Expression and Activity of Both IGF and Insulin Signaling Pathways in a Large Panel of Breast Cancer Cell Lines.

BACKGROUND: Insulin and insulin like growth factor-I (IGF-I) are key regulators in growth and metabolism. IGF-I signaling is known to be involved in tumor progression and drug resistance, and growing evidence also suggests that insulin signaling may play an important role in cancer. To gain a better understanding of these signaling pathways in human breast cancer, we measured the levels of IR and IGF-IR, and the activity of insulin and IGF-I, in a large panel of human breast cancer cell lines.METHODS: Comparative gene expression analysis was performed using publicly available gene expression data among a panel of 51 breast cancer cell lines. Q-RT-PCR was used to validate IR and IGF-IR expression differences in 19 breast cell lines. For 8 breast cancer cell lines we immunoblotted for total IR and IGF-IR levels, as well as p-Akt and p-Erk1/2 following insulin and IGF stimulation. Reverse phase protein array (RPPA) analysis was performed on MCF7 cells stimulated with increasing doses of insulin or IGF-I.RESULTS: IR and IGF-IR mRNA levels measured by microarray on breast cancer cell lines showed divergent levels with cells positive or negative for expression of each gene. There was no obvious correlation of levels with breast caner subtype. Q-RT-PCR results were generally consistent with the gene expression data with cell lines categorized into four groups: high IGF-IR and low IR e.g. MCF7 and MDA-MB-134; low IGF-IR and high IR e.g. ZR-75-1 and MDA-MB-468; intermediate levels of IGF-IR and IR e.g. MDA-MB-231 and HCC1954; and low IGF-IR and IR e.g. MDA-MB-415 and HS-578T. IGF-IR protein levels correlated with Q-RT-PCR (Spearman Rank Correlation r=0.93, p=0.0067) whereas IR didn't. We found significant activation of Akt in both insulin and IGF treated MCF7 and MDA-MB-134 cells as well as Akt activation in IGF stimulated MDA-MB-453. Interestingly, significant Erk1/2 activation was observed only in insulin and IGF stimulated MCF7 and insulin stimulated AU565 cells. Cell lines that didn't respond to insulin or IGF stimulation generally had constitutively high p-Akt and/or p-Erk1/2. From our preliminary RPPA analysis in MCF7, we observed an increase in phosphorylation of IGF-IR, Akt, Erk1/2 and p70S6K upon increasing dose of IGF stimulation and a similar effect was also observed following insulin stimulation.DISCUSSION: IGF-IR and IR mRNA and protein levels are divergent in a panel of breast cancer cell lines with cells expressing mRNA for both, one alone, or neither receptor. IGF-IR and IR protein levels indicated similar divergent expression. Interestingly, response to both IGF-IR and insulin stimulation was restricted to cells showing high levels of IGF-IR. To better understand insulin and IGF activities in breast cancer, we are currently performing RPPA on a large panel of 21 breast cancer cell lines.

Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 4168.