The knockdown of endogenous replication factor C4 decreases the growth and enhances the chemosensitivity of hepatocellular carcinoma cells.
OBJECTIVE
To identify differentially expressed genes and thereby detect potential molecular targets for future therapies directed against hepatocellular carcinoma (HCC).


METHODS
To isolate differentially expressed genes between HCC and adjacent non-cancerous liver tissues, cDNA microarray and quantitative reverse transcriptase polymerase chain reaction analyses were performed. Gene knockdown experiments in HepG2 cells were also performed using small interfering RNAs (siRNAs). Proteins were detected by immunostaining, and cell proliferation was analysed using the MTT/WST-8 assay. Apoptosis and cell cycle analyses were performed using flow cytometry.


RESULTS
After an intensive screening for differentially expressed genes in HCC tissues, we isolated 23 upregulated genes in these lesions. Among these, we focused on the replication factor C4 (RFC4) gene. The expression of endogenous RFC4 proteins in HepG2 cells was found to be significantly reduced by RFC4-specific siRNA. This inhibition of RFC4 expression correlated with a decrease in cellular proliferation, increased levels of apoptosis and a sensitizing of the cells to the DNA-damaging chemotherapeutic agents, doxorubicin and camptothecin.


CONCLUSIONS
The replication factor C4 gene may be a novel target for developing cancer therapeutics, which can enhance the antitumour activity of chemotherapeutic agents that induce DNA damage.
Arai
Masaaki
M
Department of Biochemistry, National Defense Medical College, Saitama, Japan.
Kondoh
Nobuo
N
Imazeki
Nobuo
N
Hada
Akiyuki
A
Hatsuse
Kazuo
K
Matsubara
Osamu
O
Yamamoto
Mikio
M
D000328:Adult; D000368:Aged; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism*; D049109:Cell Proliferation; D017931:DNA Primers / Q000235:genetics; D005260:Female; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics*; D006801:Humans; D008297:Male; D008875:Middle Aged; D020411:Oligonucleotide Array Sequence Analysis; D034622:RNA Interference; D051818:Replication Protein C / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D051818:Replication Protein C