Supplementarytable 1. Genes in the Mtor Pathway Examined in WCHS

SupplementaryTable 1. Genes in the mTOR pathway examined in WCHS

Gene (alias) / Function of encoded protein / Chromosome location / Number of tested SNPs
AKT1: RAC-Alpha Serine/Threonine-Protein Kinase / Regulates glucose uptake and storage; activate mTORC1 signaling by mediating phosphorylation of TSC1-TSC2 / 14q32.32-q32.33 / 13
AKT1S1: AKT1 Substrate 1 / Subunit of mTORC1; negatively regulates mTOR activity / 19q13.33 / 4
AKT2: RAC-Beta Serine/Threonine-Protein / One of the AKT kinase / 19q13.1-q13.2 / 9
CDKAL1: CDK5 Regulatory Subunit-Associated Protein 1-Like 1 / Gene mutation is associated with type II diabetes/obesity. / 6p22.3 / 1
CYB5R4: Cytochrome B5 Reductase 4 / Protects pancreatic beta-cells against oxidant stress; gene mutation is associated with diabetes/obesity. / 6q14.2 / 1
DEPTOR:DEP Domain Containing MTOR-Interacting Protein / Negatively regulate mTORC1 and mTORC2 signaling / 8q24.12 / 19
EIF4EBP1: Eukaryotic Translation Initiation Factor 4E Binding Protein 1 / Mediates the regulation of protein translation by hormones, growth factors; regulated by mTORC1 / 8q12 / 4
GIP:Gastric Inhibitory Polypeptide / Stimulates insulin secretion from pancreatic beta-cells following food ingestion and nutrient absorption. / 17q12.32 / 4
IRS1:Insulin Receptor Substrate 1 / Mediates various cellular processes by insulin; gene mutation is associated with type II diabetes. / 2q36.3 / 15
LAMTOR1: Late Endosomal/Lysosomal Adaptor, MAPK And MTOR Activator 1 / involved in amino acid sensing and activation of mTORC1 / 11q13.4 / 3
LAMTOR2:Late Endosomal/Lysosomal Adaptor, MAPK And MTOR Activator 1 / Part of the Ragulator complex composed of LAMTOR1,LAMTOR2, LAMTOR3, LAMTOR4, and LAMTOR5 / 1q22 / 1
LAMTOR3:Late Endosomal/Lysosomal Adaptor, MAPK And MTOR Activator 1 / Part of the Ragulator complex composed of LAMTOR1, LAMTOR2, LAMTOR3, LAMTOR4, and LAMTOR5 / 4q23 / 4
LRTOMT (COMT2): Leucine Rich Transmembrane And O-Methyltransferase Domain Containing / Catalyzes the O-methylation, inactivating catecholamine neurotransmitters and catechol hormones / 11q13.4 / 1
MAPKAP1: Mitogen-Activated Protein Kinase Associated Protein 1 / Subunit of mTORC2 / 9q33.3 / 19
FRAP1 (MTOR): FK506-binding protein 12-rapamycin-associated protein 1 / The gene encodes mTORC1 and mTORC2 (mTOR complex 1 and 2). / 1p36.2 / 15
PIK3CA: Phosphatidylinositol-4,5-Bisphosphate 3-Kinase, Catalytic Subunit Alpha / The gene encodes Phosphoinositide-3-kinase (PI3K), which activates AKT1. / 3q26.3 / 13
PIK3R1: Phosphoinositide-3-Kinase, Regulatory Subunit 1 (Alpha) / The gene encodesan 85 kD regulatory subunit of PI3K / 5q13.1 / 18
PRKCI: Protein Kinase C, iota / The gene encodes a member of the protein kinase C (PKC) family of serine/threonine protein kinases / 3q26.3 / 16
PRR5 (PROTOR1): Proline Rich 5 (Renal) / The gene encodes a protein with a proline-rich domain, a subunit of mTORC2 / 22q13.3 / 18
PRR5L (PROTOR2): Proline Rich 5 Like / Regulates the activity of the mTORC2 complex / 11p13 / 18
RHEB: Ras Homolog Enriched In Brain / Activates the protein kinase activity of mTORC1 / 7q36 / 15
RICTOR:RPTOR independent companion of MTOR, complex 2 / A subunit of mTORC2 / 5p13.1 / 12
RPS6KB1: ribosomal protein S6 kinase, 70kDa, polypeptide 1 / The gene encodes a protein in the ribosomal S6 kinase family of serine/threonine kinases that responds to mTOR signaling. / 17q23.1 / 6
RPTOR: Regulatory Associated Protein Of MTOR, Complex 1 / Involved in the control of the mTORC1 activity. / 17q25.3 / 116
SESN2: Sestrin 2 / A p53 target gene involving the signaling of AKT in response to energetic stress. / 1p35.3 / 6
SGK1: Serum/Glucocorticoid Regulated Kinase 1 / The gene encodes a serine/threonine protein kinase in the downstream of mTORC2 signaling. / 6q23.2 / 4
SREBF1: Sterol Regulatory Element Binding Transcription Factor 1 / Involved in lipid synthesis, which can be activated by mTORC1. / 17p11.2 / 14
TBC1D17: TBC1 Domain Family, Member 17 / The gene encodes a GTPase-activating protein involving in autophagy. / 19q13.33 / 2
TSC1: Tuberous Sclerosis 1 / Inhibits EIF4EBP1 by negatively regulating mTORC1 signaling. / 9q34 / 15
TSC2: Tuberous Sclerosis 2 / Negatively regulates mTORC1 signaling (in complex with TSC1) / 16p13.3 / 14
UBQLN4: Ubiquilin 4 / Recruits Ubiquilin 1 mediating autophagy. / 1q21 / 2

Supplementary Table 2. Association of mTOR pathway SNPs with breast cancer risk according to ER status in EA and AA women1

Gene / SNP / Genotype / European American / African American
ER+ case vs. controls / ER– case vs. controls / ER+ case vs. controls / ER– case vs. controls
OR (95% CI) / OR (95% CI) / P-heterogeneity2 / OR (95% CI) / OR (95% CI) / P-heterogeneity
FRAP1 / rs12125777 / GA/AA vs. GG / 1.84 (1.10-3.08) / 2.65 (1.21-5.77) / 0.30 / 1.06 (0.73-1.53) / 1.17 (0.72-1.91) / 0.72
RPTOR / rs9900506 / GG/GA vs. AA / 0.97 (0.72-1.29) / 0.64 (0.38-1.06) / 0.16 / 1.09 (0.83-1.43) / 0.92 (0.64-1.33) / 0.31
RPTOR / rs3817293 / AA/GA vs. GG / 0.97 (0.73-1.29) / 1.00 (0.59-1.69) / 0.99 / 0.76 (0.58-1.01) / 0.68 (0.47-0.99) / 0.45
PIK3R1 / rs7713645 / CC/AC vs. AA / 0.72 (0.54-0.97) / 0.95 (0.55-1.64) / 0.47 / 0.59 (0.32-1.09) / 2.14 (0.62-7.35) / 0.09
PRR5L / rs3740958 / GG/GA vs. AA / 1.98 (1.44-2.73) / 1.76 (1.00-3.10) / 0.91 / 0.85 (0.62-1.16) / 0.62 (0.40-0.96) / 0.37
CDKAL1 / rs9368197 / AA/AC vs. CC / 1.32 (1.01-1.74) / 0.75 (0.46-1.24) / 0.021 / 0.67 (0.45-0.99) / 0.56 (0.31-1.00) / 0.48

1 Adjusted for age at diagnosis, proportion of European ancestry, education, BMI, family history of breast cancer, history of benign breast disease, age at menarche, parity, breastfeeding status, menopausal status, and status of cigarette smoking

2Estimated from case-case only regression models comparing ER– and ER+ tumors

Supplementary Table 3. Effect modification of body size and weight gain on the association of FRAP1 rs12125777 genotype with breast cancer.1 BMI, WHR, and weight gain were not mutually adjusted.

European American / African American
GG / GA/AA / GG / GA/AA
# Case/ Control / OR (ref.) / # Case/ Control / OR (95% CI) / P-interaction / # Case/ Control / OR (ref.) / # Case/ Control / OR (95% CI) / P-interaction
Body mass index (kg/m2) / <25 / 270/260 / 1.00 / 23/24 / 0.83 (0.44-1.58) / 0.006 / 91/113 / 1.00 / 20/20 / 1.53 (0.72-3.23) / 0.28
25 - <30 / 145/167 / 1.00 / 17/8 / 2.63 (1.02-6.81) / 134/174 / 1.00 / 37/37 / 1.34 (0.78-2.28)
≥30 / 158/167 / 1.00 / 21/4 / 5.68 (1.78-18.1) / 264/323 / 1.00 / 42/57 / 0.88 (0.55-1.40)
Waist-to-hip ratio / ≤0.85 / 339/353 / 1.00 / 35/25 / 1.31 (0.74-2.32) / 0.16 / 207/260 / 1.00 / 42/53 / 1.00 (0.63-1.60) / 0.50
>0.85 / 234/241 / 1.00 / 26/11 / 2.82 (1.30-6.11) / 282/350 / 1.00 / 57/61 / 1.20 (0.79-1.80)
Weight gain from age 20 (lb.) / <102 / 134/140 / 1.00 / 13/13 / 0.98 (0.41-2.33) / 0.019 / 49/65 / 1.00 / 10/10 / 1.22 (0.41-3.60) / 0.36
10 - <25 / 96/87 / 1.00 / 9/8 / 0.96 (0.32-2.89) / 37/51 / 1.00 / 11/9 / 2.11 (0.55-8.10)
25 - <35 / 115/118 / 1.00 / 13/7 / 2.27 (0.80-6.44) / 66/109 / 1.00 / 15/21 / 1.26 (0.58-2.73)
≥35 / 228/249 / 1.00 / 26/8 / 3.29 (1.40-7.70) / 337/385 / 1.00 / 63/74 / 0.97 (0.66-1.43)

2 Including no change in weight or weight loss.

Supplementary Table 4. Effect modification of body size and weight gain on the association of FRAP1 rs12125777 genotype with breast cancer according to the estrogen receptor (ER) status among European-American women1, 3

ER+ / ER–
rs12125777GG / rs12125777GA/AA / rs12125777GG / rs12125777GA/AA
Energy balance variable / # Case/ Control / OR (ref.) / # Case/ Control / OR (95% CI) / P-interaction / # Case/ Control / OR (ref.) / # Case/ Control / OR (95% CI) / P-interaction
Body mass index (kg/m2) / <25 / 150/260 / 1.00 / 14/24 / 1.08 (0.51-2.27) / 0.039 / 35/260 / 1.00 / 3/24 / 0.86 (0.23-3.33) / 0.013
25 - <30 / 88/167 / 1.00 / 8/8 / 1.87 (0.62-5.64) / 16/167 / 1.00 / 4/8 / 5.59 (1.16-27.0)
≥30 / 104/167 / 1.00 / 12/4 / 9.05 (2.19-37.5) / 16/167 / 1.00 / 3/4 / 36.2 (3.33-393.3)
Waist-to-hip ratio / ≤0.85 / 198/353 / 1.00 / 22/25 / 1.63 (0.85-3.11) / 0.59 / 47/353 / 1.00 / 6/25 / 1.99 (0.73-5.45) / 0.20
>0.85 / 144/241 / 1.00 / 12/11 / 2.26 (0.92-5.60) / 20/241 / 1.00 / 4/11 / 4.03 (1.03-15.8)
Weight gain from age 20 (lb.) / <102 / 73/140 / 1.00 / 8/13 / 1.26 (0.45-3.54) / 0.08 / 18/140 / 1.00 / 1/13 / 0.80 (0.08-8.00) / 0.053
10 - <35 / 126/205 / 1.00 / 11/15 / 1.45 (0.59-3.54) / 25/205 / 1.00 / 5/15 / 2.91 (0.85-9.95)
≥35 / 144/249 / 1.00 / 15/8 / 3.38 (1.31-8.72) / 24/249 / 1.00 / 4/8 / 12.4 (2.17-71.1)

1 Adjusted for age at diagnosis, proportion of European ancestry, education, family history of breast cancer, history of benign breast disease, age at menarche, parity, breastfeeding status, menopausal status, and status of cigarette smoking.Models were further mutually adjusted for the body size and weight gain variables. For example, ORs for BMI were adjusted for WHR and weight gain from age 20.

2 Including no change in weight or weight loss.

3 P-interaction in case-case (ER– cancer as the outcome compared to ER+ cancer) models: 0.66 for BMI; 0.48 for waist-to-hip ratio; 0.50 for weight gain from age 20.

Supplementary Table5. Potential regulatory functionofSNPs. Data are fromHaploReg v3 (BroadInstitute),whichretrievesepigenomicdatafromtheEncyclopediaofDNAElements(ENCODE) Consortium

SNP (population)1 / Transcription factor3 / DNAse hypersensitivity cell type / Predicted motif change
FRAP1 rs12125777(EUR) / Active promoter / Mammary gland, adenocarcinoma; mammary fibroblasts / Mef2, Pou2f2, Zfp105
Tags2: / rs6678084
rs12145119 (5’UTR) / ERalpha-a_disc4
FRAP1 rs2536 (3’UTR) (EUR) / Mammary epithelial cells / HMG-IY,Ik-3
Tags: / rs743576 / Active promoter; transcription enhancer-like / Hsf
rs72870077 / Active promoter; active TSS
rs41274498 (3’-UTR); rs41274502 / Transcription enhancer-like
rs12136339; rs12133922 (synonymous); rs17552394; rs17552387 / Active promoter; active TSS / Mammary epithelial cells
rs2335276 / Strong enhancer; weak TSS; TSS: flanking more downstream
rs12137491; rs12118083 / Transcription enhancer-like
rs3737611 / – / Mammary ductal carcinoma
rs2275526; rs12141233; rs17235612; rs12122483 / Weak enhancer
rs3730381 / Strong enhancer; weak enhancer / Mammary fibroblasts
FRAP1 rs12116957 (EUR) / EBF, HDAC2, Hdx, Mef2, STAT
Tags: / rs12142442; rs12144023; rs12121277 / Strong enhancer
rs12116962; rs12124629; rs111320347;
rs201102319;
rs200433548;
rs2275942;
rs41274516;
rs4282823;
rs17036414;
rs72871462;
rs144062447;
rs78271920;
rs113604953;
rs112991345 / Weak enhancer
rs2275528; rs12130110 / Strong enhancer / Mammary epithelial cells
rs72871462 / Mammary fibroblasts
RPTOR rs9900506 (EUR) / Weak enhancer / RREB-1, Zbtb3, Zfp691
RPTOR rs3817293 (AFR) / Egr-1, Pax-5
PIK3R1 rs7713645 (AFR) / Weak enhancer / DMRT, Dbx1, Ncx_2, Sox, Zfp187
Tags: / rs706711 / Strong enhancer
rs6881033 / Transcription enhancer-like
rs7725370 / Transcription enhancer-like
PRR5L rs3740958 (EUR) / –
CDKAL1 rs9368197 (EUR) / GR,STAT,Zbtb12
Tags: / rs10806920 / Active promoter; weak TSS
rs965054 / Weak enhancer
rs58761454 / Weak enhancer
rs9368200 / Strong enhancer
rs2064321 / Strong enhancer
CDKAL1 rs9368197 (AFR) / GR, STAT, Zbtb12
Tags: / rs59621846 / Weak enhancer

TSS, transcription start site

1Associatedwithbreastcancerrisk,asshowninTable2

2LDr2≥0.8 in the populationwiththeleadSNP; only SNPs have marks in breast cell lines are shown.

3 Promotor or enhancer histone marks in mammary epithelial and myoepithelial cells