1. The role of folate in the aetiology of oesophageal lesions L Sharp, 1 A-E Carsin, 1 L Anderson, 2 H Ferguson, 2 SJ Murphy, 2 A McElholm, 2 BT Johnston, 3 RGP Watson, 3 H Comber, 1 J McGuigan, 3 JV Reynolds, 4 LJ Murray 2 1 National Cancer Registry Ireland, Cork; 2 Centre for Clinical and Population Sciences, Queen’s University, Belfast; 3 Royal Group of Hospitals, Belfast; 4 St James’ Hospital, Dublin

2. Folate B vitamin Natural sources –green vegetables –oranges –yeast –cereals and grains Synthetic form = folic acid

3. Key metabolic roles of folate FOLATE deoxyuridine dihydrofolate monophosphate (dUMP)methionine tetrahydrofolateS-adenosyl- homocysteinemethionine (SAM) 5,10-methylene 5-methyls-adenosyl tetrahydrofolate tetrahydrofolatehomocysteine (SAH) deoxythymidine monophosphate (dTMP) DNA REPAIR DNA DNA SYNTHESIS METHYLATION

4. Other factors influencing folate metabolism Efficient folate metabolism requires other B vitamins vitamin B 12 vitamin B 6 riboflavin Folate metabolism is affected by alcohol – inhibits folate absorption smoking – may impair folate utilisation

5. Oesophageal adenocarcinoma carcinogenesis ? normalinflammationspecialized dysplasia malignancy squamouscolumnar mucosaepithelium reflux Barrett’s adenocarcinoma oesophagitisoesophagus

6. FINBAR (Factors INfluencing the Barrett’s Adenocarcinoma Relationship) case-control study Aim To investigate risk factors for adenocarcinoma of the oesophagus (OAC), Barrett’s oesophagus (BO) and reflux oesophagitis (RO) Research Questions Is dietary intake of folate associated with risks of developing OAC, BO or RO? Are intakes of vitamin B 12, vitamin B 6 and riboflavin associated with risk? Are these relationship modified by alcohol intake and/or cigarette smoking? Are plasma folate, vitamin B 12 and homocysteine associated with risk?

7. FINBAR Methods 1 OAC Cases histologically confirmed disease, diagnosed May 2002-Dec 2004, < 85 years BO Cases long segment BO (≥3cm), specialised intestinal metaplasia on histology, diagnosed May 2002-Dec 2004, < 85 years RO Cases Macroscopically visible erosive oesophagitis at endoscopy, aged < 85 years Controls aged 35-84 years, no history of BO or oesophageal or other GI cancer selected from Northern Ireland GP Master Index and from GP lists in Dublin and Cork

8. FINBAR Methods 2 Assessment of dietary exposures structured interview and food frequency questionnaire (EPIC modified for dietary variations in Ireland) reference period: 5-years prior to interview Assessment of blood exposures blood sample obtained at interview plasma folate and vitamin B 12 measured by SimulTRAC-SNB radioassay total homocysteine measured by HPLC Statistical analysis estimated daily intakes of micronutrients; adjusted for total energy intake logistic regression to compute multivariate odds ratios (OR) with 95% CIs separate analyses for OAC, BO and RO

9. Characteristics of participants ControlsOACBORO No. participants260227224230 Response rate42%64%82%69% Sex% male85% 83%82% Agemedian646562 Social class% I,II, III (non-manual)53%42%41%52% BMI% ≥30kg/m 2 19%32%18%22% Regular aspirin use29%28%24%29% Frequent GOR symptoms19%49%73%39%

10. Folate intake and disease risk Folate intakeOACBORO quartile 1 OR95%CIOR95%CIOR95%CI Lowest (Q1)1.0- - - Q21.200.65-2.200.730.38-1.390.550.28-1.09 Q30.600.33-1.100.400.21-0.750.420.22-0.80 Highest (Q4)0.580.32-1.040.400.21-0.750.360.18-0.69 p (trend)0.00960.00090.0013 1 cut-points for energy-adjusted residuals: Q1: ≤318.3  g/day; Q2: 318.3-370.89  g/day; Q3: 370.90-420.00  g/day; Q4: ≥420.01  g/day 2 multivariate risk estimates

11. Plasma folate and disease risk Folate tertileOACBORO (pg/ml)OR95%CIOR95%CIOR95%CI Lowest (Q1)1.0- - - Q20.650.42-1.020.540.34-0.840.750.48-1.17 Highest (Q3)0.410.25-0.660.510.33-0.800.650.41-1.03 ORs adjusted for age, sex, BMI and, for BO and RO, waist-hip ratio Similar associations to those with dietary folate intake

12. Plasma homocysteine and disease risk homocysteine tertile OACBORO (tHcy/ml)OR95%CIOR95%CIOR95%CI Lowest (Q1)1.0- - - Q21.330.81-2.181.050.66-1.651.220.76-1.99 Highest (Q3)1.931.20-3.111.010.64-1.621.941.21-3.11 ORs adjusted for age, sex, BMI and, for BO and RO, waist-hip ratio

13. Other B vitamins Dietary vitamin B 6 Strong inverse associations with OAC, BO and RO Dietary riboflavin Inversely related to RO; not associated with OAC or BO Vitamin B 12 Dietary intake – strong positive association with OAC (OR Q4 vs Q1=3.9) and BO (OR Q4 vs Q1=2.1), but not RO Plasma levels – no associations with OAC, BO or RO

14. Folate, smoking and OAC Folate intake Smokinglow 1 high 1 statusOR 2 95% CIOR 2 95% CI never1.070.51-2.251.0- ex3.351.76-6.391.680.92-3.09 current7.283.43-15.472.211.07-4.57 p (interaction)=0.1200 1 “low” < median (Q1+Q2); “high” ≥ median (Q3+Q4) 2 multivariate ORs

15. Folate, smoking and BO Folate intake Smokinglow 1 high 1 statusOR 2 95% CIOR 2 95% CI never1.200.59-2.471.0- ex1.470.73-2.970.600.29-1.21 current2.931.24-6.930.720.30-1.72 p (interaction)=0.1154 1 “low” < median (Q1+Q2); “high” ≥ median (Q3+Q4) 2 multivariate ORs

16. Folate, alcohol and oesophageal lesions Alcohol intake (5-years before interview and at age 21) did not significantly modify associations between folate intake and OAC, BO or RO However - risk of disease was highest in those who drank alcohol at age 21 and had low folate intake (i.e. the group expected to have lowest folate status)

17. Conclusions These results suggest that folate metabolism may have a role in the aetiology of oesophageal lesions Folate may be important early in the disease process Confirmation needed of finding that low folate intake may exacerbate effects of smoking on risk Folate levels in population are potentially amenable to change (dietary change, supplement use, food fortification) Could folate have a role in prevention of OAC or management/surveillance of individuals with BO or RO?

18. Acknowledgements Northern Ireland: Liam Murray (PI); Carol Anderson; Lesley Anderson;; Geraldine Cuskelly; Anna Gavin; Heather Ferguson; Anne Hughes; Brian Johnston; Martin McAnaespie; Adrian McElholm Ann McGinty; Jim McGuigan; Damian McManus; Seamus Murphy; Peter Watson Republic of Ireland: Harry Comber; Anne-Elie Carsin; Majella Gallagher; Lisa Higgins; Dermot Kelleher; Ross McManus; Gerry O’Sullivan; John Reynolds; Siobhan Reynolds Leeds: Chris Wild; Laura Hardie Leicester: Janusz Jankowski FINBAR was funded by the Northern Ireland Research & Development Office; Health Research Board, Dublin; Ulster Cancer Foundation