Control of luminal type A intrinsic subtype enriched transcription factor network by insulin: implications of diabetes on breast cancer classification.

Abstract

Abstract Abstract #3029 Background: Luminal type A and type B represent estrogen receptor alpha (ERalpha)-positive breast cancers with luminal type A expressing higher levels of ERalpha and is associated with better prognosis. Recent studies have identified a specific functional transcription factor network comprising GATA-3, FOXA1 and ERalpha in normal luminal cells as well as in luminal type A breast cancer that dictates their hormone dependence. Signaling molecules that may disrupt this network and force these cells to acquire hormone-independence are not known. T-bet (Tbx21) has been described as a major negative regulator of GATA-3 activity. As the expression and/or activity of some of the above factors are controlled by insulin, the objective of this study was to investigate whether elevated level of insulin, as evidenced in type II diabetes, alters gene expression pattern in luminal type A breast cancers by interrupting GATA-3:FOXA1:ERalpha network and thus forcing these cancers to acquire non-luminal phenotype and/or hormone-independence.
 Methodologies: The effect of insulin on the expression of ERalpha, FOXA1, GATA-3 and T-bet was measured in ERalpha-positive MCF-7 cells by Western blot analysis. The effect of T-bet on estrogen-regulated gene expression was measured by stable overexpression of T-bet in MCF-7 cells and subsequent qRT-PCR analysis. Publicly available Oncomine database was used to determine the expression pattern of T-bet and its relation to ERalpha status in primary breast cancers. A proliferation assay was used to determine sensitivity of T-bet overexpressing cells to tamoxifen in the presence and absence of insulin.
 Results: Insulin induced the expression of T-bet, which was partially reversed by estrogen. ERalpha and GATA-3 levels were reduced in MCF-7 cells stably overexpressing T-bet suggesting that T-bet reduces GATA-3-dependent ERalpha expression. Estrogen-inducible expression of estrogen target genes GREB-1 and Myb was lower in T-bet overexpressing cells compared to parental cells, although basal expression was elevated in T-bet overexpressing cells. Treatment of T-bet overexpressing cells with insulin decreased tamoxifen sensitivity. Although T-bet expression was generally higher in ERalpha-negative breast cancers compared to ERalpha-positive breast cancers, a subpopulation of ER-positive breast cancers express elevated levels of T-bet.
 Conclusions: Insulin may change the gene expression pattern through T-bet-mediated disruption of master cell-type-specific transcriptional network including GATA-3, ERalpha and FOXA1 that dictates the phenotype of hormone-dependent luminal type A breast cancer. T-bet may serve as a marker to identify ERalpha-positive breast cancers that express low levels of GATA-3 and have progressed to hormone-independence. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3029.