The Biological Effects Of C-myc Antisense Oligodeoxynucleotide And 5-fluorouracil On Human Hepatocellular Carcinoma Cell Line HEPG-2

Malignant tumors have severly threatened the human health. At present when traditional methods shch as operation, chemotherapy, and radiotherapy have proved helpless to them, biological approaches have emerged on the atage. The gene therapy as one of them is one that obtains treating and preventing effect through some genes carried into the human body through some carrier. Liver cancer is one with most common incidence and higher motality. In China liver cancer occurs frequently with new incidence above ten thosend per year, taking the first and second position in malignancy-induced motality in the countryside and city respectively. In the world 38.16 millions patients died per year, in which China accounts for 45%. In China infection by HBV is a most related factor to liver cancer. With a low excision rate, low sensitivity to chemotherapy and radiotherapy, common trend to metastasis, and poor prognosis, liver cancer has an emergent need for new treatment methods. Recently along with the deepening researches on pathophysiologly of molecules of liver cancer, which stimulates the interest of treating it by molecule approach and will provide the theoretical evidence, a new concept of gene treatment combined with radiotherapy, chemotherapy and thermotherapy has been put forward.To investigate the biological effect of C-myc antisense oligo- deoxynucleotide and 5-fluorouracil on human hepatocellular carcinoma cell line HEPG-2 cells. HEPG-2 cells were cultured conventionally. Growth inhibition rate of HEPG-2 cells, after having been treated with C-myc ASODN and 5-fluorouracil, were detected by MTT method. Morphological change of cells were observed under a fluorescent microscope. DNA ladder was examined by DNA electrophoresis. The apoptotic rate and cell cycle phase distribution were examined by flow cytometer analysis. The expression of C-myc gene was detected by immunohistochemistry staining and RT-PCR on the mRNA or protein level respectively. The result of MTT assays showed that Both C-myc ASODN and 5-FU could inhibit the growth of HEPG-2 cells on dose and time dependent modes. Furthermore, The synergistic effect was marked between ASODN and 5-FU. Apoptotic phenomenon was certained by morphology, DNA electrophoresis and flow cytometer. C-myc ASODN could block cells cycle on G₀/G₁ phase by flow cytometer analysis. And, 5-FU could block cells cycle on S phase. We also found that both C-myc ASODN and 5-FU could downregulate the expression of C-myc. But The synergistic effect on the expression of C-myc was not marked. This finding indicated that other genes might be involved in the synergetic effect between C-myc ASODN and 5-FU on the anti-proliferation.In conclusion,â‘ C-myc ASODN could inhibit the growth, induce apoptosis and block cells cycle on G₀/G₁ phase of HEPG-2 cells.â‘¡5-FU could inhibit the growth, induce apoptosis and block cells cycle on S phase of HEPG-2 cells.â‘¢C-myc ASODN and 5-FU had significantly synergistic effect. C-myc ASODN could improve the sensitivity of HEPG-2 to 5-FU, enhance therapeutic effectiveness of 5-FU over HEPG-2 cells.â‘£Chemotherapy combined with antisensenucleic acids could reduce the dosage of those corresponding agents, suggesting that combined gene therapy does raise the effect of the chemotherapy while reduce the dosage, which subsequently reduces the side effects and drug-resistance of those agents used, and carries a promising hope to solve the problem with difficultly to thoroughly inhibit the tumor growth while with higher cost by using chemotherapy only.