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Polycomb target genes are silenced in multiple myeloma
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
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2010 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 5, no 7, p. e11483-Article in journal (Refereed) Published
Abstract [en]

Multiple myeloma (MM) is a genetically heterogeneous disease, which to date remains fatal. Finding a common mechanism for initiation and progression of MM continues to be challenging. By means of integrative genomics, we identified an underexpressed gene signature in MM patient cells compared to normal counterpart plasma cells. This profile was enriched for previously defined H3K27-tri-methylated genes, targets of the Polycomb group (PcG) proteins in human embryonic fibroblasts. Additionally, the silenced gene signature was more pronounced in ISS stage III MM compared to stage I and II. Using chromatin immunoprecipitation (ChIP) assay on purified CD138+ cells from four MM patients and on two MM cell lines, we found enrichment of H3K27me3 at genes selected from the profile. As the data implied that the Polycomb-targeted gene profile would be highly relevant for pharmacological treatment of MM, we used two compounds to chemically revert the H3K27-tri-methylation mediated gene silencing. The S-adenosylhomocysteine hydrolase inhibitor 3-Deazaneplanocin (DZNep) and the histone deacetylase inhibitor LBH589 (Panobinostat), reactivated the expression of genes repressed by H3K27me3, depleted cells from the PRC2 component EZH2 and induced apoptosis in human MM cell lines. In the immunocompetent 5T33MM in vivo model for MM, treatment with LBH589 resulted in gene upregulation, reduced tumor load and increased overall survival. Taken together, our results reveal a common gene signature in MM, mediated by gene silencing via the Polycomb repressor complex. The importance of the underexpressed gene profile in MM tumor initiation and progression should be subjected to further studies.

Place, publisher, year, edition, pages
2010. Vol. 5, no 7, p. e11483-
National Category
Hematology
Identifiers
URN: urn:nbn:se:uu:diva-133207DOI: 10.1371/journal.pone.0011483ISI: 000279715300003PubMedID: 20634887OAI: oai:DiVA.org:uu-133207DiVA, id: diva2:360368
Available from: 2010-11-03 Created: 2010-11-03 Last updated: 2017-12-12Bibliographically approved
In thesis
1. Epigenetic gene regulation in multiple myeloma and mood disorders
Open this publication in new window or tab >>Epigenetic gene regulation in multiple myeloma and mood disorders
2013 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Epigenetics continues to be redefined and new discoveries are likely to revolutionise the field still further. This thesis explores different aspects of how epigenetic regulation of gene expression contributes to human disease.

Paper I explores the function of the IKKα kinase in regulating gene expression through the nuclear retinoic acid receptor (RAR). We define a set of genes requiring IKKα for their expression and found recruitment of IKKα to the RAR dependent on structural motifs in its protein sequence. This interplay between the NFκB pathway and nuclear receptor regulated transcription is important to consider when designing therapeutic strategies.

Papers II and III focus on the plasma cell malignancy multiple myeloma (MM) and define a gene regulatory circuit defining an underexpressed gene profile in MM dependent on the Polycomb proteins. We provide proof-of-principle that the use of small chemical inhibitors may be operational in reactivating genes silenced by H3K27me3 and that this leads to decreased tumour load and increased survival in the 5T33 in vivo model of MM. We explored the genome-wide distribution of H3K27me3 and H3K4me3, and defined their association with gene expression in freshly-isolated malignant plasma cells from MM patients. Importantly, H3K27me3-marked genes in MM associated with more aggressive stages of the disease and less favourable survival. We present evidence that gene targeting by H3K27me3 is likely to not only involve a small population of tumour cells, but rather represent a common MM profile and further provide a rationale for evaluating epigenetic therapeutics in MM.

Paper IV shows that pro-inflammatory gene expression in monocytes of psychiatric patients can be induced in vitro by sodium pump inhibitors, as the steroid hormone ouabain. We suggest that the ouabain-induced gene expression is regulated by an intricate network involving microRNAs, Polycomb and the H3K27me3 demethylase JMJD3. Our data indicates that epigenetic regulators play a role in transmitting cues between intrinsic and/extrinsic stimuli and gene expression in psychiatric illness.

This thesis provides novel insights on how seemingly unrelated pathways may converge on transcriptional regulation and evidence that epigenetic modifiers contribute to the pathogenesis of human complex diseases such as multiple myeloma and mood disorders.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2013. p. 50
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 910
Keywords
IKKα, retinoic acid receptor, multiple myeloma, Polycomb, Ouabain, mood disorders
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-199494 (URN)978-91-554-8689-1 (ISBN)
Public defence
2013-06-12, Rudbecksalen, Rudbeck Laboratory, Dag Hammarskjölds väg 20, Uppsala, 13:15 (English)
Opponent
Supervisors
Available from: 2013-05-22 Created: 2013-05-06 Last updated: 2018-01-11Bibliographically approved
2. Regulation of Gene Expression in Multiple Myeloma Cells and Normal Fibroblasts: Integrative Bioinformatic and Experimental Approaches
Open this publication in new window or tab >>Regulation of Gene Expression in Multiple Myeloma Cells and Normal Fibroblasts: Integrative Bioinformatic and Experimental Approaches
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The work presented in this thesis applies integrative genomic and experimental approaches to investigate mechanisms involved in regulation of gene expression in the context of disease and normal cell biology.

In papers I and II, we have explored the role of epigenetic regulation of gene expression in multiple myeloma (MM). By using a bioinformatic approach we identified the Polycomb repressive complex 2 (PRC2) to be a common denominator for the underexpressed gene signature in MM. By using inhibitors of the PRC2 we showed an activation of the genes silenced by H3K27me3 and a reduction in the tumor load and increased overall survival in the in vivo 5TMM model. Using ChIP-sequencing we defined the distribution of H3K27me3 and H3K4me3 marks in MM patients cells. In an integrated bioinformatic approach, the H3K27me3-associated genes significantly correlated to under-expression in patients with less favorable survival. Thus, our data indicates the presence of a common under-expressed gene profile and provides a rationale for implementing new therapies focusing on epigenetic alterations in MM.

In paper III we address the existence of a small cell population in MM presenting with differential tumorigenic properties in the 5T33MM murine model. We report that the predominant population of CD138+ cells had higher engraftment potential, higher clonogenic growth, whereas the CD138- MM cells presented with less mature phenotype and higher drug resistance. Our findings suggest that while designing treatment regimes for MM, both the cellpopulations must be targeted.

In paper IV we have studied the general mechanism of differential gene expression regulation by CGGBP1 in response to growth signals in normal human fibroblasts. We found that CGGBP1 binding affects global gene expression by RNA Polymerase II. This is mediated by Alu RNAdependentinhibition of RNA Polymerase II. In presence of growth signals CGGBP1 is retained in the nuclei and exhibits enhanced Alu binding thus inhibiting RNA Polymerase III binding on Alus. Hence we suggest a mechanism by which CGGBP1 orchestrates Alu RNA-mediated regulation of RNA Polymerase II. This thesis provides new insights for using integrative bioinformatic approaches to decipher gene expression regulation mechanisms in MM and in normal cells.

Place, publisher, year, edition, pages
uppsala: Acta Universitatis Upsaliensis, 2014. p. 71
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1029
Keywords
Multiple myeloma, Integrative bioinformatics, Epigenetics, CGGBP1, RNA polymerase
National Category
Cell and Molecular Biology Genetics Bioinformatics (Computational Biology)
Research subject
Oncology; Bioinformatics
Identifiers
urn:nbn:se:uu:diva-232949 (URN)978-91-554-9045-4 (ISBN)
Public defence
2014-11-13, Rudbecksalen, Dag Hammarskjolds vag 20, Uppsala, 09:15 (English)
Opponent
Supervisors
Available from: 2014-10-22 Created: 2014-10-22 Last updated: 2018-01-11Bibliographically approved
3. Tumour Survival Signals and Epigenetic Gene Silencing in Multiple Myeloma: Implications for Biology and Therapy
Open this publication in new window or tab >>Tumour Survival Signals and Epigenetic Gene Silencing in Multiple Myeloma: Implications for Biology and Therapy
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis is focused on multiple myeloma (MM), a haematological malignancy that still remains incurable. The pathogenesis of MM is not fully understood and there is a large intra-tumour and interclonal genetic variation in MM patients. One of the most challenging areas in MM research is to find mechanisms for initiation and progression of MM, but also to overcome the arising resistance to therapy.

In paper I, a signature of under-expressed genes in MM was found to significantly correlate with already defined Polycomb target genes. In selected genes from the profile we found an enrichment of H3K27me3, a repressive mark catalysed by Polycomb repressive complex 2 (PRC2), in MM patients and MM cell lines. Treatment with LBH589 (HDAC inhibitor) and DZNep (methyltransferase inhibitor) reactivated the H3K27me3 target genes and induced apoptosis in MM cell lines. LBH589 reduced tumour load and increased overall survival in the 5T33MM mice. These results suggest an important role for Polycomb complex in MM development and highlight PRC2 as a drug target in MM.

In paper II, the insulin-like growth factor type 1 receptor tyrosine kinase (IGF-1RTK) inhibitor picropodophyllin (PPP) in combination with LBH589 synergistically inhibited cell proliferation and enhanced the apoptotic effect in MM. Since the bone marrow microenvironment has an important role in MM disease and also contributes to drug-resistance, we therefore evaluated the drug combination in the immunocompetent 5T33MM murine model. The drug combination significantly prolonged the survival of the 5T33MM mice compared to single drug treatment. We conclude that the combination of PPP and LBH589 has a therapeutic potential in MM.

In paper III, the role of the cellular inhibitor of apoptosis protein 2 (cIAP2) was evaluated in MM cells harbouring TRAF3 deletion/mutation. By overexpressing cIAP2 in these cells we found an increased resistance to proteasome inhibitors. cIAP2 over-expression by lentiviral constructs led to decreased caspase activation, activation of the canonical NF-κB pathway, and down-regulation of tumour suppressor genes and genes that contribute to apoptosis. Supporting the role of cIAP2 mediated drug-resistance, we here demonstrate that inhibiting cIAP2 using an IAP antagonist, increased the sensitivity to the proteasome inhibitor, bortezomib.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2015. p. 45
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1071
Keywords
Multiple myeloma, Polycomb, apoptosis, cIAP2, IAP-inbibitors, proteasome inhibitors, LBH589, PPP, DZNep
National Category
Hematology
Identifiers
urn:nbn:se:uu:diva-242571 (URN)978-91-554-9159-8 (ISBN)
Public defence
2015-03-25, Fåhraeussalen, Rudbeck Laboratory, Dag Hammarskjölds väg 20, Uppsala, 09:15 (English)
Supervisors
Available from: 2015-02-25 Created: 2015-01-28 Last updated: 2015-03-11

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Fryknäs, MårtenFristedt, CharlotteAgarwal, PrasoonNilsson, KennethÖberg, FredrikJernberg-Wiklund, Helena

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