Anti-cancer potentials of Gynura procumbens leaves extract against two canine mammary cancer cell lines

Background: The anti-cancer effects of Gynura procumbens leaves extract (GPE) have been reported in various human cancers. However, the anti-cancer effects and molecular mechanisms of this extract on canine mammary cancer (CMC) have not yet been elucidated. Objectives: The main goal of this study was to investigate the anti-cancer properties of GPE against two CMC cell lines (CHMp-13a and CHMp-5b). Methods: The GP leaves were extracted with 80% ethanol. Anti-cancer potentials of GPE on CHM p-13a and CHM p-5b cancer cell lines using dimethyl-2-thiazolyl-2,5-diphenyl-2H-tetrazolium bromide (MTT), wound healing, transwell migration, and caspase 3/7 activity assays were evaluated. The mRNA expression levels of two oncogenes: epidermal growth factor receptor (EGFR) and twist family bHLH transcription factor 1 (TWIST) and one tumour suppressor gene: phosphatase and tensin homolog (PTEN) in these cell lines were determined by quantitative real-time PCR (qRT-PCR). In addition, The EGFR and PTEN protein levels as well as protein kinase B (AKT) and extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation levels expression were also evaluated by western blot analysis. Results: The results showed that GPE caused a significant concentration- and time-dependent reduction in cell proliferation of both CHMp-13a and CHM p-5b cells, detected by MTT assays. This extract also significantly suppressed cancer cell migration in both cell lines, tested by wound healing and transwell migration assays. Additionally, the increase in caspase 3/7 activity observed in both CMC cell treated with GPE suggests that GPE induced caspase 3/7 dependent apoptosis. Moreover, GPE significantly decreased EGFR m RNA and protein expression levels corn pared to control in both cell lines in a dose-dependent manner. Conclusion: These findings emphasized that GPE has an in vitro anti-cancer activity against CMC by inhibiting EGFR signalling pathway. Thus, GPE may serve as an alternative therapy in CMC with high EGFR expression.