Paeoniflorin effectively inhibit melanoma growth through induce senescence and ER stress converging on the Calpain1/ERK5/P21 axis

Background: Senescence leads to permanent cell-cycle arrest and is a potential target for cancer therapy. Paeoniflorin (PAE) is a monoterpene isolated from Traditional Chinese Medicine (TCM) Paeonia Alba, which has been used as an anti-inflammatory drug in clinical practice with the potential to target senescence in recalcitrant melanoma. Purpose: To determine whether PAE can induce senescence in melanoma cells in vitro and in vivo and to elucidate the underlying mechanisms. Methods: SA-β-Gal staining was used to detect the expression of senescence-associated β-galactosidase (SA-β-Gal) in melanoma cells line. DNA damage was examined by the detection of γH2AX foci. Cancer cell proliferation was determined by colony formation and survival assay in vitro. Transmission electron microscope (TEM) was detecting endoplasmic reticulum (ER) morphology. ER stress maker expression was estimated by Western blotting. Metastatic tumor growth was determined in mouse model of intraperitoneal or intravenous injection with B16F10. In vivo tumor growth was detected by positron emission tomography/ computed tomography (PET/CT) and exterior photography of peritoneal dissection. Results: PAE-induced B16F10 and A375 cell senescence were determined by increased cell size, flattened morphology, DNA damage as well as the increased expression of senescence associated β-galactosidase. PAE inhibited cell proliferation in melanoma cells in MTT and colony formation assay. ERK5/ P21, the major cell cycle regulators and mediators of senescence, were up-regulated at the protein level in PAE-treated melanoma cell. Further studies demonstrated that PAE induced cell senescence via ER stress and Calpain1/ ERK5/ P21.