Selective inhibition of the effects of phorbol ester on doxorubicin resistance and P-glycoprotein by the protein kinase C inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H7) in multidrug-resistant MCF-7/Dox human breast carcinoma cells.
The possible regulation of the multidrug-resistant (MDR) phenotype and P-glycoprotein by protein kinase C (PKC) was investigated in the doxorubicin (Dox)-resistant MCF-7 cell line (MCF-7/Dox). In a clonogenic assay, cells exposed to 100 nM phorbol 12-myristate 13-acetate (PMA) for 1 hr were about 3-fold more resistant to Dox than were cells exposed to Dox alone. The PKC inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H7, 30 microM) completely blocked the PMA-induced effect, but did not reverse the MDR phenotype. Complete down-regulation of PKC from MCF-7/Dox cells by 24-hr preincubation with PMA did not alter the degree of Dox resistance. Intracellular accumulation of [14C]Dox decreased from a baseline of 28 pmol/10(6) cells to 15 pmol/10(6) cells in the presence of 100 nM PMA. The reduced Dox accumulation in the presence of PMA was not blocked by pretreatment of cells with H7. Following a 24-hr pretreatment with PMA, the cells accumulated almost equal amounts of [14C]Dox in the absence or presence of PMA. Cells from PMA-treated colonies showed significantly higher levels of expression of P-glycoprotein when compared with those from control colonies. H7 did not affect the basal level of P-glycoprotein in cells from control colonies or PMA-induced overexpression of P-glycoprotein in cells from PMA-treated colonies. Upon stimulation with PMA (100 nM), PKC alpha and beta translocated to the cell membrane and nucleus and PKC delta and epsilon to the perinuclear membrane and the nucleus, respectively. H7 (30 microM) completely inhibited PMA-induced translocations of PKC delta and epsilon, whereas it only partially blocked the translocations of PKC alpha and beta. These results suggest that PMA appears to alter Dox resistance and intracellular Dox accumulation in a PKC-dependent manner and to induce increased expression of P-glycoprotein in MCF-7/Dox cells. Differential effects of H7 on the PMA-induced changes suggest that different isoforms of PKC may be involved in cell growth and drug accumulation processes as well as P-glycoprotein expression.
Ahn
C H
CH
Division of Oncology, Food and Drug Administration, Rockville, MD 20857, USA.
Kong
J Y
JY
Choi
W C
WC
Hwang
M S
MS
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000187:drug effects*; D001943:Breast Neoplasms / Q000378:metabolism*; D004317:Doxorubicin / Q000494:pharmacology; D004351:Drug Resistance; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D010703:Phorbol Esters / Q000494:pharmacology*; D010879:Piperazines / Q000494:pharmacology*; D011493:Protein Kinase C / Q000378:metabolism*; D013997:Time Factors
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D010703:Phorbol Esters; D010879:Piperazines; D004317:Doxorubicin; D011493:Protein Kinase C