Deregulated miRNAs Contribute to Silencing of B-Cell Specific Transcription Factors and Activation of NF-κB in Classical Hodgkin Lymphoma.

Julia Paczkowska,Julia Bein,Natalia Rozwadowska,Marcin Skalski,Sylvia Hartmann,Maciej Giefing,Adam Ustaszewski,Martin-Leo Hansmann,Joanna Janiszewska,Agnieszka Dzikiewicz-Krawczyk

doi:10.3390/cancers13133131

Julia Paczkowska, Julia Bein + Show 8 more

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https://doi.org/10.3390/cancers13133131

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Abstract

Simple SummaryThe role of transcriptionally deregulated miRNAs (microRNAs) in classical Hodgkin lymphoma (cHL) is still not fully understood. To address this issue, we have performed global miRNA expression profiling of commonly used cHL cell lines and we present a complete cHL miRNome (microRNome). Within this group, we identify miRNAs recurrently deregulated in cHL cell lines, and compare them to non-Hodgkin lymphoma cell lines and sorted normal CD77+ germinal centre B-cells. Moreover, we show that several of the recurrently overexpressed miRNAs in cHL cell lines, and also primary microdissected HRS (Hodgkin and Reed-Sternberg) cells, target known B-cell-related transcription factors and NF-κB inhibitors. These findings provide evidence that deregulated miRNAs contribute to the loss of B-cell phenotype and NF-κB activation observed in this lymphoma.A hallmark of classical Hodgkin lymphoma (cHL) is the attenuation of B-cell transcription factors leading to global transcriptional reprogramming. The role of miRNAs (microRNAs) involved in this process is poorly studied. Therefore, we performed global miRNA expression profiling using RNA-seq on commonly used cHL cell lines, non-Hodgkin lymphoma cell lines and sorted normal CD77+ germinal centre B-cells as controls and characterized the cHL miRNome (microRNome). Among the 298 miRNAs expressed in cHL, 56 were significantly overexpressed and 23 downregulated (p < 0.05) compared to the controls. Moreover, we identified five miRNAs (hsa-miR-9-5p, hsa-miR-24-3p, hsa-miR-196a-5p, hsa-miR-21-5p, hsa-miR-155-5p) as especially important in the pathogenesis of this lymphoma. Target genes of the overexpressed miRNAs in cHL were significantly enriched (p < 0.05) in gene ontologies related to transcription factor activity. Therefore, we further focused on selected interactions with the SPI1 and ELF1 transcription factors attenuated in cHL and the NF-ĸB inhibitor TNFAIP3. We confirmed the interactions between hsa-miR-27a-5p:SPI1, hsa-miR-330-3p:ELF-1, hsa-miR-450b-5p:ELF-1 and hsa-miR-23a-3p:TNFAIP3, which suggest that overexpression of these miRNAs contributes to silencing of the respective genes. Moreover, by analyzing microdissected HRS cells, we demonstrated that these miRNAs are also overexpressed in primary tumor cells. Therefore, these miRNAs play a role in silencing the B-cell phenotype in cHL.

Highlights

Global epigenetic reprograming, mediated by alterations in DNA methylation, distinguishes classical Hodgkin lymphoma from normal germinal centre B (GCB) cells, and from other germinal centre derived B-cell lymphomas [1]
Aberrant DNA methylation is crucial for the survival of the neoplastic cells of classical Hodgkin lymphoma (cHL)—the Hodgkin and Reed–Sternberg (HRS) cells—as shown by the regression of a relapsed metastatic cHL in a patient treated with the demethylating agent 5-azacytidine for myelodysplastic syndrome, or by the enhanced efficacy of immune checkpoint inhibitors after therapy using demethylating agents [2,3]
We detected expression of 298 miRNAs in at least three out of the seven analyzed cHL cell lines (Table S3). These miRNAs present the complete cHL miRNAome established in this study, which is, to the best of our knowledge, the most detailed NGSbased miRNA profile of cHL cell lines published so far

Summary

Introduction

Global epigenetic reprograming, mediated by alterations in DNA methylation, distinguishes classical Hodgkin lymphoma (cHL) from normal germinal centre B (GCB) cells, and from other germinal centre derived B-cell lymphomas [1]. In cHL, in addition to the usual hypermethylation of TSG promoters, DNA hypermethylation was reported to contribute to the loss-of-B-cell phenotype of HRS cells by silencing the B-cell specific transcription factors [4,5]. In this context, most studies focus exclusively on DNA methylation; in contrast, the role of the other epigenetic mechanism, namely miRNA in the regulation of B-cell specific transcription factors in the pathogenesis of cHL has remained largely understudied. The importance of miRNAs in normal B-cell development shown in many studies allows us to speculate on the putative role of miRNA deregulation in silencing B-cell specific transcription factors and the consequent loss-of-B-cell identity of HRS cells [6,7,8,9]

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