ABSTRACT
As triple negative breast cancer (TNBC) lacks a specific target, exploration of abnormally expressed genes during the progression of TNBC is important for a better understanding of tumorigenesis and to find a specific target. We intended to figure out genes associated with TNBC, which can provide unique insights into gene dysregulation in TNBC while also pointing to new possible therapeutic targets for TNBC. A meta-analysis of multiple TNBC mRNA profiles was performed to identify consistently differentially expressed genes (CDGs). The pathways involved in modulating these genes were analyzed by MsigDB, and the interaction map was constructed. These CDGs were evaluated for their expression in cell lines, and drugs that could modulate the expression of CDGs were obtained using the connectivity map. CDGs were docked with doxorubicin and anethole, which is a phytocompound. The expression of selected CDGs was analyzed in MDA-MB-231 cells after treatment with doxorubicin and anethole. We found 45 CDGs, out of which 36 were upregulated and 9 were downregulated. MDA-MB-231 cell line was found to have high expression of CDGs, and drug that could modulate the expression of CDGs was doxorubicin. Docking results revealed that anethole and doxorubicin had good interaction with the CDGs especially with the genes AURKA, CDC6, DEPDC1, KIF23, KPNA2, MELK, CTNNB1, FLI1 and E2F1. Gene expression studies of the selected CDGs showed that the synergistic effect of anethole and doxorubicin effectively downregulated the expression. The CDGs identified from multiple cohorts have clinical significance and may be effectively exploited in the targeted therapy for TNBC.Communicated by Ramaswamy H. Sarma.
Subject(s)
Triple Negative Breast Neoplasms , Humans , Triple Negative Breast Neoplasms/drug therapy , Triple Negative Breast Neoplasms/genetics , Cell Line, Tumor , Doxorubicin/pharmacology , Doxorubicin/therapeutic use , Gene Expression Profiling , Gene Expression Regulation, Neoplastic , Transcriptome/genetics , Protein Serine-Threonine Kinases/metabolism , Neoplasm Proteins , GTPase-Activating Proteins/genetics , GTPase-Activating Proteins/metabolism , GTPase-Activating Proteins/therapeutic useABSTRACT
The heterogeneity and poor prognosis of triple-negative breast cancer (TNBC) have limited the treatment options and made clinical management challenging. This has nurtured a major effort to discover druggable molecular targets. Currently, chemotherapy is the primary treatment strategy for this disease. Doxorubicin is the most frequently used chemotherapeutic drug for TNBC and due to the fact that chemotherapeutic drugs have a lot of side effects, we evaluated the synergistic effect of the phytocompound anethole and doxorubicin. The cytotoxic effect of anethole in combination with doxorubicin on MDA-MB-231 cells was evaluated by various parameters, including apoptosis, cell cycle analysis, DNA damage, and cell proliferation. Furthermore, mitochondrial membranepotential (MMP), endoplasmic reticulum (ER) stress, and reactive oxygen species (ROS) levels were also evaluated in the cells treated with/without anethole and doxorubicin. Expression of the apoptotic proteins was evaluated by Western blot analysis. Initial evaluation of cytotoxicity of anethole on MDA-MB-231 cells demonstrated preferential suppression of cell proliferation and when treated along with doxorubicin it showed enhanced cytotoxicity with a synergistic effect. Cell cycle analysis revealed arrest at different stages of the cell cycle, such as sub G0-G1, G0-G1, S, and G2M in various treatment groups and apoptotic cell death was subsequently evident with propidium iodide (PI) staining. The synergistic action of anethole and doxorubicin effectively induced mitochondrial membrane potential loss, which, in turn, led to a burst of ROS production, which eventually produced unfolded protein response by damaging the ER. Synergistic anticancer effect was observed on exposure of MDA-MB-231 cells to anethole and doxorubicin in inducing cell death.
