Dataset Information

MicroRNA-1252-5p Associated with Extracellular Vesicles Enhances Bortezomib Sensitivity in Multiple Myeloma Cells by Targeting Heparanase.

ABSTRACT:

Introduction

Multiple myeloma (MM) remains an incurable disease, and patient survival requires a better understanding of this malignancy's molecular aspects. Heparanase (HPSE) is highly expressed in aggressive MM cells and related to tumor growth, metastasis, and bortezomib (BTZ) resistance. Thus, targeting HPSE seems to be a promising approach for MM treatment, and because microRNAs (miRNAs) have emerged as potential regulators of HPSE expression, the use of extracellular vesicles (EVs) can allow the efficient delivery of therapeutic miRNAs.

Methods

We used prediction algorithms to identify potential miRNAs that regulate negatively HPSE expression. RT-qPCR was performed to assess miRNAs and HPSE expression in MM lines (U266 and RPMI-8226). Synthetic miRNA mimics were electroporated in MM cells to understand the miRNA contribution in HPSE expression, glycosaminoglycans (GAGs) profile, cell proliferation, and cell death induced by BTZ. EVs derived from HEK293T cells were engineered with miRNAs to evaluate their therapeutic potential combined with BTZ.

Results

It revealed a direct association between BTZ sensitivity, HPSE, and miR-1252-5p expressions. Moreover, overexpression of miR-1252-5p significantly reduced HPSE expression and HPSE enzymatic activity in MM cells. The higher level of miR-1252-5p was correlated with a reduction of cell viability and higher sensitivity to BTZ. Further, EVs carrying miR-1252-5p increased MM cells' sensitivity to BTZ treatment.

Conclusion

These results showed that miR-1252-5p could negatively regulate HPSE in MM, indicating the use of EVs carrying miR-1252-5p as a potential novel BTZ sensitization approach in MM cells.

SUBMITTER: Rodrigues-Junior DM

PROVIDER: S-EPMC7814994 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature

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Publications

MicroRNA-1252-5p Associated with Extracellular Vesicles Enhances Bortezomib Sensitivity in Multiple Myeloma Cells by Targeting Heparanase.

Rodrigues-Junior Dorival Mendes DM Pelarin Maria Fernanda de Andrade MFA Nader Helena Bonciani HB Vettore André Luiz AL Pinhal Maria Aparecida Silva MAS

OncoTargets and therapy 20210115

<h4>Introduction</h4>Multiple myeloma (MM) remains an incurable disease, and patient survival requires a better understanding of this malignancy's molecular aspects. Heparanase (HPSE) is highly expressed in aggressive MM cells and related to tumor growth, metastasis, and bortezomib (BTZ) resistance. Thus, targeting HPSE seems to be a promising approach for MM treatment, and because microRNAs (miRNAs) have emerged as potential regulators of HPSE expression, the use of extracellular vesicles (EVs) ...[more]

PMID: 33488100

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Project description:Multiple myeloma (MM) is an incurable plasma cell malignancy with poor survival. Autophagy, a stress-responsive catabolic process mediated by lysosomal activity, plays a crucial role in the pathophysiology of MM. Growing evidence has indicated that dysregulated microRNAs (miRNAs) are associated with the aberrant autophagy in various human cancers. However, to date, few miRNAs have been reported to directly modulate autophagy in the pathobiology of MM. In this study, we investigated the role of MIR145-3p (microRNA 145-3p) in MM, with focus on cellular processes autophagy and cell death. Our results provided evidence that downregulation of MIR145-3p expression was associated with disease progression in human MM. MIR145-3p triggered autophagic flux through direct targeting of HDAC4 (histone deacetylase 4) in MM cells, leading to enhanced apoptosis. Silencing HDAC4 recapitulated the effects of MIR145-3p, whereas enforced expression of HDAC4 abrogated the effects of MIR145-3p. Furthermore, we showed that suppression of HDAC4 by MIR145-3p resulted in upregulation of the pro-apoptotic protein BCL2L11 and caused MTORC1 inactivation, which in turn led to enhanced autophagy and cell death. Importantly, we demonstrated that MIR145-3p mimic could potentiate the anti-MM activity of bortezomib in both in vitro and in vivo experiments. Overall, our findings indicate that MIR145-3p exerted a tumor suppression function in MM by inducing autophagic cell death and suggest that MIR145-3p-based targeted therapy would represent a novel strategy for MM treatment.Abbreviations: 3-MA: 3-methyladenine; 3'-UTR: 3'-untranslated region; 7-AAD: 7-aminoactinomycin D; ACTB: actin beta; ANXA5: annexin A5; ATG5: autophagy related 5; ATG7: autophagy related 7; B2M: beta-2-microglobulin; BAF: bafilomycin A1; BCL2L11: BCL2 like 11; Bort: bortezomib; CASP3: caspase 3; CCK-8: Cell Counting Kit-8; CQ: chloroquine; Ct: threshold cycle; ctrl: control; DAPI: 4',6-diamidino-2-phenylindole; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GFP: green fluorescent protein; HDAC4: histone deacetylase 4; ISS: International Staging System; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; miRNAs: microRNAs; MIR145-3p: microRNA 145-3p; MM: multiple myeloma; mRNA: messenger RNA; MTOR: mechanistic target of rapamycin kinase; MTORC1: mechanistic target of rapamycin kinase complex 1; PCs: plasma cells; PFS: progression-free survival; qRT-PCR: quantitative reverse transcription PCR; RPS6KB1: ribosomal protein S6 kinase B1; SD: standard deviation; siRNA: small interfering RNA; SQSTM1: sequestosome 1; STV: starvation; TUBB: tubulin beta class I.

Project description:The transfer of microRNAs (miRs) via extracellular vesicles (EVs) is a functionally relevant mechanism of intercellular communication that regulates both organ homoeostasis and disease development. Little is known about the packaging of miRs into EVs. Previous studies have shown that certain miRs are exported by RNA-binding proteins into small EVs, while for other miRs and for large EVs, in general, the export mechanisms remain unclear. Therefore, a proteomic analysis of endothelial cell-derived large EVs was performed, which revealed that heterogeneous nuclear ribonucleoprotein U (hnRNPU) is abundantly present in EVs. EVs were characterized by electron microscopy, immunoblotting and nanoparticle tracking analysis. Taqman microRNA array and single qPCR experiments identified specific miR patterns to be exported into EVs in an hnRNPU-dependent way. The specific role of hnRNPU for vesicular miR-sorting was confirmed independently by gain- and loss-of-function experiments. In our study, miR-30c-5p was the miR whose export was most significantly regulated by hnRNPU. Mechanistically, in silico binding analysis showed that the export of miRs into EVs depends on the binding efficiency of the respective miRs to hnRNPU. Among the exported miRs, a significant enrichment of the sequence motif AAMRUGCU was detected as a potential sorting signal. Experimentally, binding of miR-30c-5p to hnRNPU was confirmed independently by RNA-immunoprecipitation, electrophoretic mobility shift assay and reciprocally by miR-pulldown. Nuclear binding of miR-30c-5p to hnRNPU and subsequent stabilization was associated with a lower cytoplasmatic abundance and consequently reduced availability for vesicular export. hnRNPU-dependent miR-30c-5p export reduced cellular migration as well as pro-angiogenic gene expression in EV-recipient cells. In summary, hnRNPU retains miR-30c-5p and other miRs and thereby prevents their export into large EVs. The data presented provide a novel and functionally relevant mechanism of vesicular miR export.

Dataset Information

MicroRNA-1252-5p Associated with Extracellular Vesicles Enhances Bortezomib Sensitivity in Multiple Myeloma Cells by Targeting Heparanase.

Introduction

Methods

Results

Conclusion

Publications

MicroRNA-1252-5p Associated with Extracellular Vesicles Enhances Bortezomib Sensitivity in Multiple Myeloma Cells by Targeting Heparanase.

Similar Datasets

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