Breast cancer comprises about 30% of all new female cancers each year and is the most common malignant cancer in women in the United States. Breast cancer cell lines have been harnessed for many years as a foundation for in vitro analytic studies to understand the use of cancer prevention and therapy. There has yet to be a compilation of works to analyze the pitfalls, novel discoveries, and essential techniques for breast cancer cell line studies in a scientific context. In this article, we review the history of breast cancer cell lines and their origins, as well as analyze the molecular pathways that pharmaceutical drugs apply to breast cancer cell lines in vitro and in vivo. Controversies regarding the origins of certain breast cancer cell lines, the benefits of utilizing Patient-Derived Xenograft (PDX) versus Cell-Derived Xenograft (CDX), and 2D versus 3D cell culturing techniques will be analyzed. Novel outcomes from epigenetic discovery with dietary compound usage are also discussed. This review is intended to create a foundational tool that will aid investigators when choosing a breast cancer cell line to use in multiple expanding areas such as epigenetic discovery, xenograft experimentation, and cancer prevention, among other areas.

Background: Breast cancer (BC) is the second most common cancer among women in the United States. It has been estimated that one in eight women will be diagnosed with breast cancer in her lifetime. Various BC risk factors, such as age, physical inactivity, and smoking, play a substantial role in BC occurrence and development. Early life dietary intervention with plant-based bioactive compounds has been studied for its potential role in BC prevention. Sulforaphane (SFN), an isothiocyanate, is an antioxidant and anti-inflammatory agent extracted from broccoli sprouts (BSp) and other plants. Dietary supplementation of SFN suppresses tumor growth by inducing protective epigenetic changes and inhibiting cancer cell proliferation. Inulin, as a dietary fiber, has been studied for alleviating GI discomfort and weight loss by promoting the growth of beneficial bacteria in the gut. Objective: Early-life combinatorial treatment with both phytochemical SFN and potential prebiotic agent inulin at lower and safer dosages may confer more efficacious and beneficial effects in BC prevention. Methods: Transgenic mice representing estrogen receptor-negative BC were fed 26% (w/w) BSp and 2% (w/v) inulin supplemented in food and water, respectively. Results: The combinatorial treatment inhibited tumor growth, increased tumor onset latency, and synergistically reduced tumor weight. Gut microbial composition was analyzed between groups, where Ruminococcus, Muribaculaceae, and Faecalibaculum significantly increased, while Blautia, Turicibacter, and Clostridium sensu stricto 1 significantly decreased in the combinatorial group compared with the control group. Furthermore, combinatorial treatment induced a protective epigenetic effect by inhibiting histone deacetylases (HDACs) and DNA methyltransferases (DNMTs). Intermediates in the AKT/PI3K/MTOR pathway were significantly suppressed by the combinatorial treatment, including PI3K p85, p-AKT, p-PI3K p55, MTOR, and NF-κB. Cell cycle arrest and programmed cell death were induced by the combinatorial treatment via elevating the expression of cleaved-caspase 3 and 7 and inhibiting the expressions of CDK2 and CDK4, respectively. Orally administering F. rodentium attenuated tumor growth and induced apoptosis in a syngeneic triple-negative breast cancer (TNBC) mouse model. Conclusions: Overall, the findings suggest that early-life dietary combinatorial treatment contributed to BC prevention and may be a potential epigenetic therapy that serves as an adjunct to other traditional neoadjuvant therapies.

Background/Objectives: There is a clear need for more options to control the progression of breast cancer and prevent the occurrence of breast cancer in minority populations that have a higher rate of mortality due to triple-negative breast cancer (TNBC) subtypes. Prevalent nutraceuticals such as Ashwagandha (also known as the Indian Winter Cherry) have anti-inflammatory and apoptotic capabilities, as well as the ability to inhibit cancer growth. The purpose of this study is to analyze the novel combination of withaferin A (derived from the Indian Winter Cherry and known to have histone deacetylase inhibition capabilities) and sodium butyrate (a short-chain fatty acid produced from the gut microbiome and known to have DNA methyltransferase inhibition capabilities) treatment on breast cancer-derived cell lines. There is a scientific gap of possible causality of decreasing breast cancer progression when treated with sodium butyrate and withaferin A. Methods: Two in vitro cell viability assays were utilized consisting of [MTT (4,5 Dimethylthiazol-2-yl)] and the neutral red assay to analyze the impact of treatment of compounds alone and in combination on breast cancer cells for 72 h. The Highest Single Agent (HSA) combination analysis was utilized to derive combination indexes for our breast cancer cell types. Protein and gene expression was investigated for Class 1 histone deacetylases, de novo DNA methyltransferase, the p65 subunit of NF-κB, and NFκB1. Lastly, DNA methyltransferase enzymatic activity was analyzed via the Epigentek DNMT Activity/Inhibition ELISA Easy Kit. Results: Through the cell viability assay [MTT (4,5 Dimethylthiazol-2-yl)], MCF−7, MDA−MB−231, and MDA−MB−157 cells were found to have a decrease in cell viability due to combinatorial treatment with withaferin A and sodium butyrate. Western blot results depicted a decrease in protein expression levels for DNA methyltransferases due to the administration of 2.5 mM sodium butyrate and 0.2 µM withaferin A alone and in combination for breast cancer cell lines MCF−7, MDA-MB-231, and MDA−MB−157. Additionally, the combination of these two components have successfully inhibited the progression of the NFκB1 gene within analysis through the quantitative polymerase chain reaction (qPCR). Conclusions: The novel combination of withaferin A and sodium butyrate have markedly reduced the progression of breast cancer-derived cell lines for cell viability, epigenetic DNMT gene expression, as well as inhibiting NFκB1 signaling on the gene expression level.