ALTERED CELL-CYCLE DISTRIBUTION AND CYCLIN-CDK PROTEIN EXPRESSION IN A431 EPIDERMOID CARCINOMA-CELLS TREATED WITH DOXORUBICIN AND A CHIMERIC MONOCLONAL-ANTIBODY TO THE EPIDERMAL GROWTH-FACTOR RECEPTOR

C225, a chimeric monoclonal antibody recognizing the epidermal growth factor receptor (EGFR), can eliminate established A431 (human epidermo id carcinoma) xenografts in nude mice alone or in combination with DNA -damaging drugs such as doxorubicin. This report examines the in vitro effects of combining doxorubicin with C225 on cell cycle distribution and the temporal expression of several important cellular and cell cy cle-associated proteins, including the EGFR, p53, pRb, cyclins D1 and B, Cdk4, and Cdc2. Doxorubicin induced a G2/M arrest in A431 cells 24 h following treatment with the drug. At 72 h, the cell cycle distribut ion for cells treated with doxorubicin plus C225 showed an increase in a hypodiploid (<2n) peak that was suggestive of apoptosis. Morphologi cal changes for cells in the combination group included enlarged cell mass, extensive vacuolization, and the presence of multiple micronucle i. The combination regimen was also found to inhibit cell proliferatio n to a greater extent than seen for treatment with the individual agen ts. By 72 h, expression levels of p53, pRb, and EGFR were lower in cel ls treated with doxorubicin plus C225 and receptor activation (defined by phosphorylation) was substantially decreased in the combination gr oup. Down-regulation of the protein expression levels of cyclin D, Cdk 4, and Cdc2 was also observed in cells treated with doxorubicin plus C 225. In the case of Cdc2, the relative expression of the higher molecu lar weight (hyperphosphorylated) or inactive form of this molecule was increased, whereas the opposite was observed in both the control and single agent groups. These data suggest a putative model for the thera peutic effects of doxorubicin and C225 on the growth of solid tumors o bserved in preclinical animal models.