Clinical Trial in Diabetes

 A clinical study of a Ganoderma lucidum extract in patients with type II diabetes mellitus

Shufeng Zhou1, Yihuai Gao2, Guoliang Chen3, Xihu Dai4, and Jinxian Ye5

1Division of Pharmacology and Clinical Pharmacology, Faculty of Medical and Health Sciences, The University of Auckland, New Zealand; 2Landcare Research, Private Bag 92170, Auckland, New Zealand; 3Shanghai Academy of Agricultural Sciences, Shanghai, China; 4Fuzhou General Hospital of Nanjing Military Region of the Peoples’ Liberation Army, Fuzhou, China; 5Hospital of Fujian University of Traditional Chinese Medicine, Fuzhou, China.

Correspondence author: Dr Shufeng Zhou, MD, PhD.

Division of Pharmacology and Clinical Pharmacology, Faculty of Medicine and Health Science, Auckland University, Auckland, New Zealand. Ph: 0064 9 3737599 (Ex 6414); Fax: 0064 9 3737556; Email: shufeng.zhou@auckland.ac.nz

Running title: Phase I/II study of Ganopoly in patients with type II diabetes mellitus

Abstract

Animal studies have demonstrated that the polysaccharide fractions of Ganoderma lucidum have potential hypoglycemic and hypolipidemic activities. This clinical study aimed to evaluate the efficacy and safety of Ganopoly (polysaccharides fractions extracted from G. lucidum by patented technique, provided by Encore International Ltd, Auckland, NZ) in 35 patients with confirmed type II diabetes mellitus (DM). The standard oral hypoglycemic agent glibenclamide was used as positive control (n = 33). Eligibility criteria included type II DM of > 3 months’ duration for which patients were not receiving insulin; age > 18 years; normal vital signs for age and disease state; normal electrocardiogram (ECG); and fasting plasma glucose (FPG) level of 8.9?16.7 mmol/l in sulfonylurea-naﶥ patients or an FPG < 10 mmol/l before washout in sulfonylurea-treated patients. Patients were randomly grouped to be given either Ganopoly or glibenclamide. Ganopoly was given orally at 600 mg three times daily for 12 weeks. Patients underwent 4 weeks of dose adjustment followed by 8 weeks of dose maintenance for glibenclamide. Fasting and stimulated glycosylated hemoglobin (HbA1c), plasma glucose, insulin, and C-peptide were monitered at predetermined intervals. Adverse events and hypoglycemic episodes were recorded. From baseline to last visit, mean HbAlc decreased from 8.4 to 7.6% and 8.5 to 7.3% in patients treated with Ganopoly and glibenclamide respectively (P < 0.05). Mean fasting plasma glucose and post-prandial glucose decreased in patients treated with either Ganopoly or glibenclamide (P < 0.01). Concentrations of fasting and post-prandial insulin and C-peptide were lower at the last visit compared with baseline levels for patients treated with either Ganopoly or glibenclamide (P < 0.05). Overall, Ganopoly was well tolerated. This study demonstrated that Ganopoly was safe and efficacious in lowering blood glucose concentrations.

Key words: Ganoderma lucidum, Ganopoly, Glibenclamide, Diabetes.

Introduction

Lower blood glucose levels are beneficial in patients with type II diabetes mellitus (DM) (Andersson and Svardsudd, 1995; Skyler, 1996). Uncontrolled high blood glucose will result in a series of complications such as renal dysfunction, neuropathy and cardiopathy. (Andersson and Svardsudd, 1995; Ohkubo et al., 1995). These findings have stimulated the development of oral drugs, such as sulfonylureas, to treat the subpopulation of diabetic patients who respond to oral hypoglycemic agents. However, these oral hypoglycemic drugs are often long-acting agents, which are metabolized by the liver and kidneys and may cause moderate-to-severe hypoglycemia (Groop, 1996). Elderly patients are particularly susceptible to hypoglycemia induced by oral agents, especially if they miss meals or when their hepatic, renal, or cardiovascular function is impaired. Thus, safe and effective therapy approaches for type II DM are sought.

Traditional Chinese medicine has been used to treat DM for many centries. Among the potential sources of herbal medicine, Ganoderma lucidum as a medicinal mushroom has been used extensively in Asia, particularly, in China, Japan and Korea (Shiao et al., 1994; Wasser and Weis, 1999) to treat various chronic diseases, such as chronic hepatopathy, hypertension, diabetes and cancer (Gao, 2000). The polysaccharides and triterpenes isolated from G. lucidum have been shown to have beneficial effects on hyperglycemia and hyperlipidemia. For example, oxygenated lanosterol derivatives isolated from G. lucidum or their derivatives obtained by chemical conversion potently inhibited the synthesis of cholesterol from [24,25-3H]-24,25-dihydrolanosterol in the rat (Komoda et al., 1989). Administration of ߼/font>-D-glucan from the fruit body of G. lucidum increases serum, liver, and bone marrow protein synthesis in mice (Guan and Cong, 1982). All these effects of G. lucidum indicate its possible role for DM.

Many DM patients in Asian countries use G. lucidum products as an adjunct to or as the sole therapy for the treatment of DM, and some open non-randomized clinical studies have been published in Chinese or Japanese. However, no clinical trials of G. lucidum used to treat DM have been reported in English peer-reviewed journals (Gao, 2000). Ganopoly is a patented aqueous extract of G. lucidum, which has been marketed as an over-the-counter product in New Zealand, Australia, Hongkong and North America. Preliminary studies have indicated that Ganopoly has an inhibitory effect on hyperglycemia and hyperlipidemia in animal studies (Gao, 2000). Therefore, we undertook this open, non-randomized clinical study of Ganopoly as a single agent to investigate its efficacy and safety in adults with type II DM. The oral sulfonylurea agent glibenclamide, was used as a positive control.

MATERIALS AND METHODS

Patient selection

This open, multi-center and parallel-controlled trial was conducted from January 1998 to December 1999. Patients were screened for eligibility using the following assessments: medical and diabetes history; physical examination, vital signs, ECG findings, hematological profile, clinical chemistry, urinalysis, FPG, recent history of hypoglycemic events, and use of concomitant medications.

Patients were entered onto the study if they met the following eligibility criteria and did not meet any of the exclusion criteria: (1) type 2 diabetes of > 3 months’ duration for which they were not receiving insulin; (2) = 8-week interval between prior hypoglycemic therapy and entry; (3) = 18 year age; (4) body weight 90 to 150% of desirable body weight for sex, body frame, and height; normal vital signs for age and disease state; (5) normal electrocardiogram (ECG); (6) a fasting plasma glucose (FPG) concentration of 8.9?16.7 mmol/l in sulfonylurea-naive patients or an FPG < 10 mmol/1 before washout in sulfonylurea-treated patients; and (7) informed consent for participation. Ethical approval was received from the Institutional Research Ethics Committee. Exclusion criteria were: (1) severe concurrent conditions; (2) pregnant or lactating women; (3) secondary DM; and (4) patients who had taken or were taking Ganoderma preparations.

Drug treatment

After an initial screening followed by a 2-week washout period, eligible patients were randomly assigned and then to receive either Ganopoly or glibenclamide. The latter was adjusted during the first 6 weeks of treatment, followed by 12 weeks of maintenance therapy once the optimal dose was achieved. The study concluded with a 12-week follow-up period after treatment ended. Ganopoly or glibenclamide were administered as oral capsules. Patients were treated preprandially with 600 mg, three times daily before meals for 12 weeks. Each capsule contained 600-mg extract of G. lucidum (Encore International Co., Auckland, New Zealand). Glibenclamide was administered starting at 2.5 mg/day, adjusted dose for 6 weeks, then maintained the dose for 12 weeks. Use of concomitant drugs that might affect glucose metabolism, such as sulfonamides, thiazides, corticosteroids, and thyroid products, was discouraged unless the investigator determined that a patient was stable on a chronic treatment regimen. Patients were also instructed to follow the diet recommended by the Chinese Diabetes Association and taught to perform three-point in-home blood glucose monitoring.

Clinical Evaluation

Patients were monitored weekly during the treatment. At its completion, patients were assessed for vital signs; weight; concomitant medication or illness; clinical chemistry; hematological profile; lipid profile; urinalysis; serum testosterone; pregnancy test; FPG, insulin, C-peptide, and HbA1c, 2-h post-prandial glucose (PPG), insulin, and C-peptide; and hypoglycemic and adverse events. Normal reference ranges were 4.4?6.1% for HbAlc and 3.7?5.8 mmol for FPG. Patients continued treatment throughout the 12-week maintenance period at the same dose level attained in the adjustment. If the investigator considered the patient to be at risk for hypoglycemic events, the dose was adjusted accordingly.

Assessments were similar at weeks 4 and 8. At week 12, a more complete assessment was made, which included all the measurements taken at week 8 plus a physical examination and an ECG. Treatment with Ganopoly was then discontinued, and patients were prescribed treatment with another appropriate antidiabetic agent as determined by the investigator for the 12-week follow-up period (weeks 13?24). Patients were assessed for concomitant medications or illness; clinical chemistry; serum testosterone; hematological profile; urinalysis; and hypoglycemic and adverse events.

The efficacy variables included changes in HbAlc, FPG, 2-h PPG (after 100 g glucose) between week 12 and baseline, fasting insulin, 2-h postprandial insulin, and C-peptide between week 12 and baseline. The safety variables included a standard clinical laboratory screen, plasma lipids (triglycerides, total cholesterol, HDL cholesterol, and LDL cholesterol), body weight, blood pressure, adverse events, and hypoglycemic events. Clinical laboratory test results were categorized as normal, high, or low based on laboratory reference ranges.

Adverse events were defined as events reported before treatment that were exacerbated during treatment or as events reported during treatment or within 7 days after the study drug was stopped. A serious adverse event was defined as any event, whether treatment related or not, that posed a medically significant hazard to the patient; was fatal, life-threatening, or permanently disabling; or required hospitalization.

Hypoglycemic events were recorded separately from adverse events during treatment with Ganopoly. Active solicitation of symptoms that could represent hypoglycemia was made of patients during their visits. Reports of symptoms were recorded as hypoglycemia, even if confirmatory blood glucose tests were not done. Mild-to-moderate hypoglycemia was defined symptomatically as sweating, strong hunger, dizziness, or tremors or as a blood glucose value < 2.5 mmol/L. Severe hypoglycemia was defined symptomatically as severely impaired consciousness that required the assistance of another person and hospitalization.

Statistical analysis

Changes between baseline and the last treatment (last visit) were evaluated for all efficacy variables, with the last observation carried forward. Therefore, only patients with baseline and postbaseline data were included in the primary analyses. Data compiled at baseline, 8 weeks and 12 weeks were analysed by two-way analysis of variance by ranks. Median values of quantities, such as age and days since diagnosis, were compared using Wilcoxon’s rank-sum test. Test results for treatment group effects were considered statistically significant if P < 0.05. For clinical laboratory tests, an unpaired Student’s t-test examining the mean change from baseline to the end of study was performed.

RESULTS

Patient demographics

As shown in Table-1, there were 18 men and 17 women in the Ganopoly group. The median age of all patients was 57.2 yrs. In 35 patients receiving glibenclamide, there were 16 men and 17 women, with a mean age of 55.4 yrs. Eight patients were not assessable for response and adverse effects because they were lost to follow-up or refused further therapy before 12 weeks of treatment. Patients in both treatment groups were similar with respect to age, sex, and duration of diabetes. None of the patients reported severe hypoglycemia within the 3 months preceding the study or frequent moderate hypoglycemia within 1 month before treatment.

Efficacy evaluation

The progressive changes in mean HbAlc values throughout the study are shown in Figure 1. HbA1c value after 8 and 12-week treatment were significantly (P < 0.05) reduced compared to the baseline levels. The mean HbAlc decreased from 8.4 to 7.6% and 8.5 to 7.3% at 12 weeks in patients treated with Ganopoly and glibenclamide respectively. Significant changes in mean FPG and PPG levels at the last visit paralleled the changes in mean HbAlc levels. At baseline, mean FPG values were 12.0 and 12.2 mmol/ for the Ganopoly and Glibenclamide groups, respectively. At baseline, mean PPG values were 13.6 mmol/lf for the Ganopoly and 14.2 mmol/1 for the Glibenclamide group. At week 12, mean PPG values had decreased to 11.8 and 10.9 mmol/l, respectively. The between-group difference in PPG levels at week 12 was small (P > 0.05), indicating the comparable hypoglycemic effect of Ganopoly in type II DM patients as a standard oral hypoglycemic agent. Changes in fasting insulin, 2-h post-prandial insulin, fasting C-peptide, and 2h post-prandial C-peptide were consistent; the between-group differences in these end points were nonsignificant at the last visit. A substantial effect during the post-prandial period was noted. The mean total cholesterol, HDL, LDL, and triglyceride values were not significantly decreased for either group for all study weeks, and there were no between-group differences during treatment.

Adverse effects

Safety results for all patients were recorded for the 12 treatment weeks and the follow-up periods. Both drugs were well tolerated. Most of the events were mild-to-moderate adverse episodes. Patients in glibenclamide groups, but not in the Ganopoly group, reported five hypoglycemic events. The frequency of hypoglycemic events did not appear to be dose related, and none of the episodes were serious. Only one event was associated with a blood glucose value <2.5 mmol/l. Ganopoly was generally well tolerated and was not associated with hematologic or biochemical toxicity. Toxicity was generally mild (grade 1-2).

DISCUSSION

This study was designed to compare glycemic control after treatment with Ganopoly and glibenclamide in patients with type II diabetes. The results showed significant decreases in HbAlc and in FPG and PPG concentrations in patients treated with either Ganopoly or glibenclamide. During the 12-week maintenance-dose period, mean HbA1c, values decreased significantly in both group. As expected, patients who were previously treated with sulfonylureas had lower HbA1c values at baseline than did the sulfonylurea-naive patients. Five serious adverse events were reported in the glibenclamide group, but none in the Ganopoly group. These findings indicate the potential use of herbal medicine in the treatment of type II DM. Indeed, there is increasing interest in the use of alternative medicines to treat patients with type II DM (Vickers, 2000). There is also an increasing interest in identifying an anti-diabetic remedy from edible mushrooms, in particular from Ganoderma species. There are increasing investigations in the field of anti-diabetic polysaccharides and truterpenes from edible mushrooms, but reliable and objective comparative data on the anti-diabetic activity of G. lucidum preparations as over-the-counter nutritional supplements are not available.

The mechanism of anti-diabetic activity of Ganopoly is unclear. Preclinical studies indicated the anti-diabetic action of polysaccharides and triterpenes of G. lucidum. There is evidence that the ߼/font>-D-glucans induce biological response by binding to membrane complement receptor type three (CR3, aM߼/font>2 integrin, or CD11b/CD18) on immune effector cells such as macrophage (Konopski et al., 1994; Muller et al., 1996; Battle et al., 1998; Mueller et al., 2000). The ߼/font>-D-glucan binding site (lectin site) of CR3 has been mapped to a region of CD11b located C-terminal to the I-domain and its distinct metal ion-dependent adhesion site for the many protein ligands of CR3 such as iC3b, ICAM-1 and fibrinogen (Diamond et al., 1993; Lee et al., 1995; Thornton et al., 1996). There is a possibility that polysaccharides bind to ߼/font>-cells or other cells in the pancreas to stimulate the secretion of insulin and inhibition of glucagons. Other mechanisms may also be involved in the anti-diabetic activity of G. lucidum. Triterpenes may also stimulate insulin secretion in vivo. More studies are needed to identify more-active components from G. lucidum, and its mechanism of action.

REFERENCES

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Table 1. Demography of Patients enrolled.

Characteristic Ganopoly group Glibenclamide group
No. of patient 35 33
Age (yrs) 57.2 ? 8.8 (39-71) 55.4 ? 9.1 (40-74)
Gender
Men 18 (51) 16 (48)
Women 17 (49) 17 (52)
BMI (kg/m2) 29.8 ? 5.1 (20.2-38.1) 30.1 ? 6.2 (22.1-48.8)
Duration of disease (yrs) 5.8 ? 3.5 (0.6-25.2) 6.1 ? 4.5 (0.4-22.1)
Sulfonylurea-naive 4 (11) 3 (9)
Sulfonylurea-treated 31 (89) 30 (91)

Figure legend

Figure 1. Change of HBA1c in DM patients treated with either Ganopoly or glibenclamide.

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