Terminal Duct Lobular Unit Involution Research Articles

Abstract C143: Molecular drivers of terminal duct lobular unit involution: Associations with gene expression, genetic ancestry, and breast cancer disparties

Abstract Terminal duct lobular units (TDLU) are epithelial sub-structures located within breast tissue that function in lactation during child-bearing age. TDLU involution is a mechanism whereby shrinking of these units occurs once child-bearing age has been surpassed, inducing a decrease in acini sub-structures and TDLU counts in healthy breast tissues. Decreased TDLU involution has one of the largest associations with breast cancer risk, and is linked to factors such as breastfeeding, parity, hormone levels, mammographic density, age, BMI, menopausal status, and SNP risk scores. Decreased TDLU involution is seen more commonly in women of African ancestry and has been implicated to increase the risk of breast cancer development in this population. The molecular and environmental factors that influence TDLU involution have not been fully characterized. Environmentally, previous studies have shown that increased air pollution exposure with PM2.5 components is related to higher TDLU counts and lower involution. Molecularly, our previous work has shown that high TDLU counts are associated with a more pro-inflammatory immune environment and may indicate that lower TDLU involution is a result of inflammation in the breast due to persistence of at-risk epithelial tissue. TDLU involution is measured by TDLU counts, acini counts per TDLU, and TDLU diameter. Bulk RNAseq data from voluntary research biopsies of normal breast tissue donated by a cohort of 94 women (47 African American, 47 European American) was analyzed for gene expression associations with TDLU counts and self-reported race. Analysis revealed thirteen cancer- and/or immune-related genes with significant changes in expression associated with high TDLU counts indicating lower TDLU involution. For two out of three genes most relevant to mammary biology, the relationship between gene expression and increasing TDLU count was more significant among African American women than European American women. Additionally, CIBERSORTx RNAseq analysis estimated the abundance of immune cell sub- populations, and revealed significant differences between women with high and low TDLU counts. There are also differences observed in gene expression across race for women with high TDLU count that may indicate a link between lower TDLU involution and race. These initial findings provide greater insight into the molecular mechanisms that drive lower TDLU involution overall and for certain populations. Future analyses will further link these molecular factors to environmental exposures, such as air pollution, to provide context to risk factors associated with decreased TDLU involution and increased breast cancer risk. Citation Format: Jeri D. Hughes, Angel Pajimola, Brittney Davis-Lynn, Renata Cora, Alexandra Harris, Gretchen Gierach, Brittney Jenkins-Lord. Molecular drivers of terminal duct lobular unit involution: Associations with gene expression, genetic ancestry, and breast cancer disparties [abstract]. In: Proceedings of the 17th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2024 Sep 21-24; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2024;33(9 Suppl):Abstract nr C143.

Associations between quantitative measures of mammographic density and terminal ductal lobular unit involution in Chinese breast cancer patients

BackgroundHigher mammographic density (MD), a radiological measure of the proportion of fibroglandular tissue in the breast, and lower terminal duct lobular unit (TDLU) involution, a histological measure of the amount of epithelial tissue in the breast, are independent breast cancer risk factors. Previous studies among predominantly white women have associated reduced TDLU involution with higher MD.MethodsIn this cohort of 611 invasive breast cancer patients (ages 23–91 years [58.4% ≥ 50 years]) from China, where breast cancer incidence rates are lower and the prevalence of dense breasts is higher compared with Western countries, we examined the associations between TDLU involution assessed in tumor-adjacent normal breast tissue and quantitative MD assessed in the contralateral breast obtained from the VolparaDensity software. Associations were estimated using generalized linear models with MD measures as the outcome variables (log-transformed), TDLU measures as explanatory variables (categorized into quartiles or tertiles), and adjusted for age, body mass index, parity, age at menarche and breast cancer subtype.ResultsWe found that, among all women, percent dense volume (PDV) was positively associated with TDLU count (highest tertile vs. zero: Expbeta = 1.28, 95% confidence interval [CI] 1.08–1.51, ptrend = < .0001), TDLU span (highest vs. lowest tertile: Expbeta = 1.23, 95% CI 1.11–1.37, ptrend = < .0001) and acini count/TDLU (highest vs. lowest tertile: Expbeta = 1.22, 95% CI 1.09–1.37, ptrend = 0.0005), while non-dense volume (NDV) was inversely associated with these measures. Similar trend was observed for absolute dense volume (ADV) after the adjustment of total breast volume, although the associations for ADV were in general weaker than those for PDV. The MD-TDLU associations were generally more pronounced among breast cancer patients ≥ 50 years and those with luminal A tumors compared with patients < 50 years and with luminal B tumors.ConclusionsOur findings based on quantitative MD and TDLU involution measures among Chinese breast cancer patients are largely consistent with those reported in Western populations and may provide additional insights into the complexity of the relationship, which varies by age, and possibly breast cancer subtype.

Association of Genetic Ancestry With Terminal Duct Lobular Unit Involution Among Healthy Women.

Reduced age-related terminal duct lobular unit (TDLU) involution has been linked to increased breast cancer risk and triple-negative breast cancer. Associations of TDLU involution levels with race and ethnicity remain incompletely explored. Herein, we examined the association between genetic ancestry and TDLU involution in normal breast tissue donated by 2014 healthy women in the United States. Women of African ancestry were more likely than European women to have increased TDLU counts (odds ratio [OR]trend = 1.36, 95% confidence interval [CI] = 1.07 to 1.74), acini counts per TDLU (OR = 1.47, 95% CI = 1.06 to 2.03), and median TDLU span (ORtrend = 1.44, 95% CI = 1.08 to 1.91), indicating lower involution, whereas East Asian descendants were associated with decreased TDLU counts (ORtrend = 0.52, 95% CI = 0.35 to 0.78) after controlling for potential confounders. These associations are consistent with the racial variations in incidence rates of triple-negative breast cancer in the United States and suggest opportunities for future work examining whether TDLU involution may mediate the racial differences in subtype-specific breast cancer risk.

Automated Quantitative Measures of Terminal Duct Lobular Unit Involution and Breast Cancer Risk.

Manual qualitative and quantitative measures of terminal duct lobular unit (TDLU) involution were previously reported to be inversely associated with breast cancer risk. We developed and applied a deep learning method to yield quantitative measures of TDLU involution in normal breast tissue. We assessed the associations of these automated measures with breast cancer risk factors and risk. We obtained eight quantitative measures from whole slide images from a benign breast disease (BBD) nested case-control study within the Nurses' Health Studies (287 breast cancer cases and 1,083 controls). Qualitative assessments of TDLU involution were available for 177 cases and 857 controls. The associations between risk factors and quantitative measures among controls were assessed using analysis of covariance adjusting for age. The relationship between each measure and risk was evaluated using unconditional logistic regression, adjusting for the matching factors, BBD subtypes, parity, and menopausal status. Qualitative measures and breast cancer risk were evaluated accounting for matching factors and BBD subtypes. Menopausal status and parity were significantly associated with all eight measures; select TDLU measures were associated with BBD histologic subtype, body mass index, and birth index (P < 0.05). No measure was correlated with body size at ages 5-10 years, age at menarche, age at first birth, or breastfeeding history (P > 0.05). Neither quantitative nor qualitative measures were associated with breast cancer risk. Among Nurses' Health Studies women diagnosed with BBD, TDLU involution is not a biomarker of subsequent breast cancer. TDLU involution may not impact breast cancer risk as previously thought.

Outdoor air pollution and terminal duct lobular involution of the normal breast

BackgroundExposure to certain outdoor air pollutants may be associated with a higher risk of breast cancer, though potential underlying mechanisms are poorly understood. We examined whether outdoor air pollution was associated with involution of terminal duct lobular units (TDLUs), the histologic site where most cancers arise and an intermediate marker of breast cancer risk.MethodsPathologist-enumerated TDLUs were assessed in H&E (hematoxylin and eosin)-stained breast tissue sections from 1904 US women ages 18–75 who donated to the Susan G. Komen Tissue Bank (2009–2012). The 2009 annual fine particulate matter < 2.5 μm in diameter (PM2.5) total mass (μg/m3) at each woman’s residential address was estimated from the Environmental Protection Agency’s Downscaler Model combining Community Multiscale Air Quality (CMAQ) System modeling with air quality monitoring data. We secondarily considered CMAQ-modeled components of PM2.5 and gaseous pollutants. We used K-means clustering to identify groups of individuals with similar levels of PM2.5 components, selecting groups via cluster stability analysis. Relative rates (RRs) and 95% confidence intervals (95% CIs) for the association between air pollutants and TDLU counts were estimated from a zero-inflated negative binomial regression model adjusted for potential confounders.ResultsPM2.5 total mass was associated with higher TDLU counts among all women (interquartile range (IQR) increase, RR = 1.06; 95% CI: 1.01–1.11). This association was evident among both premenopausal and postmenopausal women (premenopausal RR = 1.05, 95% CI: 1.00–1.11; postmenopausal RR = 1.11, 95% CI: 1.00–1.23). We identified 3 groups corresponding to clusters that varied geographically and roughly represented high, medium, and low levels of PM2.5 components relative to population mean levels. Compared to the cluster with low levels, the clusters with both high (RR = 1.74; 95% CI: 1.08–2.80) and medium (RR = 1.82; 95% CI: 1.13–2.93) levels were associated with higher TDLU counts; although not significantly different, the magnitude of the associations was stronger among postmenopausal women.ConclusionsHigher PM2.5 levels were associated with reduced TDLU involution as measured by TDLU counts. Air pollution exposure may influence the histologic characteristics of normal tissue which could in turn affect breast cancer risk.

Polygenic risk score for the prediction of breast cancer is related to lesser terminal duct lobular unit involution of the breast

Terminal duct lobular units (TDLUs) are the predominant anatomical structures where breast cancers originate. Having lesser degrees of age-related TDLU involution, measured as higher TDLUs counts or more epithelial TDLU substructures (acini), is related to increased breast cancer risk among women with benign breast disease (BBD). We evaluated whether a recently developed polygenic risk score (PRS) based on 313-common variants for breast cancer prediction is related to TDLU involution in the background, normal breast tissue, as this could provide mechanistic clues on the genetic predisposition to breast cancer. Among 1398 women without breast cancer, higher values of the PRS were significantly associated with higher TDLU counts (P = 0.004), but not with acini counts (P = 0.808), in histologically normal tissue samples from donors and diagnostic BBD biopsies. Mediation analysis indicated that TDLU counts may explain a modest proportion (≤10%) of the association of the 313-variant PRS with breast cancer risk. These findings suggest that TDLU involution might be an intermediate step in the association between common genetic variation and breast cancer risk.

Abstract 4632: Automated quantitative measures of terminal duct lobular unit involution and breast cancer risk

Abstract Background Terminal duct lobular units (TDLUs) involute with age. Prior studies using qualitative or semi-quantitative measures of TDLU involution found inverse associations with breast cancer (BC) risk. We applied our validated deep learning computational method to yield quantitative measures of TDLU involution in normal breast tissue and then assessed their association with established BC risk factors and BC risk. Materials and Methods This study used a nested case-control design within the Nurses' Health Study cohorts. Cases and controls were diagnosed with benign breast disease (BBD) and cases subsequently developed BC (median 7.75 years later). Cases and controls were matched on year of BBD diagnosis, age at BC diagnosis (or index date for controls), and years between BBD and BC diagnosis (or index date). We applied our computational method to 3951 whole slide images of normal breast tissue from BBD biopsies (287 cases and 1083 controls; median 3 images per woman). Quantitative estimates of TDLU involution were derived for 3 standardized measures (median TDLU span, TDLU count per non-adipose tissue area, median acini count) and 5 novel measures (median TDLU area, TDLU area as a percentage of total tissue area, TDLU area as a percentage of total non-adipose tissue area, acini count per non-adipose tissue area, median acini density). TDLU involution was also manually categorized in 177 cases and 857 controls. Associations between TDLU involution measures and risk factors were evaluated among controls using Spearman's rho or Chi-squared tests. The relationship between each measure and BC risk was evaluated using logistic regression models controlling for the matching factors and BBD histological subtypes. Results All 8 TDLU measures were significantly inversely correlated with age at BBD biopsy (range: rho -0.07 to -0.42; p&amp;lt;0.05) and menopausal status (p&amp;lt;0.05). Parity was positively associated with 6 metrics (p&amp;lt;0.05); birth index was inversely associated with another 6 metrics (p&amp;lt;0.05). Select TDLU measures were also significantly associated with BBD histological subtype, BMI, age at first birth, and/or length of total breastfeeding (p&amp;lt;0.05). No metric was significantly correlated with body size at ages 5-10 years or age of menarche. No quantitative TDLU involution measure was associated with subsequent BC risk; results remained null within strata of parity and menopausal status. Qualitative categorizations of TDLU involution were also not associated with BC risk (predominant lobule type 1 no type 3 versus no type 1, adjusted OR=0.95, 95%CI 0.54-1.71). Conclusion Quantitative measures of TDLU involution were associated with age and reproductive BC risk factors. However, automated and manual assessments of TDLU involution in normal tissue were not associated with BC risk. Further work will include applying our method to assess TDLU involution and BC risk in other cohorts. Citation Format: Kevin H. Kensler, Emily Z. Liu, Suzanne C. Wetstein, Allison M. Onken, Christina I. Luffman, Gabrielle M. Baker, Laura C. Collins, Stuart J. Schnitt, Vanessa C. Bret-Mounet, Mitko Veta, Josien P. Pluim, Ying Liu, Graham A. Colditz, Rulla M. Tamimi, Yu Jing Jan Heng. Automated quantitative measures of terminal duct lobular unit involution and breast cancer risk [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4632.

Abstract A080: Associations between quantitative measures of TDLU involution and breast tumor molecular subtypes among breast cancer cases in the Black Women’s Health Study

Abstract Background: Terminal duct lobular units (TDLUs) are the structures in the breast that give rise to most breast cancers. Previous work has shown that TDLU involution is inversely associated with TDLU metrics, such as TDLU count/100mm2, TDLU span (μm), and number of acini/TDLU, and that these metrics may be elevated in the background normal breast tissue of women diagnosed with triple-negative (TN) compared with luminal A breast tumors. However, it is unknown if this relationship exists in black women, who have the highest incidence of TN breast cancer as well as the highest overall breast cancer mortality rate. We sought to determine the relationships of quantitative measures of TDLU involution with breast cancer molecular subtype among participants in the Black Women’s Health Study. Methods: We digitized hematoxylin and eosin stained normal adjacent tissues from TN (estrogen receptor negative (ER), progesterone receptor negative, and human epidermal growth factor 2 (HER2) negative; n=67) and luminal A (ER positive and HER2 negative; n=162) breast cancer cases from the Black Women’s Health Study. We used logistic regression to evaluate associations between TDLU metrics and breast cancer subtype (TN vs. luminal A), with adjustment for age and body mass index. We performed ordinal logistic regression to evaluate relationships between population and clinical characteristics and TDLU metrics. Results: Among the 229 breast cancer cases, mean age at diagnosis was 53.7 years; 68.7% of TN and 54.3% of luminal A cases were under 55 years of age. Most women had a body mass index (BMI) &amp;gt;30kg/m2, were parous, did not smoke, and did not have a family history of breast cancer. The odds of TN breast cancer were elevated for the second and third tertiles of TDLU count relative to the first tertile, with odds ratios (95% confidence interval) of 2.89 (1.11, 4.86) and 1.92 (0.93, 4.08), respectively. Similarly, the odds of TN breast cancer increased with increasing tertiles of median TDLU span, with odds ratios of 2.25 (1.06, 4.91) and 2.38 (1.14, 5.15) for the second and third tertiles, respectively, compared to the first tertile. These associations persisted even after adjustment for age and BMI. No association was observed with median acini count/TDLU and TN breast cancer. We also observed significant associations of some breast cancer risk factors with measures of TDLU involution. Higher TDLU count was associated with younger age, more physical activity, lower BMI, current use of oral contraceptives or menopausal hormones, and premenopausal status. Conclusion: The associations of TDLU metrics with breast cancer subtype observed in this population of black women are consistent with previous studies of white and Asian women, with reduced TDLU involution in TN breast cancers compared with luminal A. Further investigation is needed to understand the factors that influence TDLU involution and the mechanisms that mediate TDLU involution and breast cancer subtype. Citation Format: Brittny C Davis Lynn, Renata Cora, Ruth M Pfeiffer, Traci N Bethea, Gary Zirpoli, Julie R Palmer, Gretchen L Gierach. Associations between quantitative measures of TDLU involution and breast tumor molecular subtypes among breast cancer cases in the Black Women’s Health Study [abstract]. In: Proceedings of the Twelfth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2019 Sep 20-23; San Francisco, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(6 Suppl_2):Abstract nr A080.

Deep learning assessment of breast terminal duct lobular unit involution: Towards automated prediction of breast cancer risk.

Terminal duct lobular unit (TDLU) involution is the regression of milk-producing structures in the breast. Women with less TDLU involution are more likely to develop breast cancer. A major bottleneck in studying TDLU involution in large cohort studies is the need for labor-intensive manual assessment of TDLUs. We developed a computational pathology solution to automatically capture TDLU involution measures. Whole slide images (WSIs) of benign breast biopsies were obtained from the Nurses’ Health Study. A set of 92 WSIs was annotated for acini, TDLUs and adipose tissue to train deep convolutional neural network (CNN) models for detection of acini, and segmentation of TDLUs and adipose tissue. These networks were integrated into a single computational method to capture TDLU involution measures including number of TDLUs per tissue area, median TDLU span and median number of acini per TDLU. We validated our method on 40 additional WSIs by comparing with manually acquired measures. Our CNN models detected acini with an F1 score of 0.73±0.07, and segmented TDLUs and adipose tissue with Dice scores of 0.84±0.13 and 0.87±0.04, respectively. The inter-observer ICC scores for manual assessments on 40 WSIs of number of TDLUs per tissue area, median TDLU span, and median acini count per TDLU were 0.71, 0.81 and 0.73, respectively. Intra-observer reliability was evaluated on 10/40 WSIs with ICC scores of >0.8. Inter-observer ICC scores between automated results and the mean of the two observers were: 0.80 for number of TDLUs per tissue area, 0.57 for median TDLU span, and 0.80 for median acini count per TDLU. TDLU involution measures evaluated by manual and automated assessment were inversely associated with age and menopausal status. We developed a computational pathology method to measure TDLU involution. This technology eliminates the labor-intensiveness and subjectivity of manual TDLU assessment, and can be applied to future breast cancer risk studies.

Relationship of Serum Progesterone and Progesterone Metabolites with Mammographic Breast Density and Terminal Ductal Lobular Unit Involution among Women Undergoing Diagnostic Breast Biopsy.

The association of progesterone/progesterone metabolites with elevated mammographic breast density (MBD) and delayed age-related terminal duct lobular unit (TDLU) involution, strong breast cancer risk factors, has received limited attention. Using a reliable liquid chromatography-tandem mass-spectrometry assay, we quantified serum progesterone/progesterone metabolites and explored cross-sectional relationships with MBD and TDLU involution among women, ages 40–65, undergoing diagnostic breast biopsy. Quantitative MBD measures were estimated in pre-biopsy digital mammograms. TDLU involution was quantified in diagnostic biopsies. Adjusted partial correlations and trends across MBD/TDLU categories were calculated. Pregnenolone was positively associated with percent MBD-area (MBD-A, rho: 0.30; p-trend = 0.01) among premenopausal luteal phase women. Progesterone tended to be positively associated with percent MBD-A among luteal phase (rho: 0.26; p-trend = 0.07) and postmenopausal (rho: 0.17; p-trend = 0.04) women. Consistent with experimental data, implicating an elevated 5α-pregnanes/3α-dihydroprogesterone (5αP/3αHP) metabolite ratio in breast cancer, higher 5αP/3αHP was associated with elevated percent MBD-A among luteal phase (rho: 0.29; p-trend = 0.08), but not postmenopausal women. This exploratory analysis provided some evidence that endogenous progesterone and progesterone metabolites might be correlated with MBD, a strong breast cancer risk factor, in both pre- and postmenopausal women undergoing breast biopsy. Additional studies are needed to understand the role of progesterone/progesterone metabolites in breast tissue composition and breast cancer risk.

Using Digital Pathology to Understand Epithelial Characteristics of Benign Breast Disease among Women Undergoing Diagnostic Image-Guided Breast Biopsy.

Delayed terminal duct lobular unit (TDLU) involution is associated with elevated mammographic breast density (MD). Both are independent breast cancer risk factors among women with benign breast disease (BBD). Prior digital analyses of normal breast tissues revealed that epithelial nuclear density (END) and TDLU involution are inversely correlated. Accordingly, we examined associations of END, TDLU involution, and MD in BBD clinical biopsies. This study included digitized images of 262 representative image-guided hematoxylin and eosin-stained biopsies from 224 women diagnosed with BBD, enrolled within the cross-sectional BREAST-Stamp project that were visually assessed for TDLU involution (TDLU count/100 mm2, median TDLU span and median acini count per TDLU). A digital algorithm estimated nuclei count per unit epithelial area, or END. Single X-ray absorptiometry of prebiopsy ipsilateral craniocaudal digital mammograms measured global and localized MD surrounding the biopsy region. Adjusted ordinal logistic regression models assessed relationships between tertiles of TDLU and END measures. Analysis of covariance examined mean differences in MD across END tertiles. TDLU measures were positively associated with increasing END tertiles [TDLU count/100 mm2, ORT3vsT1: 3.42, 95% confidence interval (CI), 1.87-6.28; acini count/TDLUT3vsT1, OR: 2.40, 95% CI, 1.39-4.15]. END was significantly associated with localized, but not, global MD. Relationships were most apparent among patients with nonproliferative BBD. These findings suggest that quantitative END reflects different but complementary information to the histologic information captured by visual TDLU and radiologic MD measures and merits continued evaluation in assessing cellularity of breast parenchyma to understand the etiology of BBD.

The relationship between terminal duct lobular unit features and mammographic density among Chinese breast cancer patients.

Extensive mammographic density (MD), a well-established breast cancer risk factor, is a radiological representation of stromal and epithelial breast tissue content. In studies conducted predominantly among Caucasian women, histologic measures of reduced terminal duct lobular unit (TDLU) involution have been correlated with extensive MD, but independently associated with breast cancer risk. We therefore examined associations between TDLU measures and MD among Chinese women, a low-risk population but with high prevalence of dense breasts. Diagnostic pre-treatment digital mammograms were obtained from 144 breast cancer cases at a tertiary hospital in Beijing and scored using the Breast Imaging Reporting and Data System (BI-RADS) density classification. TDLU features were assessed using three standardized measures (count/100 mm2 , span [μm], and acini count/TDLU) in benign tissues. Associations between each of TDLU measures and MD were examined using generalized linear models for TDLU count and span and polytomous logistic regression for acini count with adjustment for potential confounders stratified by age. Among women ≥50 years, 63% had dense breasts; cases with dense breasts (BI-RADS, c-d) had greater TDLU count (21.1 [SE = 2.70] vs. 9.0 [SE = 1.83]; p = 0.0004), longer span (480.6 μm [SE = 24.6] vs. 393.8 μm [SE = 31.8]; p = 0.03), and greater acini count (ORtrend = 16.1; 95%CI = 4.08-63.1; ptrend < 0.0001) compared to those with non-dense breasts (BI-RADS, a-b). Among women <50 years, 91% had dense breasts, precluding our ability to detect associations. Our findings are consistent with previously reported associations between extensive MD and reduced TDLU involution, supporting the hypothesis that breast cancer risk associated with extensive MD may be related to the amount of "at-risk" epithelium.

Abstract A36: Serum insulin-like growth factor (IGF)-I and IGF binding protein-3 in relation to terminal duct lobular unit involution in Caucasian and African American women: The Susan G. Komen Tissue Bank

Abstract Background: Insulin-like growth factor (IGF)-I and IGF binding protein (IGFBP)-3 play important roles in carcinogenesis, particularly for breast cancer. However, little is known about whether the IGF system influences histologic characteristics of normal glandular tissue and whether relationships vary by race. Lesser degrees of age-related terminal duct lobular unit (TDLU) involution, as reflected by higher numbers of TDLUs and acini per TDLU, have been associated with higher breast cancer risk. We examined the associations of IGF measures with TDLU involution of normal breast using standardized TDLU measures. Methods: Among 715 Caucasian and 283 African American (AA) women with normal breast tissue samples from the Komen Tissue Bank, serum concentrations of IGF-I and IGFBP-3 were quantified using enzyme-linked immunosorbent assay (ELISA). Hematoxilyn and eosin-stained tissue sections were assessed for numbers of TDLUs (“TDLU count”) and acini/TDLU. Zero-inflated Poisson regression models with a robust variance estimator were used to estimate associations of IGF-I, IGFBP-3, and IGF-I:IGFBP-3 molar ratio (tertiles) with TDLU count by race and menopausal status, adjusting for potential confounders. We also tested for interactions by race using likelihood ratio tests. Results: AA (vs. Caucasian) women had higher age-adjusted mean levels of serum IGF-I (137 vs. 131 ng/mL, p=0.07) and lower levels of IGFBP-3 (4165 vs. 4684 ng/mL, p&amp;lt;0.0001); the differences persisted after adjustment for additional covariates including BMI and parity/age at first birth. Postmenopausal IGFBP-3 was inversely associated with TDLU count among both AA (RR T3vs.T1=0.49, 95% CI=0.28-0.84, p-trend=0.04) and Caucasian (RR T3vs.T1=0.63, 95% CI=0.41-0.99, p-trend=0.04) women. In premenopausal women, higher IGF-I:IGFBP-3 ratios were associated with higher TDLU count in Caucasians (OR T3vs.T1=1.33, 95% CI=1.01-1.31, p-trend=0.04) but not in AA (OR T3vs.T1=0.65, 95% CI=0.42-1.00, p-trend=0.05) women. There was no statistically significant interaction by race (p-interaction≥0.10). Conclusions: Our data suggest the potential role of the IGF system, particularly IGFBP-3, in TDLU involution of the normal breast among both Caucasian and AA women. Citation Format: Hannah OH, Ruth M. Pfeiffer, Roni T. Falk, Hisani N. Horne, Jackie Xiang, Michael Pollak, Louise A. Brinton, Anna Maria V. Storniolo, Mark E. Sherman, Gretchen L. Gierach, Jonine D. Figueroa. Serum insulin-like growth factor (IGF)-I and IGF binding protein-3 in relation to terminal duct lobular unit involution in Caucasian and African American women: The Susan G. Komen Tissue Bank [abstract]. In: Proceedings of the Tenth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2017 Sep 25-28; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2018;27(7 Suppl):Abstract nr A36.

Serum insulin-like growth factor (IGF)-I and IGF binding protein-3 in relation to terminal duct lobular unit involution of the normal breast in Caucasian and African American women: The Susan G. Komen Tissue Bank.

Lesser degrees of terminal duct lobular unit (TDLU) involution, as reflected by higher numbers of TDLUs and acini/TDLU, are associated with elevated breast cancer risk. In rodent models, the insulin-like growth factor (IGF) system regulates involution of the mammary gland. We examined associations of circulating IGF measures with TDLU involution in normal breast tissues among women without precancerous lesions. Among 715 Caucasian and 283 African American (AA) women who donated normal breast tissue samples to the Komen Tissue Bank between 2009 and 2012 (75% premenopausal), serum concentrations of IGF-I and binding protein (IGFBP)-3 were quantified using enzyme-linked immunosorbent assay. Hematoxilyn and eosin-stained tissue sections were assessed for numbers of TDLUs ("TDLU count"). Zero-inflated Poisson regression models with a robust variance estimator were used to estimate relative risks (RRs) for association of IGF measures (tertiles) with TDLU count by race and menopausal status, adjusting for potential confounders. AA (vs. Caucasian) women had higher age-adjusted mean levels of serum IGF-I (137 vs. 131 ng/mL, p = 0.07) and lower levels of IGFBP-3 (4165 vs. 4684 ng/mL, p < 0.0001). Postmenopausal IGFBP-3 was inversely associated with TDLU count among AA (RRT3vs.T1 = 0.49, 95% CI = 0.28-0.84, p-trend = 0.04) and Caucasian (RRT3vs.T1 =0.64, 95% CI = 0.42-0.98, p-trend = 0.04) women. In premenopausal women, higher IGF-I:IGFBP-3 ratios were associated with higher TDLU count in Caucasian (RRT3vs.T1 =1.33, 95% CI = 1.02-1.75, p-trend = 0.04), but not in AA (RRT3vs.T1 =0.65, 95% CI = 0.42-1.00, p-trend = 0.05), women. Our data suggest a role of the IGF system, particularly IGFBP-3, in TDLU involution of the normal breast, a breast cancer risk factor, among Caucasian and AA women.

Abstract 5290: Racial variation in terminal duct lobular unit (TDLU) involution in Chinese and Polish breast cancer patients

Abstract Introduction: Terminal duct lobular unit (TDLU) involution, an age-related physiological process, is characterized by the reduction of size and numbers. Reduced involution is associated with higher mammographic density, a strong breast cancer risk factor. Both factors predict risk of developing breast cancer among women with benign disease. Given that prior studies have been conducted mainly in White women, we compared the extent of TDLU involution among Western and Asian breast cancer cases to describe potential racial heterogeneity in breast cancer etiology. Method: We obtained three TDLU involution metrics (count/100 mm2, mean span, and mean acini count/TDLU; all inversely correlated with TDLU involution) measured in benign breast tissue sections from 379 Chinese (254 luminal A and 125 core basal phenotype [CBP] cases) and 476 Polish (407 luminal A and 69 CBP) breast cancer cases. Polytomous logistic regression was performed using tertiles of TDLU measures as ordinal dependent variable and race as an independent variable. Covariates included age, body mass index, and parity. Analyses were performed separately by age group (&amp;lt;50, ≥50) and subtype (luminal A and CBP). Results: Among luminal A cases, Chinese had significantly greater TDLU count compared with Polish in both age groups (ORtrend= 11.0; 95% CI=2.80-43.2; Ptrend=0.001 for age&amp;lt;50; ORtrend= 12.1; 95% CI=5.12-28.7; Ptrend=1E-08 for age≥50). In addition, Chinese were more likely to have greater mean span and acini count among younger cases (ORtrend=2.5; 95% CI=1.44-4.28; Ptrend=0.001 for mean span and ORtrend=7.4; 95% CI=4.13-13.4; Ptrend=3E-11 for acini count) but not among older cases. Among CBP cases, Chinese were more likely to have greater acini count compared to Polish only among younger women (ORtrend=2.6; 95% CI=1.12-6.95; Ptrend=0.03 for age&amp;lt;50). Conclusion: We found Chinese were more likely to have reduced TDLU involution compared to Polish cases after accounting for potential confounders, with greater differences for luminal A and younger women. Studies of TDLU involution in diverse populations are needed to confirm and understand whether population difference in TDLU involution metrics are related to age at breast cancer onset and risks for specific tumor subtypes. Citation Format: Hyuna Sung, Changyuan Guo, Jennifer Guida, Shan Zheng, Erni Li, Jing Li, Nan Hu, Joseph Deng, Montserrat Garcia-Closas, Jonine Figueroa, Mark Sherman, Gretchen Gierach, Ning Lu, Xiaohong R. Yang. Racial variation in terminal duct lobular unit (TDLU) involution in Chinese and Polish breast cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5290. doi:10.1158/1538-7445.AM2017-5290

Age-related terminal duct lobular unit involution in benign tissues from Chinese breast cancer patients with luminal and triple-negative tumors

BackgroundTerminal duct lobular unit (TDLU) involution is a physiological process of breast tissue aging characterized by a reduction in the epithelial component. In studies of women with benign breast disease, researchers have found that age-matched women with lower levels of TDLU involution are at increased risk of developing breast cancer. We previously showed that breast cancer cases with core basal phenotype (CBP; estrogen receptor negative [ER−], progesterone receptor-negative [PR−], human epidermal growth factor receptor 2-negative [HER2−], cytokeratins (CK 5 or CK5/6)-positive [CK5/6+] and/or epidermal growth factor receptor-positive [EGFR+]) tumors had significantly reduced TDLU involution compared with cases with luminal A (ER+ and/or PR+, HER2−, CK5/6−, EGFR−) tumors from a population-based case-control study in Poland. We evaluated the association of TDLU involution with tumor subtypes in an independent population of women in China, where the breast cancer incidence rate, prevalence of known risk factors, and mammographic breast density are thought to be markedly different from those of Polish women.MethodsWe performed morphometric assessment of TDLUs by using three reproducible semiquantitative measures that inversely correlate with TDLU involution (TDLU count/100 mm2, TDLU span in micrometer, and acini count/TDLU) by examining benign tissue blocks from 254 age-matched luminal A and 250 triple-negative (TN; ER−, PR−, HER2−, including 125 CBP) breast cancer cases treated in a tertiary hospital in Beijing, China.ResultsOverall, we found that TN and particularly CBP cases tended to have greater TDLU measures (less involution) than luminal A cases in logistic regression models accounting for age, body mass index, parity, and tumor grade. The strongest association was observed for tertiles of acini count among younger women (aged <50 years) (CBP vs. luminal A; ORtrend 2.11, 95% CI 1.22–3.67, P = 0.008).ConclusionsThese data extend previous findings that TN/CBP breast cancers are associated with reduced TDLU involution in surrounding breast parenchyma compared with luminal A cases among Chinese women, providing further support for differences in the pathogenesis of these tumor subtypes.

Abstract IA11: Life course epidemiology and breast cancer: translating risk into prevention

Abstract Molecular histology may be conceptualized as the microscopic and molecular characteristics of normal tissues that are required for physiologic function. Over the life course, the molecular histology of the breast changes in response to physiological alterations, imparting spatial and temporal heterogeneity within the breasts of individuals and among women. These transitions contribute to the enormous range and imprecisely defined limits of what pathologists consider normal. Appreciation that molecular histology may reflect the cumulative influence of prior exposures linked to breast cancer risk, and may provide information about risk of developing breast cancer in the future, has stimulated interest in this topic. Unlike the study of breast cancer or its precursors, which represents a focal pathophysiologic deviation from normal, molecular histology, if assessable, would represent the state of the entire at-risk organ, based upon examination of a small tissue sample. The adult breast is characterized by well-developed terminal duct lobular units (TDLUs), which comprise the functional unit of milk production and represent the source of nearly all breast cancer precursors. Physiological changes in human breasts are likely driven by paracrine mechanisms, suggesting that tissue context and cellular topography are critical elements in physiology and pathophysiology. The breast undergoes profound changes with completion of childbearing and aging. Age-related TDLU involution may be conceptualized as a protective mechanism that lowers breast cancer risk following completion of childbearing, and in this context, delayed or incomplete involution is a breast cancer risk factor. With aging, the percent of the breast comprised of fibroglandular tissue declines, which is associated with a reduction in mammographic density, a strong breast cancer risk factor. Mammographic density is also imperfectly correlated with epithelial content in the breast. Women with benign breast disease whose surrounding normal breast tissue does not undergo age-appropriate TDLU involution are at increased risk of developing breast cancer, and both breast density and TDLU involution are independent markers of breast cancer risk. However, an important challenge is to understand the markers and mechanisms that underlie breast involution with aging, including both the epithelial and non-epithelial components, and to learn why both density and TDLU content decline with aging as breast cancer incidence rises. The thesis of this presentation is that understanding the amount of epithelium at-risk, the insults it sustains and the mechanisms that lead to its elimination, persistence or expansion may provide a window into the development of integrative biomarkers of risk that can translate into improved screening and prevention. However, progress towards this goal is quite early. Citation Format: Mark E. Sherman. Life course epidemiology and breast cancer: translating risk into prevention. [abstract]. In: Proceedings of the AACR Special Conference: Improving Cancer Risk Prediction for Prevention and Early Detection; Nov 16-19, 2016; Orlando, FL. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2017;26(5 Suppl):Abstract nr IA11.

Association between breast cancer genetic susceptibility variants and terminal duct lobular unit involution of the breast.

Terminal duct lobular units (TDLUs) are the predominant source of future breast cancers, and lack of TDLU involution (higher TDLU counts, higher acini count per TDLU and the product of the two) is a breast cancer risk factor. Numerous breast cancer susceptibility single nucleotide polymorphisms (SNPs) have been identified, but whether they are associated with TDLU involution is unknown. In a pooled analysis of 872 women from two studies, we investigated 62 established breast cancer SNPs and relationships with TDLU involution. Poisson regression models with robust variance were used to calculate adjusted per-allele relative risks (with the non-breast cancer risk allele as the referent) and 95% confidence intervals between TDLU measures and each SNP. All statistical tests were two-sided; P < 0.05 was considered statistically significant. Overall, 36 SNPs (58.1%) were related to higher TDLU counts although this was not statistically significant (p = 0.25). Six of the 62 SNPs (9.7%) were nominally associated with at least one TDLU measure: rs616488 (PEX14), rs11242675 (FOXQ1) and rs6001930 (MKL1) were associated with higher TDLU count (p = 0.047, 0.045 and 0.031, respectively); rs1353747 (PDE4D) and rs6472903 (8q21.11) were associated with higher acini count per TDLU (p = 0.007 and 0.027, respectively); and rs1353747 (PDE4D) and rs204247 (RANBP9) were associated with the product of TDLU and acini counts (p = 0.024 and 0.017, respectively). Our findings suggest breast cancer SNPs may not strongly influence TDLU involution. Agnostic genome-wide association studies of TDLU involution may provide new insights on its biologic underpinnings and breast cancer susceptibility.

Standardized measures of lobular involution and subsequent breast cancer risk among women with benign breast disease: a nested case-control study.

Lesser degrees of terminal duct-lobular unit (TDLU) involution predict higher breast cancer risk; however, standardized measures to quantitate levels of TDLU involution have only recently been developed. We assessed whether three standardized measures of TDLU involution, with high intra/inter pathologist reproducibility in normal breast tissue, predict subsequent breast cancer risk among women in the Mayo benign breast disease (BBD) cohort. We performed a masked evaluation of biopsies from 99 women with BBD who subsequently developed breast cancer (cases) after a median of 16.9years and 145 age-matched controls. We assessed three metrics inversely related to TDLU involution: TDLU count/mm(2), median TDLU span (microns, which approximates acini content), and median category of acini counts/TDLU (0-10; 11-20; 21-30; 31-50; >50). Associations with subsequent breast cancer risk for quartiles (or categories of acini counts) of each of these measures were assessed with multivariable conditional logistic regression to estimate odds ratios (ORs) and 95% confidence intervals (CI). In multivariable models, women in the highest quartile compared to the lowest quartiles of TDLU counts and TDLU span measures were significantly associated with subsequent breast cancer diagnoses; TDLU counts quartile4 versus quartile1, OR=2.44, 95%CI 0.96-6.19, p-trend=0.02; and TDLU spans, quartile4 versus quartile1, OR=2.83, 95%CI=1.13-7.06, p-trend=0.03. Significant associations with categorical measures of acini counts/TDLU were also observed: compared to women with median category of <10 acini/TDLU, women with >25 acini counts/TDLU were at significantly higher risk, OR=3.40, 95%CI 1.03-11.17, p-trend=0.032. Women with TDLU spans and TDLU count measures above the median were at further increased risk, OR=3.75 (95%CI 1.40-10.00, p-trend=0.008), compared with women below the median for both of these metrics. Similar results were observed for combinatorial metrics of TDLU acini counts/TDLU, and TDLU count. Standardized quantitative measures of TDLU counts and acini counts approximated by TDLU span measures or visually assessed in categories are independently associated with breast cancer risk. Visual assessment of TDLU numbers and acini content, which are highly reproducible between pathologists, could help identify women at high risk for subsequent breast cancer among the million women diagnosed annually with BBD in the US.

Abstract 4283: Relationship between mammographic breast density and measures of terminal duct lobular unit involution among women diagnosed with estrogen receptor positive breast cancer

Abstract Introduction: High mammographic breast density (MD) and reduced levels of terminal duct lobular unit (TDLU) involution (the histologic source of most breast cancers) have been associated with increased risk of developing breast cancer. Data relating MD and TDLU involution to breast cancer characteristics and outcomes are sparse. Therefore, we assessed these relationships among women with invasive ER-positive breast cancer. Methods: The analysis focused on women with ER-positive breast cancers who were diagnosed at Kaiser Permanente Northwest (1990-2008) and followed through the end of 2010. Cases comprised of those who died of breast cancer (n = 54) and controls those that did not die of breast cancer (n = 180) over similar follow-up. Three reproducible measures that are inversely related to TDLU involution were evaluated in digitized hematoxilin and eosin stained sections in benign breast tissues surrounding the tumors: TDLU counts per unit area, TDLU span and median number of acini per TDLU. Percentage MD was estimated from digitized mammograms using computer-assisted thresholding software (Cumulus). Univariate associations between TDLU measurements and patient characteristics, tumor size, and disease stage at diagnosis, were calculated using Mann Whitney Wilcoxon rank test. TDLU measurements were related to baseline MD using analysis of covariance models and adjusted for age, body mass index (BMI), tumor size, stage, year of diagnosis and smoking. Results: TDLUs were observed in 95% of cases and 89% of controls. All TDLU measurements declined with age (p&amp;lt;0.001 for each TDLU measurement). Among cases, TDLU measures were not significantly associated with tumor characteristics. Controls with regional spread had greater TDLU span (p = 0.05) and median acini counts per TDLU (p = 0.03) than those with localized disease; these TDLU metrics also showed a borderline significant association with larger tumor size (&amp;gt;2cm) (p = 0.07 and p = 0.06, respectfully). All TDLU measures were associated with MD among controls (TDLU count: p = 0.04; TDLU span: p = 0.06; median acini count per TDLU: p = 0.01), whereas among cases only, TDLU span showed a significant association MD (p = 0.003). Conclusion: Preliminarily, our data suggest that among women with non-fatal ER-positive breast cancers, TDLU involution was associated with localized tumor stage, size and MD, whereas these relationships were less evident among women who died of their disease. Ongoing analyses will determine whether measures of MD and TDLU involution are independent predictors of breast cancer outcomes in this patient population. Citation Format: Maeve Mullooly, Sarah J. Nyante, Ruth M. Pfeiffer, Renata Cora, Jonine D. Figueroa, Robert N. Hoover, Andrew G. Glass, Erin J. Aiello Bowles, Louise A. Brinton, Amy Berrington de Gonzalez, Sherman E. Mark, Gretchen L. Gierach. Relationship between mammographic breast density and measures of terminal duct lobular unit involution among women diagnosed with estrogen receptor positive breast cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4283.