Effect of Zebularine on p16INK4a, p14ARF, p15INK4b, and DNA Methyltransferase 1 Gene Expression, Cell Growth Inhibition, and Apoptosis Induction in Human Hepatocellular Carcinoma PLC/PRF5 and Pancreatic Cancer PA-TU-8902 Cell Lines

Document Type: Research article

Authors

1 Research Center for Non-Communicable Diseases, Jahrom University of Medical Sciences, Jahrom, Iran.

2 Student of Research Committee, Jahrom University of Medical Sciences, Jahrom, Iran.

Abstract

Tumorigenesis must be understood as a summary of altered genetic and genomic changes
resulting in the inactivation of tumor suppressor genes (TSGs). One of the characterizations of
epigenetic alterations is DNA methylation. Epigenetic alteration of the p16INK4a, p14ARF,
p15INK4b
, and DNA methyltransferase 1 gene (DNMT1) expression occurs in hepatocellular
carcinoma (HCC) and pancreatic cancer frequently. DNA methyltransferase inhibitors (DNMTIs),
such as zebularine, play a signifcant effect on the demethylation and reactivation of TSGs. This
study aimed to investigate the effect of zebularine on p16INK4a, p14ARF, p15INK4b, and DNA
methyltransferase 1 gene expression, cell growth inhibition, and apoptosis induction in HCC
PLC/PRF5 and pancreatic cancer PA-TU-8902 cell lines. Both cell lines were cultured and treated
with zebularine at different times. The MTT assay, real-time quantitative reverse-transcription
polymerase chain reaction (qRT-PCR), and flow cytometry were used to determine cell viability,
gene expression, and apoptotic cells, respectively. The result indicated that zebularine inhibited
cell growth of both cell lines signifcantly as time- and dose-dependent manner (
P < 0.007). The
agent induced signifcant down-regulation of DNMT1 and up-regulation of p16INK4a, p14ARF,
p15INK4b (
P < 0.028). Besides, it had a signifcant apoptosis effect on both cell lines (P < 0.001).
This compound had a strong signifcant effect on PLC/PRF5 in comparison to PA-TU-8902 cells.
Concluding, zebularine inhibited PLC/PRF5 and PA-TU-8902 cell growth and induced apoptosis
in these cell lines. The most likely mechanism underlying the zebularine played its role involves
down-regulation of DNMT1 and up-regulation of p16INK4a, p14ARF, and p15INK4b genes.

Graphical Abstract

Effect of Zebularine on p16INK4a, p14ARF, p15INK4b, and DNA Methyltransferase 1 Gene Expression, Cell Growth Inhibition, and Apoptosis Induction in Human Hepatocellular Carcinoma PLC/PRF5 and Pancreatic Cancer PA-TU-8902 Cell Lines

Keywords