BRAF/EZH2 Signaling Represses miR-129-5p Inhibition of SOX4 Thereby Modulating BRAFi Resistance in Melanoma.
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ABSTRACT:
SUBMITTER: Gebhardt K
PROVIDER: S-EPMC8155874 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:PurposeThe extracellular matrix (ECM) is an intriguing, yet understudied component of therapy resistance. Here, we investigated the role of ECM remodeling by the collagenase, MT1-MMP, in conferring resistance of v-Raf murine sarcoma viral oncogene homolog B1 (BRAF)-mutant melanoma to BRAF inhibitor (BRAFi) therapy.Experimental designPublicly available RNA-sequencing data and reverse phase protein array were used to determine the relevance of MT1-MMP upregulation in BRAFi-resistant melanoma in patients, patient-derived xenografts, and cell line-derived tumors. Short hairpin RNA (shRNA)-mediated knockdown of MT1-MMP, inhibition via the selective MT1-MMP/MMP2 inhibitor, ND322, or overexpression of MT1-MMP was used to assess the role of MT1-MMP in mediating resistance to BRAFi.ResultsMT1-MMP was consistently upregulated in posttreatment tumor samples derived from patients upon disease progression and in melanoma xenografts and cell lines that acquired resistance to BRAFi. shRNA- or ND322-mediated inhibition of MT1-MMP synergized with BRAFi leading to resensitization of resistant cells and tumors to BRAFi. The resistant phenotype depends on the ability of cells to cleave the ECM. Resistant cells seeded in MT1-MMP uncleavable matrixes were resensitized to BRAFi similarly to MT1-MMP inhibition. This is due to the inability of cells to activate integrinβ1 (ITGB1)/FAK signaling, as restoration of ITGB1 activity is sufficient to maintain resistance to BRAFi in the context of MT1-MMP inhibition. Finally, the increase in MT1-MMP in BRAFi-resistant cells is TGFβ dependent, as inhibition of TGFβ receptors I/II dampens MT1-MMP overexpression and restores sensitivity to BRAF inhibition.ConclusionsBRAF inhibition results in a selective pressure toward higher expression of MT1-MMP. MT1-MMP is pivotal to an ECM-based signaling pathway that confers resistance to BRAFi therapy.
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Project description:Choline availability regulates neural progenitor cell proliferation and differentiation in the developing cerebral cortex. Here, we investigated the molecular mechanism underlying this process and demonstrated that choline regulates the transcription factor SOX4 in neural progenitor cells. Specifically, we found that low choline intake during neurogenesis reduces SOX4 protein levels, causing the downregulation of EZH2, a histone methyltransferase. Importantly, we demonstrate that low choline is not involved in SOX4 protein degradation rate and established that protein reduction is caused by aberrant expression of a microRNA (miR-129-5p). To confirm the role of miR-129-5p, we conducted gain-of-function and loss-of-function assays in neural progenitor cells and demonstrated that directly altering miR-129-5p levels could affect SOX4 protein levels. We also observed that the reduction in SOX4 and EZH2 led to decreased global levels of H3K27me3 in the developing cortex, contributing to reduced proliferation and precocious differentiation. For the first time, to our knowledge, we demonstrate that a nutrient, choline, regulates a master transcription factor and its downstream targets, providing a novel insight into the role of choline in brain development.
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Project description:Patients with advanced gastric cancer (GC) have a poor prognosis with a median overall survival of 10‑12 months. Long non‑coding RNA nicotinamide nucleotide transhydrogenase‑antisense RNA1 (NNT‑AS1) and sex‑determining region Y‑related high mobility group box 4 (SOX4) have been reported to be associated with the progression of various types of cancer; however, the regulatory mechanism between NNT‑AS1 and SOX4 in GC is not completely understood. Reverse transcription‑quantitative PCR was used to detect the expression levels of NNT‑AS1, microRNA (miR)‑142‑5p and SOX4. Western blotting was performed to assess the protein expression levels of SOX4, β‑catenin, c‑Myc, Bcl‑2 and E‑cadherin. The proliferation, apoptosis, migration and invasion of GC cells were determined using MTT, flow cytometry and Transwell assays. The relationship between miR‑142‑5p and NNT‑AS1 or SOX4 was investigated using a dual‑luciferase reporter assay. NNT‑AS1 and SOX4 were upregulated, whereas miR‑142‑5p was downregulated in GC tissues and cells compared with normal tissues and cells. Both NNT‑AS1 and SOX4 knockdown inhibited GC cell proliferation, migration and invasion, and enhanced GC cell apoptosis. Moreover, the results indicated that NNT‑AS1 modulated SOX4 expression by sponging miR‑142‑5p. In addition, SOX4 overexpression reversed NNT‑AS1 knockdown‑mediated effects on GC cell proliferation, apoptosis, migration and invasion. NNT‑AS1 knockdown blocked the Wnt/β‑catenin signaling pathway via the miR‑142‑5p/SOX4 axis. Collectively, the present study indicated that NNT‑AS1 knockdown decreased GC cell proliferation, migration and invasion, and induced GC cell apoptosis by regulating the miR‑142‑5p/SOX4/Wnt/β‑catenin signaling pathway axis.
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Project description:Emerging evidences have indicated that long non-coding RNAs (lncRNAs) are potential biomarkers, playing important roles in the development of cancer. LncRNA Activated in RCC with Sunitinib Resistance (lncARSR) is a novel lncRNA that functions as a potential biomarker and is involved in the progression of cancers. However, the clinical significance and molecular mechanism of lncARSR in bladder cancer (Bca) remains unknow. In this study, we discovered that lncARSR was significantly up-regulated in bladder cancer. In addition, increased expression of lncARSR was positively correlated with higher histological grade and larger tumor size. Further experiments demonstrated that suppression of lncARSR attenuated the proliferation, migration, invasion and epithelial-mesenchymal transition (EMT) process of Bca cells. Mechanistically, lncARSR was mainly located in the cytoplasm and acted as a miRNA sponge to positively modulate the expression of Sex-determining region Y-related high-mobility-group box transcription factor 4 (SOX4) via sponging miR-129-5p and subsequently promoted the proliferation and metastasis of Bca cells, thus playing an oncogenic role in Bca pathogenesis. In conclusion, our study indicated that lncARSR plays a critical regulatory role in Bca cells and lncARSR may serve as a potential diagnostic biomarker and therapeutic target for bladder cancer.
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Project description:Cellular heterogeneity in cancer is linked to disease progression and therapy response, although mechanisms regulating distinct cellular states within tumors are not well understood. We identified melanin pigment content as a major source of cellular heterogeneity in melanoma and compared RNAseq data from high-pigmented (HPCs) and low-pigmented melanoma cells (LPCs), suggesting EZH2 as a master regulator of these states. EZH2 protein was found to be upregulated in LPCs and inversely correlated with melanin deposition in pigmented patient melanomas. Surprisingly, conventional EZH2 methyltransferase inhibitors, GSK126 and EPZ6438, had no effect on LPC survival, clonogenicity and pigmentation, despite fully inhibiting methyltransferase activity. In contrast, EZH2 silencing by siRNA or degradation by DZNep or MS1943 inhibited growth of LPCs and induced HPCs. As the proteasomal inhibitor MG132 induced EZH2 protein in HPCs, we evaluated ubiquitin pathway proteins in HPC vs LPCs. Biochemical assays and animal studies demonstrated that in LPCs, the E2-conjugating enzyme UBE2L6 depletes EZH2 protein in cooperation with UBR4, an E3 ligase, via ubiquitination at EZH2's K381 residue, and is downregulated in LPCs by UHRF1-mediated CpG methylation. Targeting UHRF1/UBE2L6/UBR4-mediated regulation of EZH2 offers potential for modulating the activity of this oncoprotein in contexts in which conventional EZH2 methyltransferase inhibitors are ineffective.
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Project description:Glioma is a common intracranial malignant tumor with high mortality and high recurrence rate. In recent years, increasing evidence has demonstrated that circular RNAs (circRNAs) are potential biomarkers and therapeutic targets for many tumors. However, the role of circRNAs in glioma remains unclear. In this study, we found that circRNA-0002109 was highly expressed in glioma tissues and cell lines. Downregulation of circRNA-0002109 expression inhibited the proliferation, migration, and invasion of glioma cells and inhibited the malignant progression of tumors in vivo. Investigations into the relevant mechanisms showed that circRNA-0002109 upregulated the expression of EMP2 through endogenous competitive binding of microRNA-129-5P (miR-129-5P), which partially alleviated the inhibitory effect of miR-129-5P on epithelial membrane protein-2 (EMP2) and ultimately promoted the malignant development of glioma. Our results indicate that circRNA-0002109 plays an important role in the proliferation, invasion, and migration of glioma cells by regulating the miR-129-5P/EMP2 axis, which provides a new potential therapeutic target for glioma.
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Project description:Melanoma treatment with the BRAF V600E inhibitor vemurafenib provides therapeutic benefits but the common emergence of drug resistance remains a challenge. We generated A375 melanoma cells resistant to vemurafenib with the goal of investigating changes in miRNA expression patterns that might contribute to resistance. Increased expression of miR-204-5p and miR-211-5p occurring in vemurafenib-resistant cells was determined to impact vemurafenib response. Their expression was rapidly affected by vemurafenib treatment through RNA stabilization. Similar effects were elicited by MEK and ERK inhibitors but not AKT or Rac inhibitors. Ectopic expression of both miRNA in drug-naïve human melanoma cells was sufficient to confer vemurafenib resistance and more robust tumor growth in vivo Conversely, silencing their expression in resistant cells inhibited cell growth. Joint overexpression of miR-204-5p and miR-211-5p durably stimulated Ras and MAPK upregulation after vemurafenib exposure. Overall, our findings show how upregulation of miR-204-5p and miR-211-5p following vemurafenib treatment enables the emergence of resistance, with potential implications for mechanism-based strategies to improve vemurafenib responses.Significance: Identification of miRNAs that enable resistance to BRAF inhibitors in melanoma suggests a mechanism-based strategy to limit resistance and improve clinical outcomes. Cancer Res; 78(4); 1017-30. ©2017 AACR.
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Project description:BRAF inhibitors were approved for the treatment of BRAF-mutant melanoma. However, most patients acquire the resistance to BRAF inhibitors after several months of treatment. miR-524-5p is considered as a tumor suppressor in many cancers, including melanoma. In this study, we investigated the biological functions of miR-524-5p in melanoma with acquired resistance to BRAF inhibitor and evaluated the endogenous miR-524-5p expression as a biomarker for melanoma. The results showed that the expression of miR-524-5p was 0.481-fold lower in melanoma tissues (n = 117) than in nevus tissues (n = 40). Overexpression of miR-524-5p significantly reduced proliferative, anchorage-independent growth, migratory and invasive abilities of BRAF inhibitor-resistant melanoma cells. Moreover, the introduction of miR-524-5p led to a reduced development of BRAF inhibitor-resistant melanoma in vivo. Remarkably, the MAPK/ERK signaling pathway was decreased after treatment with miR-524-5p. Furthermore, next-generation sequencing analysis implied that the complement system, leukocyte extravasation, liver X receptor/retinoid-X-receptor activation, and cAMP-mediated signaling may be related to miR-524-5p-induced pathways in the resistant cells. The miR-524-5p level was higher on average in complete response and long-term partial response patients than in progressive disease and short-term partial response patients treated with BRAF inhibitors. Our results proposed that miR-524-5p could be considered as a target for treatment BRAF inhibitor-resistant melanoma and a prognostic marker in the response of patients to BRAF inhibitors for melanoma.
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