Protein kinase C delta is a prosurvival factor in human breast tumor cell lines.

Protein kinase C (PKC) promotes cell survival in response to ionizing radiation in a variety of experimental models including human carcinoma, human glioblastoma, and transformed mouse embryo fibroblast cell lines. We have introduced specific antisense oligonucleotides into human mammary tumor cell lines in vitro to analyze the role of individual PKC isoforms in radiation-induced cell death in breast cancer. MDA-MB-231 and MCF-7 cells treated with oligonucleotide directed against the PKC delta isoform exhibited impaired survival in response to 5.6 Gy gamma-radiation as measured by mitochondrial metabolism of tetrazolium dye. The role of PKC delta in the breast tumor cell lines was of particular interest, because contradictory reports exist in the literature regarding the role of PKC delta in cell survival and apoptosis. A comparison of the effects of the PKC delta antisense oligonucleotide and a nucleotide scrambled version of this nucleotide revealed only the antisense oligonucleotide decreased cell survival. The PKC delta antisense oligonucleotide decreased cell survival after exposure to low (1.5 Gy) radiation doses and in the absence of radiation insult. We found 3 micro M rottlerin, a selective PKC delta inhibitor, to reduce MCF-7 and MDA-MB-231 cell survival. Furthermore, MCF-7 cells transformed to express a dominant-negative mutant of PKC delta exhibited reduced survival. Comet analysis showed that PKC delta oligonucleotide treatment caused an accumulation of cells containing damaged DNA similar to that seen in 1.5 Gy radiation-treated cells. We conclude that PKC delta acts as a prosurvival factor in human breast tumor cells in vitro.

Control of mammary tumor cell growth in vitro by novel cell differentiation and apoptosis agents.

The use of breast tumor differentiating agents to complement existing therapies has the potential to improve breast cancer treatment. Previously we showed quinidine caused MCF-7 cells to synchronously arrest in G1 phase of the cell cycle, transition into G0 and undergo progressive differentiation. After 72-96 h cells became visibly apoptotic. Using several analogs of quinidine we determined that MCF-7 cell cycle exit and differentiation are typical of quinoline antimalarial drugs bearing a tertiary amine side chain (chloroquine, quinine, quinidine). Differentiated cells accumulated lipid droplets and mammary fat globule membrane protein. Apoptosis was assayed by a nucleosome release ELISA. Quinidine and chloroquine triggered apoptosis, but not quinine, a quinidine stereoisomer that displayed weak DNA binding. The apoptotic response to quinidine and chloroquine was p53-dependent. A 4-15-fold induction of p21(WAF1) protein was observed in cells treated with quinidine or chloroquine prior to apoptosis, but p21(WAF1) was not increased in cells that differentiated in response to quinine. Chloroquine was most active in stimulating MCF-7 apoptosis, and quinine was most active in promoting MCF-7 cell differentiation. We conclude, distinct mechanisms are responsible for breast tumor cell differentiation and activation of apoptosis by quinoline antimalarials. Alkylamino-substituted quinoline ring compounds represented by quinidine, quinine, and chloroquine will be useful model compounds in the search for more active breast tumor differentiating agents.