European Union Garlic Inflammation Study EUGIS History Garlic

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 1

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 2

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) 3

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

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 2001 2. 1 g/day 2. Statin 40 mg/day 3. Placebo Power = 0. 87 - 0. 72 for 30 - 25% CRP-reduction 5

European Union Garlic Inflammation Study (EUGIS) • Garlic powder from EU-sponsored breeding programme, Spanish Printanor year 2001 1. - all compounds present in fresh garlic / sulphurrich compounds preserved (allicin content 1. 02%) 2. 3. - compliant with European Pharmacopoeae with respect to microbial impurities - heavy metals, pesticides, aflatoxin comply 6

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) 7

European Union Garlic Inflammation Study (EUGIS) Screening -2 0 Intervention period 4 -5 11 -12 weeks Blood and urine collection 8

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) 9

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 10

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 CD 40 L (platelets) 11

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 12