Overexpression of circRNA chr7:154954255-154998784+ in cancer-associated pancreatic stellate cells promotes the growth and metastasis of pancreatic cancer by targeting the miR-4459/KIAA0513 axis.

PURPOSE

Circular RNAs (circRNAs) have been reported to act as important regulators in pancreatic cancer. Abnormal expression of circRNAs in pancreatic cancer cells (PCCs) can promote the development of pancreatic cancer; however, the role of circRNAs in cancer-associated pancreatic stellate cells (CaPSCs) remains unclear.

PATIENTS AND METHODS

In this study, we isolated CaPSCs from pancreatic cancer tissues from 5 pancreatic cancer patients and NaPSCs from normal pancreatic tissue from 5 patients with benign pancreatic disease. After the PSCs were co-cultured with the pancreatic cancer cell line PANC-1, a CCK-8 assay was used to detect PANC-1 proliferation ability, and CaPSCs, which had the strongest promoting effect on PANC-1 proliferation, and NaPSCs, which had the weakest effect, were screened. Then, the circRNA, microRNA (miRNA) and mRNA profiles between CaPSCs and NaPSCs were compared by RNA-seq. The candidate circRNA/miRNA/target protein axis was selected using bioinformatics analysis. circRNAs were silenced and miRNAs were overexpressed in CaPSCs, and the expression of circRNAs, miRNAs and target proteins were detected by qRT-PCR and Western blot, respectively. At the same time, CCK8, wound healing, and Transwell assays were used to detect the proliferation, migration and invasion of PANC-1 cells in the different co-culture groups. Moreover, a tumour xenograft model was used to observe the tumorigenic ability of PANC-1 cells in different co-culture groups. Finally, immunohistochemistry was used to detect the expression of target proteins in PDAC tissues, and the clinicopathological features and prognosis were analysed.

RESULTS

The expression of the differentially expressed RNAs identified by RNA-seq was verified by qRT-PCR, and the chr7:154954255-154998784+/miR-4459/KIAA0513 axis was selected from the candidate targets. Functional studies of PANC-1 cells after co-culture with chr7:154954255-154998784+-silenced CaPSCs showed that the proliferation, invasion and metastasis of PANC-1 cells decreased. Moreover, after chr7:154954255-154998784+ was silenced, the expression of miR-4459 in CaPSCs increased, and the expression of KIAA0513 decreased. When PANC-1 cells were co-cultured with CaPSCs with miR-4459 overexpression, they showed an increased ability to proliferate, invade and metastasize. Additionally, when miR-4459 was overexpressed in CaPSCs, the expression of chr7:154954255-154998784+ and KIAA0513 decreased. Animal experiments revealed that silencing chr7:154954255-154998784+ in CaPSCs inhibited tumour growth in nude mice inoculated with CaPSCs+PANC-1 cells. Finally, we performed immunohistochemistry and a prognostic analysis of KIAA0513 expression in paraffin tissue samples from patients with pancreatic cancer and found that high expression of KIAA0513 was associated with more aggressive clinicopathological factors. Furthermore, patients with high expression of KIAA0513 had worse disease-free survival (DFS) and overall survival (OS).

CONCLUSION

Chr7:154954255-154998784+ may promote the development of pancreatic cancer through the miR-4459/KIAA0513 axis in CaPSCs and may be an important therapeutic target for patients with pancreatic cancer in the future.