Lipid Raft Disruption Restores IGFBP7 Expression in Breast Cancer by Activating the MAPK Pathway and Hedgehog-Mediated DNMT1 and EZH2 Repression

Abstract

Abnormal chromatin (DNA and histones) alterations and disturbances in cellular signaling pathways are two of the many intricate molecular processes that cause breast cancer. These changes are essential for turning oncogenes on and suppressing tumor suppressor genes. With a particular focus on the tumor suppressor gene IGFBP7, this study examines how plasma membrane lipid rafts regulate epigenetic factors and signaling pathways in breast cancer. Several genes were identified by transcriptome analysis of publicly accessible GEO datasets, with UHRF1 standing out as a key gene associated with epigenetic DNA methylation. Methyl-β-cyclodextrin (MBCD) was shown to activate the EGFR–RAS–MAPK signaling cascade by altering the lipid raft structure in MDA-MB-231 cells, which is a triple-negative breast cancer cell line. IGFBP7 expression was reactivated as a result of this disruption, which also caused DNMT1 and EZH2 to be downregulated. Specific inhibitors that target this signaling cascade were employed to verify that the EGFR–RAS–MAPK pathway regulates IGFBP7, and the outcomes demonstrated the pathway's role in the regulation of the IGFBP7 gene. These results were corroborated by protein-protein docking simulations, which revealed positive interactions between DNMT1, EZH2, and UHRF1, indicating that these proteins constitute a coordinated repression complex. Interestingly, GLI1, one of the most important effectors of the Hedgehog (Hh) signal transduction system, was elevated upon MBCD therapy. Accordingly, it reduced DNMT1 and EZH2 and elevated IGFBP7 expression. On the contrary, these activities were reversed by inhibiting the Hedgehog axis, demonstrating its central role in regulating epigenetic suppression and achieving tumor suppressor restoration. Functional experiments showed that IGFBP7 reactivation led to decreased stemness, colony formation, and cell motility, as evidenced by a drop in OCT4 expression. These results reveal a unique mechanism that connects epigenetic regulation, Hedgehog signaling, and lipid raft stability, pointing to new therapeutic approaches for the treatment of breast cancer.