Isolation, Purification, and Antitumor Activity of a Novel Active Protein from Antrodia cinnamomea Liquid Fermentation Mycelia

Antrodia cinnamomea, a rare medicinal fungus endemic to Taiwan, contains numerous active components and displays strong antitumor and anti-inflammatory effects. We isolated and purified a novel A. cinnamomea active protein (termed ACAP) from liquid fermentation mycelia and evaluated its antitumor activity. A homogeneous protein-eluted fraction was obtained by anion exchange chromatography and gel filtration chromatography, and ACAP was identified based on the antitumor activity screening of this fraction. An in vitro assay of three tumor cell lines (HeLa, Hep G2, and Hepa 1-6) revealed significant antiproliferative effects of ACAP at low concentrations, with IC50 values of 13.10, 10.70, and 18.69 mu g/mL, respectively. Flow cytometric analysis showed that ACAP induced late apoptosis of Hep G2 cells. The apoptosis rate of 50 mu g/mL ACAP-treated cells (60%) was significantly (p < 0.01) more than that of the control. A Western blotting assay of apoptotic pathway proteins showed that ACAP significantly upregulated p53 and downregulated caspase-3 expression levels. Our findings indicate that ACAP has strong antitumor activity and the potential for development as a therapeutic agent and/or functional food.