Frequent Inactivation of Axon Guidance Molecule RGMA in Human Colon Cancer Through Genetic and Epigenetic Mechanisms

Abstract

Repulsive guidance molecule member A (RGMA) is a glycosylphosphatidylinositol-anchored glycoprotein and axon guidance molecule that signals through its receptor, neogenin (NEO1), a homologue of the deleted-in-colorectal cancer (DCC) gene. RGMA also functions as a bone morphogenetic protein (BMP) coreceptor. We studied the potential roles of RGMA and NEO1 in colorectal cancer (CRC) pathogenesis. We analyzed expression of RGMA and NEO1, as well as their epigenetic and genetic changes, in a large series of CRC samples, normal colon tissues, adenomas, and cell lines. These studies were accompanied by in vitro functional assay. RGMA and NEO1 expression were significantly down-regulated in most CRCs, adenomas, and cell lines. RGMA was frequently silenced by promoter methylation in CRCs (86.7%), adenomas (90.9%), and CRC cell lines (92.3%) but not in normal colon tissues; allelic imbalance of RGMA and NEO1 was observed in 40% and 49% of CRCs, respectively. In CRC samples, reduced RGMA levels were significantly associated with mismatch repair deficiency or mutations in KRAS or BRAF. Exposure to 5-aza-2'-deoxycytidine restored RGMA expression in CRC cell lines. Transfection of RGMA into CRC cells suppressed cell proliferation, migration, and invasion and also increased apoptosis in response to DNA-damaging agent. The frequent genetic and epigenetic inactivation of RGMA in CRCs and adenomas along with its in vitro function collectively support its role as a tumor suppressor in colon cells. These findings add to the expanding list of axon guidance molecules with disrupted function during colon carcinogenesis and create new opportunities for early detection and drug development.