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Pharmacological Values of Medicinal Mushrooms

for Prostate Cancer Therapy: The Case of
Ganoderma Lucidum

Article in Nutrition and Cancer February 2009

Impact Factor: 2.32 DOI: 10.1080/01635580802379323 Source: PubMed

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Nutrition and Cancer, 61(1), 1626
Copyright 2009, Taylor & Francis Group, LLC
ISSN: 0163-5581 print / 1532-7914 online
DOI: 10.1080/01635580802379323

Pharmacological Values of Medicinal Mushrooms for

Prostate Cancer Therapy: The Case of Ganoderma Lucidum

Jamal Mahajna
Cancer Drug Discovery Program, Migal-Galilee Technology Center, Kiryat Shmona, Israel

Nesly Dotan and Ben-Zion Zaidman

Cancer Drug Discovery Program, Migal-Galilee Technology Center, Kiryat Shmona, Israel and the
Institute of Evolution and Department of Evolutionary and Environmental Biology, Faculty of Science
and Science Education, University of Haifa, Israel

Roumyana D. Petrova
Institute of Botany, Bulgarian Academy of Sciences, Sofia, Bulgaria

Solomon P. Wasser
Institute of Evolution and Department of Evolutionary and Environmental Biology, Faculty of Science
and Science Education, University of Haifa, Israel

signal transduction pathways or molecular targets implicated in

Prostate cancer (PCa) is the most common male malignancy in
many Western countries. Primary PCa is hormone dependent and
is manageable by hormonal therapy. However, it rapidly develops
to hormone-refractory tumors due to the accumulation of muta-
tions in the androgen receptor and/or the acquisition of alternative
cellular pathways that support proliferation and inhibit apopto-
sis of prostate cancer. To date, no effective therapy is available
for clinically hormone-insensitive or hormone-refractory stages of
prostate cancer.
Whereas prostate cancer is very common in Western coun-
tries, its levels are very low in Asia, providing evidence for
a potential link between diet, environmental factors, and can-
cer incident. Natural products have been used as a source of
new pharmaceuticals including anticancer drugs. The medici-
nal properties of mushrooms have been well known in Eastern
Asia for thousands of years. Of special interest is the impli-
cation of several mushrooms in prostate cancer prevention in-
cluding the popular mushroom Ganoderma lucidum (Ling Zhi
or Reishi) that has been widely used for the general promotion
of health and longevity in Asia. Dried powder of G. lucidum
was popular as a cancer chemotherapy agent in ancient China.
The pharmaceutical activities of G. lucidum substances targeting
Submitted 27 November 2007; accepted in final form 13 April 2008.
Address correspondence to Jamal Mahajna, Cancer Drug Dis-
covery Program, Migal, P.O. Box 831, Kiryat Shmona 11016,
Israel. Phone: +972-4-6953537. Fax: +972-4-6944980. E-mail:
jamalm@migal.org.il
prostate carcinogenesis are reviewed.
OVERVIEW
For thousands of years, natural products have played an im-
portant role throughout the world in treating and preventing hu-
man diseases. Natural product medicines have originated from
various source materials including plants, fungi, microorgan-
isms, and animals. Such natural bioactive substances possess
an enormous structural and chemical diversity, unsurpassable
by any synthetic library; they are evolutionally optimized as
drug-like molecules and might be considered biologically val-
idated. Moreover, these molecules can serve as templates for
semisynthetic and fully synthetic modifications.
An analysis of the origin of drugs developed between 1981
and 2002 showed that natural products or natural product-
derived drugs comprised 28% of all novel chemical entities
(NCEs) launched onto the market (1). In addition, 24% of these
NCEs were synthetic or natural mimic compounds based on a
study of pharmacophores related to natural products. This com-
bined percentage (52% of all NCEs) suggests that natural prod-
ucts are important sources for new drugs and are also good lead
compounds suitable for further modification during drug devel-
opment (2). In the case of anticancer agents, natural products
have made significant contributions as either direct treatments
or templates for synthetic modification. In this category, there
are some 140 anticancer agents available to Western countries
and Japan; 62% of them are nonsynthetic agents.
It is estimated that there are approximately 1.5 million species
of fungi in the world, of which approximately 82,000 are

16
 MEDICINAL MUSHROOMS FOR PROSTATE CANCER THERAPY
described (3). About 20,000 of the known species belong to
macrofungi, of which about 5,000 are edible and over 2,000
safe (4). Fungi from the Basidiomycota division are of great in-
terest due to the large number of biologically active compounds
they contain (513). Fungal fruiting bodies, mycelium, or the
culture broth in which the mycelium has been cultivated are all
explored for biological activities. Consequently, approximately
650 species of higher Basidiomycetes have been found to pos-
sess antitumor activities (7).
Prostate Cancer
Prostate cancer is the most common male malignancy in
many Western countries and the third leading cause of can-
cer deaths in men worldwide. The age-standardized incidence
rate of prostate cancer is highest in the United States (137 per
100,000 in Black men), lower in European countries (28 and 31
per 100,000 in England and Denmark, respectively), and lowest
in Asia (10 and 2.3 per 100,000 in Japan and China, respectively)
(15).
The epithelium of the prostate gland is under the hormonal
control of androgens, the production of which depends on the
hypothalamic-pituitary-testicular axis. The Leydig cells of the
testes produce 95% of total androgens, and the adrenal glands
produce the remaining 5%. In the prostate, free testosterone
diffuses directly into the epithelial or stromal cells where it is
converted into the functionally active androgen, dihydrotestos-
terone (DHT), by the action of 5- reductase enzyme system
located on the nuclear membrane. Dihydrotestosterone action is
mediated by the androgen receptor (AR) (16), which functions
to preserve the normal function and structure of the prostate. The
androgen receptor is a structurally conserved member of the nu-
clear receptor superfamily of ligand-activated transcription fac-
tors. Androgen ablation therapy has been shown to produce the
most beneficial responses in patients with hormone-responsive
prostate cancer (19). Castration, either surgically or chemically,
remains the standard treatment option for most patients. Com-
bined with antiandrogens that interfere with androgen receptor
function, these methods significantly prolong the survival of
prostate cancer patients (20). Androgen deprivation therapy is
initially effective in repressing primary prostate tumors. How-
ever, progression to the clinically hormone-insensitive stage is
usually inevitable. To date, no effective therapy is available for
hormone-insensitive or hormone-refractory stages of prostate
cancer (21).
Prostate cancer has a complex etiology; currently age, ethnic-
ity, obesity, and family history are the most consistently reported
risk factors associated with the disease. Other potential risk fac-
tors, such as environmental (17) and dietary (18) factors, have
also been suggested. Conversion of a normal cell into the malig-
nant state is often linked to genetic alterations of the cell (22,23).
The known tumor-suppressor genes, Retinoblastoma (Rb) and
p53, were reported to play an important role in the progression
of prostate cancer (24). Both genes appear to be early events in
17
prostatic carcinogenesis (25,26). In addition, mutations or dele-
tions of pTEN, a tumor suppressor gene, have been found in 30%
of primary prostate cancers and 63% of malignant cases, rank-
ing it as one of the most common determinants of prostate tumor
progression (27,28,57). The abnormal expression of growth fac-
tors and their receptors including the epidermal growth factor
(EGF) (29), the transforming growth factor- (TGF-) (30), the
transforming growth factor- (TGF-) (31), HER-2/neu, and
c-erbB-3 oncogenes (32) may also contribute to the growth and
development of both local and metastatic prostate cancer. The
progression of prostate tumors to a hormone-refractory state
is frequently associated with the increased expression of the
antiapoptotic gene Bcl-2 (33), and the mutation of pTEN (34).
Androgen Receptor in Prostate Cancer
Like all steroid hormone receptors, the androgen receptor
consists of 4 distinct regions in the expressed receptor that
have specific functions (35), including the DNA-binding do-
main, which is responsible for the interaction with hormone-
responsive elements of the target gene promoters (36), and the
ligand binding region, which is located at the carboxyl termi-
nus of the receptor and contains the activation function 2 (AF2)
domain and regulates ligand-dependent receptor function. Fur-
thermore, the ligand-binding domain is thought to be important
in the function of coactivator proteins (37) and in regulating
transcription (38). The androgen receptor remains cytoplasmic
until ligand binding occurs, and the dissociation of heat-shock
proteins (HSPs) is thought to allow conformational change in
the androgen receptor and mediate translocation to the nucleus.
In the absence of ligand, the androgen receptor in the cytoplasm
is rapidly degraded. In the presence of ligand, the androgen re-
ceptor dimerizes, translocates to the nucleus, and initiates gene
transcription by binding to specific androgen responsive ele-
ments (AREs) found in androgen-responsive promoters (39).
During androgen-independent progression, prostate cancer re-
lies on various cellular pathways, some involving the androgen
receptor and others bypassing it. In the former type of path-
ways, a mutated androgen receptor may be activated by various
ligands. In addition, deregulated growth factors and cytokines
can activate the androgen receptor, usually with the help of
androgen-receptor coactivators. Most of these growth factors,
including EGF, insulin-like growth factor (IGF) and fibroblast
growth factor (FGF), are potent mitogens and are upregulated
by androgens, although the exact mechanisms are unknown. In
contrast, TGF- activity is downregulated by androgens (49).
In the pathways that bypass the androgen receptor, the loss of
pTEN reverses the inhibition of the PI3K/Akt pathway, permit-
ting activated Akt to phosphorylate Bad. This activation results
in the release of Bcl-2, which eventually leads to cell survival.
In addition, androgen-independent cells may overexpress Bcl-
2 (50). Recently, increasing evidence implicated the canoni-
cal Wnt signaling pathway in modulating the androgen signal-
ing at multiple levels (51,52). -catenin protein, a particularly
18 J. MAHAJNA ET AL.
critical molecular component of canonical Wnt signaling, is now
known to promote androgen signaling through its ability to bind
to the androgen receptor protein in a ligand-dependent fash-
ion and to enhance the ability of liganded androgen receptor to
activate transcription of androgen-regulated genes (53). Further-
more, other components of the Wnt signaling pathway also aug-
ment androgen receptor function through diverse mechanisms
(5155).
Historical and Contemporary Uses of Medicinal
Mushrooms
The medicinal use of fungi dates back thousands of years
and is recorded in the histories of both traditional Western
medicine (TWM) and traditional Chinese medicine (TCM) (72).
Although fungi have often been marginalized in traditional
Western medicine and have not been widely incorporated into
Western diets, the use of fungi has been central to the cultures
of most Asian countries since antiquity (73). In traditional Chi-
nese medicine, fungi are used in their entirety, fresh or dried,
to treat the patient (and the patients ailments) as a whole (72).
In addition, certain fungi have been used in both traditional
Western medicine and in traditional Chinese medicine to target
specific conditions. Such is the case with Fomitopsis officinalis
(Vill.) Bond. et Singer, called Agaricum by Dioscorides in his
collection De Materia Medica (60 A.D.), where it is described
as a powerful panacea, especially suited to treat intestinal ail-
ments (73). Other medicinal fungi are described in the Herbal
Classic of traditional Chinese medicine, a compilation of two
ancient bodies of writing collected between 200264 A.D., the
Sien nung Pen tsao king (or Pen king) and Ming I pie lu (or
Pie lu) (73). A number of ancient medicinal fungi discussed
are still in use. These include Ganderma lucidum, Poria cocos
(Schwein.) F.A. Wolf, Grifola umbellata (Pers.) Pilat, Calvatia
lilacina (Berk et Mont.) Lloyd, and Tremella fuciformis Berk.
Special attention is given to chi or Ling chi (zhi) varieties of
G. lucidum, said to promote well-being and immortality (73). In
Asia, this fungus has traditionally been used to treat age-related
ailments such as coronary disease, hypertension, bronchial prob-
lems, and cancer (74).
Fungal Secondary Metabolites
Secondary metabolite production in fungi is a complex pro-
cess coupled with morphological development (75). In most
cases, the function of secondary metabolites for producing fun-
gus is unknown but is inferred from several studies using mu-
tants or enzyme inhibitors. These substances have their origins
as derivatives from many intermediates in primary metabolism,
but most can be classified according to 5 main metabolic sources:
1) amino acid-derived pathways, 2) the shikimic acid pathway
for the biosynthesis of aromatic amino acids, 3) the acetate-
malonate pathway from acetyl coenzyme A, 4) the mevalonic
acid pathway from acetyl coenzyme A that functions in primary
metabolism for the synthesis of sterols, and 5) polysaccharides
and peptidopolysaccharides. The polyketide and mevalonic acid
pathways are most often involved, and they produce a greater
variety of compounds than the other pathways (11). Some of
these compounds have tremendous importance to humankind in
that they display a broad range of useful antibacterial, antivi-
ral, and pharmaceutical activities as well as less desirable toxic
activities.
Two major groups of secondary metabolites are responsible
for toxic activities (the division is rather arbitrary): mycotoxins
and mushroom poisons (76). All mycotoxins are low-molecular-
weight natural products (i.e., small molecules) produced as sec-
ondary metabolites by filamentous microfungi, whereas mush-
rooms and other macroscopic fungi produce mushroom poisons.
Depending on the definition used, and recognizing that most fun-
gal toxins occur in families of chemically related metabolites,
some 300 to 400 compounds are now recognized as mycotoxins
(77). The second group of toxic metabolites is mushroom poi-
sons. About 300 species of mushrooms are poisonous to humans.
These species produce a wide spectrum of poisons that have been
divided into the following 7 main categories (in brackets, an ex-
ample of producing fungi): amanitin (Amanita phalloides [Vaill.
: Fr.] Link, Galerina autumnalis [Peck] A.H. Sm. et Singer),
orellanine (Cortinarius orellanus Fr.), gyromitrin (Gyromitra
esculenta [Pers.] Fr.), muscarine (Clitocybe dealbata [Sowerby]
Gillet), ibotenic acid (Amanita cothurnata G.F. Atk.), psilo-
cybin (Psilocybe baeocystis Singer et A.H. Sm., Panaeolus
castaneifolius [Murrill] A.H. Sm., Conocybe cyanopus [G.F.
Atk.] Kuhner) and coprine (Coprinus atramentarius [Bull.] Fr.)
(78).
Among the fungal secondary metabolites are lectins, lac-
tones, terpenoids, alkaloids, antibiotics, and metal chelating
agents (6). Fungi also contain a number of enzymes such as lac-
case, superoxide dismutase, glucose oxidase, and peroxidases.
It has been shown that such an enzyme therapy can also play an
important role in cancer treatment preventing oxidative stress
and inhibiting cell growth (79).
Anticancer Activity of Fungal Substances
It has been demonstrated that fungal metabolites can be
used as inhibitors of molecular targets in malignant cells in
order to combat certain cancers. Fungal anticancer substances
can be roughly divided into two groups of high- and low-
molecular-weight molecules. The major difference between
these two groups is their mechanism of action. Most of the high-
molecular-weight compounds are polysaccharides or protein-
bound polysaccharides (80). It appears that these compounds
are capable of interacting nonspecifically with the immune sys-
tem to upregulate or downregulate many aspects of the host re-
sponse (81). The second group comprises low-molecular-weight
secondary compounds that can penetrate the cell membrane and
act on specific signal-transduction pathways (11). These include
mainly sesquiterpenes (which are the predominant secondary
metabolites of Basidiomycetes), triterpenes, steroids and sterols,
 MEDICINAL MUSHROOMS FOR PROSTATE CANCER THERAPY
and a few polyketides (abundantly produced by Actinomycetes).
Anticancer Activity of Ganoderma Lucidum
Ganoderma is the most popular and intensely investigated
genus among the medically active mushrooms. Plenty of its
species are famous for their antiviral, antibacterial, antifun-
gal, anticancer, and immunostimulating activities and have been
used traditionally in the folk medicine of Eastern countries for
centuries. These activities were due to the production of various
metabolites such as proteins, terpenes, sterols, and so forth.
The genus Ganoderma belongs to the class of Hymeno-
mycetes. Within the genus Ganoderma, over 250 taxonomic
names have been reported worldwide (82) including G. adsper-
sum, G. applanatum, G. australe, G. lucidum, and G. tsugae, to
name a few. However, the majority of reports in the literature
appear to refer to one species, G. lucidum (83).
Ganoderma lucidum (W. Curti: Fr.) P. Karst. (Ling Zhi or
Reishi), an oriental medical mushroom, has been used widely in
Asian countries for centuries to prevent or treat different diseases
including cancer (Fig. 1). Dried powder of G. lucidum, which
was recommended as a cancer chemotherapy agent, is currently
used popularly worldwide in the form of dietary supplements.
G. lucidum extracts were reported to possess cytotoxic ac-
tivity against various cancer cell lines including leukemia,
lymphoma, multiple myeloma (84,112), human hepatoma
PLC/PRF/5 and KB, human breast cancer MDA-MB-231 (85),
human prostate cancer PC-3 (86), human breast cancer MCF-
7 (87), human cervix uteri tumor HeLa (88), and low-grade
bladder cancer MTC-11 (89) cell lines. The cytotoxic effects of
FIG. 1. Ganoderma lucidum fruit bodies.
19
G. lucidum as demonstrated by the studies of Jiang et al. (85,86)
and Zhu et al. (88) were concentration dependent. This activity
of G. lucidum can be attributed directly to specific compounds
from experiments employing isolated and purified molecules.
However, the molecular mechanism(s) responsible for the in-
hibitory effects have not been fully elucidated.
Ganoderma Lucidum Inhibits Proliferation of Prostate
Cancer Cells
Proliferation is the multiplication or reproduction of cells
resulting in the rapid expansion of a cell population. Cell pro-
liferation is controlled by cell cycle regulatory elements.
Hsieh and Wu (92) tested the ability of extracts from indi-
vidual herbs containing the herbal mixture PC-SPES, of which
Ganoderma lucidum is one of its components, using amounts
estimated to be equivalent to that present in the herbal mixture
to suppress LNCaP, an androgen-dependent prostate cancer cell
line, growth and/or lower prostate-specific androgen (PSA) ex-
pression, compared to cells treated with PC-SPES. Treatment of
LNCaP cells with 5 microl/ml ethanol extracts of Ganoderma
lucidum showed a 63.5% reduction in cell growth and exhibited
a similar decrease in cell viability. Additional studies demon-
strated the ability of Ganoderma lucidum extracts to also inhibit
cell proliferation of AR-independent cancer cell lines such as
PC-3 in a dose- and time-dependent manner (86). Growth in-
hibition of PC-3 cells by Ganoderma lucidum was mediated
by the downregulation of expression of cyclin B and Cdc2 and
by the upregulation of p21 expression. The inhibition of cell
growth was also demonstrated by cell cycle arrest at the G2/M
20 J. MAHAJNA ET AL.
phase (86). Liu et al. (62) reported that the LNCaP growth in-
hibitory activity of G. lucidum extract is mediated by Ganoderol
B isolated from G. lucidum fruiting body extract. Liu et al. (62)
reported that Ganoderol B inhibits 5- reductase activity and
thereby causes inhibition of the proliferation of the androgen-
dependent LNCaP cell line.
Ganoderma Lucidum Induces Apoptosis in Prostate
Cancer Cells
Apoptosis is the predominant mechanism by which can-
cer cells die when subjected to chemotherapy or irradiation.
However, cancer cells develop resistance to these therapies that
may be due, at least in part, to the development of effective
antiapoptotic mechanisms (103). Another mechanism allow-
ing escape from apoptosis is the activation of survival signal
transduction pathways, including Akt-dependent (104) and Akt-
independent mechanisms (105). An Akt-independent example
includes EGF-induced survival mechanisms (31). When LNCaP
cells are treated with PI3K inhibitors and deprived of survival
factors, they spontaneously undergo apoptosis. However, treat-
ment with EGF or androgen can protect cells from apoptosis,
although Akt activity remains inhibited. It was found that EGF
can protect LNCaP cells from apoptosis induced via the mi-
tochondrial pathway but not from apoptosis induced via the
death-receptor pathway (106,107).
A large portion of prostate cancer cells contain deregulated
Akt. For example, in LNCaP prostate cancer cell line, Akt is
constitutively active as a result of a frame-shift mutation in the
pTEN tumor suppressor gene, which encodes a phosphatase
that inactivates the lipid products of PI3K. As a result, the lack
of the pTEN protein in these cells resulted in a constitutively
activated antiapoptotic NF-B pathway (108). Thus, these cells
are less sensitive to anticancer drugs whose mechanism of action
is based on the induction of apoptosis (109).
Ganoderma lucidum induced apoptosis of PC-3 cells with
a slight decrease in the expression of NF-B-regulated Bcl-2
and Bcl-xl and the upregulation of the proapoptotic Bax pro-
tein, resulting in the enhancement of the ratios of Bax/Bcl-2
and Bax/Bcl-xl (86). Bemis et al. (110) studied the bioactivity
of a unique preparation of concentrated soybean isoflavones
fermented with G. lucidum mycelia named genistein com-
bined polysaccharide (GCP). During fermentation, a concen-
trated mixture of aglycone isoflavones was produced due to the
hydrolytic cleavage of the sugar moiety from the isoflavone
via G. lucidum-derived -glycosidase. The potential utility of
GCP as a prostate cancer chemopreventative agent was ana-
lyzed in vitro and in vivo. GCP was reported to significantly
suppress LNCaP and PC-3 cell growth, which was associated
with apoptosis in LNCaP cells but not in PC-3 cells. GCP in-
duced p27 and p53 (LNCaP only) protein expression within 6
h and suppressed phosphorylated Akt in both cell lines. The
2% GCP-supplemented diet significantly slowed LNCaP tumor
growth, increasing apoptosis and decreasing proliferation over
4 wk (110). Zaidman et al. (95,139) measured 3 parameters that
indicate induction of apoptosis in LNCaP cells: PARP cleav-
age, caspase-3 activity, and annexin V-FITC staining of exter-
nalized PS membranes. G. lucidum extracts induce apoptosis
in LNCaP cell line by triggering the caspase cascade through
the extrinsic or death receptor mediated pathway that includes
the initiator caspase, caspse-8 and the effector caspase,
caspase-3.
The ability of G. lucidum extracts and compounds to induce
apoptosis was also demonstrated in several cell lines such as hu-
man leukemia, lymphoma, and multiple myeloma (112), human
breast cancer MCF-7 (87), human prostate cancer PC-3 (86),
human hepatoma HuH-7 (113), and human colonic carcinoma
HT-29 (114) cell lines. One report showed that a mixture of
extracts from G. lucidum and the herb Duchesnea chrysantha
induces apoptosis in human leukemia HL-60 cells (115). How-
ever, in some of these cell lines, it was found that G. lucidum
extracts induce apoptosis through the intrinsic or mitochondrial
pathway (87,113,115).
In a series of experiments, Sliva et al. (116,117) and Jiang
et al. (85) showed that G. lucidum extracts inhibit Akt/NF-B
signaling in prostate cancer PC-3 and breast cancer MDA-MD-
231 cells, resulting in the inhibition of proliferation, apoptosis
induction, and decreased motility.
Ganoderma Lucidum Induces Cell Cycle Arrest
in Prostate Cancer Cells
Mammalian cell growth and proliferation are mediated via
cell cycle progression (119). In each cell division cycle, chro-
mosomes are replicated once (DNA synthesis or S-phase) and
segregated to create two genetically identical daughter cells
(mitosis or M-phase). These events are spaced by intervals of
growth and reorganization (gap phases G1 and G2). Progression
through the G1 phase of the cell division cycle is a rate-limiting
step in mammalian cell proliferation and is governed by nu-
merous mitogenic pathways until the restriction point is passed.
CDK4 and CDK6 complexed with cyclin D1 are responsible
for cell cycle progression through the G1 phase (120), and the
CDK2/cyclin E complex functions in the progression of the cell
from the late G1 to the early S-phase (121). These complexes
lead to the phosphorylation of retinoblastoma gene product, a
tumor suppressor gene active in controlling the G1 phase (122).
Hyperphosphorylated pRb leads to its release from the E2 F fam-
ily of transcription factors and induces expression of a number
of genes required for S-phase transition (123).
DNA content (as an indicator of cell proliferation) analysis
showed that G. lucidum fungal extracts blocked LNCaP cell cy-
cle at the transition from the G1 to the S phase (95,139). These
results are in accordance with several recent reports showing
G1 phase arrest in a variety of cancer cell types caused by the
treatment of chemotherapeutic agents (8588,124). Cell cycle
arrest at the G1 phase was reported to be mediated through
the downregulation of cyclin D1 (85,87). In addition, other
 MEDICINAL MUSHROOMS FOR PROSTATE CANCER THERAPY
reports have demonstrated a G2 phase arrest caused by G.
lucidum fungal extracts that is mediated by the downregula-
tion of cyclin B (86,97,118). These data imply the existence
of two separate chemical moieties that regulate cyclin D1 and
cyclin B and consequently cause cell cycle arrest at G1 and G2,
respectively.
Defects in cell cycle are one of the most common features
of cancer cells. In prostate cancer, 1668% of the cases reveal
a loss of p27kip1 protein or low-grade expression (125). The
function of the p27kip1 protein is regulated mainly by posttrans-
lational, ubiquitin-mediated, proteasomal proteolysis (126). The
tumor suppressor gene p21 was also reported missing in sev-
eral solid tumors. For example, in prostate cancer PC-3, DU
145, and LNCaP cell lines, p21 was expressed at low or un-
detectable levels (127). On the other hand, overexpression of
cyclin D1 has been documented in a number of human cancers.
However, evidence suggests that this event in prostate cancer
is quite rare (128). A common polymorphism in the cyclin D1
gene is associated with the production of an alternate transcript
of cyclin D1 termed cyclin D1b. It was found that this vari-
ant actually stimulates cell-cycle progression in AR-dependent
LNCaP cells but had no effect on AR-independent PC-3
cells (129).
Additional studies by Lu et al. (118) and Hu et al. (87) re-
ported that the extract of G. lucidum inhibited cell proliferation
in vitro and induced G1 cell cycle in prostate and breast can-
cer cell lines including MCF7. Later, Zaidman et al. (95,139)
has reported that G. lucidum downregulated cyclin D1 expres-
sion leading to dephosphorylation of pRb and growth arrest
of LNCaP prostate cancer cell line. Interestingly, cyclin D1 is
controlled by the Akt and NF-B pathways (130), which are
constitutively active in LNCaP cells and can be inhibited by
G. lucidum.
Ganoderma Lucidum Inhibits Angiogenesis
in Prostate Cancer
Angiogenesis is a physiological process involving the growth
of new blood vessels from preexisting vessels. It is a normal
process in growth and development as well as in wound healing.
However, this is also a fundamental step in the transition of
tumors from a dormant state to a malignant state. Tumors induce
angiogenesis by secreting various growth factors such as VEGF
and bFGF that induce capillary growth into the tumor and allow
it to grow by supplying nutrients and oxygen and removing
waste products. Moreover, the new vessels allow tumor cells
to escape into the circulation and lodge in other organs (tumor
metastases) (135, 136).
G. lucidum extract was reported to inhibit early events in
angiogenesis, capillary morphogenesis of the human aortic en-
dothelial cells (137). The anti-angiogenic effect of G. lucidum
was mediated by the inhibition of constitutively active AP-
1 in prostate cancer cells, resulting in the downregulation of
the secretion of VEGF and TGF-1 from PC-3 cells. Inhibi-
21
tion of AP-1 activity was mediated by the inhibition of Erk
1/2 phosphorylation and Akt kinases activity in PC-3 cells
(137).
Ganoderma Lucidum Interferes With Androgen
Receptor Function
LNCaP cell lines are considered as a laboratory model for
hormone-responsive prostate cancer. These cells are sensitive to
androgen and contain high levels of mutated androgen recep-
tor, a T877A mutation (ACT GCT, Thr Ala) in the LBD
(138). This mutation renders LNCaP cells sensitive not only to
androgen but also to antiandrogens, estrogens, and progestins.
In contrast, the androgen insensitive cell lines DU 145 and PC-3
do not express the androgen receptor. DHT is an androgen re-
ceptor ligand generated from testosterone by the activity of 5-
reductase. Thus, 5- reductase inhibitors might have activity
against prostate diseases including benign prostatic hyperplasia
(BPH) and prostate cancer. Fujita et al. (140) examined the in-
hibitory effects of methanol extracts of 19 medicinal mushrooms
on 5-reductase activity including Ganoderma lucidum. The ex-
tract of Ganoderma lucidum showed the strongest 5- reductase
inhibitory activity, and it significantly inhibited testosterone-
induced growth of the ventral prostate in castrated rats
(140).
Zaidman et al. (95,139) have reported that the treatment
of LNCaP cell lines with G. lucidum organic extract resulted
in a concentration-dependent inhibition of androgen receptor
transcriptional activity, a concentration-dependent decrease
of androgen receptor-regulated PSA, abrogation of nuclear
translocation, and DNA binding activity of the androgen
receptor (95,139). Furthermore, an active fraction was isolated
from G. lucidum that competes with Dihydrotestosterone for
receptor binding and consequently interferes selectively with
androgen receptor function but not with the glucocorticoid
receptor (GR) (95,139). The authors suggested that inter-
ference with androgen receptor function might be explained
by the ability of G. lucidum to stimulate the assembly of a
transcriptionally inactive androgen receptor on DNA as sug-
gested by some other antiandrogen compounds (141) possibly
through the inhibition of the interaction with coactivators
and/or the enhancement of the interaction with corepressors
(61).
Liu et al. (62) isolated Ganoderol B from G. lucidum fruiting
body extract with 5- reductase inhibitory activity, which sup-
pressed the regrowth of the ventral prostate induced by testos-
terone in rats. In addition, Ganoderol B reduced androgen recep-
tor levels in treated animals. The downregulation of androgen
receptor signaling by Ganoderol B provided an important mech-
anism for its antiandrogenic activity. The important implication
of this study was that Ganoderol B intervention strategy could
be helpful in controlling the morbidity of prostate cancer. It was
suggested that Ganoderol B might be useful in prostate cancer
therapy through the inhibition of androgen synthesis and the
function and level of its receptor (142).
22 J. MAHAJNA ET AL.
Ganoderma Lucidum Interferes With Prostate Cancer
Invasion: The PI3K/Akt/NF-B Pathway Connection
To invade and metastasize, cancer cells must effectively de-
grade extracellular matrix (ECM) components. Plasminogen ac-
tivation has been implicated as one of the mechanisms of ECM
degradation. Mammalian cells contain two types of plasminogen
activators, the urokinase type (uPA) and the tissue type (tPA),
of which uPA is primarily involved in ECM degradation and
consequently in tumor invasion (69,99,100). The uPA system
consists of the serine proteases uPA and plasmin, their serpin
inhibitors PAI-1, PAI-2, and 2AP, as well as an uPA cell surface
receptor, uPAR (63). uPA is synthesized and released by cells
as an inactive, single-chain, proenzyme: pro-uPA. It binds to an
uPAR on the cell surface (63,64), which has an accelerating and
positioning function for both the uPA and plasminogen activa-
tion (64,65). Pro-uPA is cleaved to an active, two-chain protease
form by plasmin (66). uPA has restricted substrate specificity,
the main function of which is cleaving plasminogen to plas-
min (65), which degrades several ECM components and also
activates many promatrix metalloproteases (65,67). The plas-
minogen activation system is controlled specifically by their
serpin inhibitors PAI-1, PAI-2, and 2AP, of which PAI-1 plays
a more important role in cancer invasion, and 2AP is the main
inhibitor of plasmin (68,100). However, during cancer invasion
and metastasis, this degradation system goes out of control, al-
lowing cancer cells to cross the normal tissue boundaries. Both
the uPA secretion and the presence of receptor bound uPA at
the cell surface characterize prostate cancer having an inva-
sive phenotype (70), increased metastatic potential, and poor
survival.
The expression of uPA is regulated by the transcription fac-
tors NF-B and AP-1. The NF-B is a dimeric transcription
factor belonging to the Rel/NF-B family of transcription fac-
tors (5660). The major activator of NF-B is known as the
IB kinase complex (IKK) (58). G. lucidum extract inhibited
the constitutively active transcription factors, NF-B and AP-1,
which resulted in the inhibition of the expression of uPA and
its receptor uPAR. G. lucidum also suppressed cell adhesion
and cell migration of highly invasive prostate and breast can-
cer cells, suggesting its potency in reducing tumor invasiveness
(71). Furthermore, Jiang et al. (85) reported that Ganoderma lu-
cidum downregulated the expression of NF-B-regulated uPA
and uPAR receptor (uPAR), as well as levels of the antiapoptotic
genes, Bcl-2 and Bcl-xl, accompanied by increased expression
of the proapoptotic Bax protein, resulting in the enhancement
of the ratio of Bax/Bcl-2 and Bax/Bcl-xl. Other reports showed
that suppressing NF-B activity by G. lucidum spore and fruit
body extracts is mediated by the inhibition of the prosurvival
factor Akt in MDA-MB-231 cells (98,116,117).
Clinical Activity of Ganoderma lucidum
Ganoderma lucidum shows a very promising effect on
prostate cancer in different preclinical systems. An herbal mix-
ture called PC-SPES containing G. lucidum was prepared and
recommended as an alternative herbal therapy for prostate can-
cer (92). Ethanol extracts of the PC-SPES component were ac-
tive in inhibiting the growth of LNCaP cells. Treatment with 5
microl/ml of the individual herbal extract suppressed the growth
of LNCaP cells in the following order: Dendranthema mori-
folium (85.2% reduction), Panax pseudo-ginseng (80.9%), Gly-
cyrrhiza uralensis (73%), Rabdosia rubescens (70.8%), Scutel-
laria baicalensis (66.5%), Ganoderma lucidum (63.5%), Isatis
indigotica (50.0%), and Serenoa repens (14.5%). Analysis of
the efficacy of the individual herbs in controlling intracellu-
lar/secreted PSA levels and the expression of the androgen
receptor and PSA revealed that only Glycyrrhiza uralensis,
Scutellaria baicalensis, and Serenoa repens lowered intracel-
lular and secreted PSA, whereas the remaining herbs actually
increased PSA expression. In addition, no uniform response in
AR/PSA was observed in individual herb-treated cells, contrary
to PC-SPES, which elicited a coordinated change in AR/PSA.
Lack of concordance between changes in prostate cell growth
and prostate specific gene expression makes it unlikely that
the activity of a single herb can account for the overall effects
of PC-SPES (92). The PC-SPES was also tested in prostate
cancer patients. de la Taille et al. (93) reported two cases of
hormone-refractory prostate cancer patients who showed a fa-
vorable response to therapy with the PC-SPES herbal combi-
nation, controlling the progression of the disease. The report
included two cases of biopsy proven prostate cancer patients
with metastatic disease treated with total androgen blockade,
progressing to an androgen-independent status. PC-SPES ex-
tract decreased the PSA value for both patients from initial
values of 100 and 386 ng/ml to 24 and 114 ng/ml after 1 yr
and 4 mo, respectively. No gynecomastia or hot flashes were
observed in these patients, and the treatment was well tolerated.
PC-SPES showed strong estrogenic in vitro and in vivo activ-
ity as an alternative tool in the management of prostate cancer
patients. These cases suggest that PC-SPES might have some
potential activity in hormone-independent prostate cancers. An-
other independent study conducted by Small et al. (94) treated 33
patients having androgen-dependent prostate cancer (ADPCa)
and 37 patients having androgen-independent (AIPCa) prostate
cancer with PC-SPES at a dose of 9 capsules daily. Clinical
outcome was assessed with serial serum PSA level measure-
ment and imaging studies. As a result, 100% of ADPCa patients
experienced a PSA decline of 80%. No patient developed PSA
progression. Thirty-one patients (97%) had reductions in testos-
terone to the anorchid range. Of the 35 AIPCa patients, 19 (54%)
had a 50% PSA reduction, including 8 (50%) of the 16 patients
who had received prior ketoconazole therapy. Median time to
PSA progression was 16 wk (ranged from 2 to 69+ wk). It was
concluded that PC-SPES shows good efficacy in the treatment
of both ADPCa and AIPCa with a tolerated toxicity. At this
stage, the exact contribution of the Ganoderma lucidum extract
to the observed clinical effect is unclear; however, it is reason-
able to speculate that a kind of synergy might exist among the
 MEDICINAL MUSHROOMS FOR PROSTATE CANCER THERAPY
different herbs used that resulted in the enhanced clinical effi-
cacy of Ganoderma lucidum extract.
Ghafar et al. (111) reported the case of a patient with biopsy
proven prostate cancer showing clinical and pathologic evidence
of regression following administration of GCP, a concentrated
soybean isoflavones fermented with G. lucidum mycelia, named
genistein combined polysaccharide (GCP. The patient received
GCP for 6 wk prior to radical prostatectomy. The patients PSA
decreased from an initial value of 19.7 to 4.2 ng/ml after 44
days of low-dose GCP. No cancer was identified in the radical
prostatectomy specimen and no side-effects were observed in
this patient. This case also suggested that GCP, which had shown
potent inhibitory effects against prostate cancer cell lines in in
vitro studies, may exhibit some potential activity in the treatment
and prevention of prostate cancer.
CONCLUSIONS
Ganoderma lucidum is a popular medicinal mushroom that
has been used as a home remedy in traditional Chinese medicine
(TCM) for the prevention or treatment of a variety of diseases
including cancer. Today, G. lucidum is recognized as a dietary
supplement recommended in many countries as a cancer thera-
peutic. In addition, G. lucidum is considered by some investiga-
tors as a therapeutic biofactory that consists of diverse bioactive
molecules mediating its biological functions. This raises the
need to explore the full potential of this natural product and also
to recognize its various active substances capable of combating
a variety of diseases including cancer.
Preliminary clinical data based on a number of clinical tri-
als that were conducted have shown promising efficacies of G.
lucidum extracts or powders in cancer treatment as well as in
other indications. Some of the clinical trails were not well de-
signed and lacked appropriate controls. We believe that there is
a need to explore the full potential of the dietary supplement
of G. lucidum to assess its safety and efficacy in well-designed,
double-blinded, randomized, placebo-controlled clinical trials
using G. lucidum powders or extracts as stand-alone treatment
or in combination with other treatments. To achieve this goal,
standardization of G. lucidum is an important element. Because
the composition and amount of biologically active substances
depend on places of production, cultivation conditions, extrac-
tion procedures, and the strains of G. lucidum, standardization
will help its acceptance as a natural product suitable for cancer
treatment.
Ganoderma lucidum extracts exhibited anticancer activity
in in vitro systems against a variety of cancer cells including
leukemia, lymphoma, breast, prostate, liver, lung, and myeloma
cell lines. The anticancer activity of G. lucidum includes the
inhibition of proliferation, induction of apoptosis, induction of
cell cycle arrest, inhibition of invasive behavior, and suppression
tumor angiogenesis in many experimental systems including
prostate cancer. Studies have aimed at elucidating the mecha-
nism of action revealed that G. lucidum inhibits the function an-
23
drogen receptor and interferes with the PI3K/Akt/NF-B path-
way. The reported activity of G. lucidum is mostly reported using
crude extractions. Thus, isolating active fractions and moieties
responsible for the reported activity is an obstacle that must be
overcome to allow the structural elucidation of active moieties
and to define the exact mechanism of action of such substances.
Moreover, G. lucidum extracts that exhibited diverse pharma-
cologic functions were also shown to contain highly diverse
pharmacological moieties (4048,91). Currently, more than 100
different moieties have been reported from G. lucidum. Thus,
the immediate goal must be to identify the different chemi-
cal structures mediating the different pharmaceutical activities
aimed at improving their potency, selectivity, bioavailability, as
well as pharmacokinetics and pharmacodynamics parameters
and explore their potential synergy with other pharmaceutical
compounds available for combating different diseases including
prostate cancer.
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