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Data from multiple epidemiologic and clinical studies on immune effects of conventional cancer treatment and the clinical benefits of polysaccharide immune therapy suggest that immune function has a role in breast cancer prevention. Immune therapy utilizing the polysaccharide constituents of Trametes versicolor (Tv) as concurrent adjuvant cancer therapy may be warranted as part of a comprehensive cancer treatment and secondary prevention strategy.
Data from epidemiologic studies of African American women, immune studies of the effect of chemotherapy drugs and radiotherapy on immune status, and the Asian literature on the clinical benefit of polysaccharide immune therapy suggest that immune function has a role in primary and secondary prevention of breast cancer. High-priority research areas for breast cancer immunotherapy include clinical trials of Trametes versicolor (Tv) and the semipurified polysaccharide peptide Krestin (PSK). Two types of trials are called for; first, we need clinical trials of Tv as a concurrent adjuvant therapy along with chemotherapy, radiotherapy, and HER2/neu monoclonal antibody therapy (trastuzumab; Herceptin). Second, in keeping with its potential role in secondary prevention and common use of Tv in Asian oncology, we need clinical trials of Tv immunotherapy after completion of standard cancer treatment.
Based on what is known about the immunology of breast cancer, an effective immunomodulatory therapy would increase natural killer (NK) cell activity, alter cytokine production toward a T-helper 1 (Th1) pattern, and increase some lymphocyte populations while suppressing T regulatory cells. The underlying hypothesis of immunomodulatory therapy in breast cancer is that improving the cytotoxic activity of lymphocyte killer cells, reducing inflammation, suppressing T regulatory cells, and shifting toward a Th1 pattern of cytokine secretion will improve disease-free survival following completion of primary treatment for breast cancer.
Because of their broad immune activity, medicinal mushrooms are a rich potential source of immunoceuticals. Although other natural products from other plant species have immunologic activity (plant sterols, plant cyclooxygenase-2 inhibitors, thiol-containing allium vegetables), polysaccharides extracted from certain mushroom species have been the most thoroughly studied natural products in both preclinical and clinical studies.
More than 270 recognized species of mushrooms are known to have specific immunotherapeutic properties.1 Of these, 50 nontoxic mushrooms species have yielded potential immunoceuticals in animal models, and of these, 6 species have been studies in human cancers.2 Of these six, one medicinal mushroom, Tv, has been studied in phase I, II, and III randomized clinical trials in stomach, colorectal, esophageal, and breast cancer patients (Table 1)3–43 These Japanese and Korean clinical data provide support for the hypothesis that immunomodulation can influence the clinical course in breast cancer.
Tv, also known as Coriolus versicolor or Polyporus versicolor, has a long history of medical use in Asia, dating back hundreds of years in traditional Asian medicine. Tv belongs to the more advanced Basidiomycetes class of fungi. It grows on tree trunks throughout the world in many diverse climates, including North America. The Tv mushroom has a long history of treasured use in Asia as both a food and a medicine. In China, it is called yun zhi or cloud fungus. According to Kidd, the immunomodulatory activity of polysaccharide peptides in Tv was discovered in 1965 in Japan by a chemical engineer who observed a case of cancer remission after ingesting yun zhi.2 Subsequent research led to identification of two closely related proteoglycan constituents of Tv with anticancer activity: Krestin (PSK) and polysaccharide peptide (PSP). Krestin (PSK) has been studied most extensively and is in wide clinical use as an adjunctive and adjuvant cancer therapy in Japan and China.2,44,45 The closely related PSP was first isolated in China in 1983. Although some of the active constituents of Tv have been studied, few data exist on the anticancer activity of the whole extract despite its common use in Asia and now in the United States.
PSPs extracted from the Tv mushroom have been shown in preclinical and clinical studies to have both significant immunologic and oncologic activity in lung cancer patients,46,48 gastrointestinal cancers,5,16,21 and breast cancer.40,42,43 The M.D. Anderson Cancer Center’s Web site describes well the number and type of published studies on Tv (<http://www.mdanderson.org/departments/CIMER/display.cfm?id=3C76D2B7-11B6-11D5-810D00508B603A14&method=displayFull&pn=6EB86A59-EBD9-11D4-810100508B603A14>).
There have been many peer-reviewed publications on the Tv in cancer, including 37 in vitro articles, 55 animal studies, 43 published human clinical studies, and 11 review articles in gastrointestinal, breast, and lung cancer. In the last 2 years, five more Krestin (PSK) trials in colorectal cancer have been published,35,36,49 including one meta-analysis in 1,094 colorectal cancer patients,50 all showing a positive impact on clinical outcomes. Lindequist and colleagues and Hobbs have written excellent review articles on the pharmacologic potential of mushrooms, including Tv.51,52 Tv is standard oncologic care in mainstream modern Japanese cancer management. Krestin (PSK) was approved in 1977 as a cancer therapy by the Japanese National Health Registry and represents 25% of the total national costs of cancer care in Japan.52 Oncologists in the West have only recently begun to turn their attention to immune-based therapies and, thus, have generally not noticed this potentially important therapy. Some of the clinical trial data from the east are of high quality. Several investigator teams at several high-quality cancer centers have replicated positive clinical findings. However, although the clinical data are strong, the immunologic rationale for proceeding with trials has never been fully articulated.
The rigorous clinical evaluation of PSPs extracted from the medicinal higher fungi Tv in well-designed trials conducted in the United States is not only justified but overdue. Clinical trials hi the United States are needed. But the logic of such trials needs to be built on sound immunologic mechanisms. To proceed with clinical trials in the United States, two requirements must be met. First, there must be enough high-quality clinical trial data from Japan and China that show clinical efficacy; second, there must be plausible immunologic mechanisms to justify proceeding with expensive prospective human trials.
Clinical trial data for Krestin (PSK) from China and PSP in Japan suggest that PSP immunomodulation improves disease-free and overall survival in breast cancer. In a randomized study of 158 esophageal cancer patients, the survival of the radiochemotherapy plus Krestin (PSK) (3,000 mg/d for 12 weeks) group was significantly better than that in the group receiving radiochemotherapy alone.53 Animal studies have shown that (β-glucan extracted from yeast enhances the antitumor effects of radiation (Gary Ostroff, 2007). It has been hypothesized that Tv’s immunologic activity is the underlying mechanism responsible for its antitumor effects and its impact on survival rates.54
In the last 20 years, in vitro, animal, and human clinical studies have supported the use of commercially derived proteoglycans of Tv in the treatment of stomach, colorectal, prostate, and breast adenocarcinomas.2,44 Two specific proteoglycans (Krestin (PSK)™ and PSP) have been extracted from Tv and have been shown to have both anticancer and immunomodulatory activity in tissue culture, animal, and human studies. Of all of the medicinal mushrooms, Tv has been studied the most thoroughly in terms of analysis of active constituents. There are preliminary data to support the hypothesis that Krestin (PSK), the most widely used of the Tv derivatives, may be beneficial in the treatment of both estrogen receptor (ER)-positive and -negative breast cancers by mitigating the immunologic side effects of treatment and enhancing disease-free survival.40,41
Clinical trials of Krestin (PSK) in cancer began in the 1970s in Japan. Based on three decades of Asian clinical research, Krestin (PSK) as adjuvant therapy is indicated for cancers of the stomach, esophagus, nasopharynx, colon, rectum, lung, and breast.2,28,53–54 Most of the clinical research has focused on the effects of Tv adjuvant therapy on disease-free survival and overall survival rates. In 1984, Sugimachi’s group at Kyushu University published an uncontrolled observational retrospective analysis of breast cancer patients with recurrent disease.55 Patients had received chemotherapy with and without Krestin (PSK) immunotherapy. The survival rate after recurrence was significantly extended by Krestin (PSK) immunotherapy.
In 1992, a large randomized trial with 914 women evaluated tamoxifen as an addition to the then-conventional chemotherapy.40 Randomized subgroups received Krestin (PSK) immune therapy (3,000 mg/d for 24 months) in addition to chemotherapy. Analysis revealed that Krestin (PSK) significantly extended survival in ER-negative, stage IIA patients without lymph node involvement. Morimoto and colleagues conducted a 5-year postoperative randomized controlled trial (RCT) comparing chemotherapy with Krestin (PSK) immune therapy in 376 women with stage II ER-negative breast cancer who received either a prodrug of 5-fluorouracil or 3,000 mg/d Krestin (PSK).41 The 5-year overall and relapse-free survival rates for ER-negative patients were the same regardless of whether they had received chemotherapy alone or Krestin (PSK) alone.
A third RCT conducted in Japan evaluated the efficacy of Krestin (PSK) as an adjunctive immune therapy in addition to combination chemotherapy in 227 operable breast cancer patients with vascular invasion in the tumor and/or metastatic lymph node involvement. Patients were randomized to receive chemotherapy of 5-fluorouracil, cyclophosphamide, mitomycin C, and prednisolone (FEMP) alone, FEMP + levamisole, or FEMP + Krestin (PSK). Krestin (PSK)™ was orally administered at 3,000 mg/d for 28 days. The 5- and 10-year survival curves for the FEMP + Krestin (PSK) group were superior to either FEMP alone or FEMP + levamisole. The authors concluded that immunotherapy using Krestin (PSK) improved the prognosis for breast cancer patients whose tumors showed histopathologic evidence of vascular invasion.
Krestin (PSK) is prepared from strain CM-101 of Tv by water extraction. Analyses performed by Ikuzawa and colleagues indicate that Tv water extracts contain 62% polysaccharide and 38% protein.56 The glucan portion of Krestin (PSK) consists of β1–4 main chain and β1–3 side chains, with β1–6 side chains bound to a polysaccharide moiety through O- or N-glycosidic bonds. There is wide consensus that it is the β1–3 and β1–6 side chains that are immunologically active. Krestin (PSK) is a set of proteoglycan molecules with molecular weights ranging from 94,000 to 100,000 D and is bioavailable orally. 14C-labeled Krestin (PSK) is distributed in its full molecular weight spectrum in the bone marrow, salivary gland, brain, liver, spleen, pancreas, and tumor in mice and rabbits within 24 hours. Toxicologic assessments indicate that Krestin (PSK) has low toxicity with a high median lethal dose, with no reports of abnormalities in animals or humans following acute and chronic toxicity tests.2,54
Previously published studies analyzing the effects of PSK-induced cytokine modulation after oral administration in humans have shown that tumor necrosis factor α (TNF-α), interferon-γ (IFN-γ), and interleukin (IL)-2 and IL-8 levels are consistently affected. Krestin (PSK) has shown consistent TNF induction in human and animal models in vitro and in vivo in response to PSK.57–61 IFN-γ induction by Krestin (PSK) is more equivocal, with some published evidence that some PSK-induced immunomodulatory actions occur independently of IFN-γ.62,63 However, several studies in human cell culture systems and experimental animals show PSK-induced IFN-γ upregulation at the gene expression and/or protein production level.57,64–66
Studies of PSK-induced IL-8 and IL-12 induction support the likelihood of their Krestin (PSK) and Tv induction in human peripheral blood lymphocytes. Krestin (PSK) appears to upregulate both IL-8 gene expression and protein production in human peripheral blood mono-nuclear cells both in vitro and in vivo.58,59 Tv extracts may also induce changes in IL-12 produced by monocytes and macrophages and other antigen-presenting cells. Recent studies suggest an important role for IL-12 induced by Krestin (PSK) in upregulating a Th1 type of CD4+ T-cell response involved in immune response to infections and tumors.61,66–68
Certain PSK-induced immunomodulatory activities such as T-cell activation69 and restoration of immunodepression70 were accompanied by IL-2 upregulation. However, Krestin (PSK) appears to induce NK cell activity independently of IL-263,71,72 and regulates human NK cell transcription factors and activation pathway mediators that are distinct from those regulated by IL-2.73,74 Determining whether Tv-induced IL-2 production correlates with Tv-induced antitumor functions would serve as a first step in assessing the dependence of immunomodulatory activities on PSK-induced modulation of IL-2 expression.
Cancer immunotherapy in the twenty-first century is returning to some of the early observations of the New York surgeon William Coley, who in the 1980s proposed treating cancer by injecting streptococcal bacteria into patients to induce an antitumor immune reaction. It is the liposaccharide (a bacterial toxin) that activates the immune system via Toll-like receptors (TLRs). Currently, TLRs are the target of a new generation of cancer immune therapies and represent a new direction in immunotherapy. TLRs are transverse cell membrane proteins located on innate immune cells, such as dendritic cells and macrophages, and are the first responders to foreign invaders at the same time trigger the release of inflammatory cytokines that activate T and B cells. TLRs link innate and active immunity in a newly recognized role.75 Recent in vitro data suggest that some of Tv’s immunologic effects are mediated through the TLR. Recent data from the Bastyr University/University of Minnesota Myco-Immunology Center indicate that the stimulation of TNF secretion by Krestin (PSK) is TLR-4 dependent but dectin-1 independent.76 These new data strengthen the rationale for clinical evaluation of Tv as a cancer immunotherapy.
Preclinical data point to 1,3-β-d-glucan polysaccharide chains as the main active constituent of Tv. β-Glucans extracted from yeast with an identical β-glucan structure have been shown to have similar immunomodulatory activity to Tv. The tumor-killing mechanisms available to monoclonal antibodies (mAbs) limit efficacy. Yeast β-glucans with a 1,3 and a 1,4 linkage have been shown to function via the iC3b receptor complement receptor 3, thereby enhancing leukocyte killing of tumor cells coated with iC3b complement via naturally occurring antibodies. It has been demonstrated in a murine model that the antitumor mAb response can be improved with β-glucan. In comparison with antitumor mAb or β-glucan alone, combined treatment with mAb plus β-glucan produced greater tumor regression in all models of mammary tumors.77 Modak and colleagues at Memorial Sloan-Kettering Cancer Center showed in a mouse lymphoma model that combination treatment of the mAb drug rituximab plus 1,3-β-d-glucan improved survival without toxicities.78 These data suggest that the therapeutic efficacy of mAbs known to activate complement (eg, trasutuzumab, rituximab, and cetuximab) could be significantly enhanced if they were combined with β-glucan polysaccharides.
Recent pre-clinical pilot work conducted by collaborative work between Bastyr University and the University of Washington Tumor Vaccine Group indicates that Krestin (PSK) induced maturation of bone marrow derived DC cells in vitro and tumor regression in the neu-tg mouse, a model of ER negative, Her-2/Neu positive breast cancer (Mary L. Disis, 2007). The proposed mechanisms involve a Th1 adaptive immune response via activation of dendritic cells and thus modulate the immunosuppressive tumor microenvironment to produce an inflammatory antitumor response.
Data from epidemiologic studies of African American women, immune studies of the effect of chemotherapy drugs and radiotherapy on immune status, and the Asian literature oh the clinical benefit of polysaccharide immune therapy suggest that immune function has a role in primary and secondary prevention of breast cancer. High-priority research areas for breast cancer immunotherapy include trials of Tv and its polysaccharide peptide extract Krestin (PSK). Two types of trials are called for; first, we need clinical trials of Tv as a concurrent adjuvant therapy, along with chemotherapy, radiotherapy, and HER2/neu mAb therapy (trastuzumab). Second, in keeping with its potential role in secondary prevention and common use of Tv in Asian oncology, we need clinical trials of Tv immunotherapy after completion of standard cancer treatment.
Funding for this work came from the National Institutes of Health, National Center for Complementary and Alternative Medicine (grant no. 5U19-AT001998), and the Cancer Treatment Research Foundation (grant no. G-04-002).