Generalization of adiposity genetic loci to US Hispanic women

Abstract

BACKGROUND:Obesity is a public health concern. Yet the identification of adiposity-related genetic variants among United States (US) Hispanics, which is the largest US minority group, remains largely unknown.OBJECTIVE:To interrogate an a priori list of 47 (32 overall body mass and 15 central adiposity) index single-nucleotide polymorphisms (SNPs) previously studied in individuals of European descent among 3494 US Hispanic women in the Women's Health Initiative SNP Health Association Resource (WHI SHARe).DESIGN:Cross-sectional analysis of measured body mass index (BMI), waist circumference (WC) and waist-to-hip ratio (WHR) were inverse normally transformed after adjusting for age, smoking, center and global ancestry. WC and WHR models were also adjusted for BMI. Genotyping was performed using the Affymetrix 6.0 array. In the absence of an a priori selected SNP, a proxy was selected (r ² [= or >, slanted]0.8 in CEU).RESULTS:Six BMI loci (TMEM18, NUDT3/HMGA1, FAIM2, FTO, MC4R and KCTD15) and two WC/WHR loci (VEGFA and ITPR2-SSPN) were nominally significant (P<0.05) at the index or proxy SNP in the corresponding BMI and WC/WHR models. To account for distinct linkage disequilibrium patterns in Hispanics and further assess generalization of genetic effects at each locus, we interrogated the evidence for association at the 47 surrounding loci within 1 Mb region of the index or proxy SNP. Three additional BMI loci (FANCL, TFAP2B and ETV5) and five WC/WHR loci (DNM3-PIGC, GRB14, ADAMTS9, LY86 and MSRA) displayed Bonferroni-corrected significant associations with BMI and WC/WHR. Conditional analyses of each index SNP (or its proxy) and the most significant SNP within the 1 Mb region supported the possible presence of index-independent signals at each of these eight loci as well as at KCTD15.CONCLUSION:This study provides evidence for the generalization of nine BMI and seven central adiposity loci in Hispanic women. This study expands the current knowledge of common adiposity-related genetic loci to Hispanic women.

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Citation: Nutrition & Diabetes (2013) 3, e85; doi:10.1038/nutd.2013.26& 2013 Macmillan Publishers Limited All rights reserved 2044-4052/13

http://www.nature.com/nutd

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ORIGINAL ARTICLE

Generalization of adiposity genetic loci to US Hispanic women

M Graff1,14, L Fernndez-Rhodes1,14, S Liu2, C Carlson3, S Wassertheil-Smoller4, M Neuhouser3, A Reiner3, C Kooperberg3, E Rampersaud5, JE Manson6, LH Kuller7, BV Howard8, HM Ochs-Balcom9, KC Johnson10, MZ Vitolins11, L Sucheston12, K Monda1 and KE North1,13

BACKGROUND: Obesity is a public health concern. Yet the identication of adiposity-related genetic variants among United States (US) Hispanics, which is the largest US minority group, remains largely unknown.

OBJECTIVE: To interrogate an a priori list of 47 (32 overall body mass and 15 central adiposity) index single-nucleotide polymorphisms (SNPs) previously studied in individuals of European descent among 3494 US Hispanic women in the Womens Health Initiative SNP Health Association Resource (WHI SHARe).

DESIGN: Cross-sectional analysis of measured body mass index (BMI), waist circumference (WC) and waist-to-hip ratio (WHR) were inverse normally transformed after adjusting for age, smoking, center and global ancestry. WC and WHR models were also adjusted for BMI. Genotyping was performed using the Affymetrix 6.0 array. In the absence of an a priori selected SNP, a proxy was selected (r2X0.8 in CEU).

RESULTS: Six BMI loci (TMEM18, NUDT3/HMGA1, FAIM2, FTO, MC4R and KCTD15) and two WC/WHR loci (VEGFA and ITPR2-SSPN) were nominally signicant (Po0.05) at the index or proxy SNP in the corresponding BMI and WC/WHR models. To account for distinct linkage disequilibrium patterns in Hispanics and further assess generalization of genetic effects at each locus, we interrogated the evidence for association at the 47 surrounding loci within 1 Mb region of the index or proxy SNP. Three additional BMI loci (FANCL, TFAP2B and ETV5) and ve WC/WHR loci (DNM3-PIGC, GRB14, ADAMTS9, LY86 and MSRA) displayed Bonferroni-corrected signicant associations with BMI and WC/WHR. Conditional analyses of each index SNP (or its proxy) and the most signicant SNP within the 1 Mb region supported the possible presence of index-independent signals at each of these eight loci as well as at KCTD15. CONCLUSION: This study provides evidence for the generalization of nine BMI and seven central adiposity loci in Hispanic women. This study expands the current knowledge of common adiposity-related genetic loci to Hispanic women.

Nutrition & Diabetes (2013) 3, e85; doi:http://dx.doi.org/10.1038/nutd.2013.26

Web End =10.1038/nutd.2013.26 ; published online 26 August 2013

Keywords: obesity; Hispanic; women; genetics; generalization

INTRODUCTIONLittle is known about the etiologic factors underlying the high prevalence of obesity, particularly among United States (US) minority populations. Concurrent with the obesity epidemic, US demographics have dramatically shifted. As of 2010, US Hispanics represented approximately 16% of the nation to become its largest minority group.1 Between 2009 and 2010, 41% of US Hispanic women were overweight or obese as compared with 32% of their non-Hispanic White counterparts,2 with the most notable ethnic disparities occurring among Puerto Rican and Dominican women.3 Thus, there is a rising impetus to investigate the underlying determinants of obesity among these populations.

In the past 5 years, genome-wide association studies (GWAS) have identied nearly 50 common genetic loci associated with body mass index (BMI)46 and anthropometric measures of central adiposity (that is, waist circumference (WC) and waist-to-hip ratio (WHR))7,8 in European middle-aged adult populations from Europe, Australia, and the US recent GWAS in non-European ancestry populations have identied additional novel loci,

including four new BMI-associated loci among East Asians, of which at least two loci do not show association in individuals of European descent3,7,9 and possibly three novel loci in a GWAS of BMI in individuals of African descent completed recently.913

Targeted genotyping studies of selected variants have been undertaken in Hispanic Americans.14 However, to date the contribution of genetic variants to adiposity traits in this diverse ethnic group remain largely unknown.

We investigated the associations of adiposity measures with previously identied European descent established genetic loci for BMI, WC and WHR among 3587 self-identied Hispanic women from the Womens Health Initiative (WHI) SNP (single-nucleotide polymorphism) Health Association Resource (SHARe).

MATERIALS AND METHODSWHI SHARe participantsWHI consists of multiple components including an observational study and clinical trial cohorts of postmenopausal women in the US;15 detailed

1Department of Epidemiology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA; 2Department of Epidemiology, University of California, Los Angeles, CA, USA;

3Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, USA; 4Department of Epidemiology and Population Health, Albert Einstein College of Medicine, Bronx, NY, USA; 5Department of Human Genetics, University of Miami, Miami, FL, USA; 6Department of Medicine, Brigham and Womens Hospital, Harvard Medical School, Boston, MA, USA; 7Department of Epidemiology, University of Pittsburgh, Pittsburgh, PA, USA; 8Medstar Health Research Institute, Hyattsville, MD, USA; 9Department of Social and Preventive Medicine, University at Buffalo, Buffalo, NY, USA; 10Department of Preventive Medicine, University of Tennessee, Memphis, TN, USA; 11Department of Epidemiology and Prevention, Wake Forest School of Medicine, Winston-Salem, NC, USA; 12Department of Cancer Prevention and Control, Roswell Park Cancer Institute, Buffalo, NY, USA and 13Carolina Center for Genome Sciences, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA. Correspondence: Dr M Graff, Department of Epidemiology, University of North Carolina at Chapel Hill, 137 East Franklin Street, Bank of America, Suite 306, Campus Box 8050, Chapel Hill, NC 27514, USA.

E-mail: mailto:[email protected]

Web End [email protected]

14Co-rst authorship.

Received 11 February 2013; revised 28 June 2013; accepted 22 July 2013

Generalization of adiposity loci to Hispanic women M Graff et al

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recruitment and exclusion criteria have been described previously.16

Medical histories were updated annually or semi-annually by questionnaire or by phone. All participating institutions obtained Institutional Review Board approval. WHI SHARe included a total sample of 3642 self-identied Hispanic subjects from WHI, who had consented to genetic research.

PhenotypesAll phenotypic information (for example, covariate and outcome variables)

was obtained during the WHI baseline questionnaires and clinic examination. Weight was measured after removing shoes, heavy clothing and pocket contents using a calibrated digital scale and recorded to the nearest one-tenth of a kilogram. Height was taken using a wall-mounted stadiometer and recorded to nearest one-tenth of a centimeter. BMI was calculated from measured height and weight (kg m2) and was missing for 26 of the participants in our sample. WC was measured at the level of natural waist (narrowest part of torso, n 14 missing) and hips at top of

the iliac crest with extra layers of clothes removed (n 13 missing) and

recorded to nearest half-centimeter. WHR was then calculated as the ratio of waist to hip circumference (n 16 missing).

GenotypesAs described previously,17 DNA was extracted by the Specimen Processing

Laboratory at the Fred Hutchinson Cancer Research Center (FHCRC) using white blood cells that were collected at the time of enrollment of the subjects in WHI. Specimens were stored at a central biorepository at

80 1C until analysis. Genotyping was done at Affymetrix, Inc. on the Affymetrix 6.0 array (Santa Clara, CA, USA), using 2 mg DNA at a concentration of 100 ng ml1.

Quality controlOf the 3642 women in WHI SHARe who self-identied as Hispanic and consented for genetic testing, approximately 1% of their genetic samples could not be genotyped (n 36). We excluded samples that had call rates

below 95%, which were duplicates of subjects other than their monozygotic twins, or that appeared to include a Y chromosome (that is, representing possible sample contamination, genotyping errors or an inconsistent genotypic sex; n 19). Furthermore, SNPs that were

located on the Y chromosome or were Affymetrix QC probes (that is, not intended for analysis) were excluded (n 3280). SNPs with a call rate

below 95% or concordance rate below 98% were agged and excluded leaving 871 309 SNPs. These quality control measures left us with 3587 Hispanics and an average call rate of 99.8% across the 871 309 unagged SNPs. We also excluded one person from identied relative pairs, prioritizing for complete genotype data (n 93), leading to a nal analytic

sample of 3494 self-identied Hispanic women.

Two hundred thirty-eight (2%) additional samples were genotyped as blind duplicates. We analyzed 188 pairs of blind duplicate samples. The overall concordance rate was 99.8% (range 95100% over all samples, 98100% across 871 309 SNPs that were included after genotype cleaning).

AdmixtureEigenvectors were computed in Eigenstrat18,19 to account for global

ancestry based on 178 101 markers, excluding mitochondria and sex chromosome markers, that were in common between WHI Hispanics samples and HapMap20,21 and HGDP22 reference panels. In particular, we

excluded SNPs that were A/T or C/G, on the sex chromosomes, or in the mitochondria. Individuals included from HGDP panels were 225 East Asians and 63 Native Americans, specically 8 Surui, 22 Mayans, 13 Karitiana, 14 Pima and 6 Colombian. We also estimated proportions of European, Native American and African ancestry (Supplementary Figure 1) in the unrelated WHI SHARe sample (n 3494) using Admixture 1.22 (http://www.genetics.ucla.edu/software/admixture

Web End =http://www.

http://www.genetics.ucla.edu/software/admixture

Web End =genetics.ucla.edu/software/admixture ).

Adiposity SNP selectionOne SNP from each established adiposity locus (described as of 1 July 2012 with BMI WC or WHR in GWAS of European descent individuals) was selected. A total of 47 loci were selected; 32 loci previously associated with BMI and 15 loci previously associated with WC or WHR (Tables 2a and 3a). All selected SNPs from the original publications were those that had the lowest P-value and that met genome-wide signicance within a predened locus (typically dened as 1 Mb and r2o0.1).

GeneralizationWe assessed generalization of previously established GWAS loci using a tiered approach. All SNPs analyzed here were originally reported in populations of European descent, so we dene generalization of a genetic effect when a SNP displays a direction of effect consistent with the original report and/or in terms of statistical signicance as dened below.

First we interrogated the exact SNP from the published literature, which we dened as an index SNP. All selected index SNPs met genome-wide signicance level in prior publications. To assess the consistency of effects in our study, we accessed genome-wide publically available data from the Genetic Investigation of ANthropometric Traits (GIANT) Consortium on the risk allele and its frequency in their large sample of individuals of European descent. Loci previously described with overall or central adiposity were queried in the BMI and WHR adjusted for BMI GWAS results les, respectively. If this information was missing, then we supplemented it with the relevant publication to determine directional consistency. If the previously reported adiposity SNP was not genotyped as part of WHI SHARe, the WHI SHARe SNP in highest linkage disequilibrium (LD) with the previous reported SNP (r2X0.8 in Hap Map CEU phase II) was selected as a proxy of the index signal. Generalization of the index or proxy SNPs was declared when directional consistency and nominal statistical signicance (Po0.05) were observed.

Owing to the extensive admixture in populations of self-identied Hispanic ancestry,23,24 we also hypothesized that even if a SNP originally identied in European or East Asian ancestry populations is not associated with BMI in those within our cohort of women who report Hispanic ancestry, the locus may still show association with a different variant in the same chromosomal region. Therefore, we searched for common variants within the established loci that better captured the association of the index SNP reported in the European and Asian populations. We identied SNPs as potentially better markers of the index signal, index-dependent signals, if they were (1) within 1 Mb of the index SNP, (2) were dependent on the index SNP in the referent population (r2X0.2) and (3) were associated with the anthropometric traits in our data at a signicance level that was at least one order of magnitude greater than the index SNP or its proxy. In contrast, we also interrogated the evidence for possible index-independent signals by visual inspection of all P-values of SNP anthropometric trait associations for SNPs of interest with r2o0.2 and within the 1 Mb region of the index SNP. Index-independent signals were deemed statistically signicant if they displayed nominal signicance after correcting for the total number of regions interrogated for each phenotype of interest (BMI: P 0.05/32 and WC/WHR: P 0.05/15). Conditional

analyses were also conducted to conrm signal dependence. If adjustment for the index SNP decreased the P-value for the candidate index-independent signal, the SNPphenotype association was considered suggestive evidence for an index-independent signal, without overwhelming proof that the signal was indeed independent. Certainly these signals need to be further interrogated with much larger sample sizes and/ or ne mapping and within the different sub-populations of US Hispanic ancestry. In contrast, if the conditional P-value did not change or increased less than one order of magnitude in comparison with the unconditional P-value, then we declared this a possible index-independent signal. All conditional analyses were modeled in Stata 12 (StataCorp LP, College Station, TX, USA).

Statistical modelsAfter adjusting for age, smoking status and clinical center, BMI residuals were inverse normally transformed. WC and WHR were adjusted for age, smoking status, clinical center as well as BMI, and then the residuals were inverse normally transformed. Inverse normal transformations entail creating a modied rank variable and then computing a new transformed value for the phenotype per subject such that the distribution of the phenotype is normalized with a mean of 0 and an s.d. of 1. For each of the three inverse normalized phenotypes (that is, BMI, WC adjusted for BMI and WHR adjusted for BMI) single marker linear associations further adjusted for the top 10 principal components assuming an additive model, were run using PLINK software v1.07.25 Estimated P-values below 5 108

were considered to be genome-wide signicant.

LD assessmentWe considered signals independent if their LD was r2o0.2 in a sample of 9345 individuals of European descent (primarily non-Hispanic Whites) from the Atherosclerosis Risk in Communities Study (ARIC). If information was

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not available from ARIC then HapMap CEU data (phase II or III) were used to represent the LD structure of individuals of European descent and are specically noted in Tables 2b and 3b. In addition, estimates of LD were calculated in WHI SHARe Hispanics. LD estimates for both ARIC and WHI SHARe were calculated using the PLINK software v1.07.25

PowerWe calculated estimates of power to detect associations of similar magnitude among Hispanics as those previously described in European populations across a range of common minor allele frequencies. These calculations assumed an additive genetic model, an independent sample of 3494 women, the same Bonferroni corrections and phenotype distribution as observed in our sample of US Hispanic women. Based on effect sizes published in European populations, power to detect associations was less among measures of overall (BMI) than for central adiposity (WC and WHR; Supplementary Figure 2). For example, at the minor allele frequency and previously reported effect size of FTO (32% and beta 0.39 kg m2 change per T allele)4 we would at best have 40%

statistical power to detect this effect in our study. Similarly, power to detect associations of all other BMI loci was below 80%. Moderately common WC variants (420%) would be expected to have 480% power at mid-sized effects, which was approximately 1 cm change in WC per effect allele; whereas, most common WHR variants (45%) frequent would be expected to have 480% power at far smaller effect sizes (approximately0.011 WHR units). Power calculations were calculated using QUANTO v1.2.4 (http://hydra.usc.edu/gxe/

Web End =http://hydra.usc.edu/gxe/).

RESULTSThe nal analytic sample of self-identied Hispanic women included in this sample was 3494. As shown in Table 1, the largest percentage of the women in this sample were between 50 and 59 years of age with a high school diploma or equivalent, were married, of Mexican ancestry, overweight in the absence of abdominal obesity as dened by the World Health Organization2628

and were participants in a clinical trial from one of the Western or Southern WHI study centers.

Adiposity SNP generalizationAlthough no SNPs reached genome-wide signicance in this study, we were able to investigate the associations at 47 established obesity loci previously identied in European populations with BMI, WC and WHR in our sample of Hispanic women (Tables 2b and 3b). As summarized in Figure 1, among 16 loci with

Table 1. Characteristics of self-identied Hispanics in the WHI SHARe

Full sample (N 3587)

Analytic sample (N 3494)

Number/ range

% Number/ range

%

Gender

Women 3587 100.0% 3494 100.0% Men

Age at assessment5059 years 1803 50.3% 1751 50.1% 6069 years 1415 39.4% 1387 39.7% 7079 years 369 10.3% 356 10.2% Missing

Hispanic ethnic subgroupaMexican, Chicano,

Mexican-American

1541 43.0% 1485 42.5%

Puerto Rican 369 10.3% 361 10.3% Cuban 255 7.1% 252 7.2% Other 814 22.7% 806 23.1% No subgroup indicated 162 4.5% 159 4.6% Missing 446 12.4% 431 12.3%

Study participationObservational study only 1729 48.2% 1681 48.1% Clinical trial 1858 51.8% 1813 51.9% Hormone replacement trial 980 27.3% 953 27.3% Control arm 472 13.2% 463 13.3% Dietary modication trial 1202 33.5% 1175 33.6% Control arm 730 20.4% 715 20.5% Calcium/vitamin D trial 1057 29.5% 1025 29.3% Control arm 495 13.8% 479 13.7% Missing

US regionNortheast 448 12.5% 441 12.6% South 1459 40.7% 1422 40.7% Midwest 136 3.8% 133 3.8% West 1544 43.0% 1498 42.9% Missing

Marital statusNever married 153 4.3% 149 4.3% Divorced or separated 750 20.9% 735 21.0% Widowed 478 13.3% 463 13.3% Presently married 2076 57.9% 2020 57.8% Marriage-like relationship 85 2.4% 82 2.3% Missing 45 45

EducationNo formal to incomplete high school

816 22.7% 802 23.0%

High school diploma or equivalent

1914 53.4% 1857 53.1%

College or higher degree 802 22.4% 781 22.4% Missing 55 54

Body mass indexbMean (s.d.) 28.88(5.59)

28.87(5.59)

Full sample (N 3587)

Analytic sample (N 3494)

Number/ range

% Number/ range

%

Hip circumference (cm)

Mean (s.d.) 105.84(11.14)

105.83(11.13)

1st quartile 8598 8598 2nd quartile 99104 99104 3rd quartile 105112 105112 4th quartile 113144 113144 Missing 13 13

WHRa

Mean (s.d.) 0.82(0.07)

0.82(0.07)

No abdominal obesity (WHRp0.85)

2476 69.0% 2413 69.1%

Abdominal obesity

(WHRX0.85)

1095 30.5% 1065 30.5%

Missing 16 16

Abbreviations: WHO, World Health Organization; WHR, waist-to-hip ratio; WHI SHARe, Womens Health Initiative SNP Health Association Resource.

aThe other category may include. bOverweight and obesity as dened by the WHO expert consultation. Appropriate body mass index for Asian populations and its implications for policy and intervention strategies. The Lancet, 2004; 157163. Waist Circumference and Waist-Hip Ratio, Report of a WHO Expert Consultation". World Health Organization. 811 December 2008. Retrieved 21 March 2012.

Underweight (o18.5 kg m2) 7 0.2% 7 0.2% Normal (18.524.9 kg m2) 889 24.8% 871 24.9%

Overweight (2529.9 kg m2) 1387 38.7% 1344 38.5% Obesity (3034.9 kg m2) 828 23.1% 806 23.1% Obesity II (3539.9 kg m2) 300 8.4% 294 8.4% Extreme obesity (X40 kg m2) 150 4.2% 146 4.2%

Missing 26 26

Waist circumference (cm)

Mean (s.d.) 86.60(12.30)

86.58(12.31)

1st quartile 6278 6278 2nd quartile 7985 7985 3rd quartile 8694 8694 4th quartile 95125 95125 Missing 14 14

Table 1. (Continued)

& 2013 Macmillan Publishers Limited Nutrition & Diabetes (2013) 1 10

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Table 2a. Signicance level of loci associated with BMI and/or weight from published GWAS studies in European descent men and women and the GIANT consortium

Index or proxy SNP

In/near gene Chr BP position Phenotype GWAS P-value

Risk allele

GIANT P-value

rs2815752 NEGR1 1 72524461 BMI 1.61E22 [6] A 0.62 1.17E14 rs1514175 TNNI3K 1 74764232 BMI 8.16E14 [6] A 0.57 1.41E09 rs1555543 PTBP2 1 96717385 BMI 3.68E10 [6] C 0.58 7.81E07 rs543874 SEC16B 1 176156103 BMI 3.56E23 [6] G 0.19 1.66E13 rs6548238 TMEM18 2 624905 BMI 3.20E26 [2] C 0.88 1.02E20 rs713586 RBJ/ADCY3/POMC 2 25011512 BMI 6.17E22 [6] C 0.52 2.51E07 rs759250a FANCL 2 59182657 BMI 1.79E12 [6] A 0.32 5.34E06 rs2121279b LRP1B 2 142759755 BMI 1.35E10 [6] T 0.12 1.37E06 rs13098327c CADM2 3 85902871 BMI 3.94E11 [6] A 0.18 1.14E07 rs1516728d ETV5/SFRS10/DGKG 3 187312585 BMI and weight 7.20E11 [1] A 0.78 9.50E11 rs12641981e Gene desert; GNPDA2 4 44874640 BMI 3.78E31 [6] T 0.43 7.34E17 rs3797580f FLJ35779/HMGCR 5 75038812 BMI 2.17E13 [6] A 0.64 1.92E07 rs6864049g ZNF608 5 124358421 BMI 1.97E09 [6] G 0.43 3.73E06 rs987237h PRL 6 50911009 BMI and obesity 1.40E05 [5] G 0.09 5.97E16 rs9366426 TFAP2B 6 22172618 BMI 2.90E20 [6] C 0.60 6.00E01 rs3798560i NUDT3/HMGA1 6 34451544 BMI 3.02E08 [6] C 0.17 5.25E06 rs10508503 PTER 10 16339957 BMI and obesity 2.10E07 [5] C 0.93 6.40E01 rs10840083j RPL27A/ TUB 11 8565212 BMI 2.80E09 [6] A 0.6 1.92E07 rs10501087 BDNF/LGR4/LIN7C 11 27626684 BMI and weight 8.70E11 [1] T 0.8 1.41E12 rs3817334 MTCH2 11 47607569 BMI 1.59E12 [6] T 0.45 4.79E11 rs7138803 FAIM2 (and BCDIN3D) 12 48533735 BMI and weight 1.82E17 [6] A 0.44 3.96E11 rs7988412k MTIF3 13 26898282 BMI 9.48E10 [6] T 0.23 2.57E06 rs10151686l PRKD1 14 29536217 BMI 5.76E11 [6] A 0.03 6.62E08 rs17109221m NRXN3 14 78979872 BMI 2.75E11 [6] T 0.27 2.31E07 rs8054079n GPRC5B/IQCK 16 19882908 BMI 2.91E21 [6]ww C 0.13 2.91E21 rs8049439 SH2B1 16 28745016 BMI and weight 1.40E09 [1] C 0.37 1.48E09 rs9939609 FTO 16 52378028 BMI 4.90E74 [2] A 0.45 9.94E60 rs9921354o MAF 16 78240951 BMI and obesity 3.80E13 [5] T 0.51 2.57E01 rs1652376p NPC1 18 19363464 BMI and obesity 2.90E07 [5] G 0.51 9.14E04 rs571312 MC4R 18 55990749 BMI 6.43E42 [6] A 0.28 2.14E22 rs11084753 KCTD15 19 39013977 BMI 4.50E12 [2] G 0.63 3.62E09 rs8101149q TMEM160/ZC3H4 19 52292281 BMI 1.64E12 [6] A 0.68 2.48E06

Abbreviations: BMI, body mass index; BP, base pair; Chr, chromosome; GIANT, Genetic Investigation of ANthropometric Traits Consortium; GWAS, genome-wide association study; SNP, single-nucleotide polymorphism. wwAs data were missing for rs8054079 in the publically available sources, information on rs12444979 was extracted from Speliotes et al. In this case, because of the tight linkage disequilibrium between the two variants we inferred that the lowester frequent allele at rs80504079 would increase BMI. aProxy SNP for rs887912, r2 1; bProxy SNP for rs2890652, r2 0.8; cProxy SNP for rs13078807, r2 1; dProxy SNP for

rs7647305, r2 0.8; eProxy SNP for rs1093897, r2 1; fProxy SNP for rs2112347, r2 0.9; gProxy SNP for rs4836133, r2 1; hProxy SNP for rs4712652, r2 0.9;

iProxy SNP for rs206936, r2 1; jProxy SNP for rs4929949, r22 0.9; kProxy SNP for rs4771122, r2 0.9; lProxy SNP for rs11847697, r2 0.8; mProxy SNP for

rs10150332, r2 1; nProxy SNP for rs12444979, r2 0.9; oProxy SNP for rs1424233, r2 1; pProxy SNP for rs1805081, r2 0.9; qProxy SNP for rs3810291, r2 0.9.

Published results from studies of individuals of European decent: [1] Thorleifsson et al.; [2] Willer et al.; [3] Lindgren et al.; [4] Heard-Costa et al.; [5] Meyre et al.; [6] Speliotes et al.; [7] Heid et al.

Risk allele frequency HapMap CEU

evidence of generalization 7 were dened as index-dependent signals. Of these seven, ve loci were best represented by the index SNP (or its proxy) and two by a better marker in the region (dened as other index-dependent signal). A total of nine loci displayed, at least, suggestive evidence for index-independent signals as the SNP in these loci with the lowest P-value were in low LD with the index signals previously described among European descent individuals and remained nominally signicant after adjustment for the index SNP (or its proxy) in conditional analyses.

Among the 32 BMI index signals interrogated in this study, 25 had consistent directions of association as compared with publically available GIANT BMI results, which is more than expected by chance (binomial P 2.4 103). The ve loci

(reported above) with either evidence of generalization at the index or proxy SNP, or evidence of a better marker displayed consistent directions of effect with BMI. Among the 15 central adiposity index signals interrogated in this study, 13 had consistent directions of effects at the WC index SNP or their proxies (more than expected by chance, binomial P 3.2 103)

and all had consistent directions of effect at the WHR index SNP or its proxy (binomial P 3.1 105), as compared with publically

available GIANT WHR adjusted for BMI results. WC/WHR loci with

either evidence of generalization at the index or proxy SNP, or evidence of a better marker displayed consistent direction of effects with the central adiposity phenotype, for which they were previously reported.

Among index or proxy SNPs selected, the BMI phenotype showed the strongest association with rs9939609 in the FTO locus (beta (s.e.) 0.085 (0.026); P 0.001), followed rs11084753 in the

KCT615 locus (beta (s.e.) 0.070 (0.025); P 0.006). Other

nominally signicant (Po0.05) loci were found for MC4R (rs571312), NUDT3/HMGA1 (rs378560), FAIM2 (rs7139903) and TMEM18 (rs6548238). Again, among the index or proxy SNPs selected, the strongest association with WC and WHR was found with rs60905288 near VEGFA (WC: beta (s.e.) 0.075 (0.024);

WHR: beta (s.e.) 0.072 (0.024); P 0.002 for both). A locus near

ITPR2-SSPN showed a nominal association with WHR (rs12814794, beta (s.e.) 0.050 (0.025); P 0.04).

For 11 adiposity loci (6 BMI and 5 WC/WHR), we observed a SNP of interest with at least one order of magnitude smaller P-value than the index SNP or its proxy. These loci are displayed in Supplementary Figures 36. SNPs at two loci previously associated with BMI, NUDT3/HMGA1 (rs6925243, P 7.84 104), and MC4R

(rs1942867; P 1.95 105) were dependent on their respective

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r2 withindexSNP(in

WHISHAReHispanic

women)

l ProxySNPfor

f Proxy

e ProxySNPforrs1093897,r2 1;

Table2b.ResultsinHispanicwomenforpublishedlocifromEuropeandescentindividualsassociatedwithBMIand/orweight

Indexor

rs2815752NEGR1rs17589316172391927 G/A0.2134500.0810.0306.83E030.450.1300.174

rs1514175TNNI3Krs17095822174948453 A/G0.0134670.4030.1445.03E032.25o0.0010.004

rs1555543PTBP2rs17115529196848822 G/T0.2834650.0590.0272.94E020.330.2050.167

rs543874SEC16Brs168523251176235862G/A0.1034660.0900.0402.57E020.500.0030.002

rs6548238TMEM18rs102052042827100 C/T0.0634570.1470.0524.75E030.820.005w0.000

rs713586RBJ/ADCY3/POMCrs2384061224989124 T/C0.3234450.0730.0254.20E030.410.6990.631

rs759250a FANCLrs4672266258978503 G/A0.3434670.0930.0252.30E040.52o0.0010.002

rs2121279b LRP1Brs45959132142910831G/T0.2034680.0950.0301.90E030.530.0090.014

rs13098327c CADM2rs2875492386088900 G/A0.043458 0.1890.0653.89E03 1.050.0020.008

rs1516728d ETV5/SFRS10/DGKGrs76483363187431942A/C0.263465 0.0910.0281.14E03 0.510.001w0.011

rs12641981e Genedesert;GNPDA2rs348551444924340A/G0.0134640.4920.1602.08E032.75o0.001o0.001

rs3797580f FLJ35779/HMGCRrs16872770575020375A/G0.083360 0.1330.0442.57E03 0.750.145w0.149

rs6864049g ZNF608rs175179075124554764G/A0.183468 0.0680.0312.88E02 0.38o0.0010.017

rs987237h PRLrs2857506650908267 T/C0.0934610.0760.0427.09E020.420.0330.047

rs9366426TFAP2Brs2876611622108317A/G0.013467 0.4000.1208.54E04 2.24o0.0010.001

rs3798560i NUDT3/HMGA1rs6925243634489915G/A0.3334680.0880.0267.84E040.490.3580.745

rs10508503PTERrs47482371016091103G/C0.4934680.0680.0244.23E030.38o0.0010.001

rs10840083j RPL27A/TUBrs11041928118430591 G/C0.043457 0.1930.0611.60E03 1.080.0190.026

rs10501087BDNF/LGR4/LIN7Crs105010891127745435 A/G0.0534390.1040.0566.32E020.58o0.0010.002

rs3817334MTCH2rs108387741147812690G/A0.353466 0.0810.0261.71E03 0.450.5150.316

rs7138803FAIM2(andBCDIN3D)rs171990261248341609 A/G0.0734680.1350.0453.08E030.750.012o0.001

rs7988412k MTIF3rs104924841327130706G/A0.0334640.2290.0731.78E031.280.0010.004

rs10151686l PRKD1rs104833791429749214 T/C0.2234600.0900.0292.17E030.500.028w0.001

rs17109221m NRXN3rs80203121479223674A/T0.0134670.2990.1423.56E021.67o0.0010.000

rs8054079n GPRC5B/IQCKrs124476551620132853G/A0.093463 0.1310.0421.97E03 0.73o0.0010.006

rs8049439SH2B1rs105211451628504385A/G0.093458 0.0970.0411.71E02 0.540.0760.069

rs9939609FTOrs99413491652382989 A/G0.3234670.0890.0265.85E040.500.871w0.456

rs9921354o MAFrs99393611678220241C/G0.0134620.3580.1471.50E022.00o0.0010.006

rs1652376p NPC1rs99525921819428220A/G0.003468 0.5260.1793.34E03 2.94o0.0010.007

rs571312MC4Rrs19428671855887250 T/C0.2034540.1110.0301.95E040.620.744w0.534

rs11084753KCTD15rs81042621938895287A/G0.0134660.4240.1121.65E042.37o0.001o0.001

rs8101149q TMEM160/ZC3H4rs81053121952277204 G/A0.0234670.2050.0851.64E021.150.0050.037

Abbreviations:ARIC,AtherosclerosisRiskinCommunitiesStudy;BMI,bodymassindex;BP,basepair;Chr,chromosome;GIANT,GeneticInvestigationofANthropometricTraitsConsortium;GWAS,genome-wide

associationstudy;LD,linkagedisequilibrium;MAF,minorallelefrequency;SNP,single-nucleotidepolymorphism.*P-valuesinboldindidicateevidenceofassociationbelownominalsignicance(Po0.05)for

indexorproxySNPs(reference),andbelowaBonferronithresholdforthenumberofmostsignicantSNPstested(Po0.05/32).**MostsignicantSNPwithin500kboftheindexorproxySNP(reference).w If

pairwiseLDinformationwasunavailableinARICWhitesthenHapMap2release22orHapMap3(atMC4Ronly)datainCEUwereusedinstead.ww Asdataweremissingforrs8054079inthepublicallyavailable

r2 withindex

SNP(inARIC

Whites)

k ProxySNPforrs4771122,r2 0.9;

q ProxySNPforrs3810291,r2 0.9.

j ProxySNPforrs4929949,r2 0.9;

p ProxySNPforrs1805081,r2 0.9;

MAFNEffect

sources,informationonrs12444979wasextractedfromSpeliotesetal.Inthiscase,becauseofthetightlinkagedisequilibriumbetweenthetwovariantsweinferredthatthelowesterfrequentalleleat

rs80504079wouldincreaseBMI.a ProxySNPforrs887912,r2 1;

S.e.P-value*Estimated

effectin

kgm2

d ProxySNPforrs7647305,r2 0.8;

c ProxySNPforrs13078807,r2 1;

estimate

i ProxySNPforrs206936,r2 1;

o ProxySNPforrs1424233,r2 1;

ChrBPpositionStrandMinor/

major

allele

b ProxySNPforrs2890652,r2 0.8;

h ProxySNPforrs4712652,r2 0.9;

n ProxySNPforrs12444979,r2 0.9;

signicant

SNP**

In/nearneneMost

g ProxySNPforrs4836133,r2 1;

m ProxySNPforrs10150332,r2 1;

SNPforrs2112347,r2 0.9;

proxySNP

rs11847697,2 20.8;

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Table 3a. Signicance level of loci associated with with WC and WHR, after adjustment for BMI from published GWAS studies in European descent men and women and the GIANT consortium

Index or proxy SNP

In/near gene Chr BP position Phenotype GWAS P-value Risk allele

Risk allele frequency HapMap CEU

GIANT P-value

rs2301453a DNM3-PIGC 1 170624790 WHR 9.51E18 [7] G 0.42 2.79E10 rs2605100b LYPLAL1 1 217710847 WC and

WHR

2.55E08,6.89E21

[3,7] G 0.69 2.19E09

rs6717858c GRB14 2 165247907 WHR 2.09E24 [7] T 0.54 5.68E10 rs6784615 NISCH-STAB1 3 52481466 WHR 3.84E10 [7] T 0.95 3.18E07 rs6795735 ADAMTS9 3 64680405 WHR 9.79E14 [7] C 0.54 2.47E07 rs17695092d CPEB4 5 173270459 WHR 1.91E09 [7] G 0.29 8.27E07 rs1294421 LY86 6 6688148 WHR 1.75E17 [7] G 0.6 6.31E09 rs6905288 VEGFA 6 43866851 WHR 5.88E25 [7] A 0.58 4.72E10 rs987237 TFAP2B 6 50911009 WC and

BMI

1.87E11 [3] G 0.09 0.096

rs1936805e RSPO3 6 127493809 WHR 1.84E40 [7] T 0.55 1.28E14 rs1055144 NFE2L3 7 25837634 WHR 9.97E25 [7] T 0.18 1.49E08 rs545854ww MSRA 8 9897490 WC 8.89E09 [3] G 0.18ww 0.64 rs12814794f ITPR2-SSPN 12 26331965 WHR 1.14E17 [7] G 0.18 3.25E07 rs1822438g HOXC13 12 52628593 WHR 6.38E17 [7] A 0.2 4.70E08 rs4823006 ZNRF3-KREMEN1 22 27781671 WHR 1.10E11 [7] A 0.53 4.47E08

Abbreviations: BMI, body mass index; BP, base pair; Chr, chromosome; GIANT, Genetic Investigation of ANthropometric Traits Consortium; GWAS, genome-wide association study; SNP, single-nucleotide polymorphism; WC, waist circumference; WHR, waist-to-hip ratio. wwrs7826222 was renamed rs545854 in HapMap Build 36 and thus is not present in Build 36 imputations based on that release of HapMap (release 22). As publically available information was not available for this SNP, information on the minor allele was supplemented from Lindgren et al. aProxy SNP for rs1011731, r2 1; bIndex SNP from Lindgren et al., but proxy

SNP for rs4846567 from Heid et al. at r2 0.6; cProxy SNP for rs1095252, r2 0.9; dProxy SNP for rs6861681, r2 1; eProxy SNP for rs9491696, r2 0.9; fProxy SNP

for rs12814794, r2 1; gProxy SNP for rs1822438, r2 1. Published results from studies of individuals of European decent: [1] Thorleifsson et al.; [2] Willer et al.;

[3] Lindgren et al.; [4] Heard-Costa et al.; [5] Meyre et al.; [6] Speliotes et al.; [7] Heid et al.

index or proxy SNPs in CEU (r2X0.2), and therefore were considered to represent a better marker for Hispanics at the index signal (Table 2b, Supplementary Figures 6a and b).

Four BMI loci (FANCL, ETV5, TFAP2B and KCTD15; Supplementary Figures 3ad), and ve WC or WHR loci (DNM3-PIGC, GRB14, ADAMTS9, LY86 and MSRA; Supplementary Figures 4ad and 5ac) had low LD in HapMap CEU populations (r2o0.2; Tables 2b and 3b) and were therefore considered as possible index-independent signals. All conditional P-values for the SNP of interest phenotype association remained nominally signicant after adjustment for the index SNP or its proxy (Po0.05). One BMI locus (TFAP2B) appeared to have suggestive evidence of an index-independent signal as the P-value decreased from the unconditional analysis for the association between the SNP of interest and BMI (Table 4). However, the evidence for association at three BMI loci (near FANCL, ETV5 and KCTD15) for the SNP of interest BMI association became weaker on adjustment for the index SNP or its proxy. KCTD15 was the only locus of these nine loci to have signicant evidence of both generalization at both the index signal (P 0.006) and an independent signal (r2o0.2; Tables 2a and

2b). At two central adiposity loci (near DNM3-PIGC and MSRA), there was suggestive evidence of index-independent signals for WC (Table 4). In contrast, at three previously described WHR loci (GRB14, ADAMTS9 and LY86) there was inconsistent evidence across the central adiposity phenotypes tested (WC and WHR models adjusted for BMI).

DISCUSSIONIn this study of postmenopausal Hispanic women, we found that the majority of the 47 SNPs interrogated showed consistent direction of effect. Specically, 25 of 32 SNPs for BMI (binomial test: Po7.8E04), and 13 and 15 of 15 SNPs for WC and

WHR (binomial test: Po3.2E03 and Po3.1E05), respectively. Further, we found associations of nine loci with BMI and seven loci with waist phenotypes (WC or WHR) previously shown to be associated with these traits in European populations from Europe,

Australia and the United States. In addition, we present possible evidence for independent signals among Hispanics at nine of these loci, three of which became stronger after conditioning (locus near TFAP2B with BMI, and loci near DNM3-PIGC and MSRA with WC or WHR). As verication for the associations of these possible independent signals identied in the Hispanic women in samples of European descent individuals, we looked up the P-value in the original published results (Heid and Speliotes references) for seven of the nine SNPs that were available. None of the seven SNPs were even nominally signicant (all P40.05).

Certainly the analyses conducted here needs to be independently veried in an additional Hispanic ancestry sample. Although the exact functional variants underlying these signals still remain to be identied, it is interesting to note that TFAP2B encodes a transcription factor that has previously been associated with both BMI and type 2 diabetes in primarily non-Hispanic populations.2931 MSRA encodes a protein that is thought to repair of oxidative damage to proteins to restore biological activity.32

Deletion of this gene has been associated with insulin resistance in mice.33 Dynamin 3 (DNM3), a member of the dynamin family of enzymes, and phosphatidylinositol glycan anchor biosynthesis, class C (PIGC), are involved in cell membrane interactions and adhesion of proteins to the cell membrane.3436

Functional roles of some of the loci with possibly independent signals among Hispanics may include energy homeostasis for KCTD15 and ETV5 loci that are highly expressed in the hypothalamus,37 and insulin signaling from ADAMTS9, and GRB14 loci, particularly in muscle tissue.3841

Although the possibility of multiple signals at established GWAS loci needs to be conrmed in additional, larger samples of Hispanic ancestry, these ndings add to the growing literature that indicates multiple variants for BMI, lipids and other complex traits. Moreover, these study ndings also add to the growing literature that demonstrates suggestive generalizability of genetic loci across ancestrally distinct populations for some but not all loci. For example, for the SNPs associated with BMI in our Hispanic

Nutrition & Diabetes (2013) 1 10 & 2013 Macmillan Publishers Limited

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r2 with indexSNP

(in

Hispanic

women)

c ProxySNP

Table3b.ResultsinHispanicwomenforpublishedlocifromEuropeandescentindividualsforlociassociatedwithWCandWHR,afteradjustmentforBMI

Indexor

ChrBPpositionStrandMinor/

major

Allele

rs2301453a DNM3-PIGC1WCadjBMIrs66989871170635590 G/A0.1034010.1310.0411.47E031.6130.0570.181

WHRadjBMIrs66989871170635590 G/A0.1033990.1080.0418.24E030.0080.0570.181

rs2605100b LYPLAL11WCadjBMIrs44727631217887690C/T0.1934530.0870.0315.00E 031.065o0.0010.007

WHRadjBMIrs27859901217754055C/T0.443449 0.0730.0253.37E03 0.0050.5360.435

rs6717858c GRB142WCadjBMIrs67480912165327216G/A0.073454 0.1300.0486.09E03 1.6050.1050.134

WHRadjBMIrs67480912165327216G/A0.073452 0.1550.0479.56E04 0.0110.1050.134

rs6784615NISCH-STAB13WCadjBMIrs4687612352342972 C/T0.143452 0.0800.0352.27E02 0.9890.0080.009

WHRadjBMIrs4687612352342972 C/T0.143450 0.0880.0351.19E02 0.0060.0080.009

rs6795735ADAMTS93WCadjBMIrs17071048364522218C/T0.1334530.1100.0373.05E031.357o0.0010.083

WHRadjBMIrs4688486364557121 A/G0.343452 0.0860.0269.14E04 0.0060.0200.081

rs17695092d CPEB45WCadjBMIrs177503185173194190A/G0.313443 0.0620.0272.18E02 0.757o0.0010.006

WHRadjBMIrs29738945173187427G/A0.3134450.0730.0264.78E030.0050.0580.025

rs1294421LY866WCadjBMIrs276899766862258 C/A0.1434510.1170.0346.63E041.443o0.001o0.001

WHRadjBMIrs1714255766728239 A/T0.013451 0.3200.1062.55E03 0.0220.0050.001

rs6905288VEGFA6WCadjBMIrs6905288643866851 C/T0.403446 0.0750.0241.97E03 0.9191.000

rs1055144NFE2L37WCadjBMIrs12533343725642845G/A0.163453 0.0750.0332.45E02 0.9230.0110.002

rs1822438g HOXC1312WCadjBMIrs171019931252389169 C/T0.1234530.0870.0382.23E021.0710.005o0.001

WHRadjBMIrs171019931252389169 C/T0.1234510.1060.0384.94E030.0070.005o0.001

rs4823006ZNRF3KREMEN122WCadjBMIrs4699832227884183C/A0.123454 0.1000.0365.91E03 1.227o0.0010.003

WHRadjBMIrs37884102228000939 G/A0.443447 0.0600.0241.25E02 0.004o0.0010.002

MAFNEffect

WHRadjBMIrs1358980643872529 G/A0.503451 0.0790.0241.02E03 0.0060.6460.583

rs987237TFAP2B6WCadjBMIrs2744498650862810 C/A0.403442 0.0550.0252.58E02 0.6830.0350.139

WHRadjBMIrs9473902650795305 T/C0.003452 0.3350.1796.12E02 0.023o0.0010.003

rs1936805e RSPO36WCadjBMIrs170542046127271958 T/C0.0534240.1340.0531.16E021.6470.064w0.033

WHRadjBMIrs19309526127275973A/T0.4134150.0580.0252.01E020.0040.1240.121

WHRadjBMIrs17152367725721303C/T0.0134410.3310.1258.35E030.023o0.001o0.001

rs545854ww MSRA8WCadjBMIrs240960189771474G/C0.1134530.1170.0382.43E031.438o0.0010.008

WHRadjBMIrs484124889758513T/C0.0134510.3200.1422.42E020.022o0.0010.002

rs12814794f ITPR2-SSPN12WCadjBMIrs21709801226513365 A/T0.413449 0.0570.0252.21E02 0.6970.0070.013

WHRadjBMIrs15132211226330993 A/G0.123443 0.0910.0371.35E02 0.0060.0460.061

b IndexSNPfromLindgrenetal.,butproxySNPforrs4846567fromHeidetal.atr2 0.6;

Abbreviations:ARIC,AtherosclerosisRiskinCommunitiesStudy;BMI,bodymassindex;BP,basepair;Chr,chromosome;LD,linkagedisequilibrium;MAF,minorallelefrequency;SNP,single-nucleotide

polymorphism;WC,waistcircumference;WHR,waist-to-hipratio.*P-valuesinboldindidicateevidenceofassociationbelownominalsignicance(Po0.05)forindexorproxySNPs(reference),andbelowa

BonferronithresholdforthenumberofmostsignicantSNPstested(Po0.05/15).**MostsignicantSNPwithin500kboftheindexorproxySNP(reference).w IfpairwiseLDinformationwasunavailableinARIC

WhitesthenHapMap2release22fordatainCEUwereusedinstead.Ofnote,rs17071048ismonomorphicinCEUHapMappopulations;however,inARICWhitestherewascalculatableLDwithrs6795735.

ww rs7826222wasrenamedrs545854inHapMapBuild36andthusisnotpresentinBuild36imputationsbasedonthatreleaseofHapMap(release22).Aspublicallyavailableinformationwasnotavailableforthis

SNP,informationontheminorallelewassupplementedfromLindgrenetal.a ProxySNPforrs1011731,r2 1;

r2 with

index

SNP(in

ARIC

Whites)

effectincm

(WC)or

unitless

(WHR)

S.e.P-value*Estimated

g ProxySNPforrs1822438,r2 1.

estimate

f ProxySNPforrs12814794,r2 1;

e ProxySNPforrs9491696,r2 0.9;

In/neargeneChrPhenotypeMost

signicant

SNP**

d ProxySNPforrs6861681,r2 1;

proxySNP

forrs1095252,r2 0.9;

& 2013 Macmillan Publishers Limited Nutrition & Diabetes (2013) 1 10

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Generalization at candidate adiposity-loci?

Index-dependent signal?

Index-independent signal?

Index, or proxy SNP1

Other index-dependent signal2

SNP of interest is suggestive3

SNP of interest is possible4

BMI loci WC/WHR

loci

TMEM18 VEGFA

FAIM2 ITPR2-SSPN

FTO

BMI loci

NUDT3/HM GA1 MC4R

WC/WHR loci

TFAP2B DNM3-PIGC

MSRA

BMI loci WC/WHR

loci

FANCL GRB14

ETV5 ADAMTS9

KCTD15 LY86

BMI locus

Figure 1. Evidence for generalization of 16 previously identied obesity loci with BMI, WC and WHR in the WHI SHARe sample of Hispanic women. 1In CEU, index SNPs or proxy SNPs in LD (r2X0.8) below signicance threshold of Po0.05. 2Identied SNP of interest in 1 Mb region is in LD at r2X0.2 in CEU. P-value is Bonferroni-corrected signicant and was at least one order of magnitude smaller than the P-value of the index SNP (or its proxy). 3Identied SNP of interest in 1 Mb region is in LD at r2o0.2 in CEU. After adjustment for the index SNP (or its proxy), the P-value decreased for the SNP of interest. 4Identied SNP of interest in 1 Mb region is in LD at r2o0.2 in CEU. After adjustment for the index SNP (or its proxy), the P-value for the SNP of interest did not increase more than one order of magnitude. Abbreviations: BMI, body mass index; SNP, single nucleotide polymorphism; WC, waist circumference; WHR, waist-hip ratio.

population, using the index SNP (or proxy SNP in LD, r240.5) identied in European descent populations, six loci (TFAP2B, ETV5, TMEM18, FAIM2, FTO and MC4R) also displayed directionally consistent and statistically signicant associations with BMI in two large GWAS studies of BMI in African Americans and Asians.11,13 In addition, two other BMI loci displayed directionally consistent effect estimates for BMI in Hispanics (NUDT3 and KCTD15), but did not display statistical signicance. These study ndings demonstrate, for the rst time, a general relevance of these BMI loci across multiple ancestrally diverse US minority populations. Although we provide some evidence for generalization, our sample size is small and further verication of these ndings is necessary. Further, of note, our data demonstrate often substantial differences in allele frequencies between the reference HapMap CEU population and the female participants form the WHI SHARE study. Although this study only summarizes data from a single group of Hispanics, 43% with Mexican origins and, on average, 33% Native American ancestry (Supplementary Figure 1), these data do demonstrate the extensive diversity of the Hispanic population and the critical need for a greater focus on the genetic architecture in ethnic minority populations.

It is of interest to note that FANCL did not display any evidence for generalization in African and Asian ancestry populations. In Hispanics, we detected evidence for the proxy SNP and also for a possible independent signal, suggesting distinctions at this locus across ancestral populations.

There have been fewer genetic epidemiological studies of WC and WHR in ancestrally diverse populationsperhaps because waist traits are collected less frequently in large cohorts. One study in a sample of South Asian descent found that SNPs in LD with the identied index SNP (rs1095252) near the GRB14 loci were associated with WHR and type 2 diabetes.42 In Hispanic women, we identied a possible index-dependent signal at this locus supporting the relevance of this locus across populations.

In this study, we were able to generalize or nd evidence of association at 9 of 32 BMI loci (28%) and 7 of 15 WC/WHR loci (47%). Even among those loci that did not generalize in this study, the majority exhibited consistent directions of effect. The greater proportion of ndings at central adiposity loci may likely be due to the greater power to detect associations. As shown in Supplementary Figures 2ac, power calculations revealed disparate curves for overall (BMI) versus central adiposity

measures (WC or WHR), wherein we were underpowered (o80% power) to detect effects for BMI. Between WC and WHR, we were most powered to detect effects on WHR. Of note, these calculations were based on a range of allele frequencies and effect sizes, as well as the distribution of the phenotype in WHI SHARe. Among the three measurements, BMI had the highest level of variability (z-score 5.2), followed by WC (z-score 7.0)

and WHR (z-score 11.7), respectively. Similarly, the BMI ndings

were subjected to a higher penalty of Bonferroni correction (that is, lower alpha), because of the greater number of variants tested. We may also have had greater power to detect waist-related traits as WHI comprises women only, and we have recently established a stronger magnitude of genetic effects in women for many of the established waist variants.43

National estimates from 1982 to 1984 from the Hispanic Health and Nutritional Examination Survey44 were among the rst to show that the burden of obesity may not be similar across all adult US Hispanics. More recent data from a diverse cohort study of four US communities strongly supports the possibility that there may be disparities in obesity among this ethic group by country of origin, with Puerto Rican women have the highest prevalence of obesity (51%) followed closely by Dominican (42%), Central American (42%) and Mexican women (42%).3 Unfortunately, although the WHI Hispanic sample included in WHI SHARe roughly corresponds to the distribution of US Hispanics recorded in the 2010 Census (63% of Hispanics of Mexican descent),30 it likely

does not capture the true diversity that constitutes this ethnicity nor does it imply that these results can be generalized to all US Hispanics. Moreover, the small sample size limited our ability to assess heterogeneity of effect size by population of origin. Future research should be designed and powered to investigate genetic effects across diverse Hispanic backgrounds.

Finally, our sample of primarily postmenopausal women may have been a limitation as there is evidence that genetic effects on adiposity vary substantially across the life course.4547

In turn, this study is strengthened by a number of factors. First, obesity-related racial/ethnic- and gender disparities exist among the largest US minority groupHispanics4851 and progress in the

obesity eld will only be made when all US populations are successfully interrogated. Therefore, our interrogation of established BMI and WC/WHR loci in an ancestrally diverse population with heightened disparities in disease risk is timely and of great public health signicance. Second, to our knowledge this study constitutes

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Generalization of adiposity loci to Hispanic women M Graff et al

9

MAFNBetaS.e.P-valueNBetaS.e.P-valueTypec

the rst attempt in the scientic literature to perform a large comprehensive study of multiple adiposity phenotypes among a sample of Hispanic individuals. Although previous generalization studies have been published among Hispanics, they were largely conducted in the context of candidate gene studies and did not evaluate well established GWAS variants.

In summary, our ndings suggest similar genetic inuences on body size and shape across non-Hispanic and Hispanic descent populations, by illustrating associations at nine BMI loci and seven WC/WHR loci previously reported in European descent populations. We also provide tentative evidence that several of the BMI and WC loci harbor multiple independent signals, which has been shown to increase the heritability explained for complex traits across populations. Nonetheless, replication of these signals in larger Hispanic studies is required, as well as GWAS studies to determine if novel obesity loci can be mapped in Hispanic populations.

CONFLICT OF INTEREST

The authors declare no conict of interest.

ACKNOWLEDGEMENTS

This work was supported by the National Heart, Lung and Blood Institute of the National Institutes of Health, US Department of Health and Human Services (grants 5T32HL007055 to LF-R, 5K07CA136969 to HMO-B). WHI SHARe was also supported by the National Heart, Lung and Blood Institute through the following contracts: N01WH22110, 24152, 321002, 321056, 321089, 3211113, 32115, 321189, 32122, 4210726, 4212932 and 44221. WHI has approved the manuscript, which was prepared in collaboration with WHI investigators.

AUTHOR CONTRIBUTIONS

MG, LF-R, KM and KEN dened the study approach and aims; MG, LF-R and KM completed the data analysis; MG, LF-R and KEN contributed to the interpretation and presentation of the study ndings; all other authors reviewed the manuscript and provided their critical feedback and approval.

REFERENCES

1 Ennis SR, Ros-Vargas M, Albert NG. The Hispanic Population: 2010, in 2010

Census Briefs, US Census, Editor, 2011 US Dept. of Commerce: Washington, DC, USA.2 Flegal KM, Carroll MD, Kit BK, Ogden CL. Prevalence of obesity and trends in the distribution of body mass index among US adults, 1999-2010. JAMA 2012; 307: 491497.3 Daviglus ML, Talavera GA, Aviles-Santa ML, Allison M, Cai J, Criqui MH et al. Prevalence of major cardiovascular risk factors and cardiovascular diseases among Hispanic/Latino individuals of diverse backgrounds in the United States. JAMA 2012; 308: 17751784.4 Speliotes EK, Willer CJ, Berndt SI, Monda KL, Thorleifsson G, Jackson AU et al. Association analyses of 249 796 individuals reveal 18 new loci associated with body mass index. Nat Genet 2010; 42: 937948.5 Thorleifsson G, Walters G, Gudbjartsson D, Steinthorsdottir V, Sulem P, Helgadottir A et al. Genome-wide association yields new sequence variants at seven loci that associate with measures of obesity. Nat Genet 2009; 41: 1824.6 Willer CJ, Speliotes EK, Loos RJ, Li S, Lindgren CM, Heid IM et al. Six new loci associated with body mass index highlight a neuronal inuence on body weight regulation. Nat Genet 2009; 41: 2534.7 Heid IM, Jackson AU, Randall JC, Winkler TW, Qi L, Steinthorsdottir V et al. Meta-analysis identies 13 new loci associated with waist-hip ratio and reveals sexual dimorphism in the genetic basis of fat distribution. Nat Genet 2010; 42: 949960.8 Lindgren CM, Heid IM, Randall JC, Lamina C, Steinthorsdottir V, Qi L et al. Genome-wide association scan meta-analysis identies three Loci inuencing adiposity and fat distribution. PLoS Genet 2009; 5: e1000508.9 Okada Y, Kubo M, Ohmiya H, Takahashi A, Kumasaka N, Hosono N et al. Common variants at CDKAL1 and KLF9 are associated with body mass index in east Asian populations. Nat Genet 2012; 44: 302306.10 Wang J, Mei H, Chen W, Jiang Y, Sun W, Li F et al. Study of eight GWAS-identied common variants for association with obesity-related indices in Chinese children at puberty. Int J Obes (Lond) 2012; 36: 542547.11 Wen W, Cho YS, Zheng W, Dorajoo R, Kato N, Qi L et al. Meta-analysis identies common variants associated with body mass index in east Asians. Nat Genet 2012; 44: 307311.

Table4.ResultsforconditionalanalysesofthemostsignicantSNPassociatedwithBMI,WCandWHR,afteradjustmentforBMI,inHispanicwomenbeforeandafterconditioningontheindexSNP

publishedinEuropeandescentpopulationsoraproxySNPthereofa

BMIFANCL2rs75925059182657A/G0.18rs467226658978503G/A0.3434520.0910.0252.92E0434520.0910.0253.02E04Possible

BMIETV53rs1516728187312585A/T0.37rs7648336187431942A/C0.2634650.0910.0281.14E0334650.0900.0281.22E03Possible

BMTFAP2B6rs936642622172618C/T0.39rs287661122108317A/G0.0134640.4000.1208.49E0434640.4010.1208.31E04Suggestive

BMIKCTD1519rs1108475339013977A/G0.34rs810426238895287A/G0.0134660.4240.1121.65E0434660.4240.1121.67E04Possible

WCadjBMIDNM3-

PIGC

1rs2301453170624790G/A0.38rs6698987170635590G/A0.1033940.1090.0417.76E0333940.1240.0455.96E03Suggestive

WHRadjBMIGRB142rs6717858165247907C/T0.31rs6748091165327216G/A0.0734470.1560.0479.06E0434470.1500.0502.88E03Possible

WCadjBMIADAMTS93rs679573564680405C/T0.34rs17071048d 64522218C/T0.1334530.1100.0373.05E0334530.1170.0382.19E03Possible

WHRadjBMIrs468848664557121A/G0.343452 0.0860.0269.14E043452 0.0880.0279.59E04

WCadjBMILY866rs12944216688148A/C0.49rs27689976862258C/A0.1434510.1170.0346.63E0434510.1170.0346.75E04Possible

WHRadjBMIrs171425576728239A/T0.013451 0.3200.1062.55E033451 0.3200.1062.52E03

WCadjBMIMSRA8rs5458549897490C/G0.23rs24096019771474G/C0.1134390.1180.0382.22E0334390.1200.0391.89E03Suggestive

Abbreviations:ARIC,AtherosclerosisRiskinCommunities;BMI,bodymassindex;Chr,chromosome;SNP,single-nucleotidepolymorphisms;MAF,minorallelefrequency;WC,waistcircumference;WHR,waist-to-

hipratio.a Index(orproxy)SNPandmostsignicantSNPwereconsideredindependentperr2 o0.2betweenWHISHAReHispanicwomenandARICStudyWhites(orCEUHapMappopulations,when

appropriate).b MostsignicantSNPwithin500kboftheindexorproxySNP(reference).c NolocishowedmorethanoneorderofmagnitudeincreaseinP-valueafteradjustmentfortheindexorproxySNPand

thereforenonearecategorizedasbeingunlikelytohaveasecondarysignal.d rs17071048ismonomorphicinCEUHapMappopulations;however,inARICWhitestherewaslowlinkagedisequilibrium(r2 o0.001)

withrs6795735.

Minor/

major

Allele

Position

(inbase

pairs)

MAFMost

signicant

SNPb

BPpositionMinor/

major

allele

ChrIndexor

proxySNP

conditioned

on

PhenotypeIn/near

gene

& 2013 Macmillan Publishers Limited Nutrition & Diabetes (2013) 1 10

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