Cyclin D-1 antisense RNA destabilizes pRb and retards lung cancer cell growth

To investigate the role of cyclin D-1 in the regulation of lung cancer cell growth, we created five stably transfected cell lines carrying a cyclin D1 antisense construct. The transfected cells exhibited a marked decrease in the rate of cell growth, in contrast to the original lines (A549 and NCI-H441). The expression of several cell cycle-regulating proteins, including cyclin A, the cyclindependent kinases (cdk) 2 and cdk4, in addition to cyclin D-1 itself, was markedly decreased. The expression of one cdk inhibitor, p21(WAF1/CIP1), increased in the A549-derived cell lines. A specific target of cyclin D-1 activity, the growth-suppressing product of the retinoblastoma gene, pRb, exhibited decreased expression and a decreased level of phosphorylation in the transfected cells. Decreased expression of pRb due to a significant increase in its turnover rate suggested that the stability of the protein may depend on phosphorylation by cyclin D-1-dependent cdk activity. In addition to the impact on pRb stability, decreased expression of cyclin D-1 induced susceptibility to cell death after withdrawal of exogenous growth factors in the antisense transfected cell lines, a response that was not observed in the original cancer cell lines. We conclude that abrogation of cyclin D-1 overexpression in lung cancer cells disrupts several key pathways that are required for uncontrolled cell growth and induces those that lead to cell death after growth factor deprivation. Therefore, we speculate that use of antisense cyclin D-1 expression in appropriate gene vectors could be a useful method for retarding lung cancer cell growth in accessible tumors such as those of the lung epithelium.