Abstract

Genetic variation in ABCG2 (rs2231142, Q141K), encoding a uric acid transporter, is associated with gout in diverse populations. The aim of this study was to examine a role for ABCG2 in gout susceptibility in New Zealand Māori, Pacific Island and Caucasian samples. Patients (n = 185, 173 and 214, for Māori, Pacific Island and Caucasian, respectively) satisfied the American College of Rheumatology gout classification criteria. The comparison samples comprised 284, 129 and 562 individuals, respectively, without gout. rs2231142 was genotyped and stratification accounted for using genomic control markers. Association of the minor allele of rs2231142 with gout was observed in the Pacific Island samples (OR = 2.80, PSTRAT < 0.001 after accounting for effects of population structure), but not in the Māori samples (OR = 1.08, PSTRAT= 0.70), with heterogeneity in association evident between the Māori and Pacific Island datasets (PHET = 0.001). A similar dichotomy in association was observed when samples were stratified into Western (Tonga, Samoa, Niue, Tokelau) versus Eastern Polynesian (Māori, Cook Island) origin (OR = 2.59, PSTRAT < 0.001; OR = 1.12, PSTRAT= 0.48, respectively; PHET = 0.005). Association with gout was observed in the Caucasian samples (OR = 2.20, P = 3.2 × 10−8). Unlike SLC2A9, which is a strong risk factor for gout in both Māori and Pacific Island people, ABCG2 rs2231142 has a strong effect only in people of Western Polynesian ancestry. Our results emphasize the need to account for sub-population differences when undertaking biomedical genetic research in a group defined by a geographical region and shared ancestry but characterized by migratory events that create bottlenecks and altered genetic structure in the founder populations.

INTRODUCTION

Gout, a condition of extreme hyperuricaemia, is the most common form of inflammatory arthritis affecting men. Acute gouty arthritis results from an intense inflammatory reaction to monosodium urate crystals present in joint structures. In New Zealand (NZ), gout affects 9.3–13.9% of Māori men and 14.9% of Pacific Island men, with Māori and Pacific Island people also having high rates of severe gout, early onset, tophaceous disease and accelerated joint damage (1). Renal under-excretion of uric acid is a primary gout-determining checkpoint, particularly in Māori and Pacific Island people (2).

Recent studies have confirmed genetic regulation of renal excretion of uric acid. Genetic variation in the urate transporter SLC2A9 (GLUT9) is a risk factor for gout in sample sets drawn from Caucasian populations (3,4), NZ's Māori and Pacific Island population (3), Han Chinese and Solomon Islanders (5), but not in African-American people (4). Very recently, four further loci that influence serum urate concentration, primarily encoding uric acid transport proteins, have been confirmed (6). In addition, a missense single nucleotide polymorphism (SNP) (rs2231142; Q141K) in ABCG2 has been associated with hyperuricaemia and gout in Caucasian, Han Chinese, Japanese and African-American samples (4,7–10). ABCG2 is postulated to be a unidirectional secretory urate transporter in the proximal renal tubule (7,9), and, because of expression in the liver and intestine, is also proposed to mediate urate expression in the gut (9). The lysine allele of Q141K, which explains the genetic association in Caucasian populations, encodes a transporter with ∼50% reduced activity (7,9).

Given the very high rate of gout in NZ Māori and Pacific Island people, here we tested whether or not ABCG2 [rs2231142 (Q141K)] conferred a strong risk for gout in case–control samples drawn from these populations, and in an NZ Caucasian case–control sample set. NZ Māori are tangata whenua (‘people of the land’), settling in NZ from the ancestral homeland known as Hawaiki, which is thought to be in Eastern Polynesia, from ∼1250 AD (Fig. 1). Current evidence indicates that Polynesia was populated over a 5000 year period from Taiwan (11). The Caucasian population of NZ is derived primarily from British colonization in the 19th century, whereas the NZ Pacific Island population has largely arisen from recent immigration (since the 1950s) from Samoa, Tonga, the Cook Islands and other South Pacific nations.

Figure 1.

The Polynesian migrations. 1, voyagers migrated outside Melanesia; 2, Polynesian culture developed in situ in the Samoa/Tonga region for as much as 1500 years before voyagers settled in Eastern Polynesia ∼500 AD; 3, latest evidence suggests a multi-archipelago Polynesian homeland connected by active voyaging, a single language and culture. NZ (Aotearoa) was settled as late as 1250 AD by Māori from the ancestral homeland known as Hawaiki, which is thought to be in Eastern Polynesia; the language of NZ Māori is closely related to Cook Island Māori and Tahitian. Within the circled area ‘2’ of Western Polynesia, Tokelau lies north and Niue southeast of Samoa. Map source: http://thehonoluluadvertiser.com/dailypix/2006/Mar/10/M121533310.GIF.

Figure 1.

The Polynesian migrations. 1, voyagers migrated outside Melanesia; 2, Polynesian culture developed in situ in the Samoa/Tonga region for as much as 1500 years before voyagers settled in Eastern Polynesia ∼500 AD; 3, latest evidence suggests a multi-archipelago Polynesian homeland connected by active voyaging, a single language and culture. NZ (Aotearoa) was settled as late as 1250 AD by Māori from the ancestral homeland known as Hawaiki, which is thought to be in Eastern Polynesia; the language of NZ Māori is closely related to Cook Island Māori and Tahitian. Within the circled area ‘2’ of Western Polynesia, Tokelau lies north and Niue southeast of Samoa. Map source: http://thehonoluluadvertiser.com/dailypix/2006/Mar/10/M121533310.GIF.

RESULTS

Genotype and allele distributions of rs2231142 are shown in Table 1. There was strong evidence for association of rs2231142 with gout in both the Pacific Island and Caucasian sample sets (OR = 2.80, P=1.9 × 10−8 and OR = 2.20, P=3.2 × 10−8, respectively). Association in the Pacific Island sample set was also evident after accounting for the possible presence of stratification owing to admixture with Caucasian (PSTRAT< 0.001). In the Māori sample set, there was no evidence for association (OR = 1.08, allelic P=0.74, PSTRAT= 0.70 after accounting for possible stratification). In both the Caucasian and Pacific Island sample sets, a dominant mode of inheritance could be rejected (P< 1 × 10−4 and P=3 × 10−4, respectively), as could a recessive model (P=0.03 and P=0.01, respectively).

Table 1.

Analysis of association of rs2231142 (ABCG2: Q141K) in NZ Māori, Pacific Island and Caucasian case–control samples

 Genotype, no. (frequency)a,b
 
Minor allele, no. (frequency) Allelic OR (95% CI) Pc PSTRAT 
Cohort 1/1 1/2 2/2     
Māori 
 Case 142 (0.798) 34 (0.191) 2 (0.011) 38 (0.107)    
 Control 172 (0.811) 39 (0.184) 1 (0.005) 41 (0.097) 1.12 (0.70–1.78) 0.64  
 Control + RA 225 (0.804) 54 (0.193) 1 (0.004) 56 (0.100) 1.08 (0.70–1.66) 0.74 0.70 
Pacific Island 
 Case 58 (0.335) 78 (0.451) 37 (0.214) 152 (0.439)    
 Control 69 (0.633) 36 (0.330) 4 (0.037) 44 (0.202) 3.10 (2.09–4.59) 8.3 × 10−9  
 Control + RA 79 (0.617) 42 (0.328) 7 (0.055) 56 (0.219) 2.80 (1.94–4.03) 1.9 × 10−8 <0.001 
Caucasian 
 Case 122 (0.578) 76 (0.360) 13 (0.062) 102 (0.242)    
 Control 425 (0.762) 125 (0.224) 8 (0.014) 141 (0.126) 2.20 (1.66–2.93) 3.2 × 10−8 — 
Western Polynesiad 
 Case 29 (0.225) 66 (0.512) 34 (0.264) 134 (0.519)    
 Control 36 (0.507) 31 (0.437) 4 (0.056) 39 (0.275) 2.85 (1.83–4.44) 2.3 × 10−6  
 Control + RA 42 (0.494) 36 (0.424) 7 (0.082) 50 (0.294) 2.59 (1.72–3.91) 4.2 × 10−6 <0.001 
Eastern Polynesiad 
 Case 160 (0.796) 39 (0.194) 2 (0.010) 43 (0.107)    
 Control 173 (0.824) 36 (0.171) 1 (0.005) 38 (0.090) 1.20 (0.76–1.91) 0.43  
 Control + RA 227 (0.811) 52 (0.186) 1 (0.004) 54 (0.096) 1.12 (0.74–1.71) 0.59 0.48 
 Genotype, no. (frequency)a,b
 
Minor allele, no. (frequency) Allelic OR (95% CI) Pc PSTRAT 
Cohort 1/1 1/2 2/2     
Māori 
 Case 142 (0.798) 34 (0.191) 2 (0.011) 38 (0.107)    
 Control 172 (0.811) 39 (0.184) 1 (0.005) 41 (0.097) 1.12 (0.70–1.78) 0.64  
 Control + RA 225 (0.804) 54 (0.193) 1 (0.004) 56 (0.100) 1.08 (0.70–1.66) 0.74 0.70 
Pacific Island 
 Case 58 (0.335) 78 (0.451) 37 (0.214) 152 (0.439)    
 Control 69 (0.633) 36 (0.330) 4 (0.037) 44 (0.202) 3.10 (2.09–4.59) 8.3 × 10−9  
 Control + RA 79 (0.617) 42 (0.328) 7 (0.055) 56 (0.219) 2.80 (1.94–4.03) 1.9 × 10−8 <0.001 
Caucasian 
 Case 122 (0.578) 76 (0.360) 13 (0.062) 102 (0.242)    
 Control 425 (0.762) 125 (0.224) 8 (0.014) 141 (0.126) 2.20 (1.66–2.93) 3.2 × 10−8 — 
Western Polynesiad 
 Case 29 (0.225) 66 (0.512) 34 (0.264) 134 (0.519)    
 Control 36 (0.507) 31 (0.437) 4 (0.056) 39 (0.275) 2.85 (1.83–4.44) 2.3 × 10−6  
 Control + RA 42 (0.494) 36 (0.424) 7 (0.082) 50 (0.294) 2.59 (1.72–3.91) 4.2 × 10−6 <0.001 
Eastern Polynesiad 
 Case 160 (0.796) 39 (0.194) 2 (0.010) 43 (0.107)    
 Control 173 (0.824) 36 (0.171) 1 (0.005) 38 (0.090) 1.20 (0.76–1.91) 0.43  
 Control + RA 227 (0.811) 52 (0.186) 1 (0.004) 54 (0.096) 1.12 (0.74–1.71) 0.59 0.48 

aAll sample sets were in Hardy–Weinberg equilibrium (P > 0.05). Genotype success rates: 96.2% Māori cases, 100% Pacific Island cases, 98.6% Caucasian cases, 98.7% combined Māori and Pacific Island controls and RA cases; 99.3% Caucasian controls.

bGenotypic ORs comparing with 1/1 genotype: Caucasian 1/2, 2.12 (1.49–3.00); 2/2, 5.66 (2.29–13.97): Māori 1/2, 1.00 (0.62–1.61); 2/2, 3.17 (0.28–35.27): Pacific 1/2, 2.53 (1.53–4.19); 2/2, 7.20 (3.00–17.29).

cAllelic P-value, calculated by Pearson's method (no Yates correction).

dIndividuals of mixed Western and Eastern Polynesian ancestry were excluded.

A notable feature of the data was the apparent heterogeneity between the NZ Māori and Pacific populations at ABCG2, both Polynesian populations with a closely shared ancestry, both with a high prevalence of gout. The strongest effect was in the Pacific Island samples (OR = 2.80), with no detectable effect in the Māori samples (OR = 1.08). Also notable were a relatively large number of homozygous risk allele (2/2) individuals in the Pacific Island cases, compared with the other sample sets (Table 1; Pacific Island 21.4%; Māori 1.1%; Caucasian 6.2%). The difference in genotype distributions between the Māori and Pacific Island case samples was highly significant (P=6 × 10−19), and the Breslow–Day test revealed heterogeneity in the association with gout (PHET = 0.001; PHET = 0.001 with rheumatoid arthritis (RA) controls removed). To investigate the Māori and Pacific Island difference further, we divided the Pacific Island samples by genotype according to ancestral origin (Table 2). In the cases, all of the risk allele homozygotes (2/2) were of Western Polynesian (Samoan, Tongan, Tokelauan or Tuvaluan) ancestry, while the largest proportion of the major allele homozygotes (37%) were of Cook Island ancestry (Eastern Polynesian). There was a clear region-specific effect in genotype distribution between Western Polynesia (Samoa, Tonga, Niue) and Eastern Polynesia (Cook Islands) in the cases (P = 2 × 10−6), less so in the comparison group (P=0.008). The genotype distribution of the Cook Island samples was similar to the Māori samples (Tables 1 and 2; PCASES = 0.87, PCONTROLS = 0.52). On the basis of these observations, the Pacific Island and Māori cases and controls were divided into Western (Samoa, Tonga, Niue, Tokelau, Tuvalu) and Eastern (NZ Māori, Cook Island) Polynesian ancestry, and the association analysis was repeated (Table 1). The OR for people of Western Polynesian ancestry was 2.59 [(1.72–3.91), PSTRAT< 0.001] and 1.12 [(0.74–1.71), PSTRAT= 0.48] for people of Eastern Polynesian ancestry. The Breslow–Day test revealed heterogeneity in the association with gout between the two groups (PHET = 0.005; PHET = 0.008 with RA controls removed).

Table 2.

rs2231142 genotype and ancestry within the Pacific Island samples

Genotype Samoa Tonga Niue Cook Island Other 
1,1 
 Cases (%) 16 (29)a 6 (11) 10 (18) 21 (37) 3 (5) 
 Controls + RA (%) 36 (49) 5 (7) 11 (15) 19 (25) 3 (4) 
1,2 
 Cases (%) 43 (57) 13 (17) 9 (12) 8 (10) 3 (4) 
 Controls + RA (%) 27 (66) 7 (17) 3 (7) 2 (5) 2 (5) 
2,2 
 Cases (%) 25 (69) 8 (22) 0 (0) 0 (0) 3 (8) 
 Controls + RA (%) 4 (57) 3 (43) 0 (0) 0 (0) 0 (0) 
Genotype Samoa Tonga Niue Cook Island Other 
1,1 
 Cases (%) 16 (29)a 6 (11) 10 (18) 21 (37) 3 (5) 
 Controls + RA (%) 36 (49) 5 (7) 11 (15) 19 (25) 3 (4) 
1,2 
 Cases (%) 43 (57) 13 (17) 9 (12) 8 (10) 3 (4) 
 Controls + RA (%) 27 (66) 7 (17) 3 (7) 2 (5) 2 (5) 
2,2 
 Cases (%) 25 (69) 8 (22) 0 (0) 0 (0) 3 (8) 
 Controls + RA (%) 4 (57) 3 (43) 0 (0) 0 (0) 0 (0) 

aNumber (% of the particular genotype; rows sum to 100%). Individuals were assigned according to the self-reported ancestry of the highest proportion. Individuals were excluded if they were (i) of equal Western and Eastern Polynesian ancestry (three cases, two controls), or (ii) if no specific Pacific ancestry data were available (one case, four controls).

The urate transporter SLC2A9 also confers a strong risk for gout in those from the NZ Caucasian population (OR > 2 (3)). There was no evidence that the SLC2A9 locus (represented by rs11942223) has an epistatic relationship with rs2231142 (P=0.60). The increased risk conferred by risk allele homozygosity at both loci was OR = 4.97 (1.79–13.83) (5.3% of cases and 1.1% of controls had this genotype). We did not test for genetic interaction in the Polynesian samples owing to the extensive rs11942223 major (risk) allele homozygosity in the cases (95% of Māori cases and 99% of Pacific Island cases) (3).

Given a stronger role for rs2231142 in elevating serum urate levels in men than in women (4), we also examined the influence of gender on gout risk conferred by rs2231142 (Supplementary Material, Table S1). In the Caucasian sample set, there was significant evidence for association in both men and women (OR = 2.27, P=1 × 10−5 and OR = 1.96, P=0.04, respectively) but not in the Eastern Polynesian sample set (OR = 1.51, PSTRAT = 0.37 and OR = 0.86, PSTRAT = 0.69, respectively). Using logistic regression models, there was no evidence for a genotype by gender interaction in the Eastern Polynesian or Caucasian sample sets (P>0.14). There were insufficient Western Polynesian female cases (n = 6) to permit an equivalent analysis.

DISCUSSION

Association of rs2231142 (Q141K) with serum urate levels and gout in Caucasian and Japanese is established (4,7,9,10), and has been reported in Han Chinese and African-American sample sets (4,8). We confirmed the gout association in an NZ Caucasian sample set [Table 1; OR = 2.20 (1.66–2.93)] and found evidence for association in the Pacific Island sample set [OR = 2.80 (1.94–4.03)]. The strength of effect in Caucasian gout is higher than that in the self-reported Caucasian gout cases nested with the Framingham Heart Study, Rotterdam Heart Study and Atherosclerosis Risk in the Community studies (OR = 1.97, 1.71 and 1.68, respectively) (4), and similar to that in Han Chinese (allelic OR = 1.72) and Japanese (allelic OR = 2.23) (8,9). No significant association was found in the Māori sample set [OR = 1.08 (0.70–1.66)], which may reflect a false-negative result owing to the lack of power to detect association to a locus of weak effect, particularly since the allele frequency in the Māori non-gout samples was relatively low (0.100). It is important to note that our aim was to investigate whether or not ABCG2 conferred a strong role in gout in NZ Māori (estimated power to detect an effect OR > 1.7 was 97%); in order to determine whether or not ABCG2 confers a weaker role will require testing in a larger case–control sample set. Given the associations of ABCG2 Q141K reported in sample sets drawn from diverse populations (Caucasian, Japanese, Han Chinese, African-American, Pacific Island; Table 1) (4,7–10), the lack of association observed in NZ Māori is notable.

The different results observed between the Pacific Island and Māori cohorts were remarkable (OR = 2.80 and 1.08, respectively, Breslow–Day P=0.001) in the context of the high incidence of gout in both populations and the shared Polynesian ancestry. At SLC2A9 (rs11942223), there is a very strong effect in both Māori and Pacific Island sample sets (OR > 5) (3). At ABCG2 (rs2231142), the Pacific Island cases had a high frequency of risk allele homozygosity (0.214 compared with 0.011 in Māori). The allele and genotype distribution of the Cook Island samples was similar to that of the Māori samples, and combined association analysis emphasized lack of association of rs2231142 with gout in Eastern Polynesian samples (Table 1; OR = 1.08, PSTRAT = 0.70 in Māori; OR = 1.12, PSTRAT = 0.48 in combined Māori and Cook Island samples). These data indicate a gout-related Western versus Eastern Polynesian dichotomy at rs2231142. This is emphasized by the observation that the rs2231142 risk allele frequency is 0.294 in Western Polynesian non-gout samples compared with 0.096 in Eastern Polynesian non-gout samples. The OR decreased slightly when Eastern Polynesian (largely Cook Island) samples were removed from the Pacific Island analysis (from 2.80 to 2.59 in the Western Polynesian samples). Presumably, this reflects stratification at rs2231142 in the Pacific Island samples; a higher proportion of the controls than cases were of Eastern Polynesian ancestry. These ABCG2 data are consistent with data derived from the globin genes, which demonstrated that Polynesians could be divided into three distinct groups (NZ and Cook Island Māori, Tongans and Samoans, and Niue Islanders (12)).

Archaeological evidence demonstrates that the Polynesian culture originated ∼3000 years ago in Tonga and Samoa from the Lapita culture within the Bismarck Archipelago (13) (Fig. 1). By ∼800 years ago, the eastern region of Polynesia from Hawaii in the north to NZ in the south had been settled. The genetic structure of Polynesia is distinctive in the Oceania context, with more similarity to people of Micronesia, Taiwanese Aborigines and East Asia, than to people of Melanesia (14). Within Polynesia, there is a west to east decrease in Y-chromosome and mitochondrial DNA diversity, suggesting the occurrence of founder events (13). Such founder events are the most likely reason for generating the genetic differences underlying the differential in gout risk conferred by rs2231141 between people of Western and Eastern Polynesian ancestry living in NZ today. Our results emphasize the need to account for sub-population differences when undertaking biomedical genetic research in a group such as those of Polynesian descent, defined by a geographical region and shared ancestry but also characterized by migratory events that create bottlenecks and alter genetic structure in the founder populations. The difference between Eastern and Western Polynesia at ABCG2 is more likely to be due to neutral processes such as migration and genetic drift than to adaptation (15).

The effect of the rs2231142 risk (T) allele on serum uric acid levels is stronger in white Caucasian men than in women (16), but no gender effect is observed in African-American people (4). Previously, of the gout case–control sample sets in which interaction of rs2231142 association with gender has been tested, a significantly stronger effect in males was observed in only one out of four sample sets (4). We also found no evidence for interaction with gender in either the Caucasian or Eastern Polynesian sample sets (P > 0.14). Given that ABCG2 Q141K determines serum urate levels, where a gender effect is observed (4), and that gout is a condition of extreme serum urate levels in both men and women, a gender effect of ABCG2 in gout would not be expected since all cases, whether male or female, are already hyperuricaemic.

One issue in our study is the gender imbalance between cases and controls, with the percentage of males ranging from 31.7 in Māori to 48.9 in the Pacific Island samples. Given that female hormones protect from gout (17), the mismatch would serve to decrease the power of our study, but only marginally, and would not cause the heterogeneity observed between Māori and Pacific Island. However, a confounding issue could arise if there were an interaction between rs2231142 genotype and gender. We observed no evidence for this. In the Caucasian samples, significant association was observed in both males and females, and there was no evidence for a gender by genotype interaction in either the Eastern Polynesian or Caucasian sample sets.

SNP rs2231142 is highly likely to be the aetiological variant in Caucasian, with the risk allele encoding a urate secretory transporter with 53% reduced ability to transport urate in Xenopus oocytes (7) and HEK293 membrane vesicles (9). The simplest hypothesis is that this variant is also aetiological in people of Western Polynesian ancestry. However, this needs to be confirmed by re-sequencing ABCG2 in this population group and genotyping all variants in gout cases and controls, in a fashion analogous to that done previously in a Japanese sample set (9), in which a genetically independent variant (Q126X; rs72552713) also confers risk to gout (OR = 4.25). ABCG2 is located in the apical membrane of kidney proximal tubule cells, hepatocytes and gut enterocytes where it is hypothesized to excrete uric acid (7,9). The minor allele (141K) encodes a variant that considerably lowers the ABCG2 expression level (30–50%) in membrane vesicles from HEK293 cells (9,18), thus enhancing the susceptibility of ABCG2 to ubiquitin-mediated proteasomal degradation in the endoplasmic reticulum (19). This may account for the observation that 141K encodes a less active transporter (7,9).

ABCG2 is also known as breast cancer resistance protein, functioning as an efflux pump with an influence on the pharmacokinetic profile of a variety of drugs. Consequently, there has been considerable work undertaken studying the biochemistry and modulation of ABCG2. For example, ABCG2 mRNA levels are upregulated by statins in HepG2 cells (20). This is promising in the context of improved therapies for gout, although careful consideration will have to be given to ancestry in any clinical studies involving ABCG2 in the Pacific region.

PATIENTS AND METHODS

Patients

Subjects studied included an NZ Māori case–control sample set of 185 patients and 215 controls, an NZ Pacific Island case–control sample set of 173 patients and 109 controls and an NZ Caucasian case–control sample set of 214 patients and 562 controls. All cases (Table 3) had a confirmed diagnosis of gout by a rheumatologist according to the American College of Rheumatology (ACR) preliminary diagnostic criteria for acute gout (21). There was increased occurrence of metabolic co-morbidities in gout patients compared with controls of the same ethnicity—for example, type 2 diabetes rates were 2.5–4-fold higher in cases. Controls self-reported no history of arthritis. An additional comparison group of Māori (n = 69) and Pacific Island (n = 20) RA patients was included. All had a confirmed diagnosis of RA by a rheumatologist and were diagnosed according to the 1987 ACR criteria for RA (22). There is a widely accepted lack of a coexistent relationship between gout and RA (23). Recruitment of gout and RA patients was approved by the NZ Multi-region Ethics Committee (MEC/05/10/130 and OTA/99/02/007, respectively) and the recruitment of the controls approved by the Lower South Ethics Committee (OTA/99/11/098). All patients provided written informed consent for the collection of samples and subsequent analysis.

Table 3.

Demographic and clinical characteristics of the NZ gout patients

 Gout Māoria (n = 185) Gout Pacific Island (n = 173) Gout Caucasian (n = 214) Control Māoria (n = 284) Control Pacific Islanda (n = 129) Control Caucasian (n = 562) 
Sex, % male (patients)b 73.6 95.7 86.2 32.3 54.2 41.0 
Mean ± SD, grandparents of stated ancestryc 2.96 ± 1.03 3.60 ± 0.69 4.0 ± 0 2.71 ± 1.01 3.40 ± 0.86 4.0 ± 0 
Serum urate at recruitment, mean (range) mg dl−1 7.5 (3.5–15.3) 8.4 (2.9–12.1) 6.8 (3.0–13.4) — — — 
Age of onset, mean (range) yearsd 39.0 (10–74) 33.5 (14–70) 46.2 (4–82) 40.8 (17–75) 37.7 (18–85) 44.6 (17–95) 
Mean (range) no. of gout attacks in past year 13.1 (0 to >1/week) 18.6 (0 to >1/week) 8.5 (0 to >1/week) — — — 
% with first-degree relative with gout 71.8 62.8 51.2 — — — 
Allopurinol treatment, % 85.1 90.0 83.3 — — — 
Probenecid treatment, % 6.0 5.5 7.5 — — — 
Mean (range) BMI 34.8 (22.1–55.9) 37.8 (22.2–93.2) 31.0 (19.9–61.7) 31.8 (19.7–57.0) 32.7 (20.5–62.5) Not available 
Co-morbidities, % 
 Type 2 DM 28.6 21.4 15.7 9.3 8.2 3.5 
 Hypertension 65.5 53.6 51.6 14.7 14.0 14.9 
 Dyslipidaemia 41.1 51.8 44.9 8.5 12.5 11.6 
 Cardiovascular disease 47.9 24.2 42.1 2.4 Not available 
 Renal disease 35.0 29.7 30.0 0.8 Not available 
 Gout Māoria (n = 185) Gout Pacific Island (n = 173) Gout Caucasian (n = 214) Control Māoria (n = 284) Control Pacific Islanda (n = 129) Control Caucasian (n = 562) 
Sex, % male (patients)b 73.6 95.7 86.2 32.3 54.2 41.0 
Mean ± SD, grandparents of stated ancestryc 2.96 ± 1.03 3.60 ± 0.69 4.0 ± 0 2.71 ± 1.01 3.40 ± 0.86 4.0 ± 0 
Serum urate at recruitment, mean (range) mg dl−1 7.5 (3.5–15.3) 8.4 (2.9–12.1) 6.8 (3.0–13.4) — — — 
Age of onset, mean (range) yearsd 39.0 (10–74) 33.5 (14–70) 46.2 (4–82) 40.8 (17–75) 37.7 (18–85) 44.6 (17–95) 
Mean (range) no. of gout attacks in past year 13.1 (0 to >1/week) 18.6 (0 to >1/week) 8.5 (0 to >1/week) — — — 
% with first-degree relative with gout 71.8 62.8 51.2 — — — 
Allopurinol treatment, % 85.1 90.0 83.3 — — — 
Probenecid treatment, % 6.0 5.5 7.5 — — — 
Mean (range) BMI 34.8 (22.1–55.9) 37.8 (22.2–93.2) 31.0 (19.9–61.7) 31.8 (19.7–57.0) 32.7 (20.5–62.5) Not available 
Co-morbidities, % 
 Type 2 DM 28.6 21.4 15.7 9.3 8.2 3.5 
 Hypertension 65.5 53.6 51.6 14.7 14.0 14.9 
 Dyslipidaemia 41.1 51.8 44.9 8.5 12.5 11.6 
 Cardiovascular disease 47.9 24.2 42.1 2.4 Not available 
 Renal disease 35.0 29.7 30.0 0.8 Not available 

aFull data were available for 125 of the Māori gout patients, 129 of the Māori controls and 50 of the Pacific Island controls.

bData for total controls: Māori (n = 284), 31.7%; Pacific Island (n = 129), 48.9%.

cIn the case of the Māori and Pacific Island samples, the mean number of Māori or Pacific Island grandparents is given. [Data for total controls: Māori (n = 284), 2.18 ± 1.19; Pacific (n = 129), 2.96 ± 1.20.]

dAge of recruitment for controls. Data for total controls, mean (range) years: Māori, 41.8 (17–80); Pacific Island, 37.5 (18–85).

Genotyping and statistical analysis

All genotyping assays are detailed in Supplementary Material, Table S1. Genotyping was performed using TaqMan® assay (Applied Biosystems, Foster City, USA) using a Lightcycler® 480 Real-Time Polymerase Chain Reaction (PCR) System (Roche, Indianapolis, USA) or by PCR-restriction fragment length polymorphism with oligonucleotide primers purchased from Sigma-Aldrich. Representative Taqman® scatter plots for rs2231142 in the Māori and Pacific Island gout cases are shown in Supplementary Material, Figure S1. Allelic and genotypic frequencies were compared between case and control samples, odds ratios (OR) and adherence to Hardy–Weinberg equilibrium calculated using the SHEsis package (http://analysis.bio-x.cn/myAnalysis.php). STATA 8.0 software was used to compare logistic regression models when investigating the mode of inheritance and the influence of gender. Recessive or dominant models of inheritance with a multiplicative model were compared (where a linear correlation between number of risk alleles and effect size is assumed). Using STATA 8.0, a model assuming interaction between SLC2A9 (rs11942223) and rs2231142 was fitted against the null model by logistic regression. Meta-analysis was done by the Mantel–Haenszel test using the STATA 8.0 metan software package (http://www.stata.com). The Breslow–Day test was used to test for heterogeneity between the groups. The Caucasian sample set had 89% power to detect association (α = 0.05, allelic OR = 1.7, MAF = 0.11) (4), the Māori sample set 97% power and the Pacific Island sample set 88% power (α = 0.05, allelic OR = 1.7, MAF = 0.32; parameters estimated from the Chinese data of Wang et al. (8), the closest population ancestrally to Polynesian for which data on effect size were available).

Twenty-five biallelic markers were used as genomic controls to account for differing levels of non-Māori and non-Pacific Island ancestry between the case and control samples. The markers used were rs2075876, rs1816532, rs13419122, rs12401573, rs6945435, rs743777, rs10511216, rs12745968, rs1539438, rs729749, rs3738919, rs1130214, rs755622, rs7901695, rs7578597, rs2043211, rs10733113, rs900865, rs2059606, rs4129148, rs831628, rs7725, rs573816, rs1929480 and rs12917707 for the Māori samples, with the removal of rs13419122 and rs755622 for the Pacific Island samples. There was an average allele frequency difference of 0.230 (0.040–0.592) between the Māori cases and Caucasian controls, and a difference of 0.287 (0.091–0.683) between the Pacific Island cases and Caucasian controls. STRUCTURE (24) (http://pritch.bsd.uchicago.edu/software.html) was used to separately assign Māori and Pacific Island individuals to populations (parameters; number of populations assumed to be two, 30 000 burnin period, 1 000 000 Markov chain Monte Carlo replications after burnin). Five hundred and five of the Caucasian control individuals were included in the STRUCTURE procedure to assist in clustering, as representatives of the ancestral Caucasian population. After running STRUCTURE on the Māori samples, the proportion of membership in the inferred Caucasian cluster was 0.953 for the 505 Caucasian controls and 0.064 for the Māori samples. After running STRUCTURE on the Pacific samples, the proportions were 0.967 and 0.070, respectively. The STRUCTURE output was used to run STRAT (25) (http://pritch.bsd.uchicago.edu/software.html) to test for association in the presence of admixture (the phenotype of the 505 Caucasian individuals was set as unknown).

SUPPLEMENTARY MATERIAL

Supplementary Material is available at HMG online.

FUNDING

Supported by the Health Research Council of New Zealand and Arthritis New Zealand. J.E.H-M. was supported by a New Zealand National Heart Foundation Research Fellowship.

ACKNOWLEDGEMENTS

Gael Hewett, Jill James, Karen Lindsay, Maria Lobo, Karen Pui, Gabrielle Sexton and Chloe Waddell are thanked for assistance in recruitment.

Conflict of Interest statement. None declared

REFERENCES

1
Dalbeth
N.
Kumar
S.
Stamp
L.
Gow
P.J.
Dose adjustment of allopurinol according to creatinine clearance does not provide adequate control of hyperuricaemia in patients with gout
J. Rheumatol.
 , 
2006
, vol. 
33
 (pg. 
1646
-
1650
)
2
Simmonds
H.A.
McBride
M.B.
Hatfield
P.J.
Graham
R.
McCaskey
J.
Jackson
M.
Polynesian women are also at risk for hyperuricaemia and gout because of a genetic defect in renal urate handling
Br. J. Rheumatol.
 , 
1994
, vol. 
33
 (pg. 
932
-
937
)
3
Hollis-Moffatt
J.E.
Xu
X.
Dalbeth
N.
Merriman
M.E.
McCallum
R.
Waddell
C.
Gow
P.J.
Harrison
A.A.
Highton
J.
Jones
P.B.
, et al.  . 
A role for the urate transporter SLC2A9 gene in susceptibility to gout in New Zealand Māori, Pacific Island and Caucasian case–control cohorts
Arthritis Rheum.
 , 
2009
, vol. 
60
 (pg. 
3485
-
3492
)
4
Dehghan
A.
Köttgen
A.
Yang
Q.
Hwang
S.J.
Kao
W.H.
Rivadeneira
F.
Boerwinkle
E.
Levy
D.
Hofman
A.
Astor
B.C.
, et al.  . 
Association of three genetic loci with uric acid concentration and risk of gout: a genome-wide study
Lancet
 , 
2008
, vol. 
372
 (pg. 
1953
-
1961
)
5
Tu
H.P.
Chen
C.J.
Tovosia
S.
Ko
A.M.
Ou
T.T.
Lin
G.T.
Chang
H.C.
Chiang
S.L.
Chiang
H.C.
Cen
P.H.
, et al.  . 
Associations of a nonsynonymous variant in SLC2A9 with gouty arthritis and uric acid levels in Han Chinese and Solomon Islanders
Ann. Rheum. Dis.
 , 
2009
, vol. 
69
 (pg. 
887
-
890
)
6
van der Harst
P.
Bakker
S.J.L.
de Boer
R.A.
Wolffenbuttel
B.H.R.
Johnson
T.
Caulfield
M.J.
Navis
G.
Replication of the 5 novel loci for uric acid concentrations and potential mediating mechanisms
Hum. Mol. Genet.
 , 
2009
, vol. 
19
 (pg. 
387
-
395
)
7
Woodward
O.M.
Köttgen
A.
Coresh
J.
Boerwinkle
E.
Guggino
W.B.
Köttgen
M.
Identification of a urate transporter, ABCG2, with a common functional polymorphism causing gout
Proc. Natl Acad. Sci. USA
 , 
2009
, vol. 
106
 (pg. 
10338
-
10342
)
8
Wang
B.
Miao
Z.
Liu
S.
Wang
J.
Zhou
S.
Han
L.
Meng
D.
Wang
Y.
Li
C.
Ma
X.
Genetic analysis of ABCG2 gene C421A polymorphism with gout disease in Chinese Han male population
Hum. Genet.
 , 
2010
, vol. 
127
 (pg. 
145
-
146
)
9
Matsuo
H.
Takada
T.
Ichida
K.
Nakamura
T.
Nakayama
A.
Ikebuchi
Y.
Ito
K.
Kusanagi
Y.
Chiba
T.
Tadokoro
S.
, et al.  . 
Common defects of ABCG2, a high-capacity urate exporter, cause gout: a function-based genetic analysis in a Japanese population
Sci Transl. Med.
 , 
2010
, vol. 
1
 pg. 
5ra11
 
10
Yamagishi
K.
Tanigawa
T.
Kitamura
A.
Köttgen
A.
Folsom
A.R.
Iso
H.
The rs2231142 variant of the ABCG2 gene is associated with uric acid levels and gout among Japanese people
Rheumatology
 , 
2010
, vol. 
49
 (pg. 
1461
-
1465
)
11
Renfrew
C.
Where bacteria and languages concur
Science
 , 
2009
, vol. 
323
 (pg. 
467
-
468
)
12
Trent
R.J.
Mickleson
K.N.P.
Yakas
J.
Hertzberg
M.
Population genetics of the globin genes in Polynesians
Hemoglobin
 , 
1988
, vol. 
12
 (pg. 
533
-
537
)
13
Kayser
M.
Brauer
S.
Cordaux
R.
Casto
A.
Lao
O.
Melanesian and Asian origins of Polynesians: mtDNA and Y chromosome gradients across the Pacific
Mol. Biol. Evol.
 , 
2006
, vol. 
23
 (pg. 
2234
-
2244
)
14
Friedlaender
J.S.
Friedlaender
F.R.
Reed
F.A.
Kidd
K.K.
Kidd
J.R.
Chambers
G.K.
Lea
R.A.
Loo
J.H.
Koki
G.
Hodgson
J.A.
, et al.  . 
The genetic structure of Pacific Islanders
PLoS Genet.
 , 
2008
, vol. 
4
 pg. 
e19
 
15
Coop
G.
Pickrell
J.K.
Novembre
J.
Kudaravilli
S.
Li
J.
Absher
D.
Myers
R.M.
Cavalli-Sforza
L.L.
Feldman
M.W.
Pritchard
J.K.
The role of geography in human adaptation
PLoS Genet.
 , 
2009
, vol. 
5
 pg. 
e1000500
 
16
Kolz
M.
Johnson
T.
Sanna
S.
Teumer
A.
Vitart
V.
Perola
M.
Mangino
M.
Albrecht
E.
Wallace
C.
Farrall
M.
, et al.  . 
Meta-analysis of 28,141 individuals identifies common variants within five new loci that influence uric acid concentrations
PLoS Genet.
 , 
2009
, vol. 
5
 pg. 
e1000504
 
17
Richette
P.
Bardin
T.
Gout
Lancet
 , 
2010
, vol. 
375
 (pg. 
318
-
328
)
18
Kondo
C.
Suzuki
H.
Itoda
M.
Ozawa
S.
Sawada
J.D.
Kobayashi
D.
Ieiri
I.
Mine
K.
Ohtsubo
K.
Sugiyama
Y.
Functional analysis of SNPs variants of BCRP/ABCG2
Pharm. Res.
 , 
2004
, vol. 
21
 (pg. 
1895
-
1903
)
19
Furukawa
T.
Wakabayashi
K.
Tamura
A.
Nakagawa
H.
Morishima
Y.
Osawa
Y.
Ishikawa
T.
Major SNP (Q141K) variant of human ABC transporter ABCG2 undergoes lysosomal and proteasomal degradations
Pharm. Res.
 , 
2009
, vol. 
26
 (pg. 
469
-
479
)
20
Rodrigues
A.C.
Curi
R.
Genvigir
F.D.
Hirata
M.H.
Hirata
R.A.
The expression of efflux and uptake transporters are regulated by statins in Caco-2 and HepG2 cells
Acta Pharmacol. Sin.
 , 
2009
, vol. 
30
 (pg. 
956
-
964
)
21
Wallace
S.L.
Robinson
H.
Masi
A.T.
Decker
J.L.
McCarty
D.J.
Yu
T.F.
Preliminary criteria for the classification of the acute arthritis of primary gout
Arthritis Rheum.
 , 
1977
, vol. 
20
 (pg. 
895
-
900
)
22
Arnett
F.C.
Edworthy
S.M.
Bloch
D.A.
McShane
D.J.
Fries
J.F.
Cooper
N.S.
Healey
L.A.
Kaplan
S.R.
Liang
M.H.
Luthra
H.S.
, et al.  . 
The American Rheumatism Association 1987 revised criteria for the classification of rheumatoid arthritis
Arthritis Rheum.
 , 
1988
, vol. 
31
 (pg. 
315
-
324
)
23
Wallace
D.J.
Klinenberg
J.R.
Morhaim
D.
Berlanstein
B.
Biren
P.C.
Callis
G.
Coexistent gout and rheumatoid arthritis. Case report and literature review
Arthritis Rheum.
 , 
1979
, vol. 
22
 (pg. 
81
-
86
)
24
Pritchard
J.K.
Stephens
M.
Donnelly
P.
Inference of population structure using multilocus genotype data
Genetics
 , 
2000
, vol. 
155
 (pg. 
945
-
959
)
25
Pritchard
J.K.
Stephens
M.
Rosenberg
N.A.
Donnelly
P.
Association mapping in structured populations
Am. J. Hum. Genet.
 , 
2000
, vol. 
67
 (pg. 
170
-
181
)

Supplementary data