Laura Linville

Terminal duct lobular unit involution of the normal breast: implications for breast cancer etiology.

BACKGROUND Greater degrees of terminal duct lobular unit (TDLU) involution have been linked to lower breast cancer risk; however, factors that influence this process are poorly characterized. METHODS To study this question, we developed three reproducible measures that are inversely associated with TDLU involution: TDLU counts, median TDLU span, and median acini counts/TDLU. We determined factors associated with TDLU involution using normal breast tissues from 1938 participants (1369 premenopausal and 569 postmenopausal) ages 18 to 75 years in the Susan G. Komen Tissue Bank at the Indiana University Simon Cancer Center. Multivariable zero-inflated Poisson models were used to estimate relative risks (RRs) and 95% confidence intervals (95% CIs) for factors associated with TDLU counts, and multivariable ordinal logistic regression models were used to estimate odds ratios (ORs) and 95% CIs for factors associated with categories of median TDLU span and acini counts/TDLU. RESULTS All TDLU measures started declining in the third age decade (all measures, two-sided P trend ≤ .001); and all metrics were statistically significantly lower among postmenopausal women. Nulliparous women demonstrated lower TDLU counts compared with uniparous women (among premenopausal women, RR = 0.79, 95% CI = 0.73 to 0.85; among postmenopausal, RR = 0.67, 95% CI = 0.56 to 0.79); however, rates of age-related TDLU decline were faster among parous women. Other factors were related to specific measures of TDLU involution. CONCLUSION Morphometric analysis of TDLU involution warrants further evaluation to understand the pathogenesis of breast cancer and assessing its role as a progression marker for women with benign biopsies or as an intermediate endpoint in prevention studies.

Relationship of Terminal Duct Lobular Unit Involution of the Breast with Area and Volume Mammographic Densities

Elevated mammographic density (MD) is an established breast cancer risk factor. Reduced involution of terminal duct lobular units (TDLU), the histologic source of most breast cancers, has been associated with higher MD and breast cancer risk. We investigated relationships of TDLU involution with area and volumetric MD, measured throughout the breast and surrounding biopsy targets (perilesional). Three measures inversely related to TDLU involution (TDLU count/mm2, median TDLU span, median acini count/TDLU) assessed in benign diagnostic biopsies from 348 women, ages 40–65, were related to MD area (quantified with thresholding software) and volume (assessed with a density phantom) by analysis of covariance, stratified by menopausal status and adjusted for confounders. Among premenopausal women, TDLU count was directly associated with percent perilesional MD (P trend = 0.03), but not with absolute dense area/volume. Greater TDLU span was associated with elevated percent dense area/volume (P trend<0.05) and absolute perilesional MD (P = 0.003). Acini count was directly associated with absolute perilesional MD (P = 0.02). Greater TDLU involution (all metrics) was associated with increased nondense area/volume (P trend ≤ 0.04). Among postmenopausal women, TDLU measures were not significantly associated with MD. Among premenopausal women, reduced TDLU involution was associated with higher area and volumetric MD, particularly in perilesional parenchyma. Data indicating that TDLU involution and MD are correlated markers of breast cancer risk suggest that associations of MD with breast cancer may partly reflect amounts of at-risk epithelium. If confirmed, these results could suggest a prevention paradigm based on enhancing TDLU involution and monitoring efficacy by assessing MD reduction. Cancer Prev Res; 9(2); 149–58. ©2015 AACR.

Circulating insulin-like growth factor-I, insulin-like growth factor binding protein-3 and terminal duct lobular unit involution of the breast: a cross-sectional study of women with benign breast disease

BackgroundTerminal duct lobular units (TDLUs) are the primary structures from which breast cancers and their precursors arise. Decreased age-related TDLU involution and elevated mammographic density are both correlated and independently associated with increased breast cancer risk, suggesting that these characteristics of breast parenchyma might be linked to a common factor. Given data suggesting that increased circulating levels of insulin-like growth factors (IGFs) factors are related to reduced TDLU involution and increased mammographic density, we assessed these relationships using validated quantitative methods in a cross-sectional study of women with benign breast disease.MethodsSerum IGF-I, IGFBP-3 and IGF-I:IGFBP-3 molar ratios were measured in 228 women, ages 40-64, who underwent diagnostic breast biopsies yielding benign diagnoses at University of Vermont affiliated centers. Biopsies were assessed for three separate measures inversely related to TDLU involution: numbers of TDLUs per unit of tissue area (“TDLU count”), median TDLU diameter (“TDLU span”), and number of acini per TDLU (“acini count”). Regression models, stratified by menopausal status and adjusted for potential confounders, were used to assess the associations of TDLU count, median TDLU span and median acini count per TDLU with tertiles of circulating IGFs. Given that mammographic density is associated with both IGF levels and breast cancer risk, we also stratified these associations by mammographic density.ResultsHigher IGF-I levels among postmenopausal women and an elevated IGF-I:IGFBP-3 ratio among all women were associated with higher TDLU counts, a marker of decreased lobular involution (P-trend = 0.009 and <0.0001, respectively); these associations were strongest among women with elevated mammographic density (P-interaction <0.01). Circulating IGF levels were not significantly associated with TDLU span or acini count per TDLU.ConclusionsThese results suggest that elevated IGF levels may define a sub-group of women with high mammographic density and limited TDLU involution, two markers that have been related to increased breast cancer risk. If confirmed in prospective studies with cancer endpoints, these data may suggest that evaluation of IGF signaling and its downstream effects may have value for risk prediction and suggest strategies for breast cancer chemoprevention through inhibition of the IGF system.

Circulating Sex Hormones and Terminal Duct Lobular Unit Involution of the Normal Breast

Background: Terminal duct lobular units (TDLU) are the predominant source of breast cancers. Lesser degrees of age-related TDLU involution have been associated with increased breast cancer risk, but factors that influence involution are largely unknown. We assessed whether circulating hormones, implicated in breast cancer risk, are associated with levels of TDLU involution using data from the Susan G. Komen Tissue Bank (KTB) at the Indiana University Simon Cancer Center (2009–2011). Methods: We evaluated three highly reproducible measures of TDLU involution, using normal breast tissue samples from the KTB (n = 390): TDLU counts, median TDLU span, and median acini counts per TDLU. RRs (for continuous measures), ORs (for categorical measures), 95% confidence intervals (95% CI), and Ptrends were calculated to assess the association between tertiles of estradiol, testosterone, sex hormone–binding globulin (SHBG), progesterone, and prolactin with TDLU measures. All models were stratified by menopausal status and adjusted for confounders. Results: Among premenopausal women, higher prolactin levels were associated with higher TDLU counts (RRT3vsT1:1.18; 95% CI: 1.07–1.31; Ptrend = 0.0005), but higher progesterone was associated with lower TDLU counts (RRT3vsT1: 0.80; 95% CI: 0.72–0.89; Ptrend < 0.0001). Among postmenopausal women, higher levels of estradiol (RRT3vsT1:1.61; 95% CI: 1.32–1.97; Ptrend < 0.0001) and testosterone (RRT3vsT1: 1.32; 95% CI: 1.09–1.59; Ptrend = 0.0043) were associated with higher TDLU counts. Conclusions: These data suggest that select hormones may influence breast cancer risk potentially through delaying TDLU involution. Impact: Increased understanding of the relationship between circulating markers and TDLU involution may offer new insights into breast carcinogenesis. Cancer Epidemiol Biomarkers Prev; 23(12); 2765–73. ©2014 AACR.

Quantitative analysis of TDLUs using adaptive morphological shape techniques

Within the complex branching system of the breast, terminal duct lobular units (TDLUs) are the anatomical location where most cancer originates. With aging, TDLUs undergo physiological involution, re ected in a loss of structural components (acini) and a reduction in total number. Data suggest that women undergoing benign breast biopsies that do not show age appropriate involution are at increased risk of developing breast cancer. To date, TDLU assessments have generally been made by qualitative visual assessment, rather by objective quantitative analysis. This paper introduces a technique to automatically estimate a set of quantitative measurements and use those variables to more objectively describe and classify TDLUs. To validate the accuracy of our system, we compared the computer-based morphological properties of 51 TDLUs in breast tissues donated for research by volunteers in the Susan G. Komen Tissue Bank and compared results to those of a pathologist, demonstrating 70% agreement. Secondly, in order to show that our method is applicable to a wider range of datasets, we analyzed 52 TDLUs from biopsies performed for clinical indications in the National Cancer Institute Breast Radiology and Study of Tissues (BREAST) STAMP project and obtained 82% correlation with visual assessment. Lastly, we demonstrate the ability to uncover novel measures when researching the structural properties of the acini by applying machine learning and clustering techniques. Through our study we found that while the number of acini per TDLU increase exponentially with the TDLU diameter, the average elongation and roundness remain constant.

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.

Quantitative Analysis of TDLUs using Adaptive Morphological Shape Techniques.

Within the complex branching system of the breast, terminal duct lobular units (TDLUs) are the anatomical location where most cancer originates. With aging, TDLUs undergo physiological involution, re ected in a loss of structural components (acini) and a reduction in total number. Data suggest that women undergoing benign breast biopsies that do not show age appropriate involution are at increased risk of developing breast cancer. To date, TDLU assessments have generally been made by qualitative visual assessment, rather by objective quantitative analysis. This paper introduces a technique to automatically estimate a set of quantitative measurements and use those variables to more objectively describe and classify TDLUs. To validate the accuracy of our system, we compared the computer-based morphological properties of 51 TDLUs in breast tissues donated for research by volunteers in the Susan G. Komen Tissue Bank and compared results to those of a pathologist, demonstrating 70% agreement. Secondly, in order to show that our method is applicable to a wider range of datasets, we analyzed 52 TDLUs from biopsies performed for clinical indications in the National Cancer Institute Breast Radiology and Study of Tissues (BREAST) STAMP project and obtained 82% correlation with visual assessment. Lastly, we demonstrate the ability to uncover novel measures when researching the structural properties of the acini by applying machine learning and clustering techniques. Through our study we found that while the number of acini per TDLU increase exponentially with the TDLU diameter, the average elongation and roundness remain constant.

Quantitative Analysis of TDLUs using Adaptive Morphological Shape Techniques

Within the complex branching system of the breast, terminal duct lobular units (TDLUs) are the anatomical location where most cancer originates. With aging, TDLUs undergo physiological involution, re ected in a loss of structural components (acini) and a reduction in total number. Data suggest that women undergoing benign breast biopsies that do not show age appropriate involution are at increased risk of developing breast cancer. To date, TDLU assessments have generally been made by qualitative visual assessment, rather by objective quantitative analysis. This paper introduces a technique to automatically estimate a set of quantitative measurements and use those variables to more objectively describe and classify TDLUs. To validate the accuracy of our system, we compared the computer-based morphological properties of 51 TDLUs in breast tissues donated for research by volunteers in the Susan G. Komen Tissue Bank and compared results to those of a pathologist, demonstrating 70% agreement. Secondly, in order to show that our method is applicable to a wider range of datasets, we analyzed 52 TDLUs from biopsies performed for clinical indications in the National Cancer Institute Breast Radiology and Study of Tissues (BREAST) STAMP project and obtained 82% correlation with visual assessment. Lastly, we demonstrate the ability to uncover novel measures when researching the structural properties of the acini by applying machine learning and clustering techniques. Through our study we found that while the number of acini per TDLU increase exponentially with the TDLU diameter, the average elongation and roundness remain constant.

Abstract 4465: Breast cancer risk factor associations with breast tissue morphometry: results from the Komen for the Cure® Tissue Bank

Little is known about the effects of many recognized breast cancer risk factors on the normal breast. We hypothesize that risk factors affect the number and morphometry of terminal ductal lobular units (TDLUs), the microanatomical structure from which most breast cancers arise. To assess this hypothesis, we evaluated relationships between breast cancer risk factors and TDLU number, TDLU diameter, and acini per TDLU in breast tissue donated by healthy volunteers. Subjects included 786 healthy women ages 18-84 years without a personal history of any cancer, who provided a risk factor questionnaire and donated a 10-guage needle core from the upper outer breast quadrant. A pathologist, masked to subject data, assessed TDLU number per section, TDLU diameter and acini per TDLU using digitized images of hematoxylin and eosin stained sections of tissue. Seventy-two randomly selected images were examined by two additional pathologists. We generated ordered categorical variables for the number of TDLUs per section (0-1, 2-8, 9+), median TDLU diameter (53-262, 263-377, 378-1375 microns) and median number of acini per TDLU (2-10, 11-20, 21+). We computed Spearman correlations between measures, and intraobserver and interobserver agreement. Associations between breast cancer risk factors and TDLU measures were assessed by estimating odds ratios (OR) and 95% confidence intervals (CI) using ordinal logistic regression models, adjusted for age. TDLU diameter and acini count were highly correlated (rho =0.71); TDLU number per section was less strongly related to the diameter (rho= 0.18) and acini measures (rho=0.20). Inter and intraobserver agreement for TDLU measures were highly correlated (rho range 0.68-0.98). All TDLU measurements were significantly lower among women age 50 years and older compared with younger women. Compared with Caucasians, African American women had more TDLUs per section (OR adj =2.03, 95% CI= 1.06-3.89). Greater numbers of TDLUs were also related to having a family history of two or more relatives with breast cancer compared with none (OR adj =2.01, 95% CI=0.91-4.46); and for parous compared with nulliparous women (OR adj =1.61, 95% CI=1.16-2.25). In contrast, parity was associated with reduced median number of acini per TDLU (OR adj =0.60, 95% CI = 0.40-0.90) and median diameter (OR adj =0.64, 95% CI = 0.42-0.96) compared with nulliparity. Our data suggest that TDLUs are more frequent among women with a positive family history of breast cancer or African Americans. Parity, an established protective factor for breast cancer risk, was related to increased numbers of TDLUs, but fewer acini per TDLU, suggesting that TDLUs may increase with pregnancy and then involute. Our data, along with published results linking TDLU involution with a reduction in breast cancer risk, suggest TDLU morphometry may reflect early biologic changes marking breast cancer susceptibility. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4465. doi:1538-7445.AM2012-4465

Abstract B89: Endogenous hormone levels and lobular involution of the breast among healthy, postmenopausal women in the Komen Tissue Bank Study

Background: Terminal ductal lobular units (TDLUs), the functional units of the breast for lactation, are the structures from which most breast cancers arise. TDLU involution, reflecting atrophy and loss of TDLUs, has been related to lower subsequent breast cancer risk. The predictors of such breast tissue aging, however, are not well defined. Given that estrogens and other hormones play a central role in the etiology of postmenopausal breast cancers, we conducted a study to assess the relationship between four hormones and sex hormone-binding globulin (SHBG) with markers of TDLU involution. Methods: Subjects included healthy women who volunteered between 2009 and 2011 to participate in the Komen Tissue Bank Study. Participants completed a risk factor questionnaire, provided a blood sample, and donated breast tissue for research. Our analysis was restricted to postmenopausal women who were not currently taking menopausal hormones (n=153). A pathologist, masked to patient data, assessed tissue sections for the presence of TDLUs and, among sections containing TDLUs (n=88), measured TDLU diameters for up to ten TDLUs per section. Both presence of TDLUs and TDLU diameter are inversely related to involution. Chemiluminescent immunometric assays were used to measure serum levels of estradiol (E2), SHBG, follicle-stimulating hormone (FSH), and prolactin. Percent free E2 was calculated from measured E2 and SHBG levels. Logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) for the presence of TDLUs (none or any) associated with tertiles of serum hormone and SHBG levels. Multivariable ordinal logistic regression was used to calculate ORs, 95% CIs, and p-trend for TDLU diameter categorized as tertiles (≥218.4, 218.4-281, and ≥281 microns) with serum hormone and SHBG levels. All models were adjusted for potential confounding factors. Results: The study population was 87.6% Caucasian with a mean age of 59.5 years, and >2/3 of the women were overweight or obese. Increasing percent free E2 was significantly associated with larger TDLU diameter (OR>2.53 vs. ≥1.99: 5.58, 95% CI: 1.48-21.07; p-trend=0.01), whereas increasing SHBG was associated with smaller TDLU diameter (OR>73nmol/L vs. ≥38 nmol/L: 0.19, 95% CI: 0.05-0.74; p-trend=0.02). Total E2 (p-trend=0.49), FSH (p-trend=0.85), and prolactin (p-trend=0.58) were not associated with TDLU diameter. We did not find significant relationships between the presence of TDLUs and any of the serum levels. Our observed relationships were not modified by age or other breast cancer risk factors, including BMI, years since menopause, and history of bilateral oophorectomy. Conclusion: Based on limited data, these findings suggest that sex hormones may play a role in TDLU involution. Thus, TDLU involution may represent one mechanism by which hormone levels influence breast cancer risk. TDLU measurements may therefore be helpful in assessing breast cancer risk as a marker of breast tissue aging. Analysis of the relationships between hormones and TDLU involution among premenopausal women may provide additional insights into early events in breast carcinogenesis. Citation Format: Zeina G. Khodr, Mark E. Sherman, Ruth M. Pfeiffer, Gretchen L. Gierach, Louise A. Brinton, Roni T. Falk, Deesha Patel, Laura Linville, Susan E. Clare, Daniel Visscher, Carolyn Mies, Stephen Hewitt, Anna Maria Storniolo, Jonine D. Figueroa. Endogenous hormone levels and lobular involution of the breast among healthy, postmenopausal women in the Komen Tissue Bank Study. [abstract]. In: Proceedings of the Eleventh Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2012 Oct 16-19; Anaheim, CA. Philadelphia (PA): AACR; Cancer Prev Res 2012;5(11 Suppl):Abstract nr B89.