MiR-22 suppresses DNA ligase III addiction in multiple myeloma.
Ontology highlight
ABSTRACT:
SUBMITTER: Caracciolo D
PROVIDER: S-EPMC6365379 | biostudies-literature | 2019 Feb
REPOSITORIES: biostudies-literature
Publications
miR-22 suppresses DNA ligase III addiction in multiple myeloma.
Caracciolo Daniele D Di Martino Maria Teresa MT Amodio Nicola N Morelli Eugenio E Montesano Martina M Botta Cirino C Scionti Francesca F Talarico Daniela D Altomare Emanuela E Gallo Cantafio Maria Eugenia ME Zuccalà Valeria V Maltese Lorenza L Todoerti Katia K Rossi Marco M Arbitrio Mariamena M Neri Antonino A Tagliaferri Pierosandro P Tassone Pierfrancesco P
Leukemia 20180817 2
Multiple myeloma (MM) is a hematologic malignancy characterized by high genomic instability. Here we provide evidence that hyper-activation of DNA ligase III (LIG3) is crucial for genomic instability and survival of MM cells. LIG3 mRNA expression in MM patients correlates with shorter survival and even increases with more advanced stage of disease. Knockdown of LIG3 impairs MM cells viability in vitro and in vivo, suggesting that neoplastic plasmacells are dependent on LIG3-driven repair. To inv ...[more]
Similar Datasets
Project description:BackgroundDNA ligases are crucial for DNA repair and cell replication since they catalyze the final steps in which DNA breaks are joined. DNA Ligase III (LIG3) exerts a pivotal role in Alternative-Non-Homologous End Joining Repair (Alt-NHEJ), an error-prone DNA repair pathway often up-regulated in genomically unstable cancer, such as Multiple Myeloma (MM). Based on the three-dimensional (3D) LIG3 structure, we performed a computational screening to identify LIG3-targeting natural compounds as potential candidates to counteract Alt-NHEJ activity in MM.MethodsVirtual screening was conducted by interrogating the Phenol Explorer database. Validation of binding to LIG3 recombinant protein was performed by Saturation Transfer Difference (STD)-nuclear magnetic resonance (NMR) experiments. Cell viability was analyzed by Cell Titer-Glo assay; apoptosis was evaluated by flow cytometric analysis following Annexin V-7AAD staining. Alt-NHEJ repair modulation was evaluated using plasmid re-joining assay and Cytoscan HD. DNA Damage Response protein levels were analyzed by Western blot of whole and fractionated protein extracts and immunofluorescence analysis. The mitochondrial DNA (mtDNA) copy number was determined by qPCR. In vivo activity was evaluated in NOD-SCID mice subcutaneously engrafted with MM cells.ResultsHere, we provide evidence that a natural flavonoid Rhamnetin (RHM), selected by a computational approach, counteracts LIG3 activity and killed Alt-NHEJ-dependent MM cells. Indeed, Nuclear Magnetic Resonance (NMR) showed binding of RHM to LIG3 protein and functional experiments revealed that RHM interferes with LIG3-driven nuclear and mitochondrial DNA repair, leading to significant anti-MM activity in vitro and in vivo.ConclusionTaken together, our findings provide proof of concept that RHM targets LIG3 addiction in MM and may represent therefore a novel promising anti-tumor natural agent to be investigated in an early clinical setting.
| S-EPMC9588242 | biostudies-literature
Project description:Background: MYC is a master regulator of multiple myeloma (MM) by orchestrating several pro-tumoral pathways, including reprograming of the miRNA transcriptome. MYC is also involved in the acquirement of resistance to anti-MM drugs, including immunomodulatory imide drugs (IMiDs). Methods: In silico analysis was performed on MM proprietary and on public MMRF-CoMMpass datasets. Western blot and chromatin immunoprecipitation (ChIP) experiments were performed to validate miR-22 repression induced by MYC. Cell viability and apoptosis assays were used to evaluate lenalidomide sensitization after miR-22 overexpression. Results: We found an inverse correlation between MYC and miR-22 expression, which is associated with poor outcome in IMiD-treated MM patients. Mechanistically, we showed that MYC represses transcription of miR-22, which, in turn, targets MYC, thus establishing a feed-forward loop. Interestingly, we found that IMiD lenalidomide increases miR-22 expression by reducing MYC repression and, most importantly, that the combination of lenalidomide with miR-22 mimics results in a synergistic direct and NK-mediated cytotoxic activity. Conclusions: Taken together, our findings indicate that: (1) low miR-22 expression could represent a potential predictive biomarker of poor lenalidomide response in MM patients; and (2) miR-22 reduces MYC oncogenic activity, thus triggering a novel synthetic lethality loop, which sensitizes MM cells to lenalidomide.
| S-EPMC8431372 | biostudies-literature
Project description:The transcription factor IRF4 (interferon regulatory factor 4) is required during an immune response for lymphocyte activation and the generation of immunoglobulin-secreting plasma cells. Multiple myeloma, a malignancy of plasma cells, has a complex molecular aetiology with several subgroups defined by gene expression profiling and recurrent chromosomal translocations. Moreover, the malignant clone can sustain multiple oncogenic lesions, accumulating genetic damage as the disease progresses. Current therapies for myeloma can extend survival but are not curative. Hence, new therapeutic strategies are needed that target molecular pathways shared by all subtypes of myeloma. Here we show, using a loss-of-function, RNA-interference-based genetic screen, that IRF4 inhibition is toxic to myeloma cell lines, regardless of transforming oncogenic mechanism. Gene expression profiling and genome-wide chromatin immunoprecipitation analysis uncovered an extensive network of IRF4 target genes and identified MYC as a direct target of IRF4 in activated B cells and myeloma. Unexpectedly, IRF4 was itself a direct target of MYC transactivation, generating an autoregulatory circuit in myeloma cells. Although IRF4 is not genetically altered in most myelomas, they are nonetheless addicted to an aberrant IRF4 regulatory network that fuses the gene expression programmes of normal plasma cells and activated B cells.
| S-EPMC2542904 | biostudies-literature
Project description:BackgroundWe aimed to investigate the correlation of Circ-SMARCA5 with disease severity and prognosis in multiple myeloma (MM), and its underlying mechanisms in regulating cell proliferation and apoptosis.MethodsBone marrow samples from 105 MM patients and 36 healthy controls were collected for Circ-SMARCA5 expression measurement. And the correlation of Circ-SMARCA5 expression with patients' characteristics and survival was determined. In vitro, the effect of Circ-SMARCA5 on MM cell proliferation and apoptosis was evaluated by altering Circ-SMARCA5 expression through transfection. Rescue experiments and luciferase assay were further performed to explore the mechanism of Circ-SMARCA5 as well as its potential target miR-767-5p in regulating MM cell activity.ResultsCirc-AMARCA5 was downregulated in MM and presented a good value in distinguishing MM patients from controls and it was also negatively correlated with Beta-2-microglobulin (β2-MG) level and International Staging System (ISS) stage. Additionally, Circ-SMARCA5 high expression was associated with higher CR as well as better PFS and OS. As for in vitro experiments, Circ-SMARCA5 expression was lower in MM cell lines compared with normal cells, and Circ-SMARCA5 overexpression inhibited cell proliferation but promoted cell apoptosis in RPMI8226 cells. Rescue experiments disclosed that the effect of Circ-SMARCA5 on cell activity was attenuated by miR-767-5p, and luciferase reporter assay revealed direct binding between Circ-SMARCA5 and miR-767-5p.ConclusionsCirc-SMARCA5 is downregulated and correlated with lower β2-MG level and ISS stage as well as better prognosis in MM patients, and it inhibits proliferation but promotes apoptosis of MM cells via directly sponging miR-767-5p.
| S-EPMC6785934 | biostudies-literature
Project description:In multiple myeloma (MM), the bone marrow (BM) microenvironment may contain a myeloma cell fraction that has acquired treatment resistance by undergoing an epigenetic gene expression change. Hypoxic stress is an important factor in the BM microenvironment. Recently, we demonstrated that miR-210 was upregulated in hypoxia and downregulated IRF4, which is known as an essential factor in myeloma oncogenesis in normoxia. In the study, we demonstrated that myeloma cells still showed a strong antiapoptotic phenotype despite IRF4 downregulation, suggesting that another antiapoptotic factor might be involved under hypoxic stress. To determine the factor or factors, we conducted gene expression analysis on myeloma cells (primary samples and cell lines) that were exposed to chronic hypoxia and observed upregulation of glycolytic genes and genes encoding H3K9 demethylases in myeloma cells with hypoxia. Among these, KDM3A was most significantly upregulated in all examined cells, and its knockdown induced apoptosis of myeloma cells in chronic hypoxia. Expression of KDM3A was dependent on HIF-1?, which is a transcription factor specifically upregulated in hypoxia. We further demonstrated that an essential target of KDM3A was a noncoding gene, MALAT1, whose upregulation contributed to acquisition of an antiapoptotic phenotype by accumulation of HIF-1?, leading to upregulation of glycolytic genes under hypoxia. This process was independent from IRF4. These results led us to conclude that the hypoxia-inducible HIF-1?-KDM3A-MALAT1 axis also contributes to acquisition of the antiapoptotic phenotype via upregulation of glycolysis-promoting genes. Thus, this axis is a promising therapeutic target against myeloma cells in the BM microenvironment.
| S-EPMC5858471 | biostudies-literature
Project description:Background:Aberrant expression of long non-coding RNAs (lncRNAs) is closely associated with development and prognosis of human cancers. LncRNA SNHG16 is reportedly involved in human cancer; however, its roles in multiple myeloma (MM) remain unclear. Methods:In this study, we investigated the function and molecular mechanisms of SNHG16 in MM. MM cells were transfected with si-SNHG16 or si-NC. SNHG16 expression levels was measured by qRT-PCR. Cell proliferation was monitored using the MTS. Flow cytometry assay was performed to measure the cell cycle and apoptosis. Luciferase reporter assay were performed to confirm the sponged miRNAs of SNHG16. Results:SNHG16 expression was up-regulated in MM tissues. SNHG16 knockdown suppressed cell proliferation, arrested cell cycle transition from G1 to S phase, and promoted the apoptosis of MM cells. Moreover, SNHG16 knockdown promoted cleaved-Caspase-3, cleaved-Caspase-9, Foxa3a, and Bax expression, while markedly inhibiting CCND1, Bcl-2, Cyclin D1, PI3K, and p-AKT expression in MM cells. miR-342-3p was a direct target of SNHG16. SNHG16 knockdown significantly increased miR-342-3p expression in MM cells. Overexpression miR-342-3p markedly suppressed cell proliferation, arrested cell cycle transition from G1 to S phase, and promoted apoptosis of MM cells. Overexpression of miR-342-3p markedly promoted cleaved-Caspase-3/-9, Foxa3a, and Bax expression, and inhibited CCND1, Bcl-2, Cyclin D1, PI3K, and p-AKT expression in MM cells. Additionally, repression of miR-342-3p could rescue the effect of SNHG16 knockdown on MM cell proliferation, cycle arrest, apoptosis, and related protein expression. Conclusion:Knockdown of lncRNA SNHG16 suppresses MM cell proliferation by sponging miR-342-3p, implicating SNHG16 as a novel therapeutic target for MM.
| S-EPMC6998159 | biostudies-literature
Project description:Aberrant DNA methylation plays a relevant role in multiple myeloma (MM) pathogenesis. MicroRNAs (miRNAs) are a class of small non-coding RNAs that recently emerged as master regulator of gene expression by targeting protein-coding mRNAs. However, miRNAs involvement in the regulation of the epigenetic machinery and their potential use as therapeutics in MM remain to be investigated. Here, we provide evidence that the expression of de novo DNA methyltransferases (DNMTs) is deregulated in MM cells. Moreover, we show that miR-29b targets DNMT3A and DNMT3B mRNAs and reduces global DNA methylation in MM cells. In vitro transfection of MM cells with synthetic miR-29b mimics significantly impairs cell cycle progression and also potentiates the growth-inhibitory effects induced by the demethylating agent 5-azacitidine. Most importantly, in vivo intratumor or systemic delivery of synthetic miR-29b mimics, in two clinically relevant murine models of human MM, including the SCID-synth-hu system, induces significant anti-tumor effects. All together, our findings demonstrate that aberrant DNMTs expression is efficiently modulated by tumor suppressive synthetic miR-29b mimics, indicating that methyloma modulation is a novel matter of investigation in miRNA-based therapy of MM.
| S-EPMC3717964 | biostudies-literature
Project description:BackgroundAP4 (TFAP4) encodes a basic helix-loop-helix leucine zipper (bHLH-LZ) transcription factor and is a direct target gene of the oncogenic transcription factor c-MYC. Here, we set out to determine the relevance of AP4 in human colorectal cancer (CRC) cells.MethodsA CRISPR/Cas9 approach was employed to generate AP4-deficient CRC cell lines with inducible expression of c-MYC. Colony formation, β-gal staining, immunofluorescence, comet and homologous recombination (HR) assays and RNA-Seq analysis were used to determine the effects of AP4 inactivation. qPCR and qChIP analyses was performed to validate differentially expressed AP4 targets. Expression data from CRC cohorts was subjected to bioinformatics analyses. Immunohistochemistry was used to evaluate AP4 targets in vivo. Ap4-deficient APCmin/+ mice were analyzed to determine conservation. Immunofluorescence, chromosome and micronuclei enumeration, MTT and colony formation assays were used to determine the effects of AP4 inactivation and target gene regulation on chromosomal instability (CIN) and drug sensitivity.ResultsInactivation of AP4 in CRC cell lines resulted in increased spontaneous and c-MYC-induced DNA damage, chromosomal instability (CIN) and cellular senescence. AP4-deficient cells displayed increased expression of the long non-coding RNA MIR22HG, which encodes miR-22-3p and was directly repressed by AP4. Furthermore, Mediator of DNA damage Checkpoint 1 (MDC1), a central component of the DNA damage response and a known target of miR-22-3p, displayed decreased expression in AP4-deficient cells. Accordingly, MDC1 was directly induced by AP4 and indirectly by AP4-mediated repression of miR-22-3p. Adenomas and organoids from Ap4-deficient APCmin/+ mice displayed conservation of these regulations. Inhibition of miR-22-3p or ectopic MDC1 expression reversed the increased senescence, DNA damage, CIN and defective HR observed in AP4-deficient CRC cells. AP4-deficiency also sensitized CRC cells to 5-FU treatment, whereas ectopic AP4 conferred resistance to 5-FU in a miR-22-3p and MDC1-dependent manner.ConclusionsIn summary, AP4, miR-22-3p and MDC1 form a conserved and coherent, regulatory feed-forward loop to promote DNA repair, which suppresses DNA damage, senescence and CIN, and contributes to 5-FU resistance. These findings explain how elevated AP4 expression contributes to development and chemo-resistance of colorectal cancer after c-MYC activation.
| S-EPMC9137087 | biostudies-literature
Project description:Myeloma is consistently preceded by premalignant monoclonal gammopathy of undetermined significance (MGUS). In >5% of MGUS patients there is a second MGUS clone (biclonal gammopathy of undetermined significance; BGUS), yet, at myeloma diagnosis, presentation of biclonal gammopathy myeloma (BGMy) is considered less frequent, implying that myeloma eradicates coexisting MGUS.In the largest study of its kind, we assessed BGMy frequency amongst 6399 newly diagnosed myeloma patients enrolled in recent UK clinical trials.Compared to expected prevalence (i.e., >5% of MGUS have BGUS), only 58 of 6399 (0.91%) newly diagnosed myeloma patients had BGMy, indicating myeloma typically eliminates coexistent MGUS. In these 58 BGMy cases, the MGUS plasma cell clone was greatly suppressed in size compared to typical levels observed in conventional MGUS; contrarily, the MGUS clone did not inhibit the myeloma plasma cell clone in BGMy.Myeloma eliminates the majority of competing MGUS, and when it does not, the MGUS clone is substantially reduced in size.
| S-EPMC5589985 | biostudies-literature
Project description:BackgroundMultiple myeloma (MM) is a prevalent hematological malignancy. Long noncoding RNAs are correlated with the development of MM. In this project, the function of lncRNA opa interacting protein 5-antisense 1 (OIP5-AS1) in MM and the potential mechanistic pathway were explored.MethodsThe expression of OIP5-AS1, microRNA (miR)-27a-3p and tuberous sclerosis 1 (TSC1) was analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) assay. Cell proliferation was assessed by Cell Counting Kit-8 (CCK-8) assay, colony formation assay and Bromodeoxyuridine (BrdU) staining. And cell apoptosis was evaluated by flow cytometry assay. Cell metastasis was assessed utilizing transwell assay. Western blot analysis was employed to detect protein level. The target relation between miR-27a-3p and OIP5-AS1 or TSC1 was confirmed via dual-luciferase reporter assay and RNA immunoprecipitation assay. Tumor xenograft assay was conducted to measure the function of OIP5-AS1 in vivo.ResultsThe expression levels of OIP5-AS1 and TSC1 were decreased in MM, whereas miR-27a-3p was upregulated. High level of OIP5-AS1 could predict favourable prognosis of MM patients. Overexpression of OIP5-AS1 inhibited cell viability, colony formation ability, migration and invasion, induced cell cycle arrest in G1 phase and apoptosis of MM cells in vitro as well as repressed tumorigenesis in vivo. MiR-27a-3p was a target of OIP5-AS1, and reversed the impact of OIP5-AS1 on MM cells. MiR-27a-3p directly targeted TSC1. Silencing of miR-27a-3p repressed MM progression by elevating TSC1 expression. OIP5-AS1 upregulated TSC1 by sponging miR-27a-3p.ConclusionOIP5-AS1 repressed multiple myeloma progression by regulating miR-27a-3p/TSC1 axis.
| S-EPMC7206794 | biostudies-literature