Ganoderma Lucidum inhibits cancer cell growth and enhances natural killer cell cytotoxicity against a panel of blood and solid tumors

Taha, Hashem 1 ; Saleh, Amr 3 ; Wang, Xinghua 2 ; Keating, Armand 1, 2;

 1.  Institute of Biomaterials and Biomedical Engineering, University of Toronto;

 2.  Krembil Research Institute, Toronto Western Hospital;

 3.  Department of Biochemistry, McMaster University;

 Background: G. lucidum can inhibit cancer cell proliferation, induce apoptosis in different human tumor cell lines in vitro, and stimulate multiple system components. NK cell lines such as NK-92 and its genetically modified variants and KHYG-1 are appealing options as scalable allogeneic immunotherapies. The objective of our research is to assess the direct anticancer toxicity of G.lucidum extracts and their ability to enhance the cytotoxicity of natural killer cell lines (NK-92, KHYG-1) against a panel of blood and solid tumors in vitro.

Methods: The anticancer potency of three G. Lucidum extracts (aqueous, methanol, and ethyl acetate) was determined with administered doses of 0.1, 1, 10, 100 ug/mL against MCF 7 (breast cancer), SKOV3 (ovarian cancer), OCI AML5 (acute myeloid leukemia), K562 (chronic myelogenous leukemia). Targets were incubated for 48 hours to compare potency of extracts with published literature values. Cancer cell growth was evaluated with the MTT assay to assess cell metabolic activity and viability. Chromium release assay was used to investigate the immune enhancing function of aqueous G. Lucidum extract and an enhanced Coriolus Versicolor extract on NK-92 and KHYG-1 cytotoxicity against MCF-7, SKOV3, K562, KG1, MEC-1 RPMI 8226. Immune effectors were pretreated with mushroom extracts at 0.1, 1, 10, 100 ug/mL and incubated for 24 hours before co-incubation with cancer targets at a 5:1 effector to target ratio.

Results: G. Lucidum ethyl acetate extract completely inhibited the growth of all tested cancer targets at 100 ug/mL. The ethyl acetate fraction had maximal inhibitory effects that were observed at concentrations much lower than published IC50 values for different G. Lucidum formulations.  (Lingzhi Ethyl Acetate IC50 (MCF7) = 4-13 ug/mL; Literature IC50 range (MCF7) = 193-248 ug/mL, IC50 (SKOV3) = 650-1000 ug/mL, IC50 (leukemia) = 26-63 ug/mL).

Pre-incubation of NK-92 and KHYG-1 with the enhanced C. Versicolor extract results in a significant enhancement of cytotoxicity against all cancer lines. A 45 % increase in KHYG-1 cytotoxicity was observed against the multiple myeloma line RPMI8226 with 10 ug/mL [p<0.001 versus untreated control KHYG-1].

Pretreatment with 10 ug/mL aqueous G.Lucidum extract significantly enhanced NK-92 cytotoxicity against cancer targets with the following average killing percentage increase relative to untreated NK-92: [KG1: 12 +/- 1 %, K562: 15 +/- 2 %, MCF-7 15 +/- 2 %, MEC-1 21 +/-6 %, RPMI 25 +/- 2 %] [p<0.01at 10ug/mL].

Significance: These studies are the first to probe the anticancer effects of previously untested G. Lucidum fractions to get deeper insight about the biological activity of certain compounds present in the most potent extracts. Our research is the first preclinical study to explore the immunomodulatory effects of different mushroom extracts on NK cell lines.