Inhibition of DNA synthesis by downregulation of cyclin A but not Skp 2 overexpression in human hepatocellular carcinoma cells

Cyclin A is an S and G(2)/M phase regulatory protein and associates with Sk p 2 in many transformed cells. Our previous results showed that 12 (39%) an d 17 (55%) out of 31 hepatocellular carcinoma (HCC) patients exhibited high er protein expression levels of cyclin A and Skp 2, respectively, in their tumorous compared to non-tumorous tissues. In the present study, we used We stern blot analysis to show that 3 out of 6 HCC cell lines, HA59T, HA22T an d HCC36, exhibited overexpression of cyclin A, among which the HCC36 cell l ine also expressed a higher Skp 2 protein level. Moreover, we used the anti sense oligonucleotide phosphorothioates to down regulate the overexpression of cyclin A and Skp 2 proteins to determine whether or not these two prote ins are involved in the mitogenesis of HCC36 cells. After treatment with an tisense oligonucleotide phosphorothioates, the gene product of cyclin A or Skp 2 was suppressed dose-dependently as revealed by Western blot analyses. By [H-3]thymidine incorporation assay, we found that downregulation of cyc lin A but not Skp 2 overexpression could inhibit the DNA synthesis ability of HCC36 cells, suggesting that abnormal Skp 2 expression is not directly c orrelated with the HCC proliferation. (C) 1999 Elsevier Science Ireland Ltd . All rights reserved.