Combination of IFN-alpha and 5-fluorouracil induces apoptosis through IFN-alpha/beta receptor in human hepatocellular carcinoma cells.
OBJECTIVE
Several studies showed the effectiveness of combination therapy with IFN-alpha and 5-fluorouracil (5-FU) for advanced hepatocellular carcinoma. However, only little is known about the underlying mechanism of combination therapy. In the present study, we examined whether apoptosis through IFN-alpha/beta receptor (IFN-alpha/betaR) was associated with the effects of combination therapy.


METHODS
HuH7, PLC/PRF/5, HLE, and HLF were treated with IFN- (500 units/mL), 5-FU (0.5 microg/mL), or their combination for 10 days. In addition, IFN-alpha/betaR gene transfer with combination therapy was done.


RESULTS
Ten-day treatment by combination therapy resulted in >80% cell growth inhibition. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling analysis showed synergistic effects for combination therapy on PLC/PRF/5, HLE, and HLF. Concordant results were obtained with DNA fragmentation. Moreover, there was an evidence showing that changes in the expression of Bcl-2 family lead to apoptosis. On the other hand, the expression of IFN-alpha/betaR and up-regulation of alpha-phospho-signal transducer and activator of transcription 1, IFN regulatory factor-1 by combination therapy were observed in all cell lines. Furthermore, IFN-alpha/type 2 IFN receptor long form-transfected HuH7 cells treated with combination therapy showed strong DNA fragmentation compared with nontransfected or transfected with IFN-alpha- and 5-FU-treated HuH7.


CONCLUSIONS
Our results showed that combination of IFN-alpha plus 5-FU strongly induced cell growth inhibition of human hepatocellular carcinoma cells and indicated that one of the direct mechanisms of combination therapy may in part be attributable to alterations in induction of apoptosis through IFN-alpha/betaR.
Kondo
Motoi
M
Department of Surgery and Clinical Oncology, Graduate School of Medicine, Osaka University, 2-2 Yamada-oka E2, Suita, Osaka 565-0871, Japan.
Nagano
Hiroaki
H
Wada
Hiroshi
H
Damdinsuren
Bazarragchaa
B
Yamamoto
Hirofumi
H
Hiraoka
Nobuaki
N
Eguchi
Hidetoshi
H
Miyamoto
Atsushi
A
Yamamoto
Tameyoshi
T
Ota
Hideo
H
Nakamura
Masato
M
Marubashi
Shigeru
S
Dono
Keizo
K
Umeshita
Koji
K
Nakamori
Shoji
S
Sakon
Masato
M
Monden
Morito
M
D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000473:pathology / Q000503:physiopathology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D045304:Cytochromes c / Q000235:genetics; D053938:DNA Fragmentation / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D005472:Fluorouracil / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D050856:Interferon Regulatory Factor-1; D016898:Interferon-alpha / Q000494:pharmacology*; D008113:Liver Neoplasms / Q000235:genetics / Q000473:pathology / Q000503:physiopathology; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D017471:Receptors, Interferon / Q000235:genetics / Q000502:physiology*; D050794:STAT1 Transcription Factor; D013223:Statistics as Topic; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects / Q000235:genetics; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein
C494811:BCL2L1 protein, human; D004268:DNA-Binding Proteins; C494257:IRF1 protein, human; D050856:Interferon Regulatory Factor-1; D016898:Interferon-alpha; D010750:Phosphoproteins; D019253:Proto-Oncogene Proteins c-bcl-2; D017471:Receptors, Interferon; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D015534:Trans-Activators; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D045304:Cytochromes c; D005472:Fluorouracil