No abstract

Peroxisome proliferator-activated receptor γ (PPARγ) is an important transcription factor that modulates lipid metabolism and inflammation. However, it remains unclear whether PPARγ is involved in modulation of estrogen (E)-induced inflammation, thus affecting apoptosis of E-deprived breast cancer cells, MCF-7:5C and MCF-7:2A. Here, we demonstrated that E treatment suppressed the function of PPARγ in both cell lines, although the suppressive effect in MCF-7:2A cells was delayed owing to high PPARγ expression. Activation of PPARγ by a specific agonist, pioglitazone, selectively blocked the induction of TNFα expression by E, but did not affect other adipose inflammatory genes, such as fatty acid desaturase 1 and IL6. This suppression of TNFα expression by pioglitazone was mainly mediated by transrepression of nuclear factor-κB (NF-κB) DNA-binding activity. A novel finding was that NF-κB functions as an oxidative stress inducer in MCF-7:5C cells but an antioxidant in MCF-7:2A cells. Therefore, the NF-κB inhibitor JSH-23 displayed effects equivalent to those of pioglitazone, with complete inhibition of apoptosis in MCF-7:5C cells, but it increased E-induced apoptosis in MCF-7:2A cells. Depletion of PPARγ by siRNA or the PPARγ antagonist T0070907 accelerated E-induced apoptosis, with activation of NF-κB-dependent TNFα and oxidative stress. For the first time, we demonstrated that PPARγ is a growth signal and has potential to modulate NF-κB activity and oxidative stress in E-deprived breast cancer cell lines. All of these findings suggest that anti-PPARγ therapy is a novel strategy to improve the therapeutic effects of E-induced apoptosis in E-deprived breast cancer.

Estrogen therapy was used to treat advanced breast cancer in postmenopausal women for decades until the introduction of tamoxifen. Resistance to long-term estrogen deprivation (LTED) with tamoxifen and aromatase inhibitors used as a treatment of breast cancer inevitably occurs, but unexpectedly low-dose estrogen can cause regression of breast cancer and increase disease-free survival in some patients. This therapeutic effect is attributed to estrogen-induced apoptosis in LTED breast cancer. Here, we describe modulation of the estrogen receptor (ER) liganded with antiestrogens (endoxifen and 4-hydroxytamoxifen) and an estrogenic triphenylethylene (TPE), ethoxytriphenylethylene (EtOXTPE), on estrogen-induced apoptosis in LTED breast cancer cells. Our results show that the angular TPE estrogen (EtOXTPE) is able to induce the ER-mediated apoptosis only at a later time compared with planar estradiol in these cells. Using real-time polymerase chain reaction, chromatin immunoprecipitation, western blotting, molecular modeling, and X-ray crystallography techniques, we report novel conformations of the ER complex with an angular estrogen EtOXTPE and endoxifen. We propose that alteration of the conformation of the ER complexes, with changes in coactivator binding, governs estrogen-induced apoptosis through the protein kinase regulated by RNA-like endoplasmic reticulum kinase sensor system to trigger an unfolded protein response.

Our clinically relevant finding is that glucocorticoids block estrogen (E 2)-induced apoptosis in long-term E 2-deprived (LTED) breast cancer cells. However, the mechanism remains unclear. Here, we demonstrated that E 2 widely activated adipose inflammatory factors such as fatty acid desaturase 1 (FADS1), IL6, and TNFa in LTED breast cancer cells. Activation of glucocorticoid receptor (GR) by the synthetic glucocorticoid dexamethasone upregulated FADS1 and IL6, but downregulated TNFa expression. Furthermore, dexamethasone was synergistic or additive with E 2 in upregulating FADS1 and IL6 expression, whereas it selectively and constantly suppressed TNFa expression induced by E 2 in LTED breast cancer cells. Regarding regulation of endoplasmic reticulum stress, dexamethasone effectively blocked activation of protein kinase RNA-like endoplasmic reticulum kinase (PERK) by E 2 , but it had no inhibitory effects on inositol-requiring protein 1 alpha (IRE1a) expression increased by E 2. Consistently, results from reverse-phase protein array (RPPA) analysis demonstrated that dexamethasone could not reverse IRE1a-mediated degradation of PI3K/Akt-associated signal pathways activated by E 2. Unexpectedly, activated GR preferentially repressed nuclear factor-kB (NF-kB) DNA-binding activity and expression of NF-kB-dependent gene TNFa induced by E 2 , leading to the blockade of E 2-induced apoptosis. Together, these data suggest that trans-suppression of NF-kB by GR in the nucleus is a fundamental mechanism thereby blocking E 2-induced apoptosis in LTED breast cancer cells. This study provided an important rationale for restricting the clinical use of glucocorticoids, which will undermine the beneficial effects of E 2-induced apoptosis in patients with aromatase inhibitorresistant breast cancer.