Roberto Menéndez

Policosanol Modulates HMG-CoA Reductase Activity in Cultured Fibroblasts

BACKGROUND Cholesterol biosynthesis is strictly controlled by 3-hydroxy-3-methylglutaryl Coenzyme A (HMG-CoA) reductase. METHODS Transfer of cultured fibroblasts to a lipid-depleted medium (LDM) up-regulates the enzyme levels. This, in turn, is followed by an accelerated biosynthesis of cholesterol. RESULTS Exposure of Vero fibroblasts to LDM and policosanol (0.5-50 microg/mL), a new cholesterol-lowering drug purified from sugarcane (Saccharum officinarum L.) wax, decreased in a dose-dependent manner cholesterol biosynthesis from [14C]-acetate and 3H-water, but not from [14C]-mevalonate. CONCLUSIONS This suggests an effect on HMG-CoA reductase, the rate-controlling enzyme in cholesterol biosynthesis. When enzyme activity was measured in the presence of various concentrations of policosanol (0.5-50 microg/mL), reductase was not suppressed. Therefore, there was no evidence for a competitive or noncompetitive inhibition of enzyme activity. However, after treatment of intact cells with policosanol (50 microg/mL) in the presence of LDM, a suppressive effect on enzyme activity was observed, suggesting a modulatory effect of policosanol on reductase activity. The previous inhibition of enzyme up-regulation by policosanol suggests to date a depression of de novo synthesis of HMG-CoA reductase and/or stimulation of its degradation. However, the exact mechanism by which policosanol inhibits the activity of HMG-CoA reductase still remains unclear. Further studies are needed to clarify the precise mechanism of its inhibitory action on cholesterol biosynthesis.

A 12-month study of policosanol oral toxicity in Sprague Dawley rats.

Policosanol is a natural mixture of higher aliphatic primary alcohols. Oral toxicity of policosanol was evaluated in a 12-month study in which doses from 0.5 to 500 mg/kg were given orally to Sprague Dawley (SD) rats (20/sex/group) daily. There was no treatment-related toxicity. Thus, effects on body weight gain, food consumption, clinical observations, blood biochemistry, hematology, organ weight ratios and histopathological findings were similar in control and treated groups. This study supports the wide safety margin of policosanol when administered chronically.

Oral administration of policosanol inhibits in vitro copper ion-induced rat lipoprotein peroxidation.

Policosanol, a new cholesterol-lowering agent, is a mixture of higher aliphatic primary alcohols isolated from sugar cane (Saccharum officinarum L.) wax, which prevents the onset of espontaneously and experimentally induced atherosclerotic lesions in experimental models. Because the oxidation of low-density lipoprotein (LDL) may play a role in the pathogenesis of atherosclerosis, we investigate the effect of policosanol on copper oxidative susceptibility of rat lipoprotein fractions (VLDL + LDL). Rats fed normal diet were treated with policosanol (250-500 mg/kg/day) for up to 4 weeks. EDTA-free lipoprotein particles were oxidized in a cell-free system by the addition of copper ions, and conjugated dienes generation was monitored by changes of optical density at 234 nm. Thiobarbituric acid-reactive substances (TBARS) content and lysine-amino group reactivity were investigated. After administration, there was no change in cholesterol, triglycerides, and phospholipid content of lipoprotein fractions; however, policosanol significantly prolongs the lag time and reduces the propagation rate of diene generation. Also, policosanol reduces TBARS content and increases lysine reactivity in lipoprotein fractions treated with Cu2+. In conclusion, policosanol, in addition to its cholesterol-lowering effect, has other properties that enables it to reduce the potential of lipoprotein to undergo lipid peroxidation. Such effect can be considered of promissory value in the management of atherosclerosis.

Inhibition of human P450 enzymes by natural extracts used in traditional medicine.

Different medicinal plants are widely used in Cuba and Mexico to treat several disorders. This paper reports in vitro inhibitory effects on the P450 system of herbal products commonly used by people in Cuba and Mexico in traditional medicine for decades. Experiments were conducted in human liver microsomes. The catalytic activities of CYP1A1/2, 2D6, and 3A4 were measured using specific probe substrates. The Heliopsis longipes extract exhibited a concentration‐dependent inhibition of the three enzymes, and similar effects were produced by affinin (an alkamide isolated from the H. longipes extract) and two catalytically reduced alkamides. Mangifera indica L. and Thalassia testudinum extracts, two natural polyphenol‐rich extracts, diminished CYP1A1/2 and 3A4 activities, but not the CYP2D6 activity. These results suggest that these herbs inhibit the major human P450 enzymes involved in drug metabolism and could induce potential herbal‐drug interactions. Copyright

Effects of D-003, a new hypocholesterolaemic and antiplatelet compound, on lipid profile and lipid peroxidation in healthy volunteers

AbstractBackground: D-003 is a mixture of long-chain aliphatic primary acids purified from sugarcane wax with hypocholesterolaemic effects proven in rabbits and healthy volunteers; it lowers serum total cholesterol (TC) and low-density lipoprotein-cholesterol (LDL-C) and increases high-density lipoprotein-cholesterol (HDL-C). D-003 also prevents lipoprotein lipid peroxidation in experimental models. Objective: To investigate the effects of D-003 on lipid profile and lipid peroxidation in healthy human volunteers. Participants: Forty-six healthy volunteers (24 women, 22 men). Methods: This double-blind, randomised, placebo-controlled study investigated the effects of D-003 at 5 and 10 mg/day on the susceptibility of LDL to lipid peroxidation induced by copper ions in healthy volunteers. Forty-six individuals were randomised (1 : 2) to placebo or D-003 at 5 or 10 mg/day, the tablets being taken once a day with the evening meal for 8 weeks. Laboratory determinations and physical examination were performed at baseline and after 4 and 8 weeks of therapy, and compliance and adverse experience assessments were performed at weeks 4 and 8. Results: All groups were well matched at baseline. At study completion, D-003 at 5 and 10 mg/day significantly (p < 0.001) lowered LDL-C, the primary response variable, by 20.8% and 28.8%, respectively. In addition, D-003 at 5 and 10 mg/day reduced (p < 0.001) TC (12.7% and 17.5%, respectively), LDL-C/ HDL-C (25.9% and 36.3%, respectively) and TC/HDL-C (18.6% and 26.3%, respectively), while significantly (p < 0.01) increasing HDL-C (7.7% and 12.4%, respectively). Triglycerides were significantly (p < 0.05) reduced (8.8% and 13.1%, respectively) with respect to baseline, but not versus placebo. Responses assessed at 4 weeks showed significant reductions of LDL-C, TC and atherogenic ratios with both doses of D-003, whereas HDL-C was significantly increased. Triglycerides, however, remained unchanged. No significant changes in any lipid profile variable occurred in the placebo group. D-003 at 5 and 10 mg/day significantly (p < 0.05) increased lag time (18.3% and 32.0%, respectively) and decreased maximum rate of diene propagation (Vmax) [12.7% and 19.1%, respectively] of copper-induced LDL peroxidation. D-003 5 and 10 mg/day attenuated the reduction of the reactivity against 2, 4, 6-trinitrobenzene sulfonic acid (TNBS) by 19.9% and 32.0%, respectively. The treatment was well tolerated. Three subjects (one from each group) discontinued the study. Only one, treated with D-003 5 mg/day, discontinued because of an adverse event (gastritis). Conclusions: D-003 at 5 and 10 mg/day demonstrated dose-dependent cholesterol-lowering effects in healthy volunteers characterised by reductions in LDL-C, TC and atherogenic ratios, and increases in HDL-C. Effects on triglycerides were modest and uncertain. As expected from experimental studies, D-003 inhibited the susceptibility of LDL to lipid peroxidation assessed by three indicators lag time Vmax and reactivity versus TNBS. Further studies investigating the effect of larger doses and treatment duration must be conducted to confirm the reproducibility of the present results in different study populations.

Comparison of the Efficacy, Safety and Tolerability of Policosanol versus Fluvastatin in Elderly Hypercholesterolaemic Women

ObjectiveTo compare the efficacy and tolerability of policosanol with that of fluvastatin in older hypercholesterolaemic women.Design and SettingRandomised, single-blind, parallel-group study performed at a single centre in Cuba.Patients and Participants70 women aged 60 to 80 years with type II hypercholesterolaemia.MethodsPatients were randomised after 4 weeks’ dietary stabilisation on a step-one cholesterol-lowering diet to treatment with policosanol (10mg) or fluvastatin (20mg) tablets once daily for 8 weeks.ResultsPolicosanol significantly lowered low density lipoprotein cholesterol (LDL-C) [29.2%, p < 0.001], total cholesterol (TC) [19.3%, p <0.001], triglycerides (7%, p < 0.05) and the ratios of LDL-C (39.8%, p < 0.001) and TC (31.6%, p < 0.001) to high density lipoprotein cholesterol (HDL-C), and significantly increased HDL-C (19.8%, p < 0.001). Fluvastatin significantly lowered LDL-C (22.9%, p < 0.001), TC (16.7%, p < 0.001), triglycerides (8.2%, p < 0.05), LDL-C/HDL-C (28.4%, p < 0.001) and TC/HDL-C (22.8%, p < 0.001), and significantly increased HDL-C (9.2%, p < 0.001). Policosanol was more effective than fluvastatin in reducing LDL-C (p < 0.01), TC/HDL-C (p < 0.01) and LDL-C/HDL-C (p < 0.001) as well as in increasing HDL-C (p < 0.01). Policosanol, but not fluvastatin, significantly increased lag time for LDL lipid peroxidation (36.5%, p < 0.001) and significantly decreased the diene peroxidation rate (15.5%, p < 0.05). Both treatments were well tolerated. Five fluvastatin, but no policosanol, recipients discontinued the study, three because of adverse events (chest pain and gastric discomfort, skin rash, and dizziness). Overall, three policosanol and five fluvastatin recipients reported adverse events during the study.ConclusionsThe cholesterol-lowering effects of policosanol 10 mg/day administered for 8 weeks to older women with type II hypercholesterolaemia were slightly better than those of fluvastatin 20 mg/day with respect to the extent of the changes in LDL-C, atherogenic indices and HDL-C levels. In addition, policosanol, but not fluvastatin, significantly inhibited the susceptibility of LDL to undergo lipid peroxidation in this particular study population. Nevertheless, further studies in larger populations and with higher dosages must be conducted to corroborate the present results.

Acute and oral subchronic toxicity of D-003 in rats

D-003 is a mixture of higher aliphatic primary acids purified from sugar cane wax (Saccharum officinarum) with cholesterol-lowering and antiplatelet effects experimentally proven. The present work reports the results of two studies investigating the acute and subchronic oral toxicity of D-003 in rats. Oral acute toxicity of D-003 (2000 mg/kg) was investigated according to the Acute Toxic Class (ATC) method (an alternative for the classical LD(50) test), which was performed in Wistar rats. The results obtained in this study defined D-003 oral acute toxicity as unclassified. In the subchronic study, rats of both sexes were orally treated with D-003 at 50, 200 and 1250 mg/kg for 90 days. At this time, animals were sacrificed. No evidence of treatment-related toxicity was detected during the study. Thus, data analysis of body weight gain, food consumption, clinical observations, blood biochemical, haematology, organ weight ratios and histopathological findings did not show significant differences between control and treated groups. It is concluded that D-003 orally administered to rats was safe and that no drug-related toxicity was detected even at the highest doses investigated in both acute (2000 mg/kg) and subchronic (1250 mg/kg) studies.

Effects of policosanol on the susceptibility of low-density lipoprotein isolated from hypercholesterolemic patients at high coronary risk to in vitro copper-mediated lipid peroxidation: a randomized, double-blind pilot study

Abstract Objective: The aim of the study was to investigate the effects of policosanol administered at its starting dosage (5 mg/d) on low-density lipoprotein (LDL) peroxidation in patients with type II hypercholesterolemia and high coronary risk. Background: Oxidation of LDL has been suggested as a step in the development of atherosclerosis. Policosanol is a cholesterol-lowering drug that inhibits lipid peroxidation in experimental models and healthy volunteers. Methods: In this randomized, double-blind, placebo-controlled trial, after 6 weeks of dietary stabilization, 20 patients were randomly assigned to receive placebo or policosanol 5-mg tablets once daily for 12 weeks. LDL isolated at baseline and after 12 weeks of therapy was submitted to in vitro copper-catalyzed time-course experiments. Results: The 2 groups were statistically similar at randomization. When compared with baseline, policosanol significantly decreased ( P P P P P P P P P P Conclusions: The present study demonstrated that policosanol 5 mg/d not only lowered TC, LDL-C, and atherogenic indices, and increased HDL-C, but also significantly decreased the susceptibility of LDL to copper ion-induced lipid peroxidation in vitro in hypercholesterolemic patients at high coronary risk.

Antioxidant Effect of D-002 on Gastric Mucosa of Rats with Experimentally Induced Injury

D-002 is a natural mixture of higher aliphatic primary alcohols isolated from beeswax that has antioxidant and antiulcer properties. Because the role of lipid peroxidation in gastric damage is well recognized, this work was designed to investigate the possible effect of D-002 on lipid peroxidation in gastric mucosa in two experimental models of gastric injury in rats: (1) gastric ulcer induced by indomethacin and (2) mucosal injury induced by ischemia-reperfusion (I-R). The results demonstrated a remarkable protective effect of D-002 on lipid peroxidation in gastric ulcer induced by indomethacin and a moderate protective effect of D-002 on gastric erosions and lipid peroxidation induced by I-R.

Six-Month Toxicity Study of Oral Administration of D-003 in Sprague Dawley Rats

AbstractBackground: D-003 is a mixture of high molecular weight aliphatic primary acids purified from sugarcane wax (Saccharum officinarum) having cholesterol-lowering and antiplatelet effects. Aim: This study was undertaken to investigate the toxicity induced by long-term oral administration of D-003 for 6 months to Sprague Dawley rats of both sexes. Methods: Rats were randomly divided into four groups (20 rats of each sex/group): a control group, which received the vehicle, and three treatment groups, which received oral D-003 at doses of 250, 500 and 1000 mg/kg/day, respectively. Daily clinical observations and control of bodyweight and food consumption were conducted throughout the study period. On completion of active treatment, animals were sacrificed. Pharmacological effects associated with D-003 such as inhibition of platelet aggregation and increase in bleeding time were assessed in two satellite groups (14 animals of each sex/group): a control group and a group treated with the highest dose of D-003. Assessments of platelet aggregation to collagen were performed at baseline and at 6 months, and assessments of bleeding time were done at baseline, after 3 and 6 months of treatment, and after 30 days’ washout. Results: As expected, D-003 significantly inhibited platelet aggregation. Bleeding time was increased after 3 months of treatment with D-003; this increase was maintained at 6 months, and was reversible after washout. Coagulation factors such as prothrombin time and kaolin-activated thromboplastin-time, which were determined in eight male animals from each group, were unaffected by D-003. Data analyses of bodyweight gain, food consumption, clinical observations, blood biochemistry, haematology, organ weight ratios and histopathological findings did not show trends related to D-003 dose or significant differences between control and treated groups. Conclusion: It was concluded that the highest studied dose of D-003 (1000 mg/kg /day) represented a non-toxic dose level in the present chronic toxicity study in rats.