thiostrepton and Triple-Negative-Breast-Neoplasms

FoxM1 transcription factor contributes to tumor metastasis and poor prognosis in many cancers including triple-negative breast cancer (TNBC). In this study, we examined the effects of FoxM1 inhibitor Thiostrepton (THIO) alone or in combination with MEK inhibitor Selumetinib (SEL) on metastatic parameters in vitro and in vivo.. Cell viability was determined by MTT assay. Immunoblotting and immunohistochemistry was used to assess metastasis-related protein expressions in 4T1 cells and its allograft tumor model in BALB/c mice. In vivo uPA activity was determined by enzymatic methods.. Both inhibitors were effective on the expressions of FoxM1, ERK, p-ERK, Twist, E-cadherin, and Vimentin alone or in combination in vitro. THIO significantly decreased 4T1 cell migration and changed the cell morphology from mesenchymal-like to epithelial-like structure. THIO was more effective than in combination with SEL in terms of metastatic protein expressions in vivo. THIO alone significantly inhibited mean tumor growth, decreased lung metastasis rate and tumor foci, however, no significant changes in these parameters were observed in the combined group. Immunohistochemically, FoxM1 expression intensity was decreased with THIO and its combination with SEL in the tumors.. This study suggests that inhibiting FoxM1 as a single target is more effective than combined treatment with MEK in theTNBC allograft model. The therapeutic efficacy of THIO should be investigated with further studies on appropriate drug delivery systems.

Topics: Animals; Cell Line, Tumor; Cell Proliferation; Forkhead Box Protein M1; Humans; Mice; Mitogen-Activated Protein Kinase Kinases; Thiostrepton; Triple Negative Breast Neoplasms

The present study aimed to identify the therapeutic role of the forkhead box M1 (FOXM1)‑associated pathway in triple‑negative breast cancer (TNBC). Using a Cancer Landscapes‑based analysis, a gene regulatory network model was constructed. The present results demonstrated that FOXM1 occupies a key position in gene networks and is a critical regulatory gene in breast cancer. Using breast carcinoma gene expression data from The Cancer Genome Atlas, it was identified that FOXM1 expression was increased in the basal‑like breast cancer subtype compared with other breast cancer subtypes. RNA‑sequencing analysis of MDA‑MB‑231 cells treated with 4 and 10 µl/ml Thiostrepton identified 662 and 5,888 significantly differentially expressed genes, respectively. The Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses demonstrated that FOXM1 was highly associated with multiple biological processes and was markedly associated with metabolic pathways in TNBC. The use of Search Tool for the Retrieval of Interacting Genes/Proteins provided a critical assessment and integration of protein‑protein interactions, and demonstrated the multiple important functions of FOXM1 in TNBC. Real‑time cell analysis, reverse transcription‑quantitative polymerase chain reaction and immunofluorescence staining were used to assess the anti‑tumor activity of Thiostrepton in TNBC cells in vitro. The present results identified that suppression of FOXM1 using Thiostrepton inhibited MDA‑MB‑231 cell proliferation and the expression of cell cycle‑associated genes, including cyclin A2, cyclin B2, checkpoint kinase 1, centrosomal protein 55 and polo like kinase 1. Immunofluorescence staining analysis demonstrated that vimentin, filamentous actin and zinc finger E‑box‑binding homeobox 1 were all decreased following treatment with Thiostrepton. Furthermore, a BALB/C nude mouse subcutaneous xenograft model was used to verify the function of FOXM1 in vivo. The present results demonstrated that FOXM1 inhibition significantly suppressed MDA‑MB‑231 cell tumorigenesis in vivo. Overall, the present results suggested that FOXM1 is a key gene that serves important roles in multiple biological processes in TNBC and that it may serve as a novel therapeutic target in TNBC.

Topics: Animals; Biomarkers, Tumor; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cell Survival; Forkhead Box Protein M1; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Gene Regulatory Networks; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; Sequence Analysis, RNA; Thiostrepton; Triple Negative Breast Neoplasms; Up-Regulation

Triple-negative breast cancer (TNBC) has a relatively poor outcome. Acquired chemoresistance is a major clinical challenge for TNBC patients. Previously, we reported that kinase-dead Aurora kinase A (Aurora-A) could effectively transactivate the FOXM1 promoter. Here, we demonstrate an additional pathway through which Aurora-A stabilizes FOXM1 by attenuating its ubiquitin in TNBC. Specifically, Aurora-A stabilizes FOXM1 in late M phase and early G1 phase of the cell cycle, which promotes proliferation of TNBC cells. Knock-down of Aurora-A significantly suppresses cell proliferation in TNBC cell lines and can be rescued by FOXM1 overexpression. We observe that paclitaxel-resistant TNBC cells exhibit high expression of Aurora-A and FOXM1. Overexpression of Aurora-A offers TNBC cells an additional growth advantage and protection against paclitaxel. Moreover, Aurora-A and FOXM1 could be simultaneously targeted by thiostrepton. Combination of thiostrepton and paclitaxel treatment reverses paclitaxel resistance and significantly inhibits cell proliferation. In conclusion, our study reveals additional mechanism through which Aurora-A regulates FOXM1 and provides a new therapeutic strategy to treat paclitaxel-resistant triple-negative breast cancer.

Topics: Aurora Kinase A; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Drug Resistance, Neoplasm; Female; Forkhead Box Protein M1; Gene Expression Regulation, Neoplastic; Humans; MCF-7 Cells; Paclitaxel; Promoter Regions, Genetic; Thiostrepton; Triple Negative Breast Neoplasms

Triple-negative breast cancer (TNBC) represents a particular clinical challenge because these cancers do not respond to endocrine therapy or other available targeted agents. The lack of effective agents and obvious targets are major challenges in treating TNBC. In this study we explored the cytostatic effect of thiazole ring containing antibiotic drug thiostrepton on TNBC cell lines and investigated the molecular mechanism.. Cell viability was measured by MTT assay. Cell surface marker was monitored by FCM. Western blot was applied to assess the protein expression levels of target genes.. We found that thiostrepton remarkably suppressed the CD44+/CD24- stem-like population and sphere forming capacity of TNBC cell lines. Notably, we showed for the first time that thiostrepton exerted its pharmacological action by targeting sonic hedgehog (SHH) signaling pathway. Thiostrepton repressed SHH ligand expression and reduced Gli-1 nuclear localization in TNBC cell line. Furthermore, the downstream target of SHH signaling undergone dose-dependent, rapid, and sustained loss of mRNA transcript level after thiostrepton treatment. Finally, we showed that SHH ligand was essential for maintaining CD44+/CD24- stem-like population in TNBC cell line.. We conclude that thiostrepton suppresses the CD44+/CD24- stem-like population through inhibition of SHH signaling pathway. Our results give a new insight into the mechanism of thiostrepton anti-tumor activity and suggest thiostrepton as a promising agent that targets hedgehog signaling pathway in TNBC.

Topics: Anti-Bacterial Agents; Antineoplastic Agents; CD24 Antigen; Cell Line, Tumor; Cell Proliferation; Cell Survival; Gene Expression Regulation, Neoplastic; Hedgehog Proteins; Humans; Hyaluronan Receptors; MCF-7 Cells; Neoplastic Stem Cells; Signal Transduction; Spheroids, Cellular; Thiostrepton; Triple Negative Breast Neoplasms