Extended report c Additional data (supplementary information) are published online only at http:// ard.bmj.com/content/vol69/ issue1 1 Division of Rheumatology, Immunology, and Allergy, Brigham and Women’s Hospital and Harvard Medical School, Boston, Massachusetts, USA; 2 Harvard School of Public Health, Boston, Massachusetts, USA; 3 Immune Disease Institute, Boston, Massachusetts, USA; 4 Genzyme Corporation, Boston, Massachusetts, USA; 5 Channing Laboratory, Brigham and Women’s Hospital, Boston, Massachusetts, USA Correspondence to: E W Karlson, Brigham and Women’s Hospital, 75 Francis Street, Boston, Massachusetts 02115, USA; ekarlson@ partners.org Accepted 2 January 2009 Published Online First 16 January 2009 54 Gene–environment interaction between HLA-DRB1 shared epitope and heavy cigarette smoking in predicting incident rheumatoid arthritis E W Karlson,1 S-C Chang,2 J Cui,1 L B Chibnik,1 P A Fraser,1,3,4 I De Vivo,2,5 K H Costenbader1 ABSTRACT Background: Previous studies have reported an inter- action between ever cigarette smoking and the presence of the human leukocyte antigen (HLA)-DRB1 shared epitope (SE) genotype and rheumatoid arthritis (RA) risk. To address the effect of dosage, a case-control study nested within two prospective cohorts to determine the interaction between heavy smoking and the HLA-SE was conducted. Methods: Blood was obtained from 32 826 women in the Nurses’ Health Study and 29 611 women in the Nurses’ Health Study II. Incident RA diagnoses were validated by chart review. Controls were matched for age, menopausal status and postmenopausal hormone use. High-resolution HLA-DRB1 genotyping was performed for SE alleles. HLA-SE, smoking, HLA-SE* smoking interac- tions and RA risk, were assessed using conditional logistic regression models, adjusted for age and reproductive factors. Additive and multiplicative interactions were tested. Results: In all, 439 Caucasian matched pairs were included. Mean age at RA diagnosis was 55.2 years; 62% of cases were seropositive. A modest additive interaction was observed between ever smoking and HLA-SE in seropositive RA risk. A strong additive interaction (attributable proportion due to interaction (AP) = 0.50; p,0.001) and significant multiplicative interaction (p = 0.05) were found between heavy smoking (.10 pack-years) and any HLA-SE in seropositive RA risk. The highest risk was in heavy smokers with double copy HLA- SE (odds ratio (OR) 7.47, 95% CI 2.77 to 20.11). Conclusions: A strong gene–environment interaction was observed between HLA-SE and smoking when stratifying by pack-years of smoking rather than by ever smoking. Future studies should assess cumulative exposure to cigarette smoke when testing for gene– smoking interactions. Rheumatoid arthritis (RA), an autoimmune disease of unknown aetiology, affects approximately 1% of the adult population.1 Genetic and environmental factors are thought to interact in RA development. Epidemiological research has demonstrated a strong association between cigarette smoking and RA risk.2–14 In the Nurses’ Health Study (NHS), we have found that RA risk is significantly elevated among women with .10 pack-years of smoking, and a strong dose-response exists.12 14 The strongest genetic risk factor for RA is found within the human leukocyte antigen (HLA) com- plex (or major histocompatibility complex antigen (MHC)). Within the HLA class II region, multiple HLA-DRB1 alleles are associated with RA.15–17 In individuals of European ancestry, the associated HLA-DRB1 alleles share a region of sequence similarity or ‘‘shared epitope’’ (SE) at amino acid positions 70–74 in the third hypervariable region of the HLA-DRB1 molecule.18 Smoking and HLA shared epitope (HLA-SE) genotypes interact to increase risk of seropositive but not seronegative RA in several studies.13 19–21 However, the dose effect aspects of this interaction have not been studied. One study of three North American RA cohorts22 did not demonstrate a significant inter- action between ever smoking and HLA-SE in predicting anti-cyclic citrullinated protein (CCP) antibodies or rheumatoid factor (RF) among RA cases.19 We studied the interaction between HLA-SE alleles and smoking dose among women in a case- control study nested within two large prospective cohort studies, the Nurses’ Health Studies. We aimed to determine whether heavier smoking was associated with a stronger gene–environment interaction than was ever smoking. METHODS Study population The NHS is a prospective cohort of 121 700 female nurses, aged between 30–55 years in 1976. From 1989 to 1990, 32 826 (27%) NHS participants ages 43–70 provided blood samples. The Nurses’ Health Study II (NHSII) is a similar cohort, with 116 608 female nurses aged between 25–42 in 1989. Between 1996 and 1999, 29 611 (25%) of the women participating in NHSII cohort, aged 32–52 at that time, provided blood samples. The demo- graphics and exposure characteristics of partici- pants who provided blood samples are similar to those of the overall cohorts.23 24 All aspects of this study were approved by the Partners’ HealthCare Institutional Review Board. Identification of rheumatoid arthritis As previously described,14 we confirmed self-reports of RA based on presence of RA symptoms on a connective tissue disease screening questionnaire (CSQ),25 and, medical record review for four or more of the seven American College of Rheumatology (ACR) classification criteria for RA.26 We included a small number of subjects (n = 14) with agreement by two rheumatologist reviewers on diagnosis of RA, three documented ACR criteria for RA and a diagnosis of RA by their Ann Rheum Dis 2010;69:54–60. doi:10.1136/ard.2008.102962