Overexpression of CDHR2 inhibits proliferation of breast cancer cells by inhibiting the PI3K/Akt pathway

doi:10.12122/j.issn.1673-4254.2024.06.12

Abstract

Abstract:

Objective To investigate the mechanism by which CDHR2 overexpression inhibits breast cancer cell growth and cell cycle pragression via the PI3K/Akt signaling pathway. Methods Bioinformatic analysis was performed to investigate CDHR2 expression in breast cancer and its correlation with survival outcomes of the patients. Immunohistochemistry was used to examine CDHR2 expressions in surgical specimens of tumor and adjacent tissues from 10 patients with breast cancer. CDHR2 expression levels were also detected in 5 breast cancer cell lines and a normal human mammary epithelial cell line using qRT-PCR and Western blotting. Breast cancer cell lines MDA-MB-231 and MCF7 with low CDHR2 expression were transfected with a CDHR2-overexpressing plasmid, and the changes in cell proliferation and cell cycle were evaluated using CCK-8 assay, EdU assay, and cell cycle assay; the changes in expressions of PI3K/Akt signaling pathway and cell cycle pathway proteins were detected with Western blotting. Results Bioinformatic analysis showed low CDHR2 expression level in both breast cancer and adjacent tissues without significant difference between them (P>0.05), but breast cancer patients with a high expression of CDHR2 had a more favorable prognosis. Immunohistochemistry, qRT-PCR and Western blotting showed that the expression of CDHR2 was significantly down-regulated in breast cancer tissues and breast cancer cells (P<0.01), and its overexpression strongly inhibited cell proliferation, caused cell cycle arrest, and significantly inhibited PI3K and Akt phosphorylation and the expression of cyclin D1. Conclusion Overexpression of CDHR2 inhibits proliferation and causes cell cycle arrest in breast cancer cells possibly by inhibiting the PI3K/Akt signaling pathway.

Key words: breast cancer, CDHR2, PI3K/Akt pathway, proliferation, cell cycle

Jincun FANG, Liwei LIU, Junhao LIN, Fengsheng CHEN. Overexpression of CDHR2 inhibits proliferation of breast cancer cells by inhibiting the PI3K/Akt pathway[J]. Journal of Southern Medical University, 2024, 44(6): 1117-1125.

Figures/Tables 11

Fig.1 Analysis of CDHR2 expression levels in breast cancer and adjacent tissues based on TCGA dataset.

Fig.2 Survival outcomes of breast cancer patients with low and high CDHR2 expressions. A-D: Relationship between CDHR2 expression and prognosis of the patients with triple-negative (A), luminal A (B), luminal B (C), and Her2-overexpressing (D) breast cancer, respectively.

Fig.3 Fig. 3 Immunohistochemical staining for CDHR2 in clinical specimens of breast cancer tissues (Original magnification: ×400). A: Strong expression of CDHR2 in breast tissues. B: Weak expression of CDHR2 in breast tissues. C: Strong expression of CDHR2 in breast cancer tissues. D: Weak expression of CDHR2 in breast cancer tissues. ****P<0.0001.

Fig.4 CDHR2 mRNA expression levels in breast cancer cell lines and normal human mammary epithelial cells. **P<0.01, ***P<0.001 vs MCF10A.

Fig.5 CDHR2 protein expression levels in breast cancer cell lines and normal human mammary epithelial cells detected by Western blotting. ***P<0.001 vs MCF10A.

Fig.6 CDHR2 mRNA expression in MDA-MB-231 and MCF7 cells transfected with CDHR2-overexpressing plasmids. ***P<0.001.

Fig. 7 CCK-8 cell proliferation assay of the transfected breast cancer cell lines. **P<0.01, ***P<0.001.

Fig.8 EdU proliferation assay of the transfected breast cancer cell lines. **P<0.01.

Fig.9 Flow cytometry for cell cycle analysis of the transfected breast cancer cell lines. **P<0.01.

Fig.10 Western blotting of PI3K/Akt signaling pathway proteins and cyclin D1 in the transfected cells. ***P<0.001, ****P<0.0001 vs negetive control (NC).

Fig.11 Proteins with possible binding sites for CDHR2.

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