Apoptosis inducing effect of peptide/protein extracts from lentinus squarrosulus mont. In human h460 lung cancer cells

Abstract

In searching for the novel compounds with improving efficacy and safety profile for the treatment of lung cancer, anti-cancer activity and underlying mechanism of proteins extracted from Thai edible mushroom, Lentinus squarrosulus Mont. were firstly presented in this study. Proteins from fruiting body of L. squarrosulus Mont. were isolated by precipitation with solid ammonium sulfate at 40-80% (W/V) final concentration and further purified by ion-exchange chromatography on Diethylaminoethyl (DEAE)-cellulose, and gel filtration chromatography on Sephadex G-50 or membrane filtration, respectively. The unbound protein (F1 fraction) and the bound protein eluted with 0.3 M NaCl (F4 fraction) from ion-exchange chromatography showed the highest toxicity to human lung cancer cells with 50% inhibitory concentration (IC50) approximately 22.53 ± 0.22 µg/ml and 21.15 ± 6.14 µg/ml, respectively. The attempt to further purify to homogeneity did not succeed. So proteins from F1 and F4 fractions were used for further studies. The proteins from F4 fraction induced apoptosis cell death through the reduction of anti-apoptosis proteins including Bcl-2 and Mcl-1 which regulated mitochondrial membrane as well as the reduction of c-FLIP, an inhibitor of death receptor pathway. Meanwhile, the induction of apoptosis and the alteration of apoptosis-regulating proteins were not obviously shown in lung cancer cells incubated with proteins from F1 fraction. Although 25 µg/ml of protein from F1 and F4 fraction generated oxidative stress in human lung cancer cells, their anti-cancer activity was reactive oxygen species (ROS) independent. In summary, this study provided the novel information regarding anti-cancer activity and mechanism of proteins from fruiting body of L. squarrosulus Mont. in lung cancer cells which will be beneficial for further development as anti-cancer agent.