Down‐regulation of riα subunit of camp‐dependent protein kinase induces growth inhibition of human mammary epithelial cells transformed by c‐ha‐ ras and c‐ erb b‐2 proto‐oncogenes

MCF‐ 10A is a spontaneously immortalized, non‐transformed human mammary epithelial cell line. We have recently obtained MCF‐ 10A clones (MCF‐ 1OA HE cells) that are transformed following over‐expression of both a human point‐mutated c‐Ha‐ ras and the c‐ erb B‐2 proto‐oncogenes. Two isoforms of the cAMP‐dependent protein kinase (cAK) have been described in mammalian cells. Enhanced levels of type‐I cAK (cAKI) are generally found in tumor cells. To determine whether inhibition of cAKl expression may interfere with ras and erb B‐2 oncogene‐induced transformation of human mammary epithelial cells, we have tested the effects of 2 agents that specifically down‐regulate cAKI, such as 8‐chloro‐cAMP and an anti‐sense oligodeoxynucleotide targeted against the R1α regulatory subunit of cAKl on MCF‐10A HE cells. Treatment of MCF‐10A HE cells with 8‐chloro‐cAMP induces a dose‐dependent growth inhibition under both monolayer and soft‐agar growth conditions, that is correlated with an accumulation of MCF‐10A HE cells in G 0 /G, phases of the cell cycle and a reduction of the number of cells in S phase. In contrast, 8‐chloro‐cAMP has no effect on MCF‐10A cell growth. Furthermore, 8‐chloro‐cAMP treatment of MCF‐10A HE cells induces a 4‐ to 6‐fold reduction in p185erbB‐2 expression and brings p21 ras expression to levels comparable to those found in MCF‐10A cells. Treatment of MCF‐10A HE cells with an Rlα anti‐sense oligodeoxynucleotide determines a comparable inhibition of both anchorage‐dependent and anchorage‐independent cell growth. Our results suggest that cAKl may act as a mediator of ras and erb B‐2 oncogene action in human breast cells and that interference with cAKl action provides a potential tool for inhibiting the growth‐promoting effects of these oncogenes.