1 European Union Garlic Inflammation Study (EUGIS) History Garlic would influence lipid metabolism / atherosclerosis Proposed study: - 2 x 24 apparently healthy subjects with increased risk (mildly hypercholesterolemic, smoking) - parallel trial (24 garlic / 24 placebo) - biochemical measures lipid metabolism / inflammation

2 European Union Garlic Inflammation Study (EUGIS) Current situation: No indications that ‘garlic’ influences lipid metabolism  Human intervention study without rationale is unethical; previous proposal not feasible New situation requires evaluation; other type of intervention study

3 European Union Garlic Inflammation Study (EUGIS) Considerations: Indications from cell and animal studies that ‘garlic’ may influence inflammation parameters Cardiovascular disease (atherosclerosis) shares common features with inflammation Current treatment strategies for atherosclerosis also focus on inflammation (for example: lipid lowering drugs; statins)

4 European Union Garlic Inflammation Study (EUGIS) Primary endpoint: C-reactive protein Secondary efficacy parameters: fibrinogen, vWF, sensitivity of leukocytes to inflammatory stimulus plasma lipids blood pressure cancer biomarkers (anti-oxidant enzymes, DNA damage, antimutagenic properties) metabolites

5 European Union Garlic Inflammation Study (EUGIS) Double dummy placebo-controlled trial; 3 parallel groups (n=30); risk group (40-75 y,BMI  24.5 kg/m 2, smoking  10 cig/d, post-menopausal) 1.Spanish Printanor g/day 2.Statin40 mg/day 3.Placebo Power = for % CRP-reduction

6 European Union Garlic Inflammation Study (EUGIS) 1.Garlic powder from EU-sponsored breeding programme, Spanish Printanor year all compounds present in fresh garlic / sulphur-rich compounds preserved (allicin content 1.02%) - compliant with European Pharmacopoeae with respect to microbial impurities - heavy metals, pesticides, aflatoxin comply

7 European Union Garlic Inflammation Study (EUGIS) Recruitment: advertisements Medical screening + blood sampling Randomisation Duplicate sampling (10-14 days apart) / measurements at start, after 4 wks and at end treatment period (12 wks)

8 European Union Garlic Inflammation Study (EUGIS) Screening Intervention period weeks Blood and urine collection -2

9 European Union Garlic Inflammation Study (EUGIS) Biomarkers for inflammation C-Reactive Protein Fibrinogen Cytokines after whole blood stimulation without and with LPS (TNF-  or IL-10) von Willebrand Factor (biomarker for endothelial function)

10 European Union Garlic Inflammation Study (EUGIS) Clinical measures (ECG, HRV, BP) Routine lab (cholesterol, HDL-C, triglycerides, Hb, haematocrit, white cell count, liver/kidney/ muscle function) DNA damage in lymphocytes (comet assay) Anti-oxidant enzymes (GPX, SOD) Antimutagenic properties of urine (Ames test) Metabolites of garlic compounds in plasma and urine

11 European Union Garlic Inflammation Study (EUGIS) - Additional biomarkers (suggested (in the EU-proposal)) s-VCAM / s-ICAM / s-Selectine PAI-1 MCP-1 and other chemokines endothelin 1 and 3 SAA ox-LDL (plasma) / isoprostanes (urine) soluble CD40L (platelets) -

12 European Union Garlic Inflammation Study (EUGIS) Points to discuss Positive control (is statin the right control?; preparation placebo; costs) Mask taste/smell (DADS in placebo?) If bad compliance, reduce dosage to 1.2 g/d Costs working up blood/lymphocytes Further characterisation of garlic powder Additional measurements and financing