Insulin-like growth factor-I receptor kinase inhibitor NVP-AEW541 is active in breast cancer cells and enhances growth inhibition by herceptin through an increase in cell cycle arrest

Abstract

21077 Background: Targeting ErbB2 with the monoclonal antibody trastuzumab (Herceptin) has shown to be active in breast cancer. However, a proportion of patients do not benefit from this treatment due to primary resistance. Mechanisms proposed for this resistance include activation of other receptors involved in proliferation as is the case of the insulin-like growth factor receptor (IGF-1R).Different strategies have been proposed for the inhibition of IGF-1R such as antibodies or small tyrosine-kinase inhibitors. In this study we analyzed the mechanism of action and the antiproliferative activity of an specific IGF-1R tyrosine-kinase inhibitor termed NVP-AEW541 alone and in combination with Herceptin. Methods: MCF7, SKBR3, BT474, and MDA-MB-231 breast cancer cells were used for this study. Proliferation and apoptosis were analyzed by MTT uptake assays or Annexin-V-FITC, respectively. The levels of different signalling proteins were analyzed by Western blotting. Microarray studies were performed using the HU-133A oligonucleotide arrays (Affymetrix, Santa Clara, CA). Real time quantitative PCR was used to confirm the differentially regulated genes. Results: The IGF-1R was expressed in all the cell lines studied. Exogenous addition of IGF-1 increased the tyrosine phosphorylation of IGF-1R, and this was prevented by preincubation with NVP- AEW541. The latter also decreased MTT uptake, and provoked a slight increase in apoptosis of BT474 cells. Combination of NVP-AEW541 and Herceptin had a synergistic effect on the inhibition of BT474 proliferation. Cell cycle analyses indicated that the combined addition of NVP- AEW541 and Herceptin increased the number of cells in the G0/G1 phases. These data were complemented with an increase in p27 levels upon treatment with both compounds. Microarray analyses identified several genes whose expression was modified by each agent alone and by the combination. Conclusions: Our results indicate that the combined targeting of two receptor tyrosine kinases known to play important roles in tumor cell proliferation may be more efficient than individual targeting of these receptors. No significant financial relationships to disclose.