Crosstalk Between ER, Growth Factor Receptor Signaling Pathways
ER functions in the nucleus as a transcription factor for specific estrogen regulated genes. This activity is known as nuclear initiated steroid signaling (NISS) and is sometimes referred to as its classical or genomic activity. Many of the genes regulated by estrogen are important for cell proliferation, survival, metastasis, and angiogenesis. ER can activate transcription of genes by binding to specific estrogen response elements in the promoter of target genes and by recruiting coactivators or corepressors to this complex. Alternatively, ER can function as a coactivator itself by tethering to other transcription factors such as those regulating AP1 sites.
Tamoxifen, a selective estrogen receptor modulator (SERM), functions as an estrogen antagonist for many of these genes, although for a subset, tamoxifen has estrogen-like agonist activity. Therapies designed to reduce the level of estrogen in patients such as ovarian ablation or aromatase inhibitors function by depriving the receptor of its ligand.
Finally, a new class of endocrine agents, the selective ER downregulators (SERDs) or pure antiestrogens developed preclinically in part with our research program, is also effective even as we predicted in tamoxifen-resistant patients, and the pure antiestrogen fulvestrant is now FDA approved.
A small pool of ERs may reside in the cytoplasm or in the plasma membrane. Binding of estrogen to this receptor can directly or indirectly activate several receptor tyrosine kinases including the IGF receptor and members of the EGFR family.
In contrast to the genomic action of ER, which requires nuclear localization of the receptors and takes as usual a few hours for its significant manifestation, estrogen signaling through this non-nuclear/non-genomic activity of ER, also known as membrane initiated steroid signaling (MISS), occurs within seconds to a few minutes. Downstream kinases in these signaling pathways such as Akt, p42,44 MAPK, p38 MAPK and JNK phosphorylate and, thereby, activate ER and its coregulatory proteins. Activation of the downstream kinase pathways can also directly enhance tumor cell proliferation and survival.
Tamoxifen can also activate membrane ER just like estrogen itself. Tumors with high growth factor receptor content are those in which membrane ER signaling becomes dominant, an observation that explains why some patients with tumors overexpressing HER2 show resistance to tamoxifen. Simultaneous treatment with tamoxifen combined with specific growth factor receptor inhibitors (such as monoclonal antibodies or selective tyrosine kinase inhibitors) restores tamoxifen’s antagonist activity, a strategy that is now in clinical trial.
Our group is interested in further clarifying the role of this crosstalk in various hormone resistance and in identifying the optimal method of blocking growth factor signaling to restore tamoxifen’s antagonist activity and to optimize endocrine therapies.
Role of AIB1 in Endocrine Therapy Resistance
AIB1 is an important coregulatory protein for ER and for other transcription factors. We have reported that high expression of AIB1 together with high expression of HER2 in human tumor samples predicts for tamoxifen resistance. We have identified at least eight potential phosphorylation sites on AIB1 protein. AIB1 is a substrate for a variety of kinases including p42/44 MAPK, Akt, p38 MAPK, JNK, and IKK.
We have shown that phosphorylation of one of these sites increases the agonist activity of tamoxifen and we have shown that under circumstances of high growth factor receptor signaling, tamoxifen bound ER can recruit AIB1 and other coactivator proteins to the promoter of target genes, thereby increasing the drug’s agonist activity and resulting in tamoxifen resistance. We are interested in determining the functional consequences of phosphorylation of these various sites and to better understand the role of AIB1 in ER function and in hormone therapy resistance.
Role of FKHR in Breast Cancer Pathogenesis
FKHR is a protein important in the cell death pathway. Its function is blocked by phosphorylation induced by Akt in response to growth factor signaling.
Using a yeast two hybrid screen, we first identified FKHR as an ER interacting protein and then showed that it has corepressor functions in addition to its role in apoptosis. We then found that the gene for FKHR which is located near Rb and BRCA-2 is lost in about 50% of breast cancers suggesting the possibility that it functions in part as a tumor suppressor gene.
To further clarify the role of FKHR in mammary gland development and in breast cancer development and progression, we have developed transgenic mice overexpressing the gene in the mammary gland in an inducible manner and we have developed conditional knock out mice in which the gene is specifically lost in the mammary gland. Using these models we will test the hypothesis that overexpression of FKHR protects the mammary gland from tumorigenesis by functioning as an ER corepressor and via its role in apoptosis (tumor suppressor function). Conversely, loss of FKHR would be expected to increase tumorigenesis in the mammary gland.
Other gene networks may also contribute to hormone therapy resistance. Using our model systems and tumor specimens from patients, we are also interested in using high throughput molecular profiling of DNA, RNA, and protein to be able to predict hormone therapy resistance in patients and to help select the most appropriate treatment. These molecular profiling studies will also identify new potential treatment targets. The Breast Center microarray, pathology, and biostatic core facilities are critical for these studies.