In vitro RNA interference against beta-catenin inhibits the proliferation of pediatric hepatic tumors.
Mutations of beta-catenin have been identified in the majority of pediatric hepatic malignancies, including hepatoblastoma (HB) and hepatocellular carcinoma (HCC), suggesting its important contribution in hepatic tumorigenesis in this age group. However, the role of beta-catenin/canonical Wnt signaling pathway in the neoplastic growth of cancer cells has not been directly studied. To address beta-catenin's capability in maintaining the malignant phenotype in established pediatric HB and HCC cell lines, HuH-6 and HepG2, harboring mutated and overexpressed beta-catenin, we carried out a series of in vitro analyses through a transfection of short interfering RNAs (siRNAs) to generate a loss-of-function model. HuH-7, another HB cell line derived from a pediatric patient without a stabilizing mutation was used for comparison. RNA interference successfully manipulated the degradation of overexpressed beta-catenin. In all cell lines, beta-catenin mRNA was suppressed by 80-90% after 48 h of transfection, and a reduction of its protein expression was demonstrated. In HuH-6 and HepG2, the pre-existing beta-catenin nuclear accumulation disappeared and reductions of beta-catenin downstream target genes, c-myc and cyclinD1, were also evidenced after the treatment. The in vitro proliferation of both cell lines was transiently inhibited. In contrast, the suppression of beta-catenin in HuH-7 did not lead to a significant change in the expression of target genes or cellular proliferation. Our data indicate that beta-catenin can be considered a specific target for gene therapy in pediatric hepatic tumors with mutations and overexpression of this gene.
Sangkhathat
Surasak
S
Department of Pediatric Surgery, Graduate School of Medicine, Osaka University, Osaka, Japan.
Kusafuka
Takeshi
T
Miao
Jiangyong
J
Yoneda
Akihiro
A
Nara
Keigo
K
Yamamoto
Seiji
S
Kaneda
Yasafumi
Y
Fukuzawa
Masahiro
M
D017209:Apoptosis; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation; D002648:Child; D004252:DNA Mutational Analysis; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics / Q000473:pathology / Q000503:physiopathology; D009154:Mutation; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D014162:Transfection; D051176:beta Catenin / Q000235:genetics* / Q000378:metabolism
D034741:RNA, Small Interfering; D051176:beta Catenin