Radisky, Derek C.

Abstract Estrogen receptor (ER) β is highly expressed in normal breast epithelium and a putative tumor suppressor. Atypical hyperplasia substantially increases breast cancer risk, but identification of biomarkers to further improve risk stratification is needed. We evaluated ERβ expression in breast tissues from women with atypical hyperplasia and association with subsequent breast cancer risk. ERβ expression was examined by immunohistochemistry in a well-characterized 171-women cohort with atypical hyperplasia diagnosed 1967–1991. Nuclear ERβ percent and intensity was scored in the atypia and adjacent normal lobules. An ERβ sum score (percent + intensity) was calculated and grouped as low, moderate, or high. Competing risks regression was used to assess associations of ERβ expression with breast cancer risk. After 15-year median follow-up, 36 women developed breast cancer. ERβ expression was lower in atypia lobules in than normal lobules, by percent staining and intensity (both P < 0.001). Higher ERβ expression in the atypia or normal lobules, evaluated by percent staining, intensity or sum score, decreased the risk of subsequent breast cancer by 2-fold (P = 0.04) and 2.5-fold (P = 0.006). High normal lobule ERβ expression conferred the strongest protective effect in premenopausal women: the 20-year cumulative incidence of breast cancer was 0% for women younger than 45 years with high versus 31% for low–moderate ERβ expression (P = 0.0008). High ERβ expression was associated with a significantly decreased risk of breast cancer in women with atypical hyperplasia. These data suggest that ERβ may be a useful biomarker for risk stratification and a novel therapeutic target for breast cancer risk reduction. Cancer Prev Res; 8(11); 1084–92. ©2015 AACR.

Abstract Background Intra-tumor heterogeneity is the driving force behind disease progression and metastasis, drug resistance, and relapse in cancer patients. Defining personalized treatment regimens and increasing the arsenal of targeted therapies available will provide the best chance of prolonging survival and finding curative treatments. Recent work by our group and others has shed light on aberrant de novo lipogenesis as a feature of many cancers including kidney, breast, and lung cancer. Of the molecules involved in fatty acid metabolism, stearoyl-CoA desaturase 1 (SCD1) is frequently over-expressed. Targeted inhibition of SCD1 demonstrates a loss of tumor cell viability in a variety of cancer models. While there currently are specific small molecule inhibitors for SCD1 none are under investigation as anti-cancer therapeutics. In order to evaluate the efficacy of SCD1 inhibitors against ccRCC in both a preclinical and eventually a clinical setting we sought to generate unique compounds and test their biological efficacy. Methods Utilizing an innovative in silico approach, we designed new inhibitors for SCD1 via a scaffold hopping approach while searching for different “core” scaffolds. Critical interaction moieties from the chemical R-groups were held fixed, while generated cores were rapidly scanned for best-fit criteria (shape, docking, pharmacophore fit (QSAR)) into our Z-matrix. Following de novo compound generation and synthesis, we implemented iterative analog generation for the top hits. Using this method and diversity of chemical space, we pursued three divergent species of compounds into the nanomolar regime. Biological testing of the novel compounds included high-throughput proliferative-based screening, oleic acid rescue, and evaluation of the endoplasmic stress response. Enzymatic inhibition of SCD1 was evaluated by LC-MS. Preclinical pharmacokinetics was established using a range of doses administered intravenously and orally. Anti-tumor response was evaluated in an ectopic model of renal carcinoma, both as a single agent and in combination with SOC. Results A cohort of potential unique small molecule inhibitors against SCD1 were designed and synthesized. Of these, a select group of compounds demonstrated potent and selective enzyme inhibition and reduced tumor growth in in vitro and in vivo models of ccRCC. These novel inhibitors reliably reproduced activation of ER stress markers, authenticating this biological response as a harbinger for successful abrogation of SCD1 activity. Conclusions We have established an effective, expedited, and economical format for in silico modeling of novel agents based on a scaffold-hopping methodology. Our findings provide compelling evidence supporting the therapeutic benefit of SCD1 inhibitors for clinical use as broad-spectrum anti-tumor agents alone or in combination with SOC. Citation Format: Christina A. Von Roemeling, Thomas R. Caulfield, Derek C. Radisky, Ilah Bok, Laura A. Marlow, James Miller, Mojda Sidiqi, Anthony B. Pinkerton, Winston W. Tan, Amy L. Lane, Han W. Tun, John A. Copland. Accelerated drug discovery platform yields synthesis of novel stearoyl-CoA desaturase 1 inhibitors that demonstrate anti-tumor efficacy in several models of aggressive cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4459. doi:10.1158/1538-7445.AM2015-4459

Abstract Purpose: Breast biopsies with benign findings (benign breast disease [BBD]) outnumber biopsies revealing in-situ or invasive cancer 4:1, providing an opportunity to identify histopathological features that predict breast cancer risk. Greater degrees of involution of terminal duct lobular units (TDLUs), the structures within the breast that produce milk and the primary source of breast cancer precursors, have been inversely associated with breast cancer risk among women with BBD. We recently developed measures to quantitate levels of TDLU involution, which demonstrate high inter/intra pathologist reproducibility. Here we assessed whether TDLU counts/100mm2 and median TDLU span (microns), two measures inversely related to degree of TDLU involution, are associated with subsequent breast cancer risk among women with BBD. Methods: From the Mayo BBD cohort (n = 9,376), we evaluated benign biopsies from 99 women who later developed breast cancer (cases) and 145 age-matched controls who did not develop breast cancer. Digitized images of biopsy sections were reviewed to enumerate TDLUs/mm2 and measure median TDLU span (microns) for up to ten normal TDLUs. Breast cancer risk factors were available from questionnaires or medical records. To assess associations with breast cancer risk, subjects were categorized into quartile levels of TDLU counts and median TDLU spans, based on data from controls. Odds ratios (ORs) and 95% confidence intervals (CI) were estimated using multivariable conditional logistic regression models, adjusted for histologic type of BBD, family history of breast cancer and menopausal hormone use. Results: Compared to controls, women who developed breast cancer had higher median number of TDLU counts/100mm2 (28 vs 20, p = 0.03) and larger TDLU spans (300 vs 267 microns, p = 0.14). In multivariable models higher TDLU counts (quartile4 vs. quartile1, OR = 2.44, 95%CI = 0.96-6.19, ptrend = 0.02) and larger TDLU span measures (quartile4 vs. quartile1, OR = 2.83, 95%CI = 1.13-7.06, ptrend = 0.03) were associated with subsequent diagnosis of breast cancer. Combinatorial metrics of TDLU counts with median TDLU span measures identified women at higher risk; specifically, women above the median for both TDLU span and TDLU counts had an OR = 3.75 (95%CI = 1.40-10.00, ptrend = 0.008), compared with women below the median for TDLU span and TDLU counts. Conclusion: These data show that lack of TDLU involution, as measured by increased persistence of TDLU counts, larger median TDLU spans and cross-classification using these measures was associated with increased breast cancer risk among women with BBD, extending prior work in this cohort based on other TDLU involution metrics. Future studies to identify determinants of TDLU involution, its association with breast cancer risk and its potential as an intermediate endpoint in prevention studies warrant consideration. Citation Format: Jonine D. Figueroa, Ruth Pfeiffer, Maya Palakal, Amy C. Degnim, Derek Radisky, Lynn C. Hartmann, Marlene Frost, Melody L. Stallings Mann, Louise A. Brinton, Daphne Papathomas, Daniel Visscher, Mark E. Sherman. Standardized measures of lobular involution and subsequent breast cancer risk among women with benign breast disease. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4682. doi:10.1158/1538-7445.AM2015-4682

Abstract Introduction: The role of macrophages in the tumor microenvironment is of increasing interest for tumor progression and invasion. Here we investigated the difference in CD68+ cell density in tissue samples from women with Benign Breast Disease (BBD) and women with no clinical breast disease from the Komen Tissue Bank (KTB) at Indiana University. Method: Archived breast tissue samples from women with BBD and no clinical breast disease were obtained for this age-matched case-control study. BBD samples included cases [later got Breast Cancer (BC)] and controls (did not get BC). Samples were characterized via H&E stain for epithelial proliferation [normal (N), fibrocystic non-proliferative (NP), proliferative (P), or atypia (A)]. CD68+ cell density was obtained by digital image quantitation on a per lobule basis (Total CD68+ cells/mm2). Statistical analysis was performed with mixed effects regression. Results: Among 276 women (median age 53, range 35-79) comprising 92 age-matched triplets, 2508 lobules were evaluated [789 KTB, 839 BBD case, 880 BBD control]. 1407 (56%) lobules were normal and 1101 (44%) were fibrocystic {NP [613 (56%)], P [448 (41%)] and A [40 (4%)]}. Out of 2508 lobules analyzed, 2362 (94%) had CD68+ cells, while 146 (6%) showed zero CD68+ cells [19 (2%) BBD cases, 20 (2%) BBD controls, 107 (14%) KTB, p<0.0001]. KTB samples showed significantly lower (p<0.0001) CD68+ density (median 136 cells/mm2) as compared to BBD cases (median 313 cells/mm2) and BBD controls (median 299 cells/mm2). (See Table) For each lobule type, KTB samples had lower CD68+ density compared with the same lobule type in BBD samples (p<0.0001). Conclusion: CD68+ cells are more frequent in breast lobules from women with BBD compared to normal donors. This finding in premalignant breast tissues is consistent with inflammation as a driver of tumorigenesis. BBD case (n = 92 patients)BBD control (n = 92 patients)KTB(n = 92 patients)n lobulesMedian (IQR)n lobulesMedian (IQR)n lobulesMedian (IQR)Overall839313 (165 - 538)880299 (164 - 527)789136 (55 - 272)By Lobule TypeNormal344310 (177 - 557)434331 (177 - 567)629128 (55 - 255)All non-normal/fibrocystic lobules combined495314 (157 - 527)446274 (152 - 478)160159 (58 - 393)Fibrocystic non-proliferative231319 (153 - 543)263274 (140 - 496)119163 (71 - 359)Fibrocystic proliferative246295 (153 - 496)169277 (153 - 478)33128 (30 - 385)Atypia18484 (313 - 739)14260 (223 - 375)8216 (0 - 493)IQR - Interquartile range (25th percentile - 75th percentile) Citation Format: Muhammad A. Arshad, Daniel W. Visscher, Tanya L. Hoskin, Rushin D. Brahmbhatt, Alvaro Pena Jimenez, Melody L. Stallings Mann, Erin E. Miller, Linda M. Murphy, Jodi M. Carter, Stacey J. Winham, Keith L. Knutson, Derek C. Radisky, Amy C. Degnim. CD68+ immune cells show different infiltration patterns in tissue samples from women with no clinical breast disease and those who have benign breast disease. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2364. doi:10.1158/1538-7445.AM2015-2364

Abstract Introduction: Increased levels of circulating estrogens are associated with elevated risks of postmenopausal breast cancer. Previously, we showed that levels of sex steroid hormones in breast adipose tissue are positively correlated with circulating hormones. Further evaluation of hormone levels in breast tissues may help to explain the relationship between estrogens and breast cancer risk. Specifically, data suggest that focal breast lesions consisting of macrophages encircling dying adipocytes, referred to as “crown-like structures (CLS)”, are associated with increased aromatase activity, which could increase conversion of androgens to estrogens locally in the breast. Therefore, we assessed whether the presence of CLS affects the correlations between blood and breast adipose tissue hormone levels. Methods: Participants included 84 postmenopausal women with invasive breast cancer in the Polish Breast Cancer Study, a population based case-control study conducted from 2000 to 2003. We previously reported sex steroid hormone levels in cancer-free breast adipose tissue removed as part of breast excisions performed for cancer. Here, using immunohistochemical stains, we evaluated the number of CD68 positive macrophages and the presence of full CLS (i.e., complete adipocyte encirclement) in benign breast tissue sections prepared from surgical pathology specimens containing breast cancer. The relationship between the number of CD68 positive macrophages per unit area of fat and hormone levels was examined using Spearman rank correlations (rs). We assessed potential effect modification of the association between blood and tissue hormone levels by presence or absence of CLS by including an interaction term in linear regression models. Results: The number of CD68 positive macrophages per unit area of fat was not significantly correlated with levels of estrone, estradiol, androstenedione or testosterone measured in blood (rs = 0.001, -0.04, -0.07, and -0.09, respectively; p>0.05) or breast adipose tissue (rs = -0.15, 0.06, -0.06, and -0.08, respectively; p>0.05). Correlations between serum and adipose tissue hormones did not statistically significantly differ between women with CLS (n = 30) compared to those without CLS (n = 54). CLS were found more frequently in breast adipose tissue from overweight (i.e., women with a body mass index (BMI) between 25 and 29.9) (43%) and obese women (BMI >30) (30%) versus normal weight women, with (BMI between 18.5 and 24.9), (27%); (p = 0.04, χ2 test). The number of CD68 positive macrophages per unit area of fat was not significantly related to BMI (p = 0.31). Conclusion: Among postmenopausal women with invasive breast cancer, the presence of CLS was more common among overweight and obese women, but CLS did not influence the correlation between circulating and breast adipose tissue sex steroid hormone levels. Citation Format: Maeve Mullooly, Hannah P. Yang, Roni T. Falk, Sarah Nyante, Renata Cora, Ruth M. Pfeiffer, Derek C. Radisky, Daniel W. Visscher, Lynn C. Hartmann, Amy C. Degnim, Frank Z. Stanczyk, Jonine F. Figueroa, Montserrat Garcia-Closas, Jolanta Lissowska, Melissa T. Troester, Louise A. Brinton, Mark E. Sherman, Gretchen L. Gierach. Investigation of the relationship between crown-like structures and adipose tissue hormone levels among postmenopausal women with breast cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2767. doi:10.1158/1538-7445.AM2015-2767

Significance During branching morphogenesis of the developing lung, it is generally thought that a spatial template of biochemical cues determines the airway branching pattern. Here, however, we demonstrate that physical mechanisms can control the pattern of airway branching. Using a combination of 3D culture experiments and theoretical modeling, we show that a growth-induced physical instability initiates the formation of new epithelial branches. Tuning epithelial growth rate controls the dominant wavelength of the instability, and thereby the branching pattern. These findings emphasize the role of mechanical forces during morphogenesis and indicate that lung development is not a closed genetic system. Physical cues also regulate the spatially patterned cell behaviors that underlie organ assembly in the embryo.

Abstract Although the association between benign breast disease (BBD) and risk of subsequent breast cancer is well-established, no previous studies have compared characteristics of breast cancers in women with a history of BBD to characteristics of cancers in the general population. In this study, we compared the characteristics of 109 breast cancers that developed in a population of African-American women with a history of benign breast disease (BBD population) to 10,601 breast cancers that developed in a general population of African-American women whose cancers were recorded by the Metropolitan Detroit Cancer Surveillance System (MDCSS population). The populations were compared on demographic characteristics such as age, marital status, and previous cancers, as well as clinical characteristics such as tumor hormone receptor status, grade, stage, behavior, size, and treatment. Overall and breast cancer-specific survival were also compared between populations. Women in the BBD population developed lower grade, earlier stage cancers that were more likely to be hormone receptor-positive and less likely to exhibit invasive behavior. Women in the BBD population were also significantly less likely to die from breast cancer after ten years of follow-up. These results suggest that although women with a history of BBD are at greater breast cancer risk, their breast cancers may be more likely to be detected early, leading to better breast cancer-specific survival. Citation Format: Susanna D. Mitro, Rouba Ali-Fehmi, Sudeshna Bandyopadhyay, Baraa Alosh, Bassam Albashiti, Quratulain Ahmed, Derek C. Radisky, Marlene H. Frost, Amy C. Degnim, Julie J. Ruterbusch, Michele L. Cote. Clinical characteristics of breast cancers in African American women with benign breast disease: A comparison to the Surveillance, Epidemiology and End Results Program. [abstract]. In: Proceedings of the Sixth AACR Conference: The Science of Cancer Health Disparities; Dec 6–9, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2014;23(11 Suppl):Abstract nr B39. doi:10.1158/1538-7755.DISP13-B39

Abstract Introduction: Histologic characteristics of nonmalignant breast tissues (epithelial proliferation and involution) are associated with future breast cancer (BC) risk. We evaluated immune infiltration (lobulitis) in benign breast lobules and its association with features of BC risk. Methods: We analyzed archived nonmalignant breast tissue samples from 81 women in age-matched groups of varying cancer risk: 27 with benign breast disease (BBD) who later developed BC (cases), 27 with BBD without subsequent BC (controls), and 27 normal women without clinical breast disease from the Komen Tissue Bank at Indiana University (KTB). Up to 10 lobules in each sample were characterized by H&E for lobulitis, involution (none, partial, complete), and fibrocystic (FC) status (normal, nonproliferative, proliferative). Lobulitis was defined by H&E staining as lymphocyte nuclei in the intralobular stroma at low magnification and between adjacent acini at high magnification. Data was analyzed by mixed effects logistic regression. Results: Median age was 57 years (range 37-70). Among 81 breast tissue samples, 750 lobules were evaluated: 249 in BBD Cases, 265 in BBD Controls, and 236 in KTB donors. 56% of lobules were normal and 44% were FC, with 87% of FC lobules occurring in BBD samples. Lobulitis was present in 44% of lobules and varied according to subject age and individual lobule features. Younger women were more likely to have lobulitis (p=0.002); among women <50 the median proportion of lobules with lobulitis was 50%, versus 30% among women 50+. Lobulitis was less frequent in lobules with complete involution (37%) compared to partial (74%) or no involution (84%), p<0.0001. Lobulitis was common in normal lobules (52%) and decreased with increasing epithelial abnormality-39% of nonproliferative lobules and 27% of proliferative lobules (p<0.0001). In multivariate analysis including all 3 variables, all 3 remained significantly associated with lobulitis: age (p=0.03), proliferation (p<0.0001), and involution (p=0.0001). Lobules with lobulitis were less frequent in groups of women with increased breast cancer risk-55% of lobules in normal KTB samples, 43% in BBD controls, and 35% in BBD cases (p = 0.02). Conclusion: Lobulitis is common in normal and benign breast tissue lobules but varies with age, involution status, and epithelial abnormality. Further work is needed to understand the role of immune infiltrates in breast cancer risk. Citation Format: Rushin D. Brahmbhatt, Daniel W. Visscher, Tanya L. Hoskin, Derek C. Radisky, Linda M. Murphy, Melody L. Stallings Mann, Erin Miller, Vernon S. Pankratz, Lynn C. Hartmann, Marlene H. Frost, Amy C. Degnim. Immune infiltration of normal and benign breast lobules varies in breast tissues based on cancer risk. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1652. doi:10.1158/1538-7445.AM2014-1652

Abstract Background: Mammographic breast density (MBD) reflects the proportion of fibroglandular tissue in the breast, and is an established risk factor for breast cancer. Benign breast disease (BBD) is also a known risk factor for breast cancer. We previously showed an association between lobular involution (physiologic atrophy) and MBD in a large cohort study of women with BBD. We performed a comprehensive evaluation of the association of histologic features of BBD with MBD. Methods: The Mayo BBD cohort includes women, 18 to 85 years, who were diagnosed with BBD at Mayo Clinic in Rochester, MN between 01/01/1967 and 12/31/1991. For this study, we included women who had a mammogram within 6 months of BBD diagnosis, and diagnosed with BBD between 1985 and 1991 when parenchymal pattern of breast density was clinically recorded. Risk factor information such as age, and body mass index (BMI) was collected from medical records. Parenchymal pattern, assessed clinically and previously used in multiple studies of MBD and risk, classified breast density based on extent and type of density, into four categories: N1 (non-dense, no ducts visible); P1 (ductal prominence occupying <25% breast); P2 (prominent ductal pattern occupying >25% of the breast; DY (homogenous plaque-like areas of density). All assessments of benign breast tissue were performed by expert breast pathologists blinded to both MBD and BBD reports. Histologic characteristics included overall impression (non-proliferative disease (NPD), proliferative disease without atypia (PDWA), and atypical hyperplasia (AH)), proportion of normal lobules that were involuted (no (0%), partial (1 to 74%), or complete (≥75%)), type of AH (ADH or ALH), ductal and lobular hyperplasia, calcifications, sclerosing adenosis, columnar alteration, cyst, fibroadenoma, marked fibrosis, intra-ductal papilloma, radial scar (number and size), duct ectasia, and mucocele like tumors. Associations of parenchymal pattern with BBD characteristics were examined using multicategorical nominal logistic regression models. We first examined associations after adjustment for age and BMI. All variables statistically significant (p<0.05) in these models were then simultaneously included in a fully-adjusted model. Results: Of 2,257 women in the study, 14% were <40 years old, 52.7% between 40- 59 years, and 32.9% were ≥60 years. In this sample, 55.2% of women had NPD, 36% had PDWA, and AH in 8.8%. MBD was classified as N1 in 21.8%, P1 in 12.6%, P2 in 23.7% and DY in 41.9%. Age- and BMI-adjusted analyses showed that there was an association of parenchymal pattern with overall impression, lobular involution, ductal hyperplasia, sclerosing adenosis, columnar alteration, cyst, duct ectasia and fibrosis. Multivariate analyses found that women of younger age (p<0.001) and with lower BMI (p<0.001), presence of fibrosis (p<0.001) and no involution (p=0.016) were more likely to have P2 or DY parenchymal pattern than women without those characteristics. Conclusion: Among women with benign breast disease, higher MBD is associated with younger age, lower BMI, fibrosis, and lack of lobular involution. These findings, concordant with other studies of MBD, provide insight into underlying mechanisms by which MBD and BBD contribute to breast cancer risk. Citation Format: Karthik Ghosh, Robert A Vierkant, Ryan D Frank, Daniel W Visscher, Vernon S Pankratz, Christopher G Scott, Derek C Radisky, Marlene H Frost, Lynn C Hartmann, Amy C Degnim, Celine M Vachon. Association between mammographic breast density and histologic features of benign breast disease [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P6-10-14.