DLM: Elevated Triglycerides and Omega-3 Fatty Acids (2007)

Citation:
 
Study Design:
Class:
- Click here for explanation of classification scheme.
Quality Rating:
Research Purpose:
To evaluate the lipid-lowering efficacy of Omacor in subjects with familial combined hyperlipidemia (FCHL) as well as changes in LDL particle distribution
Inclusion Criteria:
  • Followed as an outpatient for several years at the E. Grossi Paoletti Lipid Clinic
  • Previous diagnosis of familial combined hyperlipidemia according to the following criteria
    • Primary hyperlipidemia, defined by a plasma cholesterol and/or triglyceride level exceeding the 90th percentile in the general population, adjusted for age and sex
    • Varying hyperlipidemia phenotype during at least 1-year follow-up
    • At least one first-degree relative with a hyperlipidemia phenotype different from the index patient
    • Presence of an LDL phenotype B, defined by a major LDL particle subpopulation with a diameter < 25.5 nm by non-denaturing polyacrylamide GGE
  • Following a standard low fat (30% of calories) diet for at least 6 months
Exclusion Criteria:
  • Plasma Lp(a) concentration > 30 mg/dl
  • Hormonal therapies, ß-blockers, diuretics or other products with the potential of affecting lipid metabolism
    • No lipid lowering treatments at least 3 months before the start of the study
Description of Study Protocol:

Recruitment Not described

 

Design Randomized, doubled blind, crossover trial

 

Blinding used (if applicable) not described

 

Intervention (if applicable)

  • Run in phase:
    •  Visit —2 (4 weeks prior to randomization)
    • Visit —1 (2 weeks prior to randomization)
  • Visit 0: randomized to group A or B
    • Group A:
      • Omacor first 8 weeks followed by placebo for 8 weeks
    • Group B:
      • Placebo first 8 weeks followed by Omacor for 8 weeks
  • Examined every 4 weeks
    • Visit 1: 4 weeks
    • Visit 2: 8 weeks
    • Visit 3: 12 weeks
    • Visit 4: 16 weeks
  • Visit 5: 6 weeks after the end of treatment

 

Statistical Analysis

  •  Results for normally distributed variables are given as means±SD; results for non-normally distributed variables are given as medians and ranges.
  • Individual plasma lipid and lipoprotein measures at visits —2, —1, 0 and 5 (baseline), and after 4 and 8 weeks of Omacor or placebo treatment, were averaged before comparison.
  • Changes caused by treatments were analyzed by using repeated measures ANOVA with post hoc evaluation by the Neuman-Keuls test.
  • A probability value of less than 0.05 was considered significant.
  • Correlation coefficients (r) were calcuated and the significance of the correlation determined by the F parameter; a logarithmic transformation was applied to non-normally distributed data.
  • A stepwise regression analysis was performed by entering the independent variable with the highest partial correlation coefficient at each step, until no variable remained with an F value of 4 or higher.
Data Collection Summary:

Timing of Measurements

 

  Physical exam Standard biochemical & hematological analyses Plasma lipid & apoprotein levels Blood pressure Body weight Lipoprotein characterization Food record
Adherence to drug

Visit       — 2

x   x x x      
Visit —1 x x x x x      
Visit 0 x   x x x x    
Visit 1 x   x x x   x x
Visit 2 x x x x x x   x
Visit 3 x   x x x   x x
Visit 4 x x x x x x   x
Visit 5 x x x x x x    

Dependent Variables

  • Lipid analyses:
    • Blood samples were collected after an overnight fast. Lipid analyses were performed within the same day of blood collection while lipoprotein fractionation started within 2 days.
    • Plasma total and unesterified cholesterol, triglyceride and phospholipid levels were determined with standard enzymatic techniques.
    • Plasma HDL-cholesterol levels were routinely measured after precipitation of the apo-B containing lipoproteins
    • Apo A-1 and B levels were determined by immunoturbidimetry, using commercially available polyclonal antibodies.
    • Lp(a) concentrations were measured by a sandwich-ELISA.
    • Plasma phospholipid fatty acid composition and concentrations were mesured by a gas-liquid chromatographic method.
    • Apo E phenotyping was performed by isoelectric focusing.
    • LDL subfractions
    • LDL particle size distributions were analyzed by non-denaturing polyacrylamide GGE
  • Adherence with Omacor or placebo
    • determined by pill counts
    • fatty acid composition of plasma phospholipids
  • Three day food record
    • to estimate composition of the subjects' diets
    • completed during the 4th week of each treatment period and then analyzed by an experienced dietitian
    • urged to maintain their dietary habits unless they consumed large amounts of fish, in which case, they were asked to restrict it during the study.

Independent Variables

  • Omacor: 3.4 g EPA + DHA per day (4 capsules/day)
  • Placebo

Control Variables

 

Description of Actual Data Sample:

 

Initial N: N=14 (11 males, 3 females)

  • 7 subjects were randomized to each group
  • Authors did not provide any additional information about each group

Attrition (final N):

  • All subjects completed the trial except for 1 subject (emergency hospitalization) in group B who had no data for Visit 5

Age: (mean calculated by reviewer)

  • Male: 43.5 yr (range 26-64)
  • Females: 62.7 yr (range 62-64)

Ethnicity: not described

Other relevant demographics: not described

Anthropometrics

  • Individual data for each subject at randomization was provided which included age; gender; plasma concentrations of triglyceride, total cholesterol, VLDL-C, LDL-C, HDL-C, ApoB; LDL size; and LDL flotation rate.
  • No means/medians with SD or ranges calculated
  • None of the subjects had an apo E2/E2 phenotype. All were homozygous for apo E3, except 1 with an apo E2/E3 phenotype and 1 with an apo E3/E4 phenotype.
  • None of the subjects were overweight (+10% according to the Metropolitan Height and Weight Tables)

Location: E. Grossi Paoletti Lipid Clinic, Institute of Pharmacological Sciences, University of Milano, Milan, Italy

 

Summary of Results:

 Plasma lipid and lipoprotein levels at baseline, and during placebo and Omacor treatment (mg/dl)

  Baseline Placebo Omacor
Total cholesterol 270.7±33.1 266.0±38.9 282.5±44.0
VLDL-cholesterol 39.5 (30-206) 45.5 (20-199) 32.5±(22-92)*
LDL-cholesterol 167.1±47.6 161.5±50.6 202.7±45.8§
HDL-cholesterol 41.5±11.5 39.7±10.0 42.5±14.7
Triglycerides 251.1 (141-799) 231.8 (149-1030) 183.5 (75-407)*
ApoA-I 108.8±27.2 113.5±19.6 112.1±26.8
ApoB 135.2±21.1 134.1±26.1 143.8±23.6*
*Significantly different from baseline and placebo.

§Significantly different from placebo.

 

  1. Plasma triglyceride levels were significantly lower during Omacor (—27 and —21% compared with baseline and placebo, respectively).
  2. There were no significant differences in plasma total and HDL-cholesterol among the three periods.
  3. VLDL-cholesterol was significantly lower (—18 and —29% compared with baseline and placebo, respectively).
  4. LDL-cholesterol was higher (+25% compared with placebo, P=0.032; +21% compared with baseline, P=0.054) during Omacor.
  5. No significant differences in plasma ApoA-I levels among the three periods, but there was a significant 6% increase in ApoB with Omacor compared to baseline and placebo.
  6. These changes resulted in a significant increase of the LDL-cholesterol/ApoB ratio after Omacor (1.40±0.17 vs 1.27±0.26 and 1.19±0.29 at baseline and placebo, respectively.
Correlations
  1. Highly significant correlations were found among the individual responses in various parameters to Omacor treatment, suggesting a common mechanism for the observed lipid/lipoprotein modifications.
  2. Plasma triglycerides and VLDL-cholesterol
    • The change in plasma triglycerides and VLDL-cholesterol correlated with changes in LDL-cholesterol (r=0.736 and 0.647, respectively) and in the LDL-cholesterol/ApoB ratio (r=0.791 and 0.741, respectively).
    • Individual triglyceride and LDL-cholesterol responses to Omacor treatment varied widely among the subjects (from —753 to +45 gm/dl and from —4 to +114.5 gm/dl, respectively).
    • Plasma triglyceride reductins correlated significantly with the increase in DHA (r=0.678) but not EPA content of plasma phopholipids.
    • The decrease of plasma triglycerides also correlated significantly and positively with baseline triglycerides (r=0.893) and VLDL-C (r=0.816), and negatively with baseline LDL-cholesterol (r=0.631), LDL-cholesterol/apoB ratio (r=0.826) and LDL size (r=0.744).
    • By multivariate regression analysis, baseline triglycerides were the strongest predictor of Omacor-induced triglyceride reduction (r2=0.798), but LDL size and LDL flotation rate made significant contributions to this correlation (r2=0.890 and 0.926, respectively).
      • The DHA content of plasma phospholipds did not add significantly to this correlation.
      • A combination of bseline triglycerides, LDL size and flotation rate explained more than 90% of the variation of plasma triglyceride response following Omacor treatment.
  3. Plasma LDL-cholesterol was
    • Not related to drug compliance as no significant correlation was found with the changes in EPA or DHA content of plasma phospholipids.
    • The LDL-cholesterol rise correlated significantly and positively with baseline triglycerides (r=0.571) and VLDL-C (r=0.538), and negatively with baseline LDL-cholesterol/ApoB ratio (r=0.659) and LDL size (r=0.645).
    • By multivariate regression analysis, the LDL-cholesterol/ApoB ratio was the sole predictor of LDL-cholesterol change after Omacor (r=0.434).
Percentage composition of plasma lipoproteins at baseline, and during placebo and Omacor treatment
  Unesterified cholesterol Cholesteryl esters Triglycerides Phospholipids Proteins
VLDL          
Baseline 6.45±0.5 19.01±3.44 50.51±5.24 17.33±1.13 6.68±2.07
Placebo 6.73±0.74 17.56±2.64 51.58±4.35 17.55±1.08 6.58±2.05
Omacor 6.52±0.81 18.15±3.86 47.90±5.86 18.70±1.44* 8.71±2.60*
LDL          
Baseline 8.95±0.89 39.06±4.14 9.64±4.76 18.98±1.25 23.32±1.51
Placebo 8.84±0.60 38.41±4.59 10.41±5.34 19.40±1.26 22.92±0.93
Omacor 9.45±0.92* 40.35±2.67* 8.98±2.72 18.83±1.63 22.40±1.12**
HDL          
Baseline 2.32±0.58 18.04±1.97 5.42±2.35 22.85±1.73 51.34±2.56
Placebo 2.45±0.40 18.04±2.78 5.89±2.65 22.38±2.02 51.22±2.62
Omacor 2.66±0.47* 19.59±1.98* 5.08±2.09 22.23±1.76 50.44±2.17
*Significantly different from baseline and placebo.
**Significantly different from baseline.
  1. Lipoprotein composition did not change after placebo administration but  displayed significant variations after Omacor treatment.
  2. VLDL became enriched in phospholipids and proteins, where as opposite changes were found in LDL, that also displayed a higher cholesteryl ester content.
  3. These changes were indicative of the accumulation in plasma of less buoyant and core depleted VLDL, and of more buoyant and core enriched LDL.
  4. Surface core ratio increased in VLDL from 0l44±0.06 to 0.52±0.08, and decreased in LDL from 1.06±0.06 to 1.01±0.03 (both P<0.01).
LDL Particle Distribution
  1. At baseline (expected) there was a single major LDL subpopulation of relatively small size (24.9±0.3 nm) with a very minor additional components.
  2. The diameter of the major LDL subpopulatoin correlated significantly and negatively with plasma triglycerides (r=0.549), and positively with the LDL-cholesterol/ApoB ratio (r=0.559) and the LDL flotation rate (r=0.547).
  3. The mean LDL diameter did not change signficantly after placebo (24.8±0.5 nm) or Omacor (25.0±0.3 nm).
  4. An LDL subclass pattern B persisted after treatment in all examined patients.
LDL Flotation Rate
  1. At baseline, a single polydisperse LDL component, indicative of the presence of multiple LDL subspecies, was detectable.
  2. The LDL flotation rate correlated significantly and positively with plasma HDL-cholesterol (r=0.836), ApoA-I (r=0.777), and the LDL-cholesterol/ApoB ratio (r=0.594) and the VLDL-cholesterol (r=0.599).
  3. The LDL flotation rate did not change after placebo (4.24±0.38 vs 4.11±0.27 ml), but it increased significantly after Omacor (4.43±0.38 ml) indicative of the accumulation in plasma of fast floating LDL subspecies.
  4. The plasma concentrations of IDL and of the more buoyant, fast floating LDL-1 and LDL-2 subclasses increased significantly after Omacor (see table below) whereas the concentration of the denser, slow floating LDL-3 subclass decreased, although not significantly (P=0.065).
Plasma concentration of IDL and LDL subfractions at baseline, during placebo and Omacora
  Baseline Placebo(mg/dl) Omacor
IDL 18.6±4.4 19.7±8.3 23.3±6.9*
LDL-1 14.0±3.1 15.2±7.4 20.3±10.2
LDL-2 28.5±7.5 27.6±10.5 36.0±11.5*
LDL-3 49.7±12.1 41.9±14.5 40.0±17.3
a Results are expressed as concentrations of ApoB (mg/dl).
*Significantly different from baseline and placebo.


Adherence
  1. Monitoring of drug intake, by pill count, and measurement of fatty acid profile indicated that compliance to treatment was satisfactory (see table below).
Fatty acid composition of selected plasma phospholipids during the study (mg/ml)
Fatty acids Visit 0 Placebo Omacor Visit 5
18:2  n-6 269.1±52.4 291.4±49.9 227.8±51.3* 272.3±45.1
20:3 n-5 50.1±13.0 52.9±15.0 32.3±10.3* 59.9±10.5
20:4 n-6 148.6±35.5 146.9±3.1 113.3±26.0* 161.2±34.3
20:5 n-3 11.9±7.2 10.1±4.5 65.5±25.3* 13.7±6.5
22:5 n-3 12.2±3.5 12.8±4.1 22.7±7.4* 15.0±3.3
22:6 n-3 66.0±25.1 68.0±23.3 112.2±24.9* 79.3±21.9
*Significantly different from baseline and placebo.
       
       

    Other Findings

    •  Tolerance to Omacor
      • No drug-related adverse events were reported
      • No subject discontinued treatment because of side effects
      • Omacor did not affect safety parameters including glucose, uric acid, liver enzymes, kidney function and platelet counts
    • No apparent carry-over efect of triglyceride lowering from omacor to the end of the placebo period in group A.
    • Body weight didn't change significantly during the study.
    • Blood pressure didn't change significantly during the study.
Author Conclusion:
  • Demonstrates that Omacor effectively lowers plasma triglycerides and VLDL in subjects with familial combined hyperlipidemia and causes a redistribution of LDL subclasses towards less dense lipoprotein particles. However, the abnormal LDL size is not affected by treatment.
  • Findings confirm that the LDL size is familial combined hyperlipidemia is largely controlled by genetic factors and independent of plasmalipid/lipoprotein levels.
  • The effects of Omacor on the plasma lipid/lipoprotein profile, with a decrease in the number of circulating VLDL particles and a shift from dense LDL particles to buoyant LDL (possibly indicative of a less atherogenic LDL profile), must be balanced with the intrinsic high CHD risk associated with the predominance in plasma of small LDL, which is not affected by Omacor treatment.
  • The apparent resistance of these abnormal LDL to drug-induced modifications implies that a maximal lipid-lowering effect must be achieved to reduce risk in familial combined hyperlipidemia patients. The combination of a statin with Omacor should provide the best results.
 
Funding Source:
Reviewer Comments:
No washout period between Omacor and placebo treatments. Did the investigators know in advance that one was not needed?

Why were subjects randomized to group A or B? Unclear to me whether subjects served as their own control or data were analyzed by group. No group characteristics provided at baseline just individual subject data.

What was the placebo? Could subjects differentiate the placebo from the Omacor?

Adherence: reported as "satisfactory" but "satisfactory" was not defined.

Short term study: 8 weeks of Omacor therapy

Sections of the article were poorly written and difficult to understand (especially the statistics section); tables should have mentioned whether data were means or medians; P values were inconsistently reported throughout the paper.

Supported in part by a grant from Pronova Biocare, Oslo, Norway. Pronova Biocare supplied Omacor capsules as well.
Quality Criteria Checklist: Primary Research
Relevance Questions
  1. Would implementing the studied intervention or procedure (if found successful) result in improved outcomes for the patients/clients/population group? (Not Applicable for some epidemiological studies) Yes
  2. Did the authors study an outcome (dependent variable) or topic that the patients/clients/population group would care about? Yes
  3. Is the focus of the intervention or procedure (independent variable) or topic of study a common issue of concern to dieteticspractice? Yes
  4. Is the intervention or procedure feasible? (NA for some epidemiological studies) Yes
 
Validity Questions
1. Was the research question clearly stated? Yes
  1.1. Was (were) the specific intervention(s) or procedure(s) [independent variable(s)] identified? Yes
  1.2. Was (were) the outcome(s) [dependent variable(s)] clearly indicated? Yes
  1.3. Were the target population and setting specified? Yes
2. Was the selection of study subjects/patients free from bias? Yes
  2.1. Were inclusion/exclusion criteria specified (e.g., risk, point in disease progression, diagnostic or prognosis criteria), and with sufficient detail and without omitting criteria critical to the study? N/A
  2.2. Were criteria applied equally to all study groups? Yes
  2.3. Were health, demographics, and other characteristics of subjects described? Yes
  2.4. Were the subjects/patients a representative sample of the relevant population? ???
3. Were study groups comparable? ???
  3.1. Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT) No
  3.2. Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline? ???
  3.3. Were concurrent controls or comparisons used? (Concurrent preferred over historical control or comparison groups.) ???
  3.4. If cohort study or cross-sectional study, were groups comparable on important confounding factors and/or were preexisting differences accounted for by using appropriate adjustments in statistical analysis? N/A
  3.5. If case control study, were potential confounding factors comparable for cases and controls? (If case series or trial with subjects serving as own control, this criterion is not applicable.) N/A
  3.6. If diagnostic test, was there an independent blind comparison with an appropriate reference standard (e.g., "gold standard")? N/A
4. Was method of handling withdrawals described? Yes
  4.1. Were follow-up methods described and the same for all groups? Yes
  4.2. Was the number, characteristics of withdrawals (i.e., dropouts, lost to follow up, attrition rate) and/or response rate (cross-sectional studies) described for each group? (Follow up goal for a strong study is 80%.) Yes
  4.3. Were all enrolled subjects/patients (in the original sample) accounted for? Yes
  4.4. Were reasons for withdrawals similar across groups? N/A
  4.5. If diagnostic test, was decision to perform reference test not dependent on results of test under study? N/A
5. Was blinding used to prevent introduction of bias? Yes
  5.1. In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? ???
  5.2. Were data collectors blinded for outcomes assessment? (If outcome is measured using an objective test, such as a lab value, this criterion is assumed to be met.) Yes
  5.3. In cohort study or cross-sectional study, were measurements of outcomes and risk factors blinded? N/A
  5.4. In case control study, was case definition explicit and case ascertainment not influenced by exposure status? N/A
  5.5. In diagnostic study, were test results blinded to patient history and other test results? N/A
6. Were intervention/therapeutic regimens/exposure factor or procedure and any comparison(s) described in detail? Were interveningfactors described? Yes
  6.1. In RCT or other intervention trial, were protocols described for all regimens studied? Yes
  6.2. In observational study, were interventions, study settings, and clinicians/provider described? N/A
  6.3. Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect? Yes
  6.4. Was the amount of exposure and, if relevant, subject/patient compliance measured? Yes
  6.5. Were co-interventions (e.g., ancillary treatments, other therapies) described? N/A
  6.6. Were extra or unplanned treatments described? N/A
  6.7. Was the information for 6.4, 6.5, and 6.6 assessed the same way for all groups? Yes
  6.8. In diagnostic study, were details of test administration and replication sufficient? N/A
7. Were outcomes clearly defined and the measurements valid and reliable? Yes
  7.1. Were primary and secondary endpoints described and relevant to the question? Yes
  7.2. Were nutrition measures appropriate to question and outcomes of concern? Yes
  7.3. Was the period of follow-up long enough for important outcome(s) to occur? Yes
  7.4. Were the observations and measurements based on standard, valid, and reliable data collection instruments/tests/procedures? Yes
  7.5. Was the measurement of effect at an appropriate level of precision? Yes
  7.6. Were other factors accounted for (measured) that could affect outcomes? Yes
  7.7. Were the measurements conducted consistently across groups? Yes
8. Was the statistical analysis appropriate for the study design and type of outcome indicators? Yes
  8.1. Were statistical analyses adequately described and the results reported appropriately? Yes
  8.2. Were correct statistical tests used and assumptions of test not violated? Yes
  8.3. Were statistics reported with levels of significance and/or confidence intervals? Yes
  8.4. Was "intent to treat" analysis of outcomes done (and as appropriate, was there an analysis of outcomes for those maximally exposed or a dose-response analysis)? Yes
  8.5. Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)? No
  8.6. Was clinical significance as well as statistical significance reported? Yes
  8.7. If negative findings, was a power calculation reported to address type 2 error? No
9. Are conclusions supported by results with biases and limitations taken into consideration? Yes
  9.1. Is there a discussion of findings? Yes
  9.2. Are biases and study limitations identified and discussed? No
10. Is bias due to study's funding or sponsorship unlikely? Yes
  10.1. Were sources of funding and investigators' affiliations described? Yes
  10.2. Was the study free from apparent conflict of interest? Yes