A Longitudinal Study of the Association between Mammographic Density and Gene Expression in Normal Breast Tissue

Abstract

High mammographic density (MD) is associated with a 4-6 times increase in breast cancer risk. For post-menopausal women, MD often decreases over time, but little is known about the underlying biological mechanisms. MD reflects breast tissue composition, and may be associated with microenvironment subtypes previously identified in tumor-adjacent normal tissue. Currently, these subtypes have not been explored in normal breast tissue. We obtained biopsies from breasts of healthy women at two different time points several years apart and performed microarray gene expression analysis. At time point 1, 65 samples with both MD and gene expression were available. At time point 2, gene expression and MD data were available from 17 women, of which 11 also had gene expression data available from the first time point. We validated findings from our previous study; negative correlation between RBL1 and MD in post-menopausal women, indicating involvement of the TGFβ pathway. We also found that breast tissue samples from women with a large decrease in MD sustained higher expression of genes in the histone family H4. In addition, we explored the previously defined active and inactive microenvironment subtypes and demonstrated that normal breast samples of the active subtype had characteristics similar to the claudin-low breast cancer subtype. Breast biopsies from healthy women are challenging to obtain, but despite a limited sample size, we have identified possible mechanisms relevant for changes in breast biology and MD over time that may be of importance for breast cancer risk and tumor initiation.

The use of patient‐derived breast tissue explants to study macrophage polarization and the effects of environmental chemical exposure

“…46 Furthermore, the "active" signature can change over time and location in nonaffected breast tissue, suggestive of an immune-or stromal-related change. 47 Our results suggest that perhaps IL-4 and IL-13 either directly or more likely indirectly through polarization of macrophages toward M2 alter the microenvironment to cause the "active" phenotype. Stromal responses likely perpetuate the progression of breast cancer, especially if a premalignant lesion is present, and it is therefore imperative to take advantage of complex model systems that allow us to investigate how varying stimuli can modify both stromal and epithelial responses.…”

“…More recent publications have revealed that even normal tissue from women without cancer can be classified into “active” versus “inactive” and correlates with adiposity 46 . Furthermore, the “active” signature can change over time and location in nonaffected breast tissue, suggestive of an immune‐ or stromal‐related change 47 . Our results suggest that perhaps IL‐4 and IL‐13 either directly or more likely indirectly through polarization of macrophages toward M2 alter the microenvironment to cause the “active” phenotype.…”

Re-definition of claudin-low as a breast cancer phenotype

“…Claudin-low tumors had high levels of non-tumor cell infiltration, and there was a lower prevalence of claudin-low tumors in the cohorts with a cut-off for tumor cellularity. It is also known that EMT-like gene expression features in tumors are similar to the gene expression characteristics of stromal tissue 29 , and a subset of normal breast tissue samples show marked similarities to claudin-low-like gene expression patterns 30,31 . In the context of these observations, it is pertinent to ask: How much of the claudin-low phenotype is a result of stromal infiltration, and could the claudin-low phenotype simply be an artifact of stromal infiltration?…”

“…3). This can be related to the marked stromal and immune infiltration in claudin-low tumors 12 , and to the partial overlap in gene expression features between stromal tissue and tumors with an EMT phenotype [29][30][31] . Given these challenges which arose from the unbiased approach used by Prat et al 12 , and the progress made in the understanding of claudin-low tumors 6,14,15,32 , we chose to explore a biased approach to identifying claudin-low tumors.…”

Associations of breast cancer related exposures and gene expression profiles in normal breast tissue—The Norwegian Women and Cancer normal breast tissue study

“…Such tissue samples often show more histological abnormalities when compared to tissue obtained from healthy donors, 9–11 and using different sources of control breast tissue in different studies make comparisons between studies difficult. In addition, most studies on gene expression profiles were generated from a small set of samples that were likely not representative of the general population 8,12 . Finally, a better understanding of the natural variability of gene expression in normal breast tissue would represent a significant step forward in our understanding of early disease‐related mechanisms.…”