Association Analysis of Dyslipidemia-Related Genes in Diabetic Nephropathy

Type 1 diabetes (T1D) increases risk of the development of microvascular complications and cardiovascular disease (CVD). Dyslipidemia is a common risk factor in the pathogenesis of both CVD and diabetic nephropathy (DN), with CVD identified as the primary cause of death in patients with DN. In light of this commonality, we assessed single nucleotide polymorphisms (SNPs) in thirty-seven key genetic loci previously associated with dyslipidemia in a T1D cohort using a case-control design. SNPs (n = 53) were genotyped using Sequenom in 1467 individuals with T1D (718 cases with proteinuric nephropathy and 749 controls without nephropathy i.e. normal albumin excretion). Cases and controls were white and recruited from the UK and Ireland. Association analyses were performed using PLINK to compare allele frequencies in cases and controls. In a sensitivity analysis, samples from control individuals with reduced renal function (estimated glomerular filtration rate<60 ml/min/1.73 m²) were excluded. Correction for multiple testing was performed by permutation testing. A total of 1394 samples passed quality control filters. Following regression analysis adjusted by collection center, gender, duration of diabetes, and average HbA1c, two SNPs were significantly associated with DN. rs4420638 in the APOC1 region (odds ratio [OR]  = 1.51; confidence intervals [CI]: 1.19–1.91; P = 0.001) and rs1532624 in CETP (OR = 0.82; CI: 0.69–0.99; P = 0.034); rs4420638 was also significantly associated in a sensitivity analysis (P = 0.016) together with rs7679 (P = 0.027). However, no association was significant following correction for multiple testing. Subgroup analysis of end-stage renal disease status failed to reveal any association. Our results suggest common variants associated with dyslipidemia are not strongly associated with DN in T1D among white individuals. Our findings, cannot entirely exclude these key genes which are central to the process of dyslipidemia, from involvement in DN pathogenesis as our study had limited power to detect variants of small effect size. Analysis in larger independent cohorts is required.     

Association of Levels of Mannose-Binding Lectin and the MBL2 Gene with Type 2 Diabetes and Diabetic Nephropathy

Objective

To investigate the association of Mannose-binding lectin (MBL) and the MBL2 gene with type 2 diabetes and diabetic nephropathy and the influence of MBL2 polymorphisms on serum MBL levels.

Methods

The study population included 675 type 2 diabetic patients with or without nephropathy and 855 normoglycemic controls. The single nucleotide polymorphisms (SNPs) of rs1800450, rs1800451, and rs11003125 of the MBL2 gene were determined by the Multiplex Snapshot method. Serum MBL levels were measured by enzyme-linked immune sorbent assay.

Results

Rs1800450 and rs11003125 SNPs demonstrated strong linkage disequilibrium in the study population (r² = 0.97). The haplotypes constructed from the G allele of rs1800450 and the C allele of rs11003125 increased the risk for type 2 diabetes (OR = 1.2, 95% CI = 1.1–1.4, P = 0.01). For rs1800450, GG and GA genotypes were associated with type 2 diabetes (P = 0.02, 0.01, respectively). For rs11003125, the GC genotype frequency was significantly different between patients and controls (18.1% vs. 24.9%, P = 0.001). Analyses of genotypes and allele frequency distributions among patients with normal UAE, microalbuminuria, and macroalbuminuria showed that there was no obvious evidence of association between the MBL2 gene and diabetic nephropathy. Subjects with the GG genotype of rs1800450 and the CC genotype of rs11003125 had much higher serum MBL levels.

Conclusions

The rs1800450 and rs11003125 SNPs of the MBL2 gene have strong linkage disequilibrium and are associated with type 2 diabetes in the North Chinese Han population. No association was observed between the MBL2 gene and diabetic nephropathy. Subjects with the GG genotype of rs1800450 and the CC genotype of rs11003125 had much higher serum MBL levels. An association between elevated serum MBL and diabetic nephropathy was also observed.     

Investigation of Type 2 Diabetes Risk Alleles Support CDKN2A/B,CDKAL1, and TCF7L2 As Susceptibility Genes in a Han Chinese Cohort

Background

Recent genome-wide association studies (GWASs) have reported several genetic variants to be reproducibly associated with type 2 diabetes. Additional variants have also been detected from a metaanalysis of three GWASs, performed in populations of European ancestry. In the present study, we evaluated the influence of 17 genetic variants from 15 candidate loci, identified in type 2 diabetes GWASs and the metaanalysis, in a Han Chinese cohort.

Methodology/Principal Findings

Selected type 2 diabetes–associated genetic variants were genotyped in 1,165 type 2 diabetic patients and 1,136 normoglycemic control individuals of Southern Han Chinese ancestry. The OR for risk of developing type 2 diabetes was calculated using a logistic regression model adjusted for age, sex, and BMI. Genotype-phenotype associations were tested using a multivariate linear regression model. Genetic variants in CDKN2A/B, CDKAL1, TCF7L2, TCF2, MC4R, and PPARG showed a nominal association with type 2 diabetes (P≤0.05), of whom the three first would stand correction for multiple testing: CDKN2A/B rs10811661, OR: 1.26 (1.12–1.43) P = 1.8*10⁻⁴; CDKAL1 rs10946398, OR: 1.23 (1.09–1.39); P = 7.1*10⁻⁴, and TCF7L2 rs7903146, OR: 1.61 (1.19–2.18) P = 2.3 * 10⁻³. Only nominal phenotype associations were observed, notably for rs8050136 in FTO and fasting plasma glucose (P = 0.002), postprandial plasma glucose (P = 0.002), and fasting C-peptide levels (P = 0.006) in the diabetic patients, and with BMI in controls (P = 0.033).

Conclusions/Significance

We have identified significant association between variants in CDKN2A/B, CDKAL1 and TCF7L2, and type 2 diabetes in a Han Chinese cohort, indicating these genes as strong candidates conferring susceptibility to type 2 diabetes across different ethnicities.     

Manganese Superoxide Dismutase (SOD2) Polymorphisms,Plasma Advanced Oxidation Protein Products (AOPP) Concentration and Risk of Kidney Complications in Subjects with Type 1 Diabetes

Aims

Oxidative stress is involved in the pathophysiology of diabetic nephropathy. Manganese superoxide dismutase (SOD2) catalyses the dismutation of superoxide, regulates the metabolism of reactive oxygen species in the mitochondria and is highly expressed in the kidney. Plasma concentration of advanced oxidation protein products (AOPP), a marker of oxidative stress, was found to be increased in patients with kidney disease. We investigated associations of SOD2 allelic variations, plasma SOD activity and AOPP concentration with diabetic nephropathy in type 1 diabetic subjects.

Methods

Eight SNPs in the SOD2 region were analysed in 1285 Caucasian subjects with type 1 diabetes from the SURGENE prospective study (n = 340; 10-year follow-up), GENESIS (n = 501) and GENEDIAB (n = 444) cross-sectional studies. Baseline plasma concentration of AOPP and SOD activity were measured in GENEDIAB participants. Hazard ratio (HR) and odds ratio (OR) were determined for incidence and prevalence of nephropathy. Analyses were adjusted or stratified by retinopathy stages.

Results

In the SURGENE cohort, the T-allele of rs4880 (V16A) was associated with the incidence of renal events (new cases, or the progression to a more severe stage of nephropathy; HR 1.99, 95% CI 1.24–3.12, p = 0.004) and with the decline in estimated glomerular filtration rate (eGFR) during follow-up. Similar associations were observed for rs2758329 and rs8031. Associations were replicated in GENESIS/GENEDIAB cohorts, in the subset of participants without proliferative retinopathy, and were confirmed by haplotype analyses. Risk allele and haplotype were also associated with higher plasma AOPP concentration and lower SOD activity.

Conclusions

SOD2 allelic variations were associated with the incidence and the progression of diabetic nephropathy, with a faster decline in eGFR and with plasma AOPP concentration and SOD activity in subjects with type 1 diabetes. These results are consistent with a role for SOD2 in the protection against oxidative stress and kidney disease in type 1 diabetes.     

PPARG,KCNJ11, CDKAL1, CDKN2A-CDKN2B,IDE-KIF11-HHEX,IGF2BP2 and SLC30A8 Are Associated with Type 2 Diabetes in a Chinese Population

Background

Recent advance in genetic studies added the confirmed susceptible loci for type 2 diabetes to eighteen. In this study, we attempt to analyze the independent and joint effect of variants from these loci on type 2 diabetes and clinical phenotypes related to glucose metabolism.

Methods/Principal Findings

Twenty-one single nucleotide polymorphisms (SNPs) from fourteen loci were successfully genotyped in 1,849 subjects with type 2 diabetes and 1,785 subjects with normal glucose regulation. We analyzed the allele and genotype distribution between the cases and controls of these SNPs as well as the joint effects of the susceptible loci on type 2 diabetes risk. The associations between SNPs and type 2 diabetes were examined by logistic regression. The associations between SNPs and quantitative traits were examined by linear regression. The discriminative accuracy of the prediction models was assessed by area under the receiver operating characteristic curves. We confirmed the effects of SNPs from PPARG, KCNJ11, CDKAL1, CDKN2A-CDKN2B, IDE-KIF11-HHEX, IGF2BP2 and SLC30A8 on risk for type 2 diabetes, with odds ratios ranging from 1.114 to 1.406 (P value range from 0.0335 to 1.37E-12). But no significant association was detected between SNPs from WFS1, FTO, JAZF1, TSPAN8-LGR5, THADA, ADAMTS9, NOTCH2-ADAM30 and type 2 diabetes. Analyses on the quantitative traits in the control subjects showed that THADA SNP rs7578597 was association with 2-h insulin during oral glucose tolerance tests (P = 0.0005, empirical P = 0.0090). The joint effect analysis of SNPs from eleven loci showed the individual carrying more risk alleles had a significantly higher risk for type 2 diabetes. And the type 2 diabetes patients with more risk allele tended to have earlier diagnostic ages (P = 0.0006).

Conclusions/Significance

The current study confirmed the association between PPARG, KCNJ11, CDKAL1, CDKN2A-CDKN2B, IDE-KIF11-HHEX, IGF2BP2 and SLC30A8 and type 2 diabetes. These type 2 diabetes risk loci contributed to the disease additively.     

Obesity-Related Genomic Loci Are Associated with Type 2 Diabetes in a Han Chinese Population

Background and Aims

Obesity is a well-known risk factor for type 2 diabetes. Genome-wide association studies have identified a number of genetic loci associated with obesity. The aim of this study is to examine the contribution of obesity-related genomic loci to type 2 diabetes in a Chinese population.

Methods

We successfully genotyped 18 obesity-related single nucleotide polymorphisms among 5338 type 2 diabetic patients and 4663 controls. Both individual and joint effects of these single nucleotide polymorphisms on type 2 diabetes and quantitative glycemic traits (assessing β-cell function and insulin resistance) were analyzed using logistic and linear regression models, respectively.

Results

Two single nucleotide polymorphisms near MC4R and GNPDA2 genes were significantly associated with type 2 diabetes before adjusting for body mass index and waist circumference (OR (95% CI) = 1.14 (1.06, 1.22) for the A allele of rs12970134, P = 4.75×10⁻⁴; OR (95% CI) = 1.10 (1.03, 1.17) for the G allele of rs10938397, P = 4.54×10⁻³). When body mass index and waist circumference were further adjusted, the association of MC4R with type 2 diabetes remained significant (P = 1.81×10⁻²) and that of GNPDA2 was attenuated (P = 1.26×10⁻¹), suggesting the effect of the locus including GNPDA2 on type 2 diabetes may be mediated through obesity. Single nucleotide polymorphism rs2260000 within BAT2 was significantly associated with type 2 diabetes after adjusting for body mass index and waist circumference (P = 1.04×10⁻²). In addition, four single nucleotide polymorphisms (near or within SEC16B, BDNF, MAF and PRL genes) showed significant associations with quantitative glycemic traits in controls even after adjusting for body mass index and waist circumference (all P values<0.05).

Conclusions

This study indicates that obesity-related genomic loci were associated with type 2 diabetes and glycemic traits in the Han Chinese population.     

Association analysis of canonical Wnt signalling genes in diabetic nephropathy

Aims/Hypothesis

Several studies have provided compelling evidence implicating the Wnt signalling pathway in the pathogenesis of diabetic nephropathy. Gene expression profiles associated with renal fibrosis have been attenuated through Wnt pathway modulation in model systems implicating Wnt pathway members as potential therapeutic targets for the treatment of diabetic nephropathy. We assessed tag and potentially functional single nucleotide polymorphisms (SNPs; n = 31) in four key Wnt pathway genes (CTNNB1, AXIN2, LRP5 and LRP6) for association with diabetic nephropathy using a case-control design.

Methods

SNPs were genotyped using Sequenom or Taqman technologies in 1351 individuals with type 1 diabetes (651 cases with nephropathy and 700 controls without nephropathy). Cases and controls were white and recruited from the UK and Ireland. Association analyses were performed using PLINK, to compare allele and haplotype frequencies in cases and controls. Adjustment for multiple testing was performed by permutation testing.

Results

Following logistic regression analysis adjusted by collection centre, duration of T1D, and average HbA1c as covariates, a single SNP in LRP6 (rs1337791) was significantly associated with DN (OR = 0.74; CI: 0.57–0.97; P = 0.028), although this was not maintained following correction for multiple testing. Three additional SNPs (rs2075241 in LRP6; rs3736228 and rs491347 both in LRP5) were marginally associated with diabetic nephropathy, but none of the associations were replicated in an independent dataset. Haplotype and subgroup analysis (according to duration of diabetes, and end-stage renal disease) also failed to reveal an association with diabetic nephropathy.

Conclusions/Interpretation

Our results suggest that analysed common variants in CTNNB1, AXIN2, LRP5 and LRP6 are not strongly associated with diabetic nephropathy in type 1 diabetes among white individuals. Our findings, however, cannot entirely exclude these genes or other members of the Wnt pathway, from involvement in the pathogenesis of diabetic nephropathy as our study had limited power to detect variants with small effect size.     

Association of GWAS-Identified Lung Cancer Susceptibility Loci with Survival Length in Patients with Small-Cell Lung Cancer Treated with Platinum-Based Chemotherapy

Genetic variants have been shown to affect length of survival in cancer patients. This study explored the association between lung cancer susceptibility loci tagged by single-nucleotide polymorphisms (SNPs) identified in the genome-wide association studies and length of survival in small-cell lung cancer (SCLC). Eighteen SNPs were genotyped among 874 SCLC patients and Cox proportional hazards regression was used to examine the effects of genotype on survival length under an additive model with age, sex, smoking status and clinical stage as covariates. We identified 3 loci, 20q13.2 (rs4809957G >A), 22q12.2 (rs36600C >T) and 5p15.33 (rs401681C >T), significantly associated with the survival time of SCLC patients. The adjusted hazard ratio (HR) for patients with the rs4809957 GA or AA genotype was 0.80 (95% CI, 0.66–0.96; P = 0.0187) and 0.73 (95% CI, 0.55–0.96; P = 0.0263) compared with the GG genotype. Using the dominant model, the adjusted HR for patients carrying at least one T allele at rs36600 or rs401681 was 0.78 (95% CI, 0.63–0.96; P = 0.0199) and 1.29 (95% CI, 1.08–1.55; P = 0.0047), respectively, compared with the CC genotype. Stratification analyses showed that the significant associations of these 3 loci were only seen in smokers and male patients. The rs4809957 SNP was only significantly associated with length of survival of patients with extensive-stage but not limited-stage tumor. These results suggest that some of the lung cancer susceptibility loci might also affect the prognosis of SCLC.     

A Genome-Wide Search for Linkage of Estimated Glomerular Filtration Rate (eGFR) in the Family Investigation of Nephropathy and Diabetes (FIND)

Objective

Estimated glomerular filtration rate (eGFR), a measure of kidney function, is heritable, suggesting that genes influence renal function. Genes that influence eGFR have been identified through genome-wide association studies. However, family-based linkage approaches may identify loci that explain a larger proportion of the heritability. This study used genome-wide linkage and association scans to identify quantitative trait loci (QTL) that influence eGFR.

Methods

Genome-wide linkage and sparse association scans of eGFR were performed in families ascertained by probands with advanced diabetic nephropathy (DN) from the multi-ethnic Family Investigation of Nephropathy and Diabetes (FIND) study. This study included 954 African Americans (AA), 781 American Indians (AI), 614 European Americans (EA) and 1,611 Mexican Americans (MA). A total of 3,960 FIND participants were genotyped for 6,000 single nucleotide polymorphisms (SNPs) using the Illumina Linkage IVb panel. GFR was estimated by the Modification of Diet in Renal Disease (MDRD) formula.

Results

The non-parametric linkage analysis, accounting for the effects of diabetes duration and BMI, identified the strongest evidence for linkage of eGFR on chromosome 20q11 (log of the odds [LOD] = 3.34; P = 4.4×10⁻⁵) in MA and chromosome 15q12 (LOD = 2.84; P = 1.5×10⁻⁴) in EA. In all subjects, the strongest linkage signal for eGFR was detected on chromosome 10p12 (P = 5.5×10⁻⁴) at 44 cM near marker rs1339048. A subsequent association scan in both ancestry-specific groups and the entire population identified several SNPs significantly associated with eGFR across the genome.

Conclusion

The present study describes the localization of QTL influencing eGFR on 20q11 in MA, 15q21 in EA and 10p12 in the combined ethnic groups participating in the FIND study. Identification of causal genes/variants influencing eGFR, within these linkage and association loci, will open new avenues for functional analyses and development of novel diagnostic markers for DN.     

A Single Nucleotide Polymorphism within DUSP9 Is Associated with Susceptibility to Type 2 Diabetes in a Japanese Population

Aims

The DUSP9 locus on chromosome X was identified as a susceptibility locus for type 2 diabetes in a meta-analysis of European genome-wide association studies (GWAS), and GWAS in South Asian populations identified 6 additional single nucleotide polymorphism (SNP) loci for type 2 diabetes. However, the association of these loci with type 2 diabetes have not been examined in the Japanese. We performed a replication study to investigate the association of these 7 susceptibility loci with type 2 diabetes in the Japanese population.

Methods

We genotyped 11,319 Japanese participants (8,318 with type 2 diabetes and 3,001 controls) for each of the 7 SNPs–rs5945326 near DUSP9, rs3923113 near GRB14, rs16861329 in ST6GAL1, rs1802295 in VPS26A, rs7178572 in HMG20A, rs2028299 near AP3S2, and rs4812829 in HNF4A–and examined the association of each of these 7 SNPs with type 2 diabetes by using logistic regression analysis.

Results

All SNPs had the same direction of effect (odds ratio [OR]>1.0) as in the original reports. One SNP, rs5945326 near DUSP9, was significantly associated with type 2 diabetes at a genome-wide significance level (p = 2.21×10⁻⁸; OR 1.39, 95% confidence interval [CI]: 1.24−1.56). The 6 SNPs derived from South Asian GWAS were not significantly associated with type 2 diabetes in the Japanese population by themselves (p≥0.007). However, a genetic risk score constructed from 6 South Asian GWAS derived SNPs was significantly associated with Japanese type 2 diabetes (p = 8.69×10⁻⁴, OR  = 1.06. 95% CI; 1.03−1.10).

Conclusions/interpretation

These results indicate that the DUSP9 locus is a common susceptibility locus for type 2 diabetes across different ethnicities, and 6 loci identified in South Asian GWAS also have significant effect on susceptibility to Japanese type 2 diabetes.     

Rs7206790 and rs11644943 in FTO Gene Are Associated with Risk of Obesity in Chinese School-Age Population

To evaluate the associations between candidate FTO single nucleotide polymorphisms (SNPs) and obesity, a case-control study was conducted among Chinese school-age children, which included 500 obese cases and 500 matched controls (age, gender and location). We selected 24 candidate FTO tag-SNPs via bio-informatics analysis and performed genotyping using SNPScan technology. Results indicated that rs7206790 and rs11644943 were significantly associated with obesity among school-age children in both additive and recessive models (P<0.05) after adjusting confounders. Comparing rs7206790 CC and CG genotype of carriers, those carrying the GG genotype had an increased risk of obesity (adjusted odds ratio [OR], 3.76; 95% Confidence interval [CI], 1.24–11.43). Carriers of the AA allele of rs11644943 had a lower risk of obesity (adjusted OR, 0.16; 95% CI, 0.04–0.72) compared with those of the T allele (TT and TA). These two SNPs (rs7206790 and rs11644943) were not Linkage Disequilibrium (LD) with previous reported obesity-associated SNPs. Under the recessive model adjusted for age and gender and location, rs7206790 GG allele carriers had significantly increased BMIs (P = 0.012), weight (P = 0.012), waist circumferences (WC) (P = 0.045) and hip circumferences (HC) (P = 0.033). Conversely, rs11644943 AA allele carriers had significantly decreased BMIs (P = 0.006), WC (P = 0.037) and Waist-to-height ratios (WHtR) (P = 0.012). A dose-response relationship was found between the number of risk alleles in rs7206790, rs11644943 and rs9939609 and the risk of obesity. The Genetic Risk Score (GRS) of the reference group was 3; in comparison, those of 2, 4, and ≥5 had ORs for obesity of 0.24 (95%CI, 0.05–1.13), 1.49 (95%CI, 1.10–2.01), and 5.20 (95%CI, 1.75–15.44), respectively. This study confirmed the role of FTO variation on genetic susceptibility to obesity. We reported two new obesity-related FTO SNPs (rs7206790 and rs11644943) among Chinese school-age children.     

Susceptibility Loci Associated with Specific and Shared Subtypes of Lymphoid Malignancies

The genetics of lymphoma susceptibility reflect the marked heterogeneity of diseases that comprise this broad phenotype. However, multiple subtypes of lymphoma are observed in some families, suggesting shared pathways of genetic predisposition to these pathologically distinct entities. Using a two-stage GWAS, we tested 530,583 SNPs in 944 cases of lymphoma, including 282 familial cases, and 4,044 public shared controls, followed by genotyping of 50 SNPs in 1,245 cases and 2,596 controls. A novel region on 11q12.1 showed association with combined lymphoma (LYM) subtypes. SNPs in this region included rs12289961 near LPXN, (P_LYM = 3.89×10⁻⁸, OR = 1.29) and rs948562 (P_LYM = 5.85×10⁻⁷, OR = 1.29). A SNP in a novel non-HLA region on 6p23 (rs707824, P_NHL = 5.72×10⁻⁷) was suggestive of an association conferring susceptibility to lymphoma. Four SNPs, all in a previously reported HLA region, 6p21.32, showed genome-wide significant associations with follicular lymphoma. The most significant association with follicular lymphoma was for rs4530903 (P_FL = 2.69×10⁻¹², OR = 1.93). Three novel SNPs near the HLA locus, rs9268853, rs2647046, and rs2621416, demonstrated additional variation contributing toward genetic susceptibility to FL associated with this region. Genes implicated by GWAS were also found to be cis-eQTLs in lymphoblastoid cell lines; candidate genes in these regions have been implicated in hematopoiesis and immune function. These results, showing novel susceptibility regions and allelic heterogeneity, point to the existence of pathways of susceptibility to both shared as well as specific subtypes of lymphoid malignancy.     

A Study on Genetic Variants of Fibroblast Growth Factor Receptor 2 (FGFR2) and the Risk of Breast Cancer from North India

Genome-Wide Association Studies (GWAS) have identified Fibroblast growth factor receptor 2 (FGFR2) as a candidate gene for breast cancer with single nucleotide polymorphisms (SNPs) located in intron 2 region as the susceptibility loci strongly associated with the risk. However, replicate studies have often failed to extrapolate the association to diverse ethnic regions. This hints towards the existing heterogeneity among different populations, arising due to differential linkage disequilibrium (LD) structures and frequencies of SNPs within the associated regions of the genome. It is therefore important to revisit the previously linked candidates in varied population groups to unravel the extent of heterogeneity. In an attempt to investigate the role of FGFR2 polymorphisms in susceptibility to the risk of breast cancer among North Indian women, we genotyped rs2981582, rs1219648, rs2981578 and rs7895676 polymorphisms in 368 breast cancer patients and 484 healthy controls by Polymerase chain reaction-Restriction fragment length polymorphism (PCR-RFLP) assay. We observed a statistically significant association with breast cancer risk for all the four genetic variants (P<0.05). In per-allele model for rs2981582, rs1219648, rs7895676 and in dominant model for rs2981578, association remained significant after bonferroni correction (P<0.0125). On performing stratified analysis, significant correlations with various clinicopathological as well as environmental and lifestyle characteristics were observed. It was evident that rs1219648 and rs2981578 interacted with exogenous hormone use and advanced clinical stage III (after Bonferroni correction, P<0.000694), respectively. Furthermore, combined analysis on these four loci revealed that compared to women with 0–1 risk loci, those with 2–4 risk loci had increased risk (OR = 1.645, 95%CI = 1.152–2.347, P = 0.006). In haplotype analysis, for rs2981578, rs2981582 and rs1219648, risk haplotype (GTG) was associated with a significantly increased risk compared to the common (ACA) haplotype (OR = 1.365, 95% CI = 1.086–1.717, P = 0.008). Our results suggest that intron 2 SNPs of FGFR2 may contribute to genetic susceptibility of breast cancer in North India population.     

Association between Common Germline Genetic Variation in 94 Candidate Genes or Regions and Risks of Invasive Epithelial Ovarian Cancer

Background

Recent studies have identified several single nucleotide polymorphisms (SNPs) in the population that are associated with variations in the risks of many different diseases including cancers such as breast, prostate and colorectal. For ovarian cancer, the known highly penetrant susceptibility genes (BRCA1 and BRCA2) are probably responsible for only 40% of the excess familial ovarian cancer risks, suggesting that other susceptibility genes of lower penetrance exist.

Methods

We have taken a candidate approach to identifying moderate risk susceptibility alleles for ovarian cancer. To date, we have genotyped 340 SNPs from 94 candidate genes or regions, in up to 1,491 invasive epithelial ovarian cancer cases and 3,145 unaffected controls from three different population based studies from the UK, Denmark and USA.

Results

After adjusting for population stratification by genomic control, 18 SNPs (5.3%) were significant at the 5% level, and 5 SNPs (1.5%) were significant at the 1% level. The most significant association was for the SNP rs2107425, located on chromosome 11p15.5, which has previously been identified as a susceptibility allele for breast cancer from a genome wide association study (P-trend = 0.0012). When SNPs/genes were stratified into 7 different pathways or groups of validation SNPs, the breast cancer associated SNPs were the only group of SNPs that were significantly associated with ovarian cancer risk (P-heterogeneity = 0.0003; P-trend = 0.0028; adjusted (for population stratification) P-trend = 0.006). We did not find statistically significant associations when the combined data for all SNPs were analysed using an admixture maximum likelihood (AML) experiment-wise test for association (P-heterogeneity = 0.051; P-trend = 0.068).

Conclusion

These data suggest that a proportion of the SNPs we evaluated were associated with ovarian cancer risk, but that the effect sizes were too small to detect associations with individual SNPs.     

Case-Control Analysis of SNPs in GLUT4, RBP4 and STRA6: Association of SNPs in STRA6 with Type 2 Diabetes in a South Indian Population

Background

The inverse relationship between GLUT4 and RBP4 expression is known to play a role in the pathogenesis of type 2 diabetes. Elevated levels of RBP4 were shown to cause insulin resistance in muscles and liver. Identification of STRA6 as a cell surface receptor for RBP4 provides further link in this axis and hence we analyzed SNPs in these three genes for association with type 2 diabetes in a South Indian population.

Methodology/Principal Findings

Selected SNPs in the three genes were analyzed in a total of 2002 individuals belonging to Dravidian ethnicity, South India, by Tetra Primer ARMS PCR or RFLP PCR. Allele frequencies and genotype distribution were calculated in cases and controls and were analyzed for association by Chi-squared test and Logistic regression. Haplotype analysis was carried out for each gene by including all the markers in a single block. We observed a significant association of three SNPs, rs974456, rs736118, and rs4886578 in STRA6 with type 2 diabetes (P = 0.001, OR 0.79[0.69–0.91], P = 0.003, OR 0.81[0.71–0.93], and P = 0.001, OR 0.74[0.62–0.89] respectively). None of the SNPs in RBP4 and GLUT4 showed any association with type 2 diabetes. Haplotype analysis revealed that two common haplotypes H1 (111, P = 0.001, OR 1.23[1.08–1.40]) and H2 (222, P = 0.002 OR 0.73[0.59–0.89]) in STRA6, H6 (2121, P = 0.006, OR 1.69[1.51–2.48]) in RBP4 and H4 (2121, P = 0.01 OR 1.41[1.07–1.85]) in GLUT4 were associated with type 2 diabetes.

Conclusion

SNPs in STRA6, gene coding the cell surface receptor for RBP4, were significantly associated with type 2 diabetes and further genetic and functional studies are required to understand and ascertain its role in the manifestation of type 2 diabetes.     

CDKAL1-Related Single Nucleotide Polymorphisms Are Associated with Insulin Resistance in a Cross-Sectional Cohort of Greek Children

Five novel loci recently found to be associated with body mass in two GWAS of East Asian populations were evaluated in two cohorts of Swedish and Greek children and adolescents. These loci are located within, or in the proximity of: CDKAL1, PCSK1, GP2, PAX6 and KLF9. No association with body mass has previously been reported for these loci in GWAS performed on European populations. The single nucleotide polymorphisms (SNPs) with the strongest association at each loci in the East Asian GWAS were genotyped in two cohorts, one obesity case control cohort of Swedish children and adolescents consisting of 496 cases and 520 controls and one cross-sectional cohort of 2293 nine-to-thirteen year old Greek children and adolescents. SNPs were surveyed for association with body mass and other phenotypic traits commonly associated with obesity, including adipose tissue distribution, insulin resistance and daily caloric intake. No association with body mass was found in either cohort. However, among the Greek children, association with insulin resistance could be observed for the two CDKAL1-related SNPs: rs9356744 (β = 0.018, p = 0.014) and rs2206734 (β = 0.024, p = 0.001). CDKAL1-related variants have previously been associated with type 2 diabetes and insulin response. This study reports association of CDKAL1-related SNPs with insulin resistance, a clinical marker related to type 2 diabetes in a cross-sectional cohort of Greek children and adolescents of European descent.     

Physical Activity Attenuates the Genetic Predisposition to Obesity in 20,000 Men and Women from EPIC-Norfolk Prospective Population Study

Background

We have previously shown that multiple genetic loci identified by genome-wide association studies (GWAS) increase the susceptibility to obesity in a cumulative manner. It is, however, not known whether and to what extent this genetic susceptibility may be attenuated by a physically active lifestyle. We aimed to assess the influence of a physically active lifestyle on the genetic predisposition to obesity in a large population-based study.

Methods and Findings

We genotyped 12 SNPs in obesity-susceptibility loci in a population-based sample of 20,430 individuals (aged 39–79 y) from the European Prospective Investigation of Cancer (EPIC)-Norfolk cohort with an average follow-up period of 3.6 y. A genetic predisposition score was calculated for each individual by adding the body mass index (BMI)-increasing alleles across the 12 SNPs. Physical activity was assessed using a self-administered questionnaire. Linear and logistic regression models were used to examine main effects of the genetic predisposition score and its interaction with physical activity on BMI/obesity risk and BMI change over time, assuming an additive effect for each additional BMI-increasing allele carried. Each additional BMI-increasing allele was associated with 0.154 (standard error [SE] 0.012) kg/m² (p = 6.73×10⁻³⁷) increase in BMI (equivalent to 445 g in body weight for a person 1.70 m tall). This association was significantly (p _interaction = 0.005) more pronounced in inactive people (0.205 [SE 0.024] kg/m² [p = 3.62×10⁻¹⁸; 592 g in weight]) than in active people (0.131 [SE 0.014] kg/m² [p = 7.97×10⁻²¹; 379 g in weight]). Similarly, each additional BMI-increasing allele increased the risk of obesity 1.116-fold (95% confidence interval [CI] 1.093–1.139, p = 3.37×10⁻²⁶) in the whole population, but significantly (p _interaction = 0.015) more in inactive individuals (odds ratio [OR] = 1.158 [95% CI 1.118–1.199; p = 1.93×10⁻¹⁶]) than in active individuals (OR = 1.095 (95% CI 1.068–1.123; p = 1.15×10⁻¹²]). Consistent with the cross-sectional observations, physical activity modified the association between the genetic predisposition score and change in BMI during follow-up (p _interaction = 0.028).

Conclusions

Our study shows that living a physically active lifestyle is associated with a 40% reduction in the genetic predisposition to common obesity, as estimated by the number of risk alleles carried for any of the 12 recently GWAS-identified loci. Please see later in the article for the Editors'' Summary     

Genetic Variations in the Flanking Regions of miR-101-2 Are Associated with Increased Risk of Breast Cancer

Genetic variants in human microRNA (miRNA) genes may alter mature miRNA processing and/or target selection, and likely contribute to cancer susceptibility and disease progression. Previous studies have suggested that miR-101 may play important roles in the development of cancer by regulating key tumor-associated genes. However, the role of single nucleotide polymorphisms (SNPs) of miR-101 in breast cancer susceptibility remains unclear. In this study, we genotyped 11 SNPs of the miR-101 genes (including miR-101-1 and miR-101-2) in a case-control study of 1064 breast cancer cases and 1073 cancer-free controls. The results revealed that rs462480 and rs1053872 in the flank regions of pre-miR-101-2 were significantly associated with increased risk of breast cancer (rs462480 AC/CC vs AA: adjusted OR = 1.182, 95% CI: 1.030–1.357, P = 0.017; rs1053872 CG/GG vs CC: adjusted OR = 1.179, 95% CI: 1.040–1.337, P = 0.010). However, the remaining 9 SNPs were not significantly associated with risk of breast cancer. Additionally, combined analysis of the two high-risk SNPs revealed that subjects carrying the variant genotypes of rs462480 and rs1053872 had increased risk of breast cancer in a dose-response manner (P _trend = 0.002). Compared with individuals with “0–1” risk allele, those carrying “2–4” risk alleles had 1.29-fold risk of breast cancer. In conclusion, these findings suggested that the SNPs rs462480 and rs1053872 residing in miR-101-2 gene may have a solid impact on genetic susceptibility to breast cancer, which may improve our understanding of the potential contribution of miRNA SNPs to cancer pathogenesis.     

Identification of a Functional Genetic Variant at 16q12.1 for Breast Cancer Risk: Results from the Asia Breast Cancer Consortium

Genetic factors play an important role in the etiology of breast cancer. We carried out a multi-stage genome-wide association (GWA) study in over 28,000 cases and controls recruited from 12 studies conducted in Asian and European American women to identify genetic susceptibility loci for breast cancer. After analyzing 684,457 SNPs in 2,073 cases and 2,084 controls in Chinese women, we evaluated 53 SNPs for fast-track replication in an independent set of 4,425 cases and 1,915 controls of Chinese origin. Four replicated SNPs were further investigated in an independent set of 6,173 cases and 6,340 controls from seven other studies conducted in Asian women. SNP rs4784227 was consistently associated with breast cancer risk across all studies with adjusted odds ratios (95% confidence intervals) of 1.25 (1.20−1.31) per allele (P = 3.2×10⁻²⁵) in the pooled analysis of samples from all Asian samples. This SNP was also associated with breast cancer risk among European Americans (per allele OR  = 1.19, 95% CI  = 1.09−1.31, P = 1.3×10⁻⁴, 2,797 cases and 2,662 controls). SNP rs4784227 is located at 16q12.1, a region identified previously for breast cancer risk among Europeans. The association of this SNP with breast cancer risk remained highly statistically significant in Asians after adjusting for previously-reported SNPs in this region. In vitro experiments using both luciferase reporter and electrophoretic mobility shift assays demonstrated functional significance of this SNP. These results provide strong evidence implicating rs4784227 as a functional causal variant for breast cancer in the locus 16q12.1 and demonstrate the utility of conducting genetic association studies in populations with different genetic architectures.     

Differential effects of MYH9 and APOL1 risk variants on FRMD3 Association with Diabetic ESRD in African Americans

Single nucleotide polymorphisms (SNPs) in MYH9 and APOL1 on chromosome 22 (c22) are powerfully associated with non-diabetic end-stage renal disease (ESRD) in African Americans (AAs). Many AAs diagnosed with type 2 diabetic nephropathy (T2DN) have non-diabetic kidney disease, potentially masking detection of DN genes. Therefore, genome-wide association analyses were performed using the Affymetrix SNP Array 6.0 in 966 AA with T2DN and 1,032 non-diabetic, non-nephropathy (NDNN) controls, with and without adjustment for c22 nephropathy risk variants. No associations were seen between FRMD3 SNPs and T2DN before adjusting for c22 variants. However, logistic regression analysis revealed seven FRMD3 SNPs significantly interacting with MYH9—a finding replicated in 640 additional AA T2DN cases and 683 NDNN controls. Contrasting all 1,592 T2DN cases with all 1,671 NDNN controls, FRMD3 SNPs appeared to interact with the MYH9 E1 haplotype (e.g., rs942280 interaction p-value = 9.3E⁻⁷ additive; odds ratio [OR] 0.67). FRMD3 alleles were associated with increased risk of T2DN only in subjects lacking two MYH9 E1 risk haplotypes (rs942280 OR = 1.28), not in MYH9 E1 risk allele homozygotes (rs942280 OR = 0.80; homogeneity p-value = 4.3E⁻⁴). Effects were weaker stratifying on APOL1. FRMD3 SNPS were associated with T2DN, not type 2 diabetes per se, comparing AAs with T2DN to those with diabetes lacking nephropathy. T2DN-associated FRMD3 SNPs were detectable in AAs only after accounting for MYH9, with differential effects for APOL1. These analyses reveal a role for FRMD3 in AA T2DN susceptibility and accounting for c22 nephropathy risk variants can assist in detecting DN susceptibility genes.