A screening system for anti-metastatic small-molecule compounds based on perinucleolar compartment prevalence in liver cancer cells / 中华肝脏病杂志

ABSTRACT

Objective@#To establish a screening system for anti-metastatic small-molecule compounds based on perinucleolar compartment (PNC) prevalence in liver cancer cells and to investigate its validity.@*Methods@#Polypyrimidine tract-binding protein (PTB) monoclonal antibody was used to measure the PNC prevalence in HepG2, HepG2M, and Huh7 cells, and wound healing assay and transwell assay were used to analyze the migration and invasion abilities of hepatoma cells. HepG2M cells were used as the model for the screening of anti-metastatic small-molecule compounds, and after the treatment with the compounds A1, A4, and E696, qPCR was used to measure the expression of metastasis-related miRNAs (miR-141 and miR-200c). A one-way analysis of variance was used for comparison of data between multiple groups.@*Results@#PTB immunofluorescence assay showed that HepG2M cells had the highest PNC prevalence, followed by Huh7 and HepG2 cells, and PNC prevalence was positively correlated with the metastasis and invasion abilities of hepatoma cells. The PNC prevalence of HepG2M cells was reduced to 22.88% ±4.61% by A1, 14.22% ± 3.05% by A4, and 26.12% ± 4.94% by E696. Wound healing assay showed that the 48-hour scratch ratio increased from 17.70% ± 3.34% to 64.50% ± 2.65%, 83.40% ± 5.10%, and 57.20% ± 3.06% (F = 171.1, P < 0.01), respectively. Transwell assay showed that the number of invasive cells was reduced from 264.33 ± 30.50 to 104.33 ± 13.50, 58.00 ± 11.00, and 111.33 ± 19.50 (F = 59.87, P < 0.01), respectively. The anti-metastatic effect of these three compounds was positively correlated with their ability to destroy PNC. A4 upregulated the expression of miR-141 and miR-200c in a dose-dependent manner, and after HepG2M cells were treated with A4 at a concentration of 5 μM, 10 μM, or 20 μM, the level of miR-141 was increased to 3.61 ± 0.78, 8.12 ± 1.15, and 18.24 ± 2.44 folds (F = 88.01, P < 0.01), respectively, and that of miR-200c was increased to 2.82 ± 0.43, 4.82 ± 0.89, and 10.74 ± 1.22 folds (F = 87.94, P < 0.01), respectively.@*Conclusion@#The screening system for anti-metastatic small-molecule compounds based on PNC prevalence can provide an effective technical platform for research and development of anti-metastatic drugs for liver cancer.