Statins for Oxidative Stress and Mitochondrial Function in Diabetic Polyneuropathy

Oxidative Stress Clinical Trial

Official title:

Ezetimibe/Simvastatin and Rosuvastatin for Oxidative Stress and Mitochondrial Function in Diabetic Polyneuropathy: a Randomized, Double Blinded, Placebo Controlled Clinical Trial

Clinical Trial Details — Status: Completed

Administrative data

• NCT number: NCT02129231
• Other study ID #: STAT/DPN_2014
• Status: Completed
• Phase: Phase 2
• First received: April 30, 2014
• Last updated: May 5, 2014
• Start date: February 2012
• Est. completion date: April 2014

Study information

• Verified date: May 2014
• Source: University of Guadalajara
• Contact: n/a
• Is FDA regulated: No
• Health authority: Mexico: Ethics Committee
• Study type: Interventional

Clinical Trial Summary

Aims: To evaluate the effect of ezetimibe/simvastatin and rosuvastatin on oxidative stress and mitochondrial function in patients with DPN.

Methods: We performed a randomized, double-blinded, placebo-controlled phase II clinical trial in adult patients with type 2 Diabetes Mellitus (T2DM) who had Diabetic Polyneuropathy (DPN) evaluated by composite scores and nerve conduction studies (NCS), HBA1C <12% (108 mmol/mol), previous exclusion of other neuropathies. Ninety-eight persons with T2DM were allocated 1:1:1 to either placebo, ezetimibe/simvastatin 10/20 mg or rosuvastatin 20 mg for 16 weeks, and healthy controls (not randomized) were included for comparisons. Primary outcomes were lipid peroxidation (LPO), nitric oxide (NO), and total antioxidant capacity (TAC); secondary were clinical, NCS and metabolic parameters. Results were expressed as mean ± standard deviation (SD) or standard error of the mean (SEM), frequencies and percentages. Non-parametric analysis was used.

Description:

Introduction Nerve dysfunction system in patients with diabetes is known as diabetic neuropathy and is considered as the most prevalent microvascular complication -up to 60%- in type 2 Diabetes Mellitus (T2DM) subjects. Diabetic polyneuropathy (DPN) comprise ≈70% of all cases. Its diagnosis is established by means of validated scores based on clinical features and abnormal nerve conduction studies (NCS). Pathophysiologic findings include loss of multifocal and focal nerve fibers secondary to axonal degeneration an segmental demyelinization, basically due to oxidative stress induced by chronic hyperglycemia, which leads to neural apoptosis. Other mechanisms involved in peripheral nerve injure is nitrosative stress induced by nitric oxide (NO).

Ezetimibe diminishes cholesterol esters content in chylomicrons by reducing liver cholesterol intake which in consequence increases LDL uptake and plasma depuration; as monotherapy it reduces LDL-C by 17%. When combined with simvastatin, cholesterol reduction is potentially increased; furthermore, pleiotropic effects of statins include an increase on nuclear factor kappa B activity and amelioration of superoxide ions after 12 week treatment. Another hydroxy-methyl-glutaryl coenzyme A inhibitor, rosuvastatin, has an antioxidant effect by acting as free radical carrier diminishing mitochondrial and cellular lipid peroxidation (LPO) production.

We conducted this study to evaluate the effect of ezetimibe/simvastatin and rosuvastatin on oxidative stress in patients with DPN.

Methods Study design A randomized, double blinded, placebo controlled phase II clinical trial was performed in the Clinic and Experimental Therapeutics Institute, University of Guadalajara, Mexico. Subjects were assigned to three group treatments in blocks with a parallel sequence 1:1:1 through a randomized computer-based list, generated by a different researcher unaware of drugs given. Patients were divided to: control group that received placebo, ezetimibe/simvastatin and rosuvastatin as a daily single dose for 16 weeks. The selection period was performed from February 2012 to January 2013. We selected 5 non-randomized healthy subjects (HS) from a blood bank to compare the oxidative stress status.

Study population Inclusion criteria were ≥18 years old, T2DM according to American Diabetes Association and DPN by Dyck et. al.3 criteria, HbA1c <12%, and informed consent signed. They were excluded if renal or hepatic failure, pregnant or breastfeeding, other neuropathies (alcohol-induced, radiculopathy, autoimmune, cancer-related), and eliminated if lack treatment adherence (<80% of drug intake), severe adverse drug reaction and/or serious health illness. Patients were selected by invitation in forums; outpatients recruited from primary care clinics; and database collected previously by our Institute from February 2010 to 2012. Patients were instructed to take their drugs only by night at the same time every day, as follows: placebo 100 mg, ezetimibe/simvastatin 10/20 mg, and rosuvastatin 20 mg. All drugs were similar in physical characteristics and presented in dark vials, carefully filled by another group researcher, who placed a respective tag with the patient code. Also, patients were provided with a diary where they would write down the date and time of drug administration, and drug adverse reactions felt. Such information was collected and registered every 4 weeks. Primary outcomes were oxidative stress markers LPO, NO, and TAC before and after 16 week intervention. Secondary outcomes were clinical, NCS and metabolic [fasting glucose, HbA1c, total cholesterol, high and low density lipoproteins (HDL, LDL), and triglycerides] parameters. Safety profile was assessed with drug adverse reactions, renal (urea, creatinin), and hepatic [(alamin- and aspartate-aminotransferase, gama-glutamyltransferase, bilirubins and phosphokinase] laboratory variables.

Oxidative stress and mitochondrial function markers LPO was measured according to kit specifications (Oxford Biomedical Research Inc., FR12), 200 μL of serum where processed with a chromogen substance that reacts with malondialdehyde (MDA) and 4-hydroxy-alkenals (HNA), the absorbance measured at 586 nm, and results expressed in nmol/mL.

Previous deproteinization of the samples, we performed a colorimetric for determining the concentration of NO with 85 µL of serum (Nitric Oxide Assay Kit, User protocol 482650), with results expressed as pmol/mL.

Total antioxidant capacity (TAC) was realized with 200 µL of serum, to obtain values of millimole (mM) equivalent of uric acid (Total Antioxidant Power Kit, No. 02090130, Oxford Biomedical Research®).

Clinical and nerve conduction variables Neuropathic symptoms (NSS) and disability scores (NIS) described by Dyck, et. al. were obtained by physical examination and anamnesis. We also measured the latency, duration, amplitude and motor nerve conduction velocity from fibula, tibial, median and ulnae nerves, and sensitivity parameters from sural, median and ulnae nerves, as required by the American Association of Electrodiagnostic Medicine.

Ethical considerations The study was approved by the Research and Ethics Committee of the Health Science University Center, University of Guadalajara, Mexico. Identification codes were assigned to each participant to guarantee patient confidentiality, and an informed consent form was signed before entering the protocol, according to national and international laws, and also as stipulated by the Helsinki Statements (http://www.wma.net/es/30 publications/ 10 policies/b3/17c.pdf, accessed January 2011).

Statistical analysis The sample size was obtained by a clinical study design formula taking in account a difference change of 0.05 nmol/mL in LPO, 95% confidence interval, 80% potency, and two-tailed p<0.05, which resulted in 21 for each group. Quantitative variables were expressed as mean ± standard deviation. Kolmogorov-Smirnov and Shapiro-Wilk tests were performed to determine the non-parametric distribution of variables. Friedman and Wilcoxon tests were realized for before and after measurements, and Kruskal-Wallis with Mann-Whitney´s U as post-hoc analysis for between group comparisons. Qualitative variables were expressed as frequencies and percentages. Chi square test was used to evaluate differences in dichotomy variables before and after treatment, between group comparisons were determined by Fisher´s exact test and χ2 as needed. Significance level was established with p value <0.05.

Recruitment information / eligibility

• Status: Completed
• Enrollment: 74
• Est. completion date: April 2014
• Est. primary completion date: January 2014
• Accepts healthy volunteers: Accepts Healthy Volunteers
• Gender: Both
• Age group: 18 Years to 75 Years

Eligibility

Inclusion Criteria:

• 18 years old and older

• Type 2 Diabetes Mellitus according to American Diabetes Association

• Diabetic Polyneuropathy by Dyck et. al. criteria

• HbA1c <12%

• Informed consent signed

Exclusion Criteria:

• Renal or hepatic failure

• Pregnant or breastfeeding

• Other neuropathies (alcohol-induced, radiculopathy, autoimmune, cancer-related)

Study Design

Allocation: Randomized, Endpoint Classification: Safety/Efficacy Study, Intervention Model: Parallel Assignment, Masking: Double Blind (Subject, Caregiver, Investigator), Primary Purpose: Treatment

Related Conditions & MeSH terms

• Diabetic Neuropathies
• Diabetic Polyneuropathy
• Oxidative Stress
• Polyneuropathies

Intervention

Drug:
• calcined magnesia
We ensured the patient took the drug at night before meals
• Ezetimibe/simvastatin
We ensured the patient took the drug at night before meals
• Rosuvastatin
We ensured the patient took the drug at night before meals

Locations

Country	Name	City	State
Mexico	Cardiovascular Research Unit	Guadalajara	Jalisco

Sponsors (1)

Lead Sponsor	Collaborator
University of Guadalajara

Country where clinical trial is conducted

Mexico, 

References & Publications (1)

Hernández-Ojeda J, Cardona-Muñoz EG, Román-Pintos LM, Troyo-Sanromán R, Ortiz-Lazareno PC, Cárdenas-Meza MA, Pascoe-González S, Miranda-Díaz AG. The effect of ubiquinone in diabetic polyneuropathy: a randomized double-blind placebo-controlled study. J Diabetes Complications. 2012 Jul-Aug;26(4):352-8. doi: 10.1016/j.jdiacomp.2012.04.004. Epub 2012 May 16. — View Citation

Outcome

Type	Measure	Description	Time frame	Safety issue
Other	Creatinin	We measured creatinin to evaluate renal function.	16 weeks	Yes
Other	aspartate aminotransferase	We measured aspartate aminotransferase regarding hepatic function.	16 weeks	Yes
Other	alanine aminotransferase	We measured alanine aminotransferase regarding hepatic function.	16 weeks	Yes
Primary	lipid peroxidation	LPO was measured according to kit specifications (Oxford Biomedical Research Inc., FR12), 200 µL of serum where processed with a chromogen substance that reacts with malondialdehyde (MDA) and 4-hydroxy-alkenals (HNA), the absorbance measured at 586 nm, and results expressed in nmol/mL.	16 weeks	No
Primary	Nitric oxide	Previous deproteinization of the samples, we performed a colorimetric for determining the concentration of NO with 85 µL of serum (Nitric Oxide Assay Kit, User protocol 482650), with results expressed as pmol/mL.	16 weeks	No
Primary	Total antioxidant capacity	Total antioxidant capacity (TAC) was realized with 200 µL of serum, to obtain values of mM equivalent of uric acid (Total Antioxidant Power Kit, No. 02090130, Oxford Biomedical Research®).	16 weeks	No
Secondary	neuropathic symptoms score	Symptoms referred by the patient as pain and discomfort	16 weeks	No
Secondary	neuropathic impairment score	Neuropathic signs assessed by the physician through neurological techniques previously published by Dyck et. al.	16 weeks	No
Secondary	nerve conduction studies	Latency, duration, amplitude and motor nerve conduction velocity from fibula, tibial, median and ulnae nerves, and sensitivity parameters from sural, median and ulnae nerves, as required by the American Association of Electrodiagnostic Medicine.	16 weeks	No