A NONANTISENSE SEQUENCE-SELECTIVE EFFECT OF A PHOSPHOROTHIOATE OLIGODEOXYNUCLEOTIDE DIRECTED AGAINST THE EPIDERMAL GROWTH-FACTOR RECEPTOR IN A431 CELLS

The overexpression of epidermal growth factor receptor (EGFr) has been implicated as a causative factor and a poor prognostic marker in a nu mber of carcinomas. Therefore, strategies that down-regulate EGFr expr ession may be therapeutically useful. We designed antisense ODNs compl ementary to the initiation codon region of the EGFr mRNA and evaluated their efficacy in several tumor-derived cells, including the A431 cel l line, that express amplified levels of EGFr. A 15-mer phosphorothioa te (PS) antisense ODN (erbB1AS15) induced a concentration-dependent re duction in proliferation that was accompanied by a change in the morph ology of A431 cells into more tightly clustered and discrete colonies. A 15-mer sense (PS) control oligodeoxynucleotide (ODN) and a phosphod iester (PO) version of erbB1AS15 had little or no effect on cell numbe r or morphology, and erbB1AS15 (PS) did not induce these effects in co ntrol cell lines expressing lower levels of EGFr. The effects of erbB1 AS15 (PS) on A431 cells were not mediated by a true antisense mechanis m in that there was no reduction in the level of EGFr mRNA or protein over a 24-hr period, as determined by Northern and Western blotting, r espectively. However, autophosphorylation of the receptor was signific antly reduced by erbB1AS15 (PS) and not by control ODNs. The results o f further studies suggested that this effect was mediated by a direct, dose-dependent inhibition of the EGFr tyrosine kinase enzyme and was not due to impairment of either ligand-binding or receptor dimerizatio n. These data suggest that erbB1AS15 (PS) can inhibit proliferation an d alter the morphology of A431 cells by a sequence-selective, but nona ntisense, mechanism affecting receptor tyrosine kinase activity.