Transfection of the breast cancer cell line MDA-468 with antisense RNA to P21 CIP1 in order to investigate the mechanism of EGF-mediated G1 arrest in these cells

Paquin, Andre (2000) Transfection of the breast cancer cell line MDA-468 with antisense RNA to P21 CIP1 in order to investigate the mechanism of EGF-mediated G1 arrest in these cells. Masters thesis, Memorial University of Newfoundland.

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Abstract

The human breast cancer cell line MDA-468 is characterized by an overexpression of the epidermal growth factor (EGF) receptor. As a result of this overexpression, MDA-468 cells have the unique characteristic of undergoing arrest of cell cycle in G₁ when treated with pharmacological concentrations of EGF. While the ability of EGF to cause arrest of this cancer cell line is well documented, the effectors that mediate this arrest remain unknown. Since the tumour suppressor gene, p21Cipl, is a potent inhibitor of cyclin dependent kinases with a well-defined ability to confer G₁ arrest on tumour cells, it is proposed that p21Cipl may play a role in the EGF-mediated arrest of these cells. An antisense against p21cipI expression was transfected into MDA-468 cells to test this hypothesis. Single-clone cell lines were derived and examined for (i) an elimination of p21Cipl mRNA and protein, and (ii) a reduction of the ability of these cells to undergo G₁ arrest when treated with EGF. Three antisense-transfected clones demonstrated that antisense expression caused a reduction of p21Cipl with a significantly decreased capacity to undergo G₁ arrest In these cell lines an absence of p21Cipl reduced the ability of EGF-treatment to cause growth arrest This correlation may indicate that p21Cipl is an important player in the EGF-mediated growth arrest of MDA-468 cells.

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