Paclitaxel induces programmed cell death in MDA-MB-468 human breast cancer cells.

The ability of paclitaxel, one of the most active chemotherapeutic agents against breast cancer, to induce programmed cell death in hormone-independent MDA-MB-468 human breast cancer cells was assessed. Treatment of MDA-MB-468 cells led to growth inhibition, high-molecular-weight and oligonucleosomal DNA fragmentation, and apoptosis-associated morphological changes after either 3- or 24-h exposure to paclitaxel concentrations >/=10 nM. Additionally, cleavage products of poly(ADP-ribose) polymerase and lamin B1, two proteins that are cleaved early in the execution phase of programmed cell death, were detected. Quantitative studies indicated that exposure to paclitaxel for 24 h resulted in more DNA fragmentation than did 3-h exposure. Rapid induction of the early-response gene c-jun but not c-myc was associated with paclitaxel treatment. The ability of paclitaxel to induce high-molecular-weight DNA fragmentation and apoptosis-associated morphological changes in three other breast cancer cell lines was also established. These data suggest that paclitaxel, an agent known to stabilize microtubules and prevent cell division but not to act directly on DNA, induces programmed cell death in breast cancer cells.

Growth inhibition of hormone-responsive and -resistant human breast cancer cells in culture by N1, N12-bis(ethyl)spermine.

Previous studies have documented differential sensitivity of human lung cancer and melanoma cell lines to the cytotoxic effects of N1, N12-bis(ethyl)spermine (BESpm). We show here that BESpm can significantly inhibit the growth of six human breast cancer cell lines with 50% inhibitory concentration in the microM range. The degree of inhibition does not correlate with estrogen receptor status. Detailed studies with estrogen receptor-positive MCF-7 and estrogen receptor- negative Hs578t cells show a similar dose-response curve with concentrations of 1-10 microM resulting in maximal growth inhibition. Growth inhibition in both lines is associated with an 8-12-fold induction of the polyamine catabolic enzyme, spermidine/spermine N1-acetyltransferase, and a progressive decrease in intracellular polyamine levels over 6 days even though steady-state levels of BESpm are achieved within 24 h. Similar studies on WTMCF7 and AdrRMCF7 cells show that the acquisition of resistance to hormonal or doxorubicin therapy is not associated with resistance to the growth-inhibitory effects of BESpm. These results suggest that BESpm exerts similar growth-inhibitory effects against both hormone-responsive and -unresponsive human breast cancer cells, a finding which has significance for the potential use of polyamine analogues in treating human breast cancer.

Induction of jun gene family members by transforming growth factor alpha but not 17 beta-estradiol in human breast cancer cells.

To investigate whether estrogen treatment of hormone-responsive human breast cancer cells was associated with activation of members of the jun family of immediate early response genes, the expression of these oncogenes in human breast cancer cells was examined. 17 beta-Estradiol had little effect on expression of c-jun, jun B, jun D, or c-fos mRNA by MCF-7 cells over 12 h, although it stimulated c-myc expression 4-fold within 30 min. In contrast, several peptide growth factors, including transforming growth factor-alpha (TGF-alpha), rapidly and transiently induced expression of c-jun, jun B, and c-fos mRNA 4- to 10-fold over control. A similar pattern of expression was seen in two other estrogen-responsive human breast cancer cell lines, ZR-75-1 and T47D. Inhibition of protein synthesis by cycloheximide did not abrogate induction of c-jun or jun B mRNA by TGF-alpha in MCF-7 cells, suggesting that new protein synthesis was not required. In addition, nuclear runoff transcription analysis demonstrated that increased expression of c-jun and jun B mRNA after TGF-alpha treatment of MCF-7 cells was regulated at least in part at the transcriptional level. Chronic exposure of MCF-7 cells to 17 beta-estradiol for 24-48 h was associated with decreased expression of jun B mRNA only, while similar treatment with TGF-alpha did not change mRNA expression of any jun family member. Thus, expression of jun family members is induced by peptide growth factors like TGF-alpha but not 17 beta-estradiol in human breast cancer cells. These results suggest that these nuclear protooncogenes play different roles in modulating gene expression by MCF-7 cells after exposure to TGF-alpha or 17 beta-estradiol.

Differential effects of bryostatin 1 and phorbol ester on human breast cancer cell lines.

The effects of the protein kinase C (PKC) activators, phorbol ester 12-O-tetradecanoyl-13-phorbol acetate (TPA) and the marine natural product, bryostatin 1, on the growth and morphology of human breast cancer cell lines were examined. TPA (1 to 100 nM) inhibited growth of four of six cell lines by up to 75% in 5-day cultures. Bryostatin 1 inhibited growth of only MCF-7 cells and only at a high dose (100 nM). However, bryostatin 1 completely antagonized the growth inhibition and morphological changes induced by TPA in MCF-7 cells. The divergent effects of these two agents are associated with differing effects on PKC activity and isoform expression in MCF-7 cells. TPA induced rapid translocation of the PKC-alpha isozyme and PKC activity to the membrane fraction of MCF-7 cells. In contrast, bryostatin 1 treatment resulted in the loss of the PKC-alpha isozyme and PKC activity from both cytosolic and membrane compartments within 10 min of treatment. In coincubation assays the bryostatin 1 effect was dominant over that of TPA. Similar effects on PKC-alpha isozyme and PKC activity were seen in a second cell line whose growth was inhibited by TPA but not by bryostatin 1, MDA-MB-468. In contrast, in the T47D cell line, where TPA was not growth inhibitory, TPA failed to induce translocation of PKC-alpha to the cell membrane. Bryostatin, however, still caused loss of PKC-alpha isozyme and PKC activity from cytosolic and membrane fractions. Thus, differential actions of bryostatin 1 and TPA on PKC activity and alpha-isoform level in the membrane-associated fraction of MCF-7 and MDA-MB-468 cells may account for the divergent effects of these two agents on cell growth and morphology. These results suggest that the PKC-alpha isoform may specifically play a role in inhibiting growth of human breast cancer cells.