Analysis of ecDNA and Heterogeneity in SUM159PT as a Model System for TNBC

Triple negative breast cancer (TNBC) is the most lethal subtype of breast cancer, exhibiting frequent metastasis, strong resistance to traditional therapies, and rapid recurrence. Heterogeneity contributes to the aggressive nature of this disease and presents significant obstacles to successful treatment. Extrachromosomal DNA (ecDNA) elements are acentromeric episomes that randomly segregate into daughter cells and are known drivers of heterogeneity and gene amplification in human cancers. Currently, few cancer cell lines have been classified for the presence of ecDNA. Prediction software suggests that many TNBC cell lines contain ecDNA, but due to limited efficacy of these programs, experimental validation is still needed. Here we confirmed the presence of ecDNA that encodes the oncogenic transcription factor MYC in SUM159PT, a TNBC cell line. To further characterize the cell line and examine how ecDNA impacts drug response, SUM159 cells were treated with JQ1, a small molecule inhibitor which prevents activation of MYC by inhibiting BRD4, and imaged using Fluorescence in situ Hybridization (FISH). Integrating real time cell analysis (RTCA) growth monitoring, quantitative polymerase chain reaction (qPCR), and FISH data, we demonstrated JQ1 treatment resulted in decreased cell growth and expression of target oncogenes which was associated with changes in ecDNA. The total number of ecDNA, particularly MYC positive ecDNA, significantly decreased after JQ1 treatment while the number of ecDNA that amplified other genes did not significantly change compared to the control condition. These results provide useful insight into how ecDNA may contribute to heterogeneity in cancers like TNBC.