C 4.4 A is associated with tumor budding and epithelial–mesenchymal transition of colorectal cancer

C 4.4 A is a glycolipid‐anchored membrane protein expressed in several human malignancies. The aim of this study was to explore the association between C 4.4 A expression at the invasion front of colorectal cancer ( CRC ) and tumor budding, a putative hallmark of cell invasion of CRC . Advanced CRC s ( T 2–4, n  = 126) had a budding count of 3.66 ± 5.66, which was significantly higher than that of T 1 early CRC s (1.75 ± 2.78, n  = 87). C 4.4 A ‐positive CRC specimens showed a larger budding cell number than C 4.4 A ‐negative CRC specimens in T 1 CRC s, and especially advanced CRC s (9.45 ± 5.83 vs 1.60 ± 3.93). Furthermore, we found a correlation between the percentage of C 4.4 A ‐positive cases and budding count in advanced CRC . Multivariate analysis for patients' survival showed that C 4.4 A was superior to tumor budding as a prognostic factor. With si RNA treatment, C 4.4 A levels were associated with cell invasion, but not with proliferation, in HCT 116 and DLD 1 cell lines. An immunohistochemical study in a subset of CRC s showed no relationship between C 4.4 A and K i‐67 proliferation marker. In vitro assays using HCT 116 indicated that C 4.4 A levels correlated well with epithelial–mesenchymal transition ( EMT ) with regard to cell morphology and alterations of EMT markers including E ‐cadherin, vimentin, and partially N ‐cadherin. We also found that C 4.4 A expression was significantly associated with loss of E ‐cadherin and gain of β‐catenin in clinical CRC tissue samples. These findings suggest that a tight association between C 4.4 A and tumor budding may, in part, be due to C 4.4 A promoting EMT at the invasive front of CRC . ( Cancer Sci 2012; 103: 1155–1164)