Genetic and Clinical Predictors for Breast Cancer Risk Assessment and Stratification Among Chinese Women

Background Most of the genetic variants identified from genome-wide association studies of breast cancer have not been validated in Asian women. No risk assessment model that incorporates both genetic and clinical predictors is currently available to predict breast cancer risk in this population. Me...

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Published inJNCI : Journal of the National Cancer Institute Vol. 102; no. 13; pp. 972 - 981
Main Authors Zheng, Wei, Wen, Wanqing, Gao, Yu-Tang, Shyr, Yu, Zheng, Ying, Long, Jirong, Li, Guoliang, Li, Chun, Gu, Kai, Cai, Qiuyin, Shu, Xiao-Ou, Lu, Wei
Format Journal Article
LanguageEnglish
Published Cary, NC Oxford University Press 07.07.2010
Oxford Publishing Limited (England)
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Online AccessGet full text
ISSN0027-8874
1460-2105
1460-2105
DOI10.1093/jnci/djq170

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Abstract Background Most of the genetic variants identified from genome-wide association studies of breast cancer have not been validated in Asian women. No risk assessment model that incorporates both genetic and clinical predictors is currently available to predict breast cancer risk in this population. Methods We analyzed 12 single-nucleotide polymorphisms (SNPs) identified in recent genome-wide association studies mostly of women of European ancestry as being associated with the risk of breast cancer in 3039 case patients and 3082 control subjects who participated in the Shanghai Breast Cancer Study. All participants were interviewed in person to obtain information regarding known and suspected risk factors for breast cancer. The c statistic, a measure of discrimination ability with a value ranging from 0.5 (random classification) to 1.0 (perfect classification), was estimated to evaluate the contribution of genetic and established clinical predictors of breast cancer to a newly established risk assessment model for Chinese women. Clinical predictors included in the model were age at menarche, age at first live birth, waist-to-hip ratio, family history of breast cancer, and a previous diagnosis of benign breast disease. The utility of the models in risk stratification was evaluated by estimating the proportion of breast cancer patients in the general population that could be accounted for above a given risk threshold as predicted by the models. All statistical tests were two-sided. Results Eight SNPs (rs2046210, rs1219648, rs3817198, rs8051542, rs3803662, rs889312, rs10941679, and rs13281615), each of which reflected a genetically independent locus, were found to be associated with the risk of breast cancer. A dose–response association was observed between the risk of breast cancer and the genetic risk score, which is an aggregate measure of the effect of these eight SNPs (odds ratio for women in the highest quintile of genetic risk score vs those in the lowest = 1.85, 95% confidence interval = 1.58 to 2.18, Ptrend = 2.5 × 10−15). The genetic risk score, the waist-to-hip ratio, and a previous diagnosis of benign breast disease were the top three predictors of the risk of breast cancer, each contributing statistically significantly (P < .001) to the full risk assessment model. The model, with a c statistic of 0.6295 after adjustment for overfitting, showed promise for stratifying women into different risk groups; women in the top 30% risk group accounted for nearly 50% of the breast cancers diagnosed in the general population. Conclusion A risk assessment model that includes both genetic markers and clinical predictors may be useful to classify Asian women into relevant risk groups for cost-efficient screening and other prevention programs.
AbstractList Most of the genetic variants identified from genome-wide association studies of breast cancer have not been validated in Asian women. No risk assessment model that incorporates both genetic and clinical predictors is currently available to predict breast cancer risk in this population. A risk assessment model that includes both genetic markers and clinical predictors may be useful to classify Asian women into relevant risk groups for cost-efficient screening and other prevention programs.
Most of the genetic variants identified from genome-wide association studies of breast cancer have not been validated in Asian women. No risk assessment model that incorporates both genetic and clinical predictors is currently available to predict breast cancer risk in this population. We analyzed 12 single-nucleotide polymorphisms (SNPs) identified in recent genome-wide association studies mostly of women of European ancestry as being associated with the risk of breast cancer in 3039 case patients and 3082 control subjects who participated in the Shanghai Breast Cancer Study. All participants were interviewed in person to obtain information regarding known and suspected risk factors for breast cancer. The c statistic, a measure of discrimination ability with a value ranging from 0.5 (random classification) to 1.0 (perfect classification), was estimated to evaluate the contribution of genetic and established clinical predictors of breast cancer to a newly established risk assessment model for Chinese women. Clinical predictors included in the model were age at menarche, age at first live birth, waist-to-hip ratio, family history of breast cancer, and a previous diagnosis of benign breast disease. The utility of the models in risk stratification was evaluated by estimating the proportion of breast cancer patients in the general population that could be accounted for above a given risk threshold as predicted by the models. All statistical tests were two-sided. Eight SNPs (rs2046210, rs1219648, rs3817198, rs8051542, rs3803662, rs889312, rs10941679, and rs13281615), each of which reflected a genetically independent locus, were found to be associated with the risk of breast cancer. A dose-response association was observed between the risk of breast cancer and the genetic risk score, which is an aggregate measure of the effect of these eight SNPs (odds ratio for women in the highest quintile of genetic risk score vs those in the lowest = 1.85, 95% confidence interval = 1.58 to 2.18, P(trend) = 2.5 x 10(-15)). The genetic risk score, the waist-to-hip ratio, and a previous diagnosis of benign breast disease were the top three predictors of the risk of breast cancer, each contributing statistically significantly (P < .001) to the full risk assessment model. The model, with a c statistic of 0.6295 after adjustment for overfitting, showed promise for stratifying women into different risk groups; women in the top 30% risk group accounted for nearly 50% of the breast cancers diagnosed in the general population. A risk assessment model that includes both genetic markers and clinical predictors may be useful to classify Asian women into relevant risk groups for cost-efficient screening and other prevention programs.
Most of the genetic variants identified from genome-wide association studies of breast cancer have not been validated in Asian women. No risk assessment model that incorporates both genetic and clinical predictors is currently available to predict breast cancer risk in this population.BACKGROUNDMost of the genetic variants identified from genome-wide association studies of breast cancer have not been validated in Asian women. No risk assessment model that incorporates both genetic and clinical predictors is currently available to predict breast cancer risk in this population.We analyzed 12 single-nucleotide polymorphisms (SNPs) identified in recent genome-wide association studies mostly of women of European ancestry as being associated with the risk of breast cancer in 3039 case patients and 3082 control subjects who participated in the Shanghai Breast Cancer Study. All participants were interviewed in person to obtain information regarding known and suspected risk factors for breast cancer. The c statistic, a measure of discrimination ability with a value ranging from 0.5 (random classification) to 1.0 (perfect classification), was estimated to evaluate the contribution of genetic and established clinical predictors of breast cancer to a newly established risk assessment model for Chinese women. Clinical predictors included in the model were age at menarche, age at first live birth, waist-to-hip ratio, family history of breast cancer, and a previous diagnosis of benign breast disease. The utility of the models in risk stratification was evaluated by estimating the proportion of breast cancer patients in the general population that could be accounted for above a given risk threshold as predicted by the models. All statistical tests were two-sided.METHODSWe analyzed 12 single-nucleotide polymorphisms (SNPs) identified in recent genome-wide association studies mostly of women of European ancestry as being associated with the risk of breast cancer in 3039 case patients and 3082 control subjects who participated in the Shanghai Breast Cancer Study. All participants were interviewed in person to obtain information regarding known and suspected risk factors for breast cancer. The c statistic, a measure of discrimination ability with a value ranging from 0.5 (random classification) to 1.0 (perfect classification), was estimated to evaluate the contribution of genetic and established clinical predictors of breast cancer to a newly established risk assessment model for Chinese women. Clinical predictors included in the model were age at menarche, age at first live birth, waist-to-hip ratio, family history of breast cancer, and a previous diagnosis of benign breast disease. The utility of the models in risk stratification was evaluated by estimating the proportion of breast cancer patients in the general population that could be accounted for above a given risk threshold as predicted by the models. All statistical tests were two-sided.Eight SNPs (rs2046210, rs1219648, rs3817198, rs8051542, rs3803662, rs889312, rs10941679, and rs13281615), each of which reflected a genetically independent locus, were found to be associated with the risk of breast cancer. A dose-response association was observed between the risk of breast cancer and the genetic risk score, which is an aggregate measure of the effect of these eight SNPs (odds ratio for women in the highest quintile of genetic risk score vs those in the lowest = 1.85, 95% confidence interval = 1.58 to 2.18, P(trend) = 2.5 x 10(-15)). The genetic risk score, the waist-to-hip ratio, and a previous diagnosis of benign breast disease were the top three predictors of the risk of breast cancer, each contributing statistically significantly (P < .001) to the full risk assessment model. The model, with a c statistic of 0.6295 after adjustment for overfitting, showed promise for stratifying women into different risk groups; women in the top 30% risk group accounted for nearly 50% of the breast cancers diagnosed in the general population.RESULTSEight SNPs (rs2046210, rs1219648, rs3817198, rs8051542, rs3803662, rs889312, rs10941679, and rs13281615), each of which reflected a genetically independent locus, were found to be associated with the risk of breast cancer. A dose-response association was observed between the risk of breast cancer and the genetic risk score, which is an aggregate measure of the effect of these eight SNPs (odds ratio for women in the highest quintile of genetic risk score vs those in the lowest = 1.85, 95% confidence interval = 1.58 to 2.18, P(trend) = 2.5 x 10(-15)). The genetic risk score, the waist-to-hip ratio, and a previous diagnosis of benign breast disease were the top three predictors of the risk of breast cancer, each contributing statistically significantly (P < .001) to the full risk assessment model. The model, with a c statistic of 0.6295 after adjustment for overfitting, showed promise for stratifying women into different risk groups; women in the top 30% risk group accounted for nearly 50% of the breast cancers diagnosed in the general population.A risk assessment model that includes both genetic markers and clinical predictors may be useful to classify Asian women into relevant risk groups for cost-efficient screening and other prevention programs.CONCLUSIONA risk assessment model that includes both genetic markers and clinical predictors may be useful to classify Asian women into relevant risk groups for cost-efficient screening and other prevention programs.
Background Most of the genetic variants identified from genome-wide association studies of breast cancer have not been validated in Asian women. No risk assessment model that incorporates both genetic and clinical predictors is currently available to predict breast cancer risk in this population. Methods We analyzed 12 single-nucleotide polymorphisms (SNPs) identified in recent genome-wide association studies mostly of women of European ancestry as being associated with the risk of breast cancer in 3039 case patients and 3082 control subjects who participated in the Shanghai Breast Cancer Study. All participants were interviewed in person to obtain information regarding known and suspected risk factors for breast cancer. The c statistic, a measure of discrimination ability with a value ranging from 0.5 (random classification) to 1.0 (perfect classification), was estimated to evaluate the contribution of genetic and established clinical predictors of breast cancer to a newly established risk assessment model for Chinese women. Clinical predictors included in the model were age at menarche, age at first live birth, waist-to-hip ratio, family history of breast cancer, and a previous diagnosis of benign breast disease. The utility of the models in risk stratification was evaluated by estimating the proportion of breast cancer patients in the general population that could be accounted for above a given risk threshold as predicted by the models. All statistical tests were two-sided. Results Eight SNPs (rs2046210, rs1219648, rs3817198, rs8051542, rs3803662, rs889312, rs10941679, and rs13281615), each of which reflected a genetically independent locus, were found to be associated with the risk of breast cancer. A dose–response association was observed between the risk of breast cancer and the genetic risk score, which is an aggregate measure of the effect of these eight SNPs (odds ratio for women in the highest quintile of genetic risk score vs those in the lowest = 1.85, 95% confidence interval = 1.58 to 2.18, Ptrend = 2.5 × 10−15). The genetic risk score, the waist-to-hip ratio, and a previous diagnosis of benign breast disease were the top three predictors of the risk of breast cancer, each contributing statistically significantly (P < .001) to the full risk assessment model. The model, with a c statistic of 0.6295 after adjustment for overfitting, showed promise for stratifying women into different risk groups; women in the top 30% risk group accounted for nearly 50% of the breast cancers diagnosed in the general population. Conclusion A risk assessment model that includes both genetic markers and clinical predictors may be useful to classify Asian women into relevant risk groups for cost-efficient screening and other prevention programs.
Background Most of the genetic variants identified from genome-wide association studies of breast cancer have not been validated in Asian women. No risk assessment model that incorporates both genetic and clinical predictors is currently available to predict breast cancer risk in this population. Methods We analyzed 12 single-nucleotide polymorphisms (SNPs) identified in recent genome-wide association studies mostly of women of European ancestry as being associated with the risk of breast cancer in 3039 case patients and 3082 control subjects who participated in the Shanghai Breast Cancer Study. All participants were interviewed in person to obtain information regarding known and suspected risk factors for breast cancer. The c statistic, a measure of discrimination ability with a value ranging from 0.5 (random classification) to 1.0 (perfect classification), was estimated to evaluate the contribution of genetic and established clinical predictors of breast cancer to a newly established risk assessment model for Chinese women. Clinical predictors included in the model were age at menarche, age at first live birth, waist-to-hip ratio, family history of breast cancer, and a previous diagnosis of benign breast disease. The utility of the models in risk stratification was evaluated by estimating the proportion of breast cancer patients in the general population that could be accounted for above a given risk threshold as predicted by the models. All statistical tests were two-sided. Results Eight SNPs (rs2046210, rs1219648, rs3817198, rs8051542, rs3803662, rs889312, rs10941679, and rs13281615), each of which reflected a genetically independent locus, were found to be associated with the risk of breast cancer. A dose-response association was observed between the risk of breast cancer and the genetic risk score, which is an aggregate measure of the effect of these eight SNPs (odds ratio for women in the highest quintile of genetic risk score vs those in the lowest = 1.85, 95% confidence interval = 1.58 to 2.18, P sub(trend) = 2.5 10 super(-15)). The genetic risk score, the waist-to-hip ratio, and a previous diagnosis of benign breast disease were the top three predictors of the risk of breast cancer, each contributing statistically significantly (P < .001) to the full risk assessment model. The model, with a c statistic of 0.6295 after adjustment for overfitting, showed promise for stratifying women into different risk groups; women in the top 30% risk group accounted for nearly 50% of the breast cancers diagnosed in the general population. Conclusion A risk assessment model that includes both genetic markers and clinical predictors may be useful to classify Asian women into relevant risk groups for cost-efficient screening and other prevention programs.
Author Shyr, Yu
Li, Chun
Cai, Qiuyin
Lu, Wei
Zheng, Wei
Shu, Xiao-Ou
Wen, Wanqing
Long, Jirong
Zheng, Ying
Li, Guoliang
Gu, Kai
Gao, Yu-Tang
AuthorAffiliation Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
AuthorAffiliation_xml – name: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
Author_xml – sequence: 1
  givenname: Wei
  surname: Zheng
  fullname: Zheng, Wei
  email: wei.zheng@vanderbilt.edu
  organization: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
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  givenname: Wanqing
  surname: Wen
  fullname: Wen, Wanqing
  organization: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
– sequence: 3
  givenname: Yu-Tang
  surname: Gao
  fullname: Gao, Yu-Tang
  organization: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
– sequence: 4
  givenname: Yu
  surname: Shyr
  fullname: Shyr, Yu
  organization: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
– sequence: 5
  givenname: Ying
  surname: Zheng
  fullname: Zheng, Ying
  email: wei.zheng@vanderbilt.edu
  organization: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
– sequence: 6
  givenname: Jirong
  surname: Long
  fullname: Long, Jirong
  organization: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
– sequence: 7
  givenname: Guoliang
  surname: Li
  fullname: Li, Guoliang
  organization: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
– sequence: 8
  givenname: Chun
  surname: Li
  fullname: Li, Chun
  organization: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
– sequence: 9
  givenname: Kai
  surname: Gu
  fullname: Gu, Kai
  organization: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
– sequence: 10
  givenname: Qiuyin
  surname: Cai
  fullname: Cai, Qiuyin
  organization: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
– sequence: 11
  givenname: Xiao-Ou
  surname: Shu
  fullname: Shu, Xiao-Ou
  organization: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
– sequence: 12
  givenname: Wei
  surname: Lu
  fullname: Lu, Wei
  organization: Affiliations of authors: Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Vanderbilt-Ingram Cancer Center (WZ, WW, JL, GL, QC, X-OS) and Department of Biostatistics (YS, CL), Vanderbilt University School of Medicine, Nashville, TN; Department of Epidemiology, Shanghai Cancer Institute, Shanghai, China (Y-TG); Shanghai Center for Disease Prevention and Control, Shanghai Institute of Preventive Medicine, Shanghai, China (YZ, KG, WL)
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IsDoiOpenAccess true
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Issue 13
Keywords Human
Breast disease
Asiatic
Genotype
Breast cancer
Malignant tumor
Epidemiology
Mammary gland diseases
Cancerology
Statistical model
Risk factor
Genetics
Adult
Chinese
Female
Single nucleotide polymorphism
Public health
Cancer
Language English
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Snippet Background Most of the genetic variants identified from genome-wide association studies of breast cancer have not been validated in Asian women. No risk...
Most of the genetic variants identified from genome-wide association studies of breast cancer have not been validated in Asian women. No risk assessment model...
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StartPage 972
SubjectTerms Adult
Age of Onset
Asian Continental Ancestry Group - genetics
Asian Continental Ancestry Group - statistics & numerical data
Asian people
Biological and medical sciences
Breast cancer
Breast Neoplasms - epidemiology
Breast Neoplasms - etiology
Breast Neoplasms - genetics
Case-Control Studies
China - epidemiology
European Continental Ancestry Group - genetics
Female
Genetic diversity
Genetic Predisposition to Disease
Genome-Wide Association Study
Genomics
Genotype
Gynecology. Andrology. Obstetrics
Health risk assessment
Humans
Live Birth
Mammary gland diseases
Medical sciences
Menarche
Middle Aged
Models, Statistical
Polymorphism, Single Nucleotide
Predictive Value of Tests
Risk Assessment
Risk Factors
Tumors
Waist-Hip Ratio
Women
Title Genetic and Clinical Predictors for Breast Cancer Risk Assessment and Stratification Among Chinese Women
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https://pubmed.ncbi.nlm.nih.gov/PMC2897876
https://www.ncbi.nlm.nih.gov/pmc/articles/2897876
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