Regulation of protumorigenic pathways by insulin like growth factor binding protein2 and its association along with ß-catenin in breast cancer lymph node metastasis.

ABSTRACT

BACKGROUND: Insulin like growth factor binding proteins modulate the mitogenic and pro survival effects of IGF. Elevated expression of IGFBP2 is associated with progression of tumors that include prostate, ovarian, glioma among others. Though implicated in the progression of breast cancer, the molecular mechanisms involved in IGFBP2 actions are not well defined. This study investigates the molecular targets and biological pathways targeted by IGFBP2 in breast cancer. METHODS: Transcriptome analysis of breast tumor cells (BT474) with stable knockdown of IGFBP2 and breast tumors having differential expression of IGFBP2 by immunohistochemistry was performed using microarray. Differential gene expression was established using R-Bioconductor package. For validation, gene expression was determined by qPCR. Inhibitors of IGF1R and integrin pathway were utilized to study the mechanism of regulation of ß-catenin. Immunohistochemical and immunocytochemical staining was performed on breast tumors and experimental cells, respectively for ß-catenin and IGFBP2 expression. RESULTS: Knockdown of IGFBP2 resulted in differential expression of 2067 up regulated and 2002 down regulated genes in breast cancer cells. Down regulated genes principally belong to cell cycle, DNA replication, repair, p53 signaling, oxidative phosphorylation, Wnt signaling. Whole genome expression analysis of breast tumors with or without IGFBP2 expression indicated changes in genes belonging to Focal adhesion, Map kinase and Wnt signaling pathways. Interestingly, IGFBP2 knockdown clones showed reduced expression of ß- catenin compared to control cells which was restored upon IGFBP2 re-expression. The regulation of ß-catenin by IGFBP2 was found to be IGF1R and integrin pathway dependent. Furthermore, IGFBP2 and ß-catenin are co-ordinately overexpressed in breast tumors and correlate with lymph node metastasis. CONCLUSION: This study highlights regulation of ß-catenin by IGFBP2 in breast cancer cells and most importantly, combined expression of IGFBP2 and ß-catenin is associated with lymph node metastasis of breast tumors.