PFAS Levels, Early Life Factors, and Mammographic Breast Density in Premenopausal Women

Background: Mammographic breast density (MBD) is a strong risk factor and an intermediate phenotype for breast cancer, yet there are limited studies on how environmental pollutants are associated with MBD. Objective: We investigated associations of perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), and perfluorohexane sulfonate (PFHxS) levels with measures of MBD and evaluated if early life factors modified any associations. Methods: Metabolon performed metabolomics analysis using ultrahigh-performance liquid chromatography/tandem accurate mass spectrometry in fasting blood from 705 premenopausal women completing their annual screening mammogram in St. Louis, Missouri. We calculated least square means (LSM) of mammographic volumetric percent density (VPD), dense volume (DV), and nondense volume (NDV) by quartiles (Q) of PFOS, PFOA, and PFHxS from multivariable linear regression modeling overall and stratified by recruitment period, race, age at menarche, and body shape at age 10. Models were adjusted for age, age at menarche, body fat percentage, race, family history of breast cancer, oral contraceptive use, alcohol consumption, parity/age at first birth, and body shape at age 10. Results: PFOS, PFOA, and PFHxS were not significantly associated with VPD or NDV. PFHxS was significantly positively associated with DV (Q1=67.64 cm3, Q2=69.91 cm3, Q3=69.06 cm3, Q4=75.79 cm3; p-trend=0.03). PFOS was positively associated with DV (Q1=65.45 cm3, Q2=70.74 cm3, Q3=73.31 cm3, Q4=73.52 cm3; p-trend=0.06) with DV being 8.1%, 12%, and 12.3% higher in Q2, Q3, and Q4 compared to Q1. Among women who were underweight/normal weight at age 10, PFOS was positively associated with VPD (Q1=9.02%, Q2=9.11%, Q3=9.48%, Q4=9.92%; p-trend=0.04) while there was an inverse association among women who were overweight/obese at age 10 (Q1=7.46%, Q2=6.94%, Q3=6.78%, Q4=5.47%; p-trend=0.005) (p-interaction=0.04). Discussion: We report novel associations of PFHxS and PFOS with DV in premenopausal women. PFOS, PFOA, and PFHxS were not associated with VPD and NDV. In addition, body shape at age 10 may modify the associations of PFOS with MBD. Further studies are needed to validate our findings and to evaluate the associations of other per- and polyfluoroalkyl substances (PFAS), as well as mixtures of PFAS, with MBD. https://doi.org/10.1289/EHP14065


Table of Contents
Table S1.Measures of mammographic breast density in premenopausal women recruited during annual screening mammogram at Washington University School of Medicine, St. Louis, MO stratified by recruitment period (n = 705).Table S2.Peak area values of PFOS, PFOA, and PFHxS in 705 premenopausal women recruited during annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022.Table S3.Peak area values of PFOS, PFOA, and PFHxS in 705 premenopausal women recruited during annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022 stratified by body shape at age 10.Table S4.Peak area values of PFOS, PFOA, and PFHxS in 705 premenopausal women recruited during annual screening mammogram at Washington University School of Medicine, St. Louis, stratified by recruitment period.
Table S5.Spearman's Correlation Coefficients for PFOS, PFOA, and PFHXS.Table S6.PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density in 617 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022 with complete covariate data.
Table S7.PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density stratified by body shape at age 10 (underweight/normal weight vs. overweight/obese) in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022.Table S8.PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density stratified by age at menarche (≤13 years old vs >13 years old) in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022.Table S9.PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density stratified by race (non-Hispanic black vs. non-Hispanic white) in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022.Table S10.PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density stratified by recruitment period (December 2015 to October 2016 vs September 2020 to February 2022) in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO.

Figure S1 .
Figure S1.PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density stratified by age at menarche (≤13 years old vs >13 years old) in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022.

Figure S2 .
Figure S2.PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density stratified by race (non-Hispanic black vs. non-Hispanic white) in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022.

Figure S3 .
Figure S3.PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density stratified by recruitment period (December 2015 to October 2016 vs September 2020 to February 2022) in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO.

Figure S4 .
Figure S4.Unadjusted cubic spline regression of PFOS, PFOA, and PFHxS levels and mammographic breast density in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO Between December 2015 to October 2016 and September 2020 to February 2022.

Figure S5 .
Figure S5.PFOS, PFOA, and PFHxS levels and covariate-adjusted mammographic breast density in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO Between December 2015 to October 2016 and September 2020 to February 2022 estimated using univariate Bayesian kernel machine regression (estimates and 95% CI).
Figure S1: PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density stratified by age at menarche (≤13 years old vs >13 years old) in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022

Table S1 :
Measures of mammographic breast density in premenopausal women recruited during annual screening mammogram at Washington University School of Medicine, St. Louis, MO stratified by recruitment period (n = 705)

Table S2 :
Peak area values of PFOS, PFOA, and PFHxS in 705 premenopausal women recruited during annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022.

Table S3 :
Peak area values of PFOS, PFOA, and PFHxS in 705 premenopausal women recruited during annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022 stratified by body shape at age 10

Table S6 :
PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density in 617 premenopausal women who completed their annual screening mammogram at Washington University School ofMedicine, St. Louis, MO between December 2015to October 2016 and September 2020 to February 2022 with complete covariate data Note: Multivariable linear regression models were performed to calculate adjusted least square means of MBD (VPD, NDV, DV) by quartile of PFAS (PFOS, PFOA, PFHxS), and p for trend values are presented to assess significance across the quartiles.Measures of MBD were log 10 transformed and coefficients back-transformed (10^β).Participants with missing covariate data were excluded from this analysis.

Table S7 :
PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density stratified by body shape at age 10 (underweight/normal weight vs. overweight/obese) in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022 Multivariable linear regression models were performed to calculate adjusted least square means of MBD (VPD, NDV, DV) by quartile of PFAS (PFOS, PFOA, PFHxS) separately for women who reported an underweight/normal weight body shape at age 10 (1-4) and women who reported an overweight/obese body shape at age 10 (5-9).P for trend values are presented to assess significance across the quartiles within each group and p-interaction to see if the relationship between PFAS and MBD within each strata were different.Measures of MBD were log10 transformed and coefficients back-transformed (10^β). Note:

Table S8 :
PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density stratified by age at menarche (≤13 years old vs >13 years old) in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022 Multivariable linear regression models were performed to calculate adjusted least square means of MBD (VPD, NDV, DV) by quartile of PFAS (PFOS, PFOA, PFHxS) separately for women who reported age at menarche ≤ the median (13 years old) and women who reported age at menarche > the median (>13 years old).P for trend values are presented to assess significance across the quartiles within each group and p-interaction to see if the relationship between PFAS and MBD within each strata were different. Note:

Table S9 :
PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density stratified by race (non-Hispanic black vs. non-Hispanic white) in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St.

Table S10 :
PFOS, PFOA, and PFHxS levels and covariate-adjusted least square means of mammographic breast density stratified by recruitment period (December 2015 to October 2016 vs September 2020 to February 2022) in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St.
: Multivariable linear regression models were performed to calculate adjusted least square means of MBD (VPD, NDV, DV) by quartile of PFAS (PFOS, PFOA, PFHxS) separately for women who who were recruited between December 2015 to October 2016 and women who were recruited between September 2020 and February 2022.P for trend values are presented to assess significance across the quartiles within each group and p-interaction to see if the relationship between PFAS and MBD within each strata were different.Measures of MBD were log10 transformed and coefficients back-transformed (10^β). Note

Table S12 :
P-values of likelihood ratio test of covariate-adjusted restricted cubic spline regressions of PFOS, PFOA, and PFHxS and mammographic breast density in 700 premenopausal women who completed their annual screening mammogram at Washington University School of Medicine, St. Louis, MO between December 2015 to October 2016 and September 2020 to February 2022 Models were adjusted for age (continuous), age at menarche (continuous), body fat % (continuous), race (non-Hispanic white, non-Hispanic black, other), family history of breast cancer (yes, no