In vitro and in vivo anti-cancer effects of hibernating common carp ( Cyprinus carpio ) plasma on metastatic triple-negative breast cancer

Uncontrollable proliferation is a hallmark of cancer cells. Cell proliferation and migration are significantly depressed during hibernation state. Many studies believe some factors in the plasma of hibernating animals cause these effects. This study aimed to assess the anti-cancer effects of hibernating common carp ( Cyprinus carpio ) plasma on 4T1 cancer cells in vitro and in vivo. The effect of hibernating plasma on cell viability, morphology, migration, apoptosis rate, and cell cycle distribution of 4T1 cells was investigated in vitro and in vivo. Hibernating plasma at a concentration of 16 mg/ml significantly reduced the viability of 4T1 cancer cells, without any toxicity on L929 normal fibroblast cells. It could change the morphology of cancer cells, induced apoptosis and cell cycle arrest at the G2/M phase, and inhibited migration. Furthermore, intratumoral injection of hibernating plasma (200 µl, 16 mg/ml) in the tumor-bearing mice caused a significant inhibition of 4T1 breast tumors volume (46.9%) and weight (58.8%) compared with controls. A significant decrease in the number of metastatic colonies at the lungs (80%) and liver (52.8%) of hibernating plasma-treated animals was detected which increased the survival time (21.9%) compared to the control groups. Immunohistochemical analysis revealed a considerable reduction in the Ki-67-positive cells in the tumor section of the hibernating plasma-treated animals compared with controls. Taken together, the SDS-PAGE and mass spectrometry analysis indicated the alpha-2-macroglobulin level in the hibernating fish plasma was significantly increased. It could exert an anti-cancer effect on breast cancer cells and suggested as a novel cancer treatment strategy.