Fish Oils and Adipose Inflammation Reduction
Condition: Obesity
Study type: Interventional
Phase: Phase 4

Summary: 
      This study is a clinical trial designed to assess whether fish oil treatments are effective
      in the prevention of obesity-related fat tissue (adipose) inflammation. Specifically it
      addresses the hypothesis that fish oils treatments will reduce signaling by chemokine
      pathways (fractalkine and MCP-1) important in adipose tissue for the recruitment and
      activation of certain white blood cells (macrophages). The study is a double-blind
      placebo-controlled trial of the fish oil Lovaza from GlaxoSmithKline (omega-3-acid ethyl
      esters; a combination of ethyl esters of eicosapentaenoic acid (EPA) and docosahexaenoic acid
      (DHA)) in obese non-diabetic adults, to determine if Lovaza decreases markers of inflammation
      and macrophage activation in adipose and blood and understand the mechanism by which fish
      oils affect inflammation.
    

Detailed description: 
      We propose to conduct a parallel arm, double blind, placebo-controlled trial to understand
      the anti-inflammatory and metabolic effects of marine derived omega-3 fatty acids (EPA + DHA)
      on obesity-related inflammation and chemokine signaling in humans. Obese (BMI>30 kg/m2) but
      non-diabetic young males and females (n~44; 25 to 50 years of age) taking no omega-3 fatty
      acid (EPA + DHA) supplements and consuming a low fish diet (defined as usual intake of high
      omega-3 fish (tuna and other non-fried fish) < 3 to 4 servings per month) will be included. A
      low habitual fish intake is important to control EPA + DHA intake during the supplement
      period (see exclusion criteria below). Subjects also will be required to maintain their
      current fish intake throughout the treatment period.

      There will be two treatment groups; (1) placebo and (2) omega-3 fatty acids 4 grams/day. The
      trial will be randomized, double blind, placebo-controlled, with parallel treatment groups;
      treatment with omega-3 fatty acids 8 weeks. We will enroll participants until at least 22
      subjects per study-arm complete all study visits (i.e., at least 44 participants).

      Prescription Lovaza (omega-3-acid ethyl esters) from GlaxoSmithKline at 4 grams day (2 1-gram
      capsules twice daily, with food) will be used. We expect this dose to be safe because it is a
      dose that is recommended, and routinely used, for management of moderate or worse
      hypertriglyceridemia (levels > 400 mg/dL). We will use matching placebo capsules so that all
      participants are taking 2 capsules twice daily - with food as recommended in clinical
      practice.

      The study duration was chosen in order to provide adequate time for potential
      anti-inflammatory effects of treatments without requiring maximum lipid modifying effects
      while also reducing the potential for drop-out and loss to follow up that is associated with
      longer study durations.

      Participants will be recruited from the Preventive Cardiology Research databases, comprised
      of healthy subjects who have participated in previous research studies and who have asked to
      be contacted for future studies. Potential participants will also be recruited via local
      IRB-approved flyers, internet, and newspaper advertisements (see flyer and advertisement
      attached) targeting the Delaware Valley region.

      STUDY PROCEDURES

      Informed Consent Before study initiation, the clinical protocol, the Informed Consent Form
      (ICF), and any advertisements for subject recruitment will be submitted for review and
      approval to the Institutional Review Board (IRB). The investigator, or his designated
      research staff, will obtain written informed consent from each subject enrolled in the study,
      in accordance with the U.S. Food and Drug Administration (FDA) regulations 21 CFR parts
      50.20-50.27. It is the responsibility of the investigator to ensure that informed consent is
      obtained from the subject or his/her guardian or legal representative before any activity or
      treatment is undertaken which is not part of routine care.

      Safety Assessments Safety will be evaluated by the incidence, severity, and relationship to
      treatment agent of adverse events.

      Monitoring for Adverse Events: Details of all AEs that occur after the first dose of study
      drug will be collected as indicated in Section 12. Any illness or injury that occurs before
      the first dose of a study drug will be recorded under Medical History and evaluated to
      determine if the occurrence affects the patient's eligibility to participate in the study.
      Subjects will be asked the following standard questions by the designated clinical evaluator:
      At clinic check-ins:

        1. "Have you had any medical problems since your last visit?"

        2. "Have any medical problems present at your last visit changed, i.e., stopped, worsened,
           or improved?"

        3. "Have you taken any medicines, other than study drug, since your last visit?" Any
           spontaneous AE information provided by the patient will be recorded. NOTE: An AE may be
           a new disease(s), any untoward event(s), or an exacerbation of a pre existing condition.
           See Section 12.0 for complete details.

      Adipose biopsy: Approximately 500-1000 mg of subcutaneous adipose tissue will be biopsied
      from the gluteal region, processed, weighed and frozen until analyzed. Adipose tissue will be
      obtained through a small (~5mm) incision under local anesthesia (2% Lidocaine without
      epinephrine), as has been previously described13-15. A portion (200 mg) of the samples will
      be flash frozen and analyzed for gene expression and protein assays. Another portion (500-800
      mg) will be placed in sterile media and undergo digestion with collagenase, filtration, and
      centrifugation for separation of the stromal vascular fraction (SVF). SVF cells will be
      stained with antibodies for flow cytometer analysis. Finally, remaining tissue will be fixed
      in paraffin for immunohistochemistry.

      6.5 Subgingival analysis

      Saliva samples will be collected using the OMNIgene DISCOVER (OM-505) kit (DNA Genotek), an
      all-in-one kit which collects and stabilizes both microbial DNA and RNA in saliva. Samples
      will be stored at -80C prior to extraction. Nucleic acids will be extracted using the QIAGEN
      QIAamp MinElute Virus Spin Kit (Qiagen). Subgingival plaque samples will be obtained from 3
      oral cavity regions in each subject (2 samples per region) using a soft-bristled microbrush
      (DenTek). Specimens will be collected from the same anatomical location in all subjects. DNA
      will be isolated using the PSP Spin Stool DNA kit (Invitec). DNA samples will be amplified
      using V1-V2 region primers targeting bacterial 16S genes and sequenced using IlluminaHiSeq or
      454 Pyrosequencing technology.

      Specific Study Procedures (by visit)

      Screening (Visit 1) Potential participants will be screened first by a telephone or e-mail
      interview with the research coordinator (see Appendix B for email interview form; for
      telephone interviews this information will be verbally exchanged). Participants who meet
      initial study requirements will be invited for a screening visit (visit 1) at the CTRC. Prior
      to this visit, participants will receive a detailed health questionnaire (see Appendix C)
      that has been used in numerous previous studies that addresses questions regarding past
      medical history, current use of medications and dietary supplements, exercise and social
      habits. In addition, participants will be asked about fish consumption in order to ensure
      recruitment of subjects with lower EPA + DHA intake during the study period. Participants
      will be asked to complete these questionnaires at home and bring the completed forms with
      them to their screening visit. During the screening visit, they will meet with research staff
      to review and sign an IRB-approved consent form. Research staff will review the health
      questionnaire with the participant as well as measure weight, height, waist and hip
      circumference, sitting blood pressure and heart rate. A brief physical exam will be performed
      as well as an electrocardiogram (ECG). In addition, participants will have blood drawn (after
      a 12 hour fast) for evaluating fasting lipids, glucose, serum chemistries and liver function
      and a complete blood count. Female participants will also be asked to provide a urine sample
      for a pregnancy test. These laboratory parameters are included to assure the participant
      meets eligibility criteria and that he/she is generally in good health. Once all clinical
      data has been reviewed, study personnel will contact each study volunteer to let them know if
      they are eligible.

      Randomization visit (Visit 2) One to four weeks after the screening Visit 1, eligible
      participants will return to the CTRC after a 12 hour fast for a 2 hour visit. Participants
      will be asked to brush their teeth after their last meal the day before the visit, or at the
      start of the 12-hour fasting period, and to refrain from additional brushing or use of
      mouthwash until after the study visit. Participants will meet with a research nurse /
      coordinator who will assess any change in medical history and medication use, as well as
      review any adverse events. Blood pressure, heart rate, height and weight will be measured,
      blood and urine will be collected for pre-randomization baseline efficacy measures and a
      urine pregnancy test performed for all females. The simple fish consumption questionnaire
      will be administered to assess any changes to the level of omega-3 intake in the diet.

      They will also be asked to record all food and beverages consumed on dietary records on 3
      days prior to the visit, as well as completing a digestive health questionnaire and diet
      history questionnaire. All nutrient data collected from 3-day dietary records will be
      analyzed by Food Processor 8.1 (06/03, Escha). Participants will be asked to complete these
      questionnaires at home and bring the completed forms with them to their study visit. If a
      subject is unable to print out or complete the forms at home, they will be completed at the
      visit.

      Saliva and subgingival plaque samples will be taken for assessment of oral microbiome. Saliva
      samples will be collected using a standardized kit. Under the supervision of study personnel,
      subjects will be given the kit, which consists of a funnel attached to a tube, and instructed
      to spit into the funnel until the amount of liquid reaches a fill line located on the outside
      of the attached tube. Once the proper amount has been collected, the lid of the funnel will
      be closed and the funnel removed from the tube. The tube is then capped and shaken for about
      10 seconds. Subgingival plaque samples will be collected by swabbing several regions of the
      buccal mucosa using microbrushes. A separate brush will be used for each swab and two samples
      will be collected from each region.

      In order to establish effects of treatments on adipose inflammatory parameters, we will
      obtain gluteal region subcutaneous adipose tissue biopsies under local anesthesia (2%
      Lidocaine). A detailed description of the adipose collection procedures is given in Section
      6.4. Participants will then be randomized to drug or placebo group. Supplies of study drug
      will be provided to last 8 weeks (4 pills/day of either Lovaza or placebo).

      Participants will be provided verbal and written instruction to take the study drugs as
      follows; 2 fish oil/placebo with food in the morning, 2 fish oil/placebo with food in the
      evening. A pill diary will be given to all subjects.

      Telephone / E-mail Follow up (Visit 3) Participants will be contacted by telephone or e-mail
      approximately four weeks (visit 3) post-randomization to review medication compliance and
      pill diary, adverse events and any changes in fish consumption. Participants with <80%
      apparent adherence to study medication will be counseled to increase compliance so that their
      adherence at 8 weeks will be ≥80%.

      Completion Visit (Visit 4) Participants will come to the CTRC after a 12 hour fast.
      Participants will meet with a study coordinator who will assess changes to the medical
      history and medication use. They will also be asked to record all food and beverages consumed
      on dietary records on 3 days prior to the visit. Any adverse events will be recorded, pill
      diary reviewed, and study medications will be counted (pill count) Weight and height will be
      recorded and BMI calculated. ECG will be performed. Blood will be drawn for post-drug
      screening of liver functions and chemistries as well as for all outcome-related measures
      (lipids, insulin/glucose, inflammatory markers). Urine for efficacy measures will also be
      collected. Urine pregnancy test will be performed in all females. Saliva and subgingival
      plaque samples will be taken to assess for any change in oral microbiome by the treatment. An
      adipose biopsy will be performed as above.
    

Intervention name(s): Lovaza; Placebo
Intervention type(s): Drug; Drug
Intervention description(s): Lovaza 4 grams per day; Placebo pill
Allocation: Randomized
Intervention model: Parallel Assignment
Masking: Triple (Participant, Investigator, Outcomes Assessor)
Primary purpose: Treatment