) HepG2 and (B) Hep3B, were transfected with non-targeting control siRNA or siSTEAP1. Cells were stained with propidium iodide 72 h after siRNA transfection. Subsequently, cell cycles were analyzed by flow cytometry. The percentage of G0/1 cells transfected with siSTEAP1 was compared to that of cells transfected with non-targeting control siRNA. Data are presented as the mean of triplicate measurements ± SD. **P<0.01. (C) Two different liver cancer cell lines, HepG2 and Hep3B, were transfected with non-targeting control siRNA or siSTEAP1. Cell cycle-associated proteins were analyzed by western blotting. siRNA, small interfering RNA; STEAP1, six-transmembrane epithelial antigen of the prostate 1.