Ganoderma lucidum: A Review with Special Emphasis on the Treatment of Various Cancer

Ganoderma lucidum, commonly referred to as Lingzhi or Reishi, is a basidiomycete rot fungus which has been used for centuries in East Asia for promotion of good health and longevity. The main bioactive components of G. lucidum can be broadly grouped into polysaccharides and triterpenes. The anticancer properties of G. lucidum have been proved in both in vitro and in vivo studies using human and murine cell lines. Various pharmacological activities have been reported such as, hepatoprotective, anti-diabetic, anti-hypertensive, cardioprotective, immune modulatory, antioxidant, anticancer, etc. Regardless of polysaccharides and triterpenes have been used for treatment of different types of cancers the mechanism by which they exert their anticancer effect remains undefined. The aim of this paper is to summarise the treatment of various cancer with respect to various mechanisms that have been suggested for the anticancer properties of polysaccharides and triterpenes extracted from G. lucidum.


Introduction
Ganoderma lucidum is a basidiomycete white rot fungus [1] that belongs to the family Polyporaceae (or Ganodermataceae) of Aphyllophorales [2]. It is commonly known as ''Lingzhi'' in Chinese, ''Reishi'' in Japanese and ''Youngzhi'' in Korean [3]. Its medicinal values has been documented in the Chinese literature which can be dated back nearly two thousand years to the Shen Nong Materia Medica (102-200AD). It is regarded as a symbol of happiness, good fortune, good health and even immortality in Chinese traditional culture [4].
More than 120 species of Ganoderma have been reported in the world out of which 98 species were found in China. However, only two species of Ganoderma (Figure 1) i.e., G. lucidum (Leyss.ex Fr.) Karst. and Ganoderma sinense Zhao, Xu et Zhang, are documented in Chinese Pharmacopoeia (2010) as Lingzhi [5]. The fungus is commonly used in East Asian countries for the promotion of health and longevity and as a remedy for illness. Ganoderma is consumed for its medicinal value rather than nutritional value. Lingzhi has been reckoned to extend the life span and to increase youthful vigour and vitality [6].
In this article, we laid emphasis on the treatment of various cancers with G. lucidum as well the bioactive pathways which might be associated with the anticancer activity.

Effect of G. lucidum on human colorectal cancer cells
Zengenni et al. reported that cell viability on HCT-116 cells was reduced by GLP in a time-and dose-dependent manner which in turn induced cell apoptosis. Apoptosis was characterized by morphological changes, DNA fragmentation, mitochondrial membrane potential decrease, S phase population increase, and caspase-3 and -9 activation. GLP-induced apoptosis was further decreased by inhibition of c-Jun N-terminal kinase (JNK) by SP600125. Western blot analysis revealed that GLP influenced the expression of Bax/Bcl-2, caspase-3 and poly (ADP-ribose) polymerase (PARP). It has been reported that activation of mitochondrial and mitogen-activated protein kinase (MAK) pathways proved the apoptosis stimulated by GLP in human colorectal cancer cells [40].

Effect of G. lucidum on mouse hepatoma, sarcoma S-180 and reticulocyte sarcoma L-II cells
Xin Liu et al. showed the inhibitory effects of dormant spores, the germinating spores, the sporoderm-broken germinating spores (SBGS) and the lipids extracted from the germinating spores of G. lucidum on the growth of mouse hepatoma, sarcoma S-180 and reticulocyte sarcoma L-II cells. The sporoderm-broken spores indicated much higher bioactivities than the whole spores. The bioactivities of the spores were enhanced by germinating the dormant spores. The lipids extracted from germinating spores and the sporoderm-broken germinating spores of G. lucidum inhibited three tumors in dosedependent manner with an inhibition of 80-90% [28].

Effect of G. lucidum on Lewis lung carcinoma bearing mice
Shiu-Nan et al. showed the effect of mushroom β-glucans (MBGS) by analyzing size of primary tumor and rate of metastasis in Lewis lung carcinoma (LLC) bearing mice (C57BL/6). MBGS was derived from solid culture of Ganodermalucidum and was administered orally along with radiation therapy. MBGS enhances NK cell-mediated cytotoxicity in mice without LLC bearing mice. When MBGS is administered in conjugation with radiation therapy it serves as a protective factor for the hair loss and wounds due to the overgrowth of primary tumor in LLC bearing mice. It is also effective in controlling tumor growth and rate of metastasis [41].

Effect of G. lucidum on colitis associated carcinogenesis in mice
Daniel et al. demonstrated anticancer and anti-inflammatory activity of triterpene extract isolated from G. lucidum. It was reported that mice exposed to PhIP/DSS was treated with G. lucidum triterpene (GLT) which suppressed focal hyperplasia, aberrant crypt foci (ACF) formation and tumor formation in mice. Further decreased staining with Ki-67 in colon tissues confirmed the anti-proliferative effect of GLT. PhIP/DSS-induced colon inflammation in mice was proved by shortening of the large intestine and macrophage infiltrations. GLT treatment forbade the shortening of colon length and reduced macrophage infiltration. It also decreased PhIP/DSS-dependent expression of cyclin D1, COX-2, CYP1A2 and CYP3A4 in colon tissues [38].

Effect of G. lucidum on inflammatory breast cancer
Ivette et al. showed the mechanism of G. lucidum concentrating on the phosphoinositide-3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) pathway. Early treatment of Inflammatory breast cancer (IBC SUM-149) with Reishi reduces expression of mTOR, reduced eIF4G level coupled with increased levels of eIF4E bound to 4E-BP as well as consequential protein synthesis reduction. Reishi treated severe combined immunodeficient mice injected with IBC cells for 13 weeks showed reduction in tumor growth and weight by 50%. It also showed reduction in the expression of E-cadherin, mTOR, eIF4G and p70S6K and activity of extracellular regulated kinase (ERK1/2). Thus it is evidenced that Reishi suppresses protein synthesis and tumor growth by affecting survival and proliferative signalling pathways [42].
transcriptional activity in breast cancer MDA-kb2 cells. It also suppressed androgen receptor (AR) protein level in LNCaP and MDA-kb2 cells in a dose-dependent manner [43]. Sun et al. reported the effect SCGLP1 on human osteosarcoma MG63 cell line. The treatment with SCGLP1 showed inhibitory effect on cell proliferation and cell viability of MG63 cells in a dosedependent manner. It also caused apoptotic death in MG63 cells through an increase in G0/G1 phase arrest. SCGLP1 induced apoptosis was related with protein expression of pro-apoptotic Bax and Bad, decreased expression of anti-apoptotic Bcl-2 and Bcl-XL, loss of mitochondrial membrane potential, the release of mitochondrial cytochrome C to cytosol, and cleavage of caspase-9, caspase-3, and poly (ADP ribose) polymerase (PARP). Pre-treatment with pancaspase inhibitor had blocked SCGLP1 induced apoptosis in MG63 cells. Hence it was suggested that SCGLP1 induced apoptosis was related to caspase-3 and caspase-9-dependent apoptotic pathway [44]. Tong et al. reported that morphological changes and apoptosis were observed in J558 cells when treated with G. lucidum extract at a dose of 150 µg/mL. But when the dose was increased to 200 and 400 µg/mL there was no significant reduction in cell viability. Necrosis occurred which was characterized by small fragments with uniformly stained red nuclei when the dose was increased to 400 µg/mL. After treatment the viable cells decreased by 45.6% whereas the apoptotic and necrotic cells increased by 16.5 and 29.1% respectively. But there was no changes observed in 3T3 cells. Thus it was confirmed various necrotic and apoptotic changes of cells by scanning electron microscopy and transmission electron microscopy [45]. Wu et al. reported that miR-378 cells were transfected into tumor cells which acquired aggressive properties of cancer cells. Enhanced cell survival and colony formation was observed due to over expression of miR-378 cells which led to multiple drug resistance. To accelerate the death of miR-378-transfected cells higher concentration of chemotherapeutic agents were required rather than control cells. The active ingredient from G. lucidum was purified and isolated as ergosterol peroxide which increased the death of miR-378 cells than GFP cells. Thus from here it was shown that lower concentration of ergosterol peroxide was required to enhance death of miR-378transfected cells compared to chemotherapeutic agents. Thus it serves as a promising new agent which can overcome the resistance of chemotherapeutic agents in cancer cells [46]. It leads to inhibition of carcinogen 4-aminobiphenyl induced migration in HUC-PC and MTC-11 cells. The expression of matrix metalloproteinase-2 and focal adhesion kinase were unchanged which suggests that other mechanism may be involved [47]. Sun et al. reported that plasma-induced suppression of lymphocytes in lung cancer patients can be treated with G. lucidum polysaccharides (Gl-PS). Various immunosuppressive mediators such as, PGE2, TGF-β, IL-10 and VEGF are released by cancer cells to inhibit the immune response. Gl-PS antagonises the immune inhibition to facilitate tumor control in animal model. Hence, with the treatment of Gl-PS it was possible to supress proliferation, CD69 expression, and perforin and granzyme B production in lymphocytes which was activated by Phytohemagglutinin (PHA) in the plasma of lung cancer patients. An observation was made that Gl-PS can fully or partially reverse those effects [48]. Oliveira et al. reported the inhibition of the growth of a gastric cancer cell line (AGS) by interfering with cellular autophagy and cell cycle. G. lucidum extract was found to possess antitumor activity. The phytochemical constituent of the various extract of G. lucidum from fruiting body and spores were investigated [49].

Current Scenario and Future perspectives
G. lucidum has been used all over the world for centuries as a source of health food supplements and nutraceuticals. It has various pharmacological benefits, such as, hepatoprotective, ant diabetic, antimicrobial, neuroprotective, antihypertensive, cardioprotective, anti-oxidant, anti-HIV-1 activity, immune-modulating, anticancer, etc. The effectiveness of G. lucidum reckons mainly on its chemical constituents, namely, polysaccharides and triterpenes that make up the fruiting body, mycelium or spores.
Polysaccharides have been shown to activate immune effector cells and supress the growth of several cancer cells in vivo, enhance the host's immune response by stimulating the production of macrophages, NK cells and T-lymphocytes, etc. It precludes tumor metastasis by various mechanisms such as inhibition of c-Jun Nterminal kinase (JNK) by SP600125, inhibition of VEGF expression, reduced expression of mTOR, reduced eIF4G level, suppressed androgen receptor (AR) protein level; inhibit immunosuppressive mediators such as, PGE2, TGF-β, IL-10, etc.
Triterpenes have shown to inhibit eukaryotic DNA polymerases, thromboxane A2-signaling pathways in human platelets, inhibition of tumor invasion in vitro and in vivo, cytotoxicity to several cancer cells in vitro, anti-human immunodeficiency virus-1 protease activity and regulation of osteoclastogenesis. It also prevented tumor metastasis by inhibiting the expression of cyclin D1, COX-2, CYP1A2 and CYP3A4, regulating MMp and IL-8, suppressed inflammatory cytokine secretion in macrophage cells, down regulation of cyclin D1, etc.