Association Of Obesity And Single Nucleotide Polymorphisms In Fatty Acid Binding Protein 4 With Breast Cancer

BackgroudBreast cancer is the most common cancer among women worldwidely.As is shown in Globocan reports,an estimated 187 thousand new breast cancer cases were diagnosed in China in 2012,accounting for nearly 12%of the whole world new breast cancer cases.Moreover,there has been an ever-increasing incidence of breast cancer in the past two decades,at rate of 3%-4%per year,nearly twice the global rate.In consideration of the huge population of 1.34 billion,China is facing increasing enormous social and economic burden of breast cancer.Understanding the risk factors,especially manageable factors,is supposed to be the most fundamental and effective ways to suppress the increasing trend.Extensive epidemiological evidence supports a close association between breast cancer and obesity.Obesity was supposed to be present in up to 50%of all breast cancer cases in older women,and it was estimated that the incidence of breast cancer could be reduced by half in the European Union.However,there are still some controversies.Firstly,the association between obesity and breast cancer may be different across races.As is shown in a meta-analysis,obesity was negatively associated with premenopausal breast cancer risk among white and black women,but positively among Asian women.However,our previous study showed no correlation between body mass index(BMI)and premenopausal breast cancer risk.Secondly,the association of central obesity and general obesity with breast cancer may be different.Previously,BMI was more commonly employed as an indicator of obesity,however waist-hip-ratio(WHR)or waist circumference have gained more focus since 2007.Though the results were far from agreement and little was known among Chinese females.Thirdly,the association between obesity and breast cancer subtypes defined by estrogen receptor(ER)status may be different.It was indicated that obesity is merely asscociated with ER-positive breast cancer,but no asscociation with ER-negative breast cancer.And lastly,the potential mechanism or interactions between obesity and breast cancer is unclear.Besides classic estrogen related pathway,inflammation,adopokine,insulin resistance etc were also supposed to be involved in the interaction.Elevated in breast cancer patients,fatty acid binding protein 4(FABP4),a kind of proinflammatory cytokines,was believed to be related to breast cancer.The association between single nucleotide polymorphism(SNP)of FABP4 gene(also known as FABP4)and breast cancer is,yet,unknown.PurposeBased on the above background and a multi-center case control study,three parts of investigations were conducted:1.To clarify the association between obesity and breast cancer risk.2.To clarify the association between obesity types(centrol obesity or general obesity)and risk of breast cancer subtypes defined by ER status.3.To clarify the association between SNPs of FABP4 and breast cancer risk.Methods1.SubjectsFrom June 2012 to June 2013,inpatients with breast cancer were recruited from 21 hospitals in 11 provinces or municipalities(Shandong Province,Hebei Province,Jiangsu Province,Henan Province,Shanxi Province,Liaoning Province,Jilin Province,Heilongjiang Province,Anhui Province,Tianjin municipality,and Beijing municipality)in Northern and Eastern China.Han Chinese females aged 25-70 years,who were newly histopathologically confirmed as primary breast cancer were included in the study.And controls were randomly selected from healthy outpatients who visited the centers for regular physical examinations.Controls were matched I:1 with cases from the same hospital based on the following criteria:same age(±3 years);and similar visiting period(±2 months).All controls were confirmed as having no diagnosis of breast cancer or other malignancies.2.Data collectionAll participants completed face-to-face interviews based on a self-designed structured questionnaire and clinical breast examination,including both physical and imaging examination.For breast cancer cases,medical and pathology records were documented and reviewed to obtain information on ER and PR statuses and other pathological results.Fasting blood samples for SNPs genotype tests were also collected for all participants.?The questionnaire covered the following factors:demographic characteristics,female physiological and reproductive factors,medical and family history,dietary habits,lifestyle habits,and breast cancer-related knowledge.?Both physical and imaging examination,were also documented in the form of tables,including height,weight,waist circumference,hip circumference,blood pressure,fasting glucose,breast ultrasonograpy,mammography,pathology test and etc.The Both ER and PR status were primarily determined immunohistochemically,and following ASCO/CAP(2010)recommendations,ER-positivity was defined as>1%of tumor cells with positive staining.?Genotype tests of SNPs located in FABP4.Based on the Hapmap database(Han Chinese in Bejing,China;CHB)and literature review,4 SNPs loci located in FABP4(rs1054135?rs2290201?rs16909192 and rs77878271)were genotyped with Sequenom MassARRAY(?).3.Statistical analysisThe database was established using EpiData 3.1 and was subsequently converted into SPSS.21.0 format.All statistical tests were based on two-sided probabilities with p<0.05 considered significant.The frequencies and percentages of risk variables at baseline were calculated.Analysis of variance and Pearson's ?2 tests were used to compare differences in frequency distributions between case and control groups and across case subgroups.Univariate and multivariate logistic regression analysis was used to calculate odds ratios(OR)and corresponding 95%confidence intervals(CI)for exposures variables in relation to breast cancer.Breast cancer cases were grouped into four categories according to the joint ER and PR statuses:ER+/PR+,ER+/PR-,ER-/PR-,and ER-/PR+.Multiple polytomous unconditional logistic regression analysis was used to calculate OR and corresponding 95%Cl for body-size indicators(height,weight,waist circumference,hip circumference,WHR,BMI,etc)in relation to breast cancer case subtype(ER+/PR+,ER+/PR-and ER-/PR-);ER-/PR+ cases were excluded from the final OR analysis because of the limited number of cases(n=21).Tests for trends were carried out by fitting ordinal values.Pearson's ?2 tests were used to compare differences in frequency distributions of SNPs genotypes between case and control groups and across subgroups.Logistic regression analysis,based on dominant model and additive model respectively,was used to calculate OR and corresponding 95%CI for genotypes in relation to breast cancer.ResultsIn total,1613 pairs of breast cancer cases and healthy controls were recruited.1489 breast cancer cases and 1489 matched controls were included for final analysis,among whom 1,316 breast cases were with clear joint ER/PR statuses.SNPs of FABP4 were successfully genotyped for 1016 breast cancer cases and 1028 controls.The main findings of the studies were as following.Part 1.Association of manageable risk factors with breast caner1.Factors related to breast cancer risk on univariate analysisSignificant differences between the case and control groups were observed in the following factors:residence,classroom period,education degree,family income,economic status,social status,menopausal status,body mass index,waist hip ratio,times of giving birth,times of misbirth,history of hypertension,soy products,red meat,dairy products,vegetable and fruit,quality of sleep,insomnia,early awake,sleep late,smoking,physical activity,family history of breast cancer,family history amongst secondary degree of relatives,healthy behavior and prevention,awareness of breast cancer,current quality of life satisfaction.2.Factors related to breast cancer risk on multivariate analysisCox-regression models were carried out to analyze risk factors for breast cancer for variables with p values less than 0.05 on Pearson's chi-square tests and univariate analysis.Ten factors were significantly related to breast cancer,for which the ORs and 95%CIs were:Residence,1.269(0.984-1.638,p=0.067);economic status,1.237(1.019-1.501,p=0.032),family history of breast cancer,2.418(1.361-4.294,p=0.003);Waist Hip Ratio,1.329(0.983-1.797,p=0.065);menopausal status,1.982(1.360-2.888,p=0.000);quality of sleep,1.412(1.140-1.749,p=0.002);current quality of life satisfaction,1.852(1.436-2.390,p=0.000);dairy products,0.813(0.716-0.923,p=0.001);awareness,0.685(0.517-0.907,p=0.008),healthy behavior and prevention,0.675(0.520-0.876,p=0.003).Compared to the previous population-based case-control study,consistent association with family history of breast cancer and current quality of life satisfaction were respectively observed,while contrary association with healthy behavior and prevention were revealed;obesity was consistently related to breast cancer,though different indicators(BMI or WHR)were used in the two case control studies.3.Factors related to breast cancer risk by menopausal status on multivariate analysisFour factors,including dairy products,family history of breast cancer,current quality of life satisfaction and healthy behavior and prevention,were correlated with both premenopausal and postmenopausal breast cancer risk.Social status,times of misbirth and BMI were merely related with premenopausal breast cancer risk,while WHR,age at menarche and quality of sleep related with post menopausal breast cancer risk.Part 2.Association of body size-related factors with risk of breast cancer according to joint estrogen and progestogen receptor status1.Baseline characteristicsAmong the 1,316 cases with clear joint ER/PR statuses,885(67.2%)were ER+/PR+,112(8.5%)were ER+/PR-,298(22.6%)were ER-/PR-,and 21(1.6%)were ER-/PR+.Compared with the controls,more breast cancer cases were from urban areas(75.8%vs 62.1%),were postmenopausal(34.1%vs 29.0%),and had an earlier age at menarche(14.00 vs 14.59 years).Regarding body-size related variables,cases tended to be lighter(61.50 kg vs 61.64 kg),but have a higher BMI(24.03 kg/m2 vs 23.99 kg/m2),a larger WC(80.00 cm vs 78.43 cm)and a higher WHR(0.85 vs 0.84).2.Associations between body size-related factors and breast cancer according to joint ER/PR statusCompared with women with a body weight<62.0 kg,women weighing>62.0 kg had a 21%increased risk(OR=1.21,95%CI:1.02,1.45)of ER+/PR+ breast cancer and a 34%increased risk(OR=1.34,95%CI:1.03,1.73)of ER-/PR-breast cancer.BMI was positively associated with both ER+/PR+ and ER-/PR-breast cancer,although trend tests were only significant for ER+/PR+ subtype(p trend for ER-/PR-subtype=0.093).WHR was only positively correlated with ER-/PR-subtype,with a 64%increase(OR=1.64,95%CI:1.23,2.18)in risk in women with WHR>0.85.WC was positively associated with risks of ER+/PR+(p trend=0.000)and ER-/PR-(p trend=0.004)breast cancer.While,HC was only positively associated with ER+/PR+subtype(p trend=0.027).3.Associations between body size-related factors and breast cancer according to joint ER/PR status in relation to menopausal statusAmongst premenopausal women,body weight,BMI,and waist circumference were positively associated with ER+/PR+ and ER-/PR-subtypes.Compared with women with a BMI<24.0 kg/m2,there was a 121%increase(OR=2.21,95%CI:1.52,3.21)in risk of ER+/PR+ breast cancer and a 105%increase(OR=2.21,95%CI:1.18,3.56)in risk of ER-/PR-breast cancer among women with a BMI>28.0 kg/m2.Amongst postmenopausal women,WHR>0.85 was associated with a 125%increased risk of ER-/PR-breast cancer(OR=2.25,95%CI:1.34,3.80).4.Associations between body size-related factors and breast cancer according to joint ER/PR status after adjusting for BMIThere were no differences in risks associated with WC,HC,or WHR across the three analyzed subgroups(ER+/PR+?ER+/PR-and ER-/PR-)regardless of BMI.However,higher WHR was associated with an increased risk of ER-/PR-breast cancer,independent of BMI.Part 3.Associations of SNP genotypes in FABP4 with breast cancer risk1.Genotyping results of case and control groupsThe call rates of rs77878271,rs 16909192 and rs2290201 were 99.7%(case group 99.8%and control group 99.6%),99.2%(case group 98.2%and control group 99.5%),90.3%(case group 90.7%and control group 89.9%)respectively.While the call rate of rs1054135 was lower,at 43.4%in case group and 44.8%in control group respectively.Two genotypes(AA/AC)were indentified as to rs16909192,and the C-Allele frequency was substantially higher in control group(p=0.007).Three genotypes were indentified,respectively,as to rs1054135(CC/CT/TT)and rs2290201(AA/AG/GG),and no difference between the two groups was observed.And as to rs77878271,only AA genotype was identified.2.Associations between genotypes and breast cancer riskrs16909192 AC genotype was correlated to a 76%decrease(OR=0.24,95%CI:0.08-0.70)in overall breast cancer risk,and a 85%decrease(OR=0.15,95%CI:0.04-0.64)in ER positive breast cancer,but no correlation with ER negative breast cancer was observed.Neither rs1054135 nor rs2290201 genotypes were statistically associated with overall breast cancer,ER positive breast cancer and ER negative breast cancer risks.3.Associations between genotypes and breast cancer risk adjusting for BMI and/or WHRrs16909192 AC genotype was correlated to a 80%decrease(OR=0.20,95%CI:0.04-0.89)in overall breast cancer risk among nonobese women(BMI?24.0kg/m2),and a non-statistically significant lower breast cancer risk(OR=0.32,95%CI:0.10-1.02)among women with WHR>0.85 as well.However,for nonobese women with higher WHR(BMI?24.0kg/m2&WHR>0.85),rs 16909192 AC genotype was correlated to a 88%decrease(OR=0.12,95%C1:0.02-0.96)in overall breast cancer risk.4.Associations between genotypes and risks of breast cancer subtypes adjusting for BMI and/or WHRAmong women with WHR>0.85,rs 16909192 AC genotype was associated with an 88%decrease(OR=0.12,95%CI:0.12-0.95)in risk of ER positive breast cancer.While rs2290201 mutant genotype(GG+AG)was associated with a 29%decrease(OR=0.71,95%CI:0.50-1.00)in ER positive breast cancer risk among women with WHR>0.85,and 25%decrease(OR=0.755 95%CI:0.55-1.00)among women with BMI>24.0kg/m2,respectively.rs2290201 AG genotype was correlated to a 41%decrease(OR=0.59,95%CI:0.36-0.95)in ER positive breast cancer risk.However,no correlation between rs 1054135,rs2290201 and rs16909192 genotypes and ER negative breast cance risks were observed in all subgroups.Conclusion1.Obesity and quality of life satisfaction are constantly associated with female breast cancer risk,though BMI and WHR may contribute differently to breast cancer risk.2.General obesity,indicated by BMI,and central obesity,indicated by WHR,contributed differently to risk of breast cancer according to ER and PR status.BMI is more strongly associated with ER+/PR+ subtype,while WHR is more specific for ER-/PR-subtype,independent of BMI.3.rs16909192,a SNP in FABP4,is a potential protective loci for ER positive breast cancer,especially for women with normal BMI but higher WHR(BMI?24.0kg/m2&WHR>0.85).Orignality1.We clarified that the association between obesity and breast cancer subtypes defined by estrogen receptor status were different among Chinese women.2.We indentified a potential protective SNP loci associated with breast cancer susceptibility.3.The results indicated the complicated underlying interaction between obesity and breast cancer,and concerns towards central obesity should be raised.