Elevated miR-29c-5p Expression in Nipple Aspirate Fluid Is Associated with Extremely High Mammographic Breast Density

SIMPLE SUMMARY: High mammographic density is a known risk factor for breast cancer. However, the underlying mechanisms of high mammographic density development and breast cancer are unknown. MicroRNAs are potential biomarkers indicative of carcinogenesis and can be assessed in nipple aspirate fluid. We used nipple aspirate fluid from women with very low and extremely high mammographic density to examine differences in expression of multiple miRNAs between both extremes in the spectrum of mammographic density. We found that hsa-miR-29c-5p was upregulated in an extremely high mammographic density context and potential targets were identified that might provide clues of the relationship between high mammographic density and breast cancer risk. Understanding the relationship between high mammographic density and breast cancer is of great value for early breast cancer diagnosis and treatment. With our research we provide new insight into this relationship and further research could determine the effects of dysregulated hsa-miR-29c-5p on the identified candidate targets. ABSTRACT: High mammographic density (MD) is associated with an increased risk of breast cancer, however the underlying mechanisms are largely unknown. This research aimed to identify microRNAs (miRNAs) that play a role in the development of extremely dense breast tissue. In the discovery phase, 754 human mature miRNAs were profiled in 21 extremely high MD- and 20 very low MD-derived nipple aspirate fluid (NAF) samples from healthy women. In the validation phase, candidate miRNAs were assessed in a cohort of 89 extremely high MD and 81 very low MD NAF samples from healthy women. Independent predictors of either extremely high MD or miRNA expression were identified by logistic regression and linear regression analysis, respectively. mRNA targets and pathways were identified through miRTarBase, TargetScan, and PANTHER pathway analysis. Statistical analysis identified four differentially expressed miRNAs during the discovery phase. During the validation, linear regression (p = 0.029; fold change = 2.10) and logistic regression (p = 0.048; odds ratio = 1.38) showed that hsa-miR-29c-5p was upregulated in extremely high MD-derived NAF. Identified candidate mRNA targets of hsa-miR-29c-5p are CFLAR, DNMT3A, and PTEN. Further validation and exploration of targets and downstream pathways of has-miR-29c-5p will provide better insight into the processes involved in the development of high MD and in the associated increased risk of breast cancer.