scholarly journals Genome-Wide CRISPR-Cas9 Screening Reveals a Role for TRAF2 in Resistance to IMiDs in Multiple Myeloma

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 1917-1917
Author(s):  
Jiye Liu ◽  
Wenrong Zhou ◽  
Lijie Xing ◽  
Liang Lin ◽  
Kenneth Wen ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Molecular Mechanisms ◽  
Lentiviral Vector ◽  
Protein Family ◽  
Signal Pathways ◽  
Tnf Receptor ◽  
Associated Factor ◽  
Cytokine Analysis ◽  
Genome Wide ◽  
Equity Ownership

Abstract Major improvements in patient outcome have resulted from the development of novel agents targeting multiple myeloma (MM) cells in the bone marrow (BM) microenvironment. Immunomodulatory drugs (IMiDs) including lenalidomide and pomalidomide bind to the CRL4CRBN ubiquitin ligase and promote proteasomal degradation of IKZF1/3 followed by induction of growth inhibition in MM cells. Although IMiDs-based treatments have achieved significant responses and improved outcomes in MM patients, acquired resistance to IMiDs commonly underlies relapse. Although the molecular mechanisms regulating sensitivity to IMiDs have not been fully defined, cereblon (CRBN) is considered to be the primary binding target of IMiDs. However, some MM cells show resistance to IMiDs despite harboring high CRBN expression levels. In this study, we show that TRAF2, a member of TNF receptor associated factor protein family, represents a novel regulator of IMiDs sensitivity in MM cells. To study the molecular mechanisms underlying IMiDs resistance, we first performed genome-wide knockout screening in IMiDs-sensitive MM.1S cells using a CRISPR-Cas9 GeCKOv2 library containing 6 unique sgRNAs against each of 19,050 genes and 4 sgRNAs against each of 1,864 miRNAs Twenty-eight genes and one miRNA were identified which were associated with resistance to IMiDs. Of note, all six sgRNAs targeting CRBN were identified, consistent with previous studies. Among these novel genes, we found that three different sgRNAs targeting TRAF2 were enriched after IMiDs selection. Therefore, we next individually cloned the sgRNAs of TRAF2 into the Cas9 lentiviral vector, and then re-introduced them into MM.1S cells. Importantly, TRAF2 knockout (KO) MM.1S cells acquire significant resistance to pomalidomide and lenalidomide treatments. To examine whether TRAF2 KO-induced IMiDs resistance was CRBN-pathway dependent, we assessed CRBN and its downstream protein levels. TRAF2 KO showed no effect on CRBN expression; IMiDs can still induce IKZF1/3 degradation, associated with downregulation of IRF4 in TRAF2 KO cells. Taken together, these data suggest that TRAF2 mediates sensitivity of IMiDs, independent of CRBN-IKZF1/3 axis. Since TRAF2 is a member of the TNF receptor associated factor (TRAF) protein family required for activation of several signal pathways including NF-ĸB and JNK, we next examined signaling cascades modulated by TRAF2 KO. Importantly, TRAF2-KO MM cells revealed increased processing of p52 (NF-ĸB2) from its precursor p100, resulting in hyperactivation of the non-canonical NF-κB pathway. The soluble factors (ie, cytokines) secreted by MM cells have important paracrine role on MM cell growth and drug resistance. We therefore performed cytokine analysis of proteins secreted by TRAF2 KO cells, and found that chemokine (C-C motif) ligand 5 (CCL5), a target gene of NF-kB pathway, was upregulated in TRAF2 KO cells. Moreover, upregulation of CCL5 was also observed in acquired IMiDs resistant MM1.S cells, which was confirmed by quantitative real-time PCR. In addition, exogenous CCL5 in the culture medium partially downregulated sensitivity to IMiDs treatment. In conclusion, we here identify and validate TRAF2 as a novel regulator of IMiDs sensitivity, independent of CRBN. Our mechanistic studies provide the preclinical rationale for combination treatment strategies with non-canonical NF-kB inhibitors to overcome IMiDs resistance in MM. Disclosures Zhou: WuXi App Tec Inc: Employment. Anderson:Oncopep: Equity Ownership; C4 Therapeutics: Equity Ownership; Celgene: Consultancy; Takeda Millennium: Consultancy; Bristol Myers Squibb: Consultancy; Gilead: Membership on an entity's Board of Directors or advisory committees.

scholarly journals Genome-wide discovery and characterization of long noncoding RNAs in patients with multiple myeloma

2019 ◽  
Vol 12 (1) ◽  
Author(s):  
Minqiu Lu ◽  
Ying Hu ◽  
Yin Wu ◽  
Huixing Zhou ◽  
Yuan Jian ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Molecular Mechanisms ◽  
Malignant Tumors ◽  
Coiled Coil ◽  
Noncoding Rnas ◽  
Long Noncoding Rnas ◽  
Aberrant Expression ◽  
Genome Wide ◽  
Wide Range ◽  
Whole Transcriptome

Abstract Background Long noncoding RNAs (lncRNAs) are involved in a wide range of biological processes in tumorigenesis. However, the role of lncRNA expression in the biology, prognosis, and molecular classification of human multiple myeloma (MM) remains unclear, especially the biological functions of the vast majority of lncRNAs. Recently, lncRNAs have been identified in neoplastic hematologic disorders. Evidence has accumulated on the molecular mechanisms of action of lncRNAs, providing insight into their functional roles in tumorigenesis. This study aimed to characterize potential lncRNAs in patients with MM. Methods In this study, the whole-transcriptome strand-specific RNA sequencing of samples from three newly diagnosed patients with MM was performed. The whole transcriptome, including lncRNAs, microRNAs, and mRNAs, was analyzed. Using these data, MM lncRNAs were systematically analyzed, and the lncRNAs involved in the occurrence of MM were identified. Results The results revealed that MM lncRNAs had distinctive characteristics different from those of other malignant tumors. Further, the functions of a set of lncRNAs preferentially expressed in MM were verified, and several lncRNAs were identified as competing endogenous RNAs. More importantly, the aberrant expression of certain lncRNAs, including maternally expressed gene3, colon cancer–associated transcript1, and coiled-coil domain-containing 26, as well as some novel lncRNAs involved in the occurrence of MM was established. Further, lncRNAs were related to some microRNAs, regulated each other, and participated in MM development. Conclusions Genome-wide screening and functional analysis enabled the identification of a set of lncRNAs involved in the occurrence of MM. The interaction exists among microRNAs and lncRNAs.

scholarly journals Molecular Mechanisms of Cereblon-Interacting Small Molecules in Multiple Myeloma Therapy

2021 ◽  
Vol 11 (11) ◽  
pp. 1185
Author(s):  
Matteo Costacurta ◽  
Jackson He ◽  
Philip E. Thompson ◽  
Jake Shortt
Keyword(s):  
Multiple Myeloma ◽  
Small Molecules ◽  
Molecular Mechanisms ◽  
Rational Drug Design ◽  
Genome Wide ◽  
Rational Drug ◽  
Anti Cancer ◽  
Cellular Machinery ◽  
Endogenous Substrates ◽  
New Generation

Thalidomide analogues (or immunomodulatory imide drugs, IMiDs) are cornerstones in the treatment of multiple myeloma (MM). These drugs bind Cereblon (CRBN), a receptor for the Cullin-ring 4 ubiquitin-ligase (CRL4) complex, to modify its substrate specificity. IMiDs mediate CRBN-dependent engagement and proteasomal degradation of ‘neosubstrates’, Ikaros (IKZF1) and Aiolos (IKZF3), conveying concurrent antimyeloma activity and T-cell costimulation. There is now a greater understanding of physiological CRBN functions, including endogenous substrates and chaperone activity. CRISPR Cas9-based genome-wide screening has further elucidated the complex cellular machinery implicated in IMiD sensitivity, including IKZF1/3-independent mechanisms. New-generation IMiD derivatives with more potent anti-cancer properties—the CELMoDs (Cereblon E3 ligase modulators)—are now being evaluated. Rational drug design also allows ‘hijacking’ of CRL4CRBN utilising proteolysis targeting chimeras (PROTACs) to convey entirely distinct substrate repertoires. As all these chemotypes—thalidomide, IMiDs, CELMoDs and PROTACs—engage CRBN and modify its functions, we describe them here in aggregate as ‘CRBN-interacting small molecules’ (CISMs). In this review, we provide a contemporary summary of the biological consequences of CRBN modulation by CISMs. Detailed molecular insight into CRBN–CISM interactions now provides an opportunity to more effectively target previously elusive cancer dependencies, representing a new and powerful tool for the implementation of precision medicine.

scholarly journals Genome Wide Transcriptomic Analysis Identifies Dysregulated Splicing Factor Profile with Molecular and Functional Role in Multiple Myeloma

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 361-361 ◽  
Author(s):  
Anil Aktas Samur ◽  
Mehmet Kemal Samur ◽  
Michael A Lopez ◽  
Sanika Derebail ◽  
Kenneth C. Anderson ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Alternative Splicing ◽  
Research Funding ◽  
Functional Role ◽  
Molecular Mechanisms ◽  
Differentially Expressed ◽  
Alternate Splicing ◽  
Genome Wide ◽  
Splicing Pattern

Alternative splicing (AS) is a critical post-transcriptional event, which affects the number of cellular processes. Aberrant splicing of some genes has been reported in multiple myeloma (MM). However, to date, whole-transcriptome-wide AS study has not been performed. We used deep RNA-sequencing data from 16 normal plasma cells (NPC) and 360 newly-diagnosed MM patients to describe the landscape of the alternative splicing events and the molecular mechanisms driving aberrant AS in MM. Global splicing analysis showed that mutually exclusive exon (MXE) (n=510) and Skipped Exon (SE) (n=417) are the most frequent splicing events in MM compared to NPC. Among these events, ~54% were observed in genes which are not differentially expressed between MM and NPC and 46% of the AS events (SE, MXE, retained intron, alternative 3'/5' sites) were observed in differentially expressed genes targeting 203 unique genes. AS affected RNA transcription regulation genes such as IKZF1, IKZF3, and key regulatory elements in MM including, IRF3, IRF4, or key transcription factors such as MEF2C, XPB1, STAT2, and ILF3. In general, AS targetted DNA replication, cell cycle, and apoptosis pathways. MM subgroups showed a heterogeneity for AS events. Monosomy 14, t(4;14), del1p and del17p had the highest number of unique (not observed in other subgroups) AS events compared to NPC.To understanding the molecular mechanisms driving aberrant alternate splicing we next investigated115 splicing factors (SF) in MM and associated them with AS events. We observed that ~40% of SF were dysregulated (dysregulated expression and/or copy number alteration) in MM compared to NPC, including SRSF, PCBP and RBM families. To understand the key binding regions, we have performed SF binding motif enrichment analysis around AS events and found that SRFS1, SRSF9, and PCB1 motifs to be enriched among the splicing events. Importantly, SRSF1 expression was linked with survival in two independent MM datasets.We therefore explored functional role of SRSF1 in MM with perturbation studies. While upregulation of SRSF1 expression significantly increased the cell growth and survival, conversely downregulation of SRSF1 inhibited the both. To dissect the mechanisms of SRSF1-mediated MM growth induction, we utilized SRSF1 mutants lacking either of the 2 RNA-recognition motifs or the serine/argine-rich C-terminal domain involved in protein-protein interactions, and recruitment of spliceosome components. We also utilized a C-terminal fusion of SRSF1 with the nuclear-retention signal of SRSF2 (NRS1 mutant), to force SRSF1 retention in the nucleus and assess the role of its nuclear versus cytoplasmic functions. These studies suggested that SRSF1-regulated AS effects MM cell proliferation. We surprisingly also found that even NRS1 mutant failed to promote MM growth, suggesting an important role of cytoplasmic SRSF1 in promoting MM cells proliferation.We next investigated alternative splicing pattern changes induced by SRSF1 knock down.When analyzing cellular functions of SRSF1-regulated splicing events, we found that SRSF1 knock down affect's genes in the RNA processing pathway as well as genes involved in cancer-related functions such as mTOR, E2F and MYC-related pathways. Splicing analysis was corroborated with immunoprecipitation (IP) followed by mass spectrometry (MS) analysis of T7-tagged SRSF1 MM cells.Finally, using genome wide chromatin and transcription landscape mapping techniques, we have found SRSF1 to be under the transcriptional control of oncogenic E2F1 in MM cells. Consistent with these findings, we observed greater in vitro loss of viability in a large panel of MM cell lines compared with PBMCs from healthy volunteers, following exposure to the splicing modulator pladeniolide. In summary, this study for the first time reports a detailed splicing landscape in myeloma and highlights the biological and clinical importance of alternative splicing events. Moreover, these results indicate a functional role and clinical significance of a gene involved in regulation of alternate splicing in MM, highlighting the need to further understand the splicing pattern in myeloma initiation and progression. Disclosures Anderson: Takeda: Consultancy, Speakers Bureau; Celgene: Consultancy, Speakers Bureau; Amgen: Consultancy, Speakers Bureau; Janssen: Consultancy, Speakers Bureau; Oncopep: Other: Scientific Founder; Sanofi-Aventis: Other: Advisory Board; Bristol-Myers Squibb: Other: Scientific Founder. Avet-Loiseau:takeda: Consultancy, Other: travel fees, lecture fees, Research Funding; celgene: Consultancy, Other: travel fees, lecture fees, Research Funding. Munshi:Adaptive: Consultancy; Abbvie: Consultancy; Takeda: Consultancy; Janssen: Consultancy; Oncopep: Consultancy; Amgen: Consultancy; Celgene: Consultancy.

scholarly journals Genome-Wide Analysis of Protein Family Diversity Provides Insights into Species-Specific Protein Family Expansions in Insects

2021 ◽  
Author(s):  
Mehmet DAYI
Keyword(s):  
Copy Number ◽  
Molecular Mechanisms ◽  
Specific Protein ◽  
Protein Family ◽  
Insect Species ◽  
Protein Families ◽  
Family Diversity ◽  
Genome Wide ◽  
Wide Range ◽  
Species Specific

Abstract Insects are one of the earliest land animals with more than 400 million years old history on Earth, and they compose more than 80% of species. Insects invade a wide range of ecosystems and are considered one of the most evolutionary successful organism groups. Today, many insect species’ genomes have been sequenced to encode molecular mechanisms behind this magnificent evolutionary plasticity. However, only limited genome-wide studies have been carried out to compare protein family diversity in insects. A total of 20 insect species belonging to seven insect orders and two morphogenesis groups were investigated for evolutionary relationships and to uncover protein family diversity in the present study. The phylogenetic analysis inferred from a total of 530 one-to-one single-copy ortholog genes were separated insects into two evolutionary clades based on morphogenesis. Protein family analyses showed that insects share core protein families that perform essential tasks in development and metabolic processes, such as Pkinase and Zinc Finger, cellular signaling and odorant perception (7tm), digestion of food molecules (Trypsin), and detoxification (p450) with copy number expansion compared to other protein families. Additionally, species-specific protein family expansion was observed in various protein families. This study provided insights into protein family diversity and variation among insects and highlights high copy number variation in protein families species-wide.

CS1 Promotes Multiple Myeloma Cell Adhesion, Clonogenic Growth, and Tumorigenicity Via C-Maf-Mediated Interactions with Bone Marrow Stromal Cells (BMSCs)

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 840-840
Author(s):  
Yu-Tzu Tai ◽  
Ender Soydan ◽  
Mariateresa Fulciniti ◽  
Weihua Song ◽  
Xian-Feng Li ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Cell Adhesion ◽  
Membrane Protein ◽  
Molecular Mechanisms ◽  
Target Genes ◽  
Lentiviral Vector ◽  
Myeloma Cell ◽  
Differentially Expressed ◽  
Parp Cleavage ◽  
Cyclin D2

Abstract Cell membrane protein CS1 is highly expressed by tumor cells from the majority of multiple myeloma (MM) patients (>95%) regardless of cytogenetic abnormalities and response to current treatments. Furthermore, CS1 is detected in MM patient sera and correlates with active MM. However, its role in MM pathophysiology is undefined. In the present study, we first generated CS1 null OPM2 MM cells using lentiviral CS1 short interfering RNA. Specific CS1 knockdown was confirmed by depletion of CS1 mRNA and membrane protein, whereas CS1 was expressed in parental OPM2 and OPM2 cells infected with control lentiviral vector (cntOPM2). Immunoblotting of phopho-site of multiple kinase screen analysis showed decreased phosphorylation of ERK1/2, AKT, and STAT3 in CS1null OPM2 cells vs. cntOPM2 cells. Serum deprivation markedly blocked survival at earlier time points in CS1null OPM2 cells vs. cntOPM2 cells. Earlier apoptosis in CS1null OPM2 cells correlated with earlier activation of caspases, PARP cleavage, and increased proapoptotic proteins BNIP3, BIK. CS1 knockdown further delayed development of OPM2 tumor and prolonged survival in mice. CS1null OPM2 cells failed to grow tumors in the majority of mice (n=8) at 5 weeks after cell inoculation, whereas cntOPM2 cells formed tumors within 1.5 weeks in all animals (n=8). Interestingly, CS1 was expressed in tumors that developed late in mice injected with CS1null OPM2 cells. Concomitantly, we overexpressed CS1 in CS1-low expressing U266 cells by transfecting an expressing plasmid pflagCS1 or control vector. Enforced CS1 expression enhanced U266 cell growth and survival. In contrast to the majority of U266 cells (>95%) that grow in suspension in standard tissue culture flasks, all U266CS1 cells exhibited adherent growth and homotypic adhesion. Importantly, overexpressed CS1 increased adhesion of U266 and MM1S cells to BMSCs. Furthermore, U266CS1 cells formed more and larger colonies in methylcellulose than U266 cells. Interestingly, tumors that developed in mice injected with U266 cells expressed significantly higher levels of CS1 than injected U266 cells; moreover, exercised tumors grew in an adherent manner in vitro. Overlapping differentially expressed genes in U266CS1 vs. U266 and CS1null OPM2 vs. cntOPM2 was next analyzed by gene expression profiling. Importantly, c-maf pathway was significantly upregulated in U266CS1 vs. U266 cells and downregulated in CS1null OPM2 vs. cntOPM2 cells, as evidenced by differentially expressed c-maf and its target genes, i.e., cyclin D2, integrin αE/β7 at both mRNA and protein levels. Myeloma cell adhesion-induced VEGF secretion by BMSCs was greater with U266CS1 than U266 cells. Finally, immunoblotting showed upregulation of c-maf and cyclin D2 in U266 tumors overexpressing CS1. These studies provide direct evidence of the role of CS1 in myeloma pathogenesis, define molecular mechanisms regulating its effects, and further support novel therapies targeting CS1 in MM.

scholarly journals Characterization of Lentiviral Vector Derived Anti-Bcma CAR T Cells Reveals Key Parameters for Robust Manufacturing of Cell-Based Gene Therapies for Multiple Myeloma

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3243-3243
Author(s):  
Graham Lilley ◽  
Alden Ladd ◽  
Daniel Cossette ◽  
Laura Viggiano ◽  
Gregory Hopkins ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
T Cells ◽  
T Cell ◽  
Lentiviral Vector ◽  
Employment Equity ◽  
Cell Product ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T ◽  
Equity Ownership

Abstract T cells engineered with chimeric antigen receptors (CAR) specific to CD19 have caused rapid and durable clinical responses in ~90% of patients with acute lymphoblastic leukemia. These data support the development of additional CAR T cell products for the treatment of other hematological malignancies. Recently, B cell maturation antigen (BCMA) expression has been proposed as a marker for identification of malignant plasma cells in patients with multiple myeloma (MM). Nearly all MM and some non-Hodgkin's lymphoma tumor cells express BCMA, while normal tissue expression is restricted to plasma cells and a subset of mature B cells. Therefore, BCMA is an attractive CAR T cell target to treat patients with MM and some B cell lymphomas. To this end, using lentiviral vector technology, we successfully generated CAR T cells specific to BCMA that exhibit potent anti-tumor activity to both multiple myeloma and Burkitt's lymphoma in animal models. Manufacture of CAR T cells for individual patient treatment requires the establishment of a robust and reproducible process - since variability in manufacturing could impact the potency of each cell product. To begin to understand the parameters of the manufacturing process that might contribute to the activity of the final product, we first tested the impact of lentiviral vector (LVV) multiplicity of infection (MOI) on CAR T cell phenotype and function. Using a broad range of MOIs (0.625 to 40) across multiple independent PBMC donors we observed no differences in population doubling or cell size throughout the ~10 day manufacturing process, irrespective of the MOI used. As expected, the number of anti-BCMA CAR expressing cells, the level of CAR expression per cell and the average vector copy number (VCN) in the cell product increased proportionally with MOI. Similarly, T cell function, as determined by an IFNg cytokine release assay in response to BCMA-expressing K562 target cells, was also correlated with the LVV MOI. Notably, increased IFNg expression was readily observable at MOIs as low as 1.25 and reached a plateau with T cells generated using an MOI of 20 or more - highlighting the sensitivity of this functional assay. Analogous data demonstrating MOI dependent in vitro killing activity were obtained using a BCMA-expressing tumor cell cytotoxicity assay. Varying the LVV MOI used during transduction simultaneously alters both the amount of anti-BCMA CAR molecules expressed per cell as well as the number of T cells in the cell product that express anti-BCMA CAR. To evaluate each variable in isolation we generated T cell products containing the same frequency of anti-BCMA CAR T cells (26 ± 4% CAR+ T cells) but different levels of anti-BCMA expression per cell by diluting T cell products made with MOIs from 5 to 40 with donor-matched untransduced cells. While these populations had markedly different levels of CAR surface expression per cell (based on anti-BCMA CAR MFI levels measured by flow cytometry) both low and high expressing anti-BCMA CAR T cell products exhibited identical levels of cytotoxicity against BCMA-expressing tumor cells. These data suggest it is the number of CAR expressing cells that is the critical driver of higher functional activity (perhaps due to the efficiency of LVV mediated anti-BCMA CAR expression per transduced cell). Finally, using this information the variability in manufacturing of anti-BCMA CAR T cells was evaluated across 11 independent normal PBMC donors. All 11 products demonstrated very similar properties with respect to cell growth (population doublings, cell volume), and VCN. Importantly, using our standard MOI we obtained a consistent and high level of anti-BCMA CAR expressing T cells that resulted in robust IFNg cytokine release when co-cultured with BCMA-expression cells. Together, our data highlight the frequency of anti-BCMA CAR T cells per cell product as a key parameter for anti-tumor activity in vitro. Moreover, these data suggest that our LVV driven T cell engineering process can reproducibly generate robust anti-BCMA CAR expressing T cell products in a donor independent manner. A phase I clinical trial to evaluate this technology as a cell-based gene therapy for MM is under development. Disclosures Lilley: bluebird bio, Inc: Employment, Equity Ownership. Ladd:bluebird bio, Inc: Employment, Equity Ownership. Cossette:bluebird bio, Inc: Employment, Equity Ownership. Viggiano:bluebird bio, Inc: Employment, Equity Ownership. Hopkins:bluebird bio, Inc: Employment, Equity Ownership. Evans:bluebird bio, Inc: Employment, Equity Ownership. Li:bluebird bio, Inc: Employment, Equity Ownership. Latimer:bluebird bio: Employment, Equity Ownership. Miller:bluebird bio: Employment, Equity Ownership. Kuczewski:bluebird bio: Employment, Equity Ownership. Bakeman:bluebird bio, Inc: Employment, Equity Ownership. MacLeod:bluebird bio, Inc: Employment, Equity Ownership. Friedman:bluebird bio: Employment, Equity Ownership. Maier:bluebird bio, Inc: Employment, Equity Ownership. Paglia:bluebird bio, Inc: Employment, Equity Ownership. Morgan:bluebird bio: Employment, Equity Ownership. Angelino:bluebird bio, Inc: Employment, Equity Ownership.

scholarly journals Genome-Wide Circular RNA Expression Patterns Reflect Resistance to Immunomodulatory Drugs in Multiple Myeloma Cells

Cancers ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 365 ◽  
Author(s):  
Theresa Jakobsen ◽  
Mette Dahl ◽  
Konstantinos Dimopoulos ◽  
Kirsten Grønbæk ◽  
Jørgen Kjems ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Molecular Mechanisms ◽  
Drug Exposure ◽  
Cpg Island ◽  
Expression Patterns ◽  
Acquired Resistance ◽  
Circular Rnas ◽  
Immunomodulatory Drugs ◽  
Genome Wide

Immunomodulatory drugs (IMiDs), such as lenalidomide and pomalidomide, may induce significant remissions in multiple myeloma (MM) patients, but relapses are frequently observed and the underlying molecular mechanisms for this are not completely understood. Circular RNAs (circRNAs) constitute an emerging class of non-coding RNAs with important roles in cancer. Here, we profiled genome-wide expression patterns of circRNAs in IMiD-sensitive MM cells and their resistant counterparts as well as in IMiD-resistant cells treated with specific epigenetic drugs alone or in combination. We found that genome-wide circRNA expression patterns reflect IMiD sensitivity and ciRS-7 (also known as CDR1as) was the most downregulated circRNA upon acquired resistance. The depletion of ciRS-7 correlated with increased methylation levels of the promoter CpG island of its host gene, LINC00632. Expression of LINC00632 and ciRS-7 was partly restored by treatment with a combination of an EZH2 inhibitor (EPZ-6438) and a DNA methyl transferase inhibitor (5-azacytidine), which also restores the IMiD sensitivity of the cells. However, knockdown of ciRS-7 did not affect IMiD sensitivity and we found that ciRS-7 also becomes epigenetically silenced after prolonged cell culture without drug-exposure. In conclusion, we found that genome-wide circRNA expression patterns reflect IMiD sensitivity in an in vitro model of acquired resistance.

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