Data Availability StatementThe raw data used and analyzed in the current study are available from the corresponding author upon a reasonable request. of miR-21 significantly abolished the antimetastatic effects of CASC2 on PANC-1 cells. Moreover, the downregulation of PTEN significantly abolished the antimetastatic effects of CASC2. Conclusion CASC2 functions as a tumor suppressor in pancreatic cancer cells to inhibit tumor cell migration and invasion. Our work revealed a novel regulatory mechanism of the CASC2/miR-21/PTEN axis that may be important in pancreatic cancer. test and one-way analysis of variance (ANOVA) with Tukeys post hoc test. P-values less than 0.05 were considered statistically significant. Results Expression levels of CASC2 are low in pancreatic cancer cells, and CASC2 suppresses cell migration and invasion The expression levels of CASC2 in human pancreatic cancer cell lines CAPAN-1, BxPC-3, JF305, PANC-1 and SW1990 and in normal human pancreatic HPDE6-C7 cells were assayed (Fig.?1a). The qRT-PCR analysis results showed that the levels of CASC2 in the pancreatic cancer cell lines were significantly MS-275 kinase inhibitor lower than that in the normal human pancreatic cells (P? ?0.01). Open in a separate window Fig.?1 CASC2 suppressed metastasis of the PANC-1 pancreatic carcinoma cell. a Levels of CASC2 expression are low in the pancreatic carcinoma cells. Expression of CASC2 in the human pancreatic cancer cell lines and normal pancreatic HPDE6-C7 cells was detected by qRT-PCR. **P? ?0.01 vs. HPDE6-C7. bCd CASC2 sequences were ligated into the pEX-2 vector (pEX-CASC2). An empty pEX-2 vector was used as a negative control (pEX). Pancreatic carcinoma cells were transfected with the CASC2-expressing vector (pEX-CASC2) or the corresponding negative control (pEX) for 48?h. The cells without transfection were used as a control (CT). b Expression of CASC2 in the cells. c MS-275 kinase inhibitor Cell migration and d invasion were assessed by the transwell assay (n?=?3; 10 random fields were counted). ***P? ?0.001. Scale bar: 100?m To detect whether CASC2 regulated cell migration MS-275 kinase inhibitor and invasion in the pancreatic cancer cells, CASC2 was overexpressed by pEX-CASC2 in PANC-1 cells (Fig.?1b, P? ?0.001). The overexpression of CASC2 significantly inhibited the migration of PANC-1 cells (P? ?0.001). Similar to migration, the overexpression of CASC2 significantly inhibited the invasion of PANC-1 cells (P? ?0.001). Thus, these data suggest that CASC2 plays an antimetastatic role in PANC-1 cells. CASC2 inhibits the migration and invasion of pancreatic cancer cells by directly targeting miR-21 To test whether lncRNA CASC2 acts as a ceRNA via sponging miR-21, we detected the levels of miR-21 in CAPAN-1, BxPC-3, JF305, PANC-1 and SW1990 cells and in normal human pancreatic HPDE6-C7 cells (Fig.?2a), as well MS-275 kinase inhibitor as in the pEX-CASC2-transfected PANC-1 cells (Fig.?2b). The qRT-PCR results showed that levels of miR-21 in the pancreatic cancer cell lines were significantly higher than those in the HPDE6-C7 cells (P? ?0.01, Fig.?2a). The overexpression of CASC2 significantly downregulated the expression of miR-21 (P? ?0.001, Fig.?2b). Moreover, the CASC2-wt or CASC2-mut vectors were WASF1 cotransfected with miR-21 mimics or miR-21 mimic NC into the cells. Cotransfection of miR-21 mimics and CASC2-wt significantly decreased the luciferase activity (P? ?0.001, Fig.?2c); however, the cotransfection of miR-21 and CASC2-mut did not change luciferase activity. These results suggested that miR-21 is a direct target of CASC2. MiR-21 mimics significantly increased the miR-21 levels in the pEX CASC2 transfected PANC-1 cells, while pEX CASC2 significantly downregulated the expression of miR-21 (P? ?0.01, Fig.?2d). MiR-21 mimics significantly promoted cell migration and invasion and significantly reversed the suppression of migration and invasion induced by CASC2 in the PANC-1 MS-275 kinase inhibitor cells, suggesting that the overexpression of miR-21 significantly abolished the antimetastatic activity of CASC2 in PANC-1 cells (P? ?0.001, Fig.?2e, f). Thus, these results suggest that CASC2 inhibited cell metastasis through the negative regulation of miR-21. Open in a separate window Fig.?2 MiR-21 overexpression reversed the role of CASC2 in PANC-1 pancreatic carcinoma cells. a Expression of miR-21 in the human pancreatic cancer cell lines and normal pancreatic HPDE6-C7 cells. b PANC-1 cells were transfected with the CASC2-expressing vector (pEX-CASC2) or.