Eaves, Connie J.

Abstract A problematic feature of many human cancers is a lack of understanding of mechanisms controlling organ-specific patterns of metastasis, despite recent progress in identifying many mutations and transcriptional programs shown to confer this potential. To address this gap, we developed a methodology that enables different aspects of the metastatic process to be comprehensively characterized at a clonal resolution. Our approach exploits the application of a computational pipeline to analyze and visualize clonal data obtained from transplant experiments in which a cellular DNA barcoding strategy is used to distinguish the separate clonal contributions of two or more competing cell populations. To illustrate the power of this methodology, we demonstrate its ability to discriminate the metastatic behavior in immunodeficient mice of a well-established human metastatic cancer cell line and its co-transplanted LRRC15 knockdown derivative. We also show how the use of machine learning to quantify clone-initiating cell (CIC) numbers and their subsequent metastatic progeny generated in different sites can reveal previously unknown relationships between different cellular genotypes and their initial sites of implantation with their subsequent respective dissemination patterns. These findings underscore the potential of such combined genomic and computational methodologies to identify new clonally-relevant drivers of site-specific patterns of metastasis.

Macrophages help mediate hormone-controlled changes in the mouse mammary gland

Abstract Background: Accumulating epidemiological studies positively associate healthy body mass index (BMI) and higher physical activity with lowered risk of postmenopausal breast cancer. Many of the proposed biomarkers underlying these associations, such as insulin, inflammatory markers, steroid hormones, and adipokines, are produced or regulated by adipose tissue. However, the biological impact of lifestyle factors at the level of the breast tissue, particularly adipose tissue in the breast, is unclear and may play a role in the etiology of breast cancer. Methods: Using a cross-sectional design, we examined the impact of demographic and lifestyle factors on histological features of breast adipose tissue. Women (age) undergoing reduction mammoplasty surgery were consented to collection of their cancer-free breast tissue at the time of surgery. An adipose rich section of the sample was dissected under sterile conditions and formalin fixed. Samples were sectioned at 7 µm thickness and stained with hematoxylin and eosin. The stained sections were imaged and mean adipocyte size was determined as diameter (µm) from 3 randomly selected areas at 10X magnification and each sample was scored by two independent assessors using Image J software. BMI status documented at the time of surgery was abstracted form medical records. A subset of women completed an additional visit where percent total body fat was measured by air displacement plethysmography and aerobic fitness (VO2peak) was measured by a maximal graded exercise test with expired gas collection. The association between adipocyte size and demographic/lifestyle factors was examined using multivariate linear regression adjusted for age and menopausal status. Results: Participants (n=42) were primarily Caucasian (77%) and pre/peri-menopausal (62%), with a mean age of 44.6±12.9 years (range 19-70) and mean BMI of 27.7±5.1 kg/m2 (range 19.1-37.4). Adipocyte size was associated with BMI (per 5 kg/m2 increase, β = 5.6±2.15, p=0.01) and number of pregnancies (β = 6.04±2.34, p=0.02), but not age (β = 0.1±0.3, p=0.81). In the subset of participants who completed an additional study visit (n=9), there was a trend towards an inverse association between adipocyte size and aerobic fitness (per 2 ml/kg/min of O2 consumption, β = -3.54±1.52, p=0.07) and percent body fat (per 2% increase, β = 1.94±1.28, p=0.17) in unadjusted analyses. Conclusion: Higher BMI is associated with larger adipocyte size in adipose tissue taken from the breast, suggesting a biological role for body composition in influencing gross histological features of adipocytes and its behavior that would impact the health of the mammary gland. A better understanding of the biological mechanisms underlying the observed epidemiological associations are needed to guide the development of intervention strategies and the most effective public health messages for breast cancer prevention. Citation Format: Kristin L. Campbell, Nagarajan Kannan, Sarah E. Neil-Sztramko, Connie J. Eaves, Jonathan P. Little, Ilona Csizmadi, David Zhu, Sarah Sayyari, Kelcey Bland, James D. Johnson. Influence of lifestyle factors on adipocyte size in human breast tissue [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 4258. doi:10.1158/1538-7445.AM2017-4258

Abstract BRCA1 deficiency alters the relative proportions of progenitor cells in preneoplastic mammary tissue, and typically associates with breast cancers characterized by genomic instability and a basal-like cell phenotype. Oriented division of progenitor cells is one mechanism these cells use to maintain tissue homeostasis, and to suppress tumor formation. We now show that shRNA-mediated reduction of BRCA1 levels in non-tumorigenic and immortalized or freshly isolated, normal human mammary cells alters their plane of division with graded consequences that include the induction of aneuploidy in progeny cells, perturbation of polarity in spheroid cultures, and inhibition of clonal growth with favored expression of basal features. We also demonstrate a requirement for BRCA1 in establishing cortical asymmetry of NUMA-dynein complexes. Mutation of a single BRCA1 allele (BRCA1 185delAG/+) altered the division axis of isolated cells but their deficient spindle positioning was supervised by CDH1-positive adherens, which sustained oriented divisions and produced colonies with luminal features. These findings reveal a previously unrecognized consequence of mutant BRCA1 on the cell division axis, post-mitotic integrity and phenotype control in normal human mammary epithelial cells. Note: This abstract was not presented at the meeting. Citation Format: Zhengcheng He, Oksana Nemirovsky, Nagarajan Kannan, Connie Eaves, Christopher A. Maxwell. BRCA1 controls the cell division axis and governs ploidy and phenotype in human mammary cells [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 3459. doi:10.1158/1538-7445.AM2017-3459