A kinase inhibitor screen reveals MEK1/2 as a novel therapeutic target to antagonize IGF1R-mediated antiestrogen resistance in ERα-positive luminal breast cancer.

Antiestrogen resistance of breast cancer has been related to enhanced growth factor receptor expression and activation. We have previously shown that ectopic expression and subsequent activation of the insulin-like growth factor-1 receptor (IGF1R) or the epidermal growth factor receptor (EGFR) in MCF7 or T47D breast cancer cells results in antiestrogen resistance. In order to identify novel therapeutic targets to prevent this antiestrogen resistance, we performed kinase inhibitor screens with 273 different inhibitors in MCF7 cells overexpressing IGF1R or EGFR. Kinase inhibitors that antagonized antiestrogen resistance but are not directly involved in IGF1R or EGFR signaling were prioritized for further analyses. Various ALK (anaplastic lymphoma receptor tyrosine kinase) inhibitors inhibited cell proliferation in IGF1R expressing cells under normal and antiestrogen resistance conditions by preventing IGF1R activation and subsequent downstream signaling; the ALK inhibitors did not affect EGFR signaling. On the other hand, MEK (mitogen-activated protein kinase kinase)1/2 inhibitors, including PD0325901, selumetinib, trametinib and TAK-733, selectively antagonized IGF1R signaling-mediated antiestrogen resistance but did not affect cell proliferation under normal growth conditions. RNAseq analysis revealed that MEK inhibitors PD0325901 and selumetinib drastically altered cell cycle progression and cell migration networks under IGF1R signaling-mediated antiestrogen resistance. In a group of 219 patients with metastasized ER + breast cancer, strong pMEK staining showed a significant correlation with no clinical benefit of first-line tamoxifen treatment. We propose a critical role for MEK activation in IGF1R signaling-mediated antiestrogen resistance and anticipate that dual-targeted therapy with a MEK inhibitor and antiestrogen could improve treatment outcome.

Classifying the adverse mitogenic mode of action of insulin analogues using a novel mechanism-based genetically engineered human breast cancer cell panel.

Insulin analogues are widely used in clinical practice. Modifications on the insulin molecular structure can affect the affinity and activation towards two closely related receptor tyrosine kinases: the insulin receptor (INSR) and the insulin-like growth factor 1 receptor (IGF1R). A switch towards higher IGF1R affinity is likely to emphasize mitogenesis rather than glucose metabolism. Relevant well-validated experimental tools to address the insulin analogue activation of either INSR or IGF1R are missing. We have established a panel of human MCF-7 breast cancer cell lines either ectopically expressing the INSR (A or B isoform) in conjunction with a stable knockdown of the IGF1R or ectopically expressing the IGF1R in conjunction with a stable knockdown of the INSR. In these cell lines, we systematically evaluated the INSR and IGF1R receptor activation and downstream mitogenic signalling of all major clinical relevant insulin analogues in comparison with insulin and IGF1R. While most insulin analogues primarily activated the INSR, the mitogenic activation pattern of glargine was highly similar to IGF1 and insulin AspB10, known to bind IGF1R and induce carcinogenesis. Yet, in a long-term proliferation assay, the proliferative effect of glargine was not much different from regular insulin or other insulin analogues. This was caused by the rapid enzymatic conversion into its two metabolic active metabolites M1 and M2, with reduced mitogenic signalling through the IGF1R. In summary, based on our new cell models, we identified a similar mitogenic potency of insulin glargine and AspB10. However, rapid enzymatic conversion of glargine precludes a sustained activation of the IGF1R signalling pathway.