FOXD3 Regulates CSC Marker, DCLK1-S, and Invasive Potential: Prognostic Implications in Colon Cancer

The 5' (α)-promoter of the human doublecortin-like kinase 1 (DCLK1) gene becomes epigenetically silenced during colon carcinogenesis, resulting in loss of expression of the canonical long(L)-isoform1 (DCLK1-L) in human colon adenocarcinomas (hCRCs). Instead, hCRCs express a short(S)-isoform2 (DCLK1-S) from an alternate (β)-promoter of DCLK1. The current study, examined if the transcriptional activity of the (β)-promoter is suppressed in normal versus cancerous cells. On the basis of in silico and molecular approaches, it was discovered that FOXD3 potently inhibits the transcriptional activity of the (β)-promoter. FOXD3 becomes methylated in human colon cancer cells (hCCC), with loss of FOXD3 expression, allowing expression of the DCLK1(S) variant in hCCCs/hCRCs. Relative levels of FOXD3/DCLK1(S/L) were measured in a cohort of CRC patient specimens (n = 92), in relation to overall survival (OS). Patients expressing high DCLK1(S), with or without low FOXD3, had significantly worse OS compared with patients expressing low DCLK1(S). The relative levels of DCLK1-L did not correlate with OS. In a pilot retrospective study, colon adenomas from high-risk patients (who developed CRCs in <15 years) demonstrated significantly higher staining for DCLK1(S) + significantly lower staining for FOXD3, compared with adenomas from low-risk patients (who remained free of CRCs). Latter results strongly suggest a prognostic value of measuring DCLK1(S)/FOXD3 in adenomas. Overexpression of DCLK1(S), but not DCLK1(L), caused a significant increase in the invasive potential of hCCCs, which may explain worse outcomes for patients with high DCLK1-S-expressing tumors. On the basis of these data, FOXD3 is a potent repressor of DCLK1-S expression in normal cells; loss of FOXD3 in hCCCs/hCRCs allows upregulation of DCLK1-S, imparting a potent invasive potential to the cells. Mol Cancer Res; 15(12); 1678-91. ©2017 AACR.