Differential epigenetic regulation between the alternative promoters, PRDM1α and PRDM1β, of the tumour suppressor gene PRDM1 in human multiple myeloma cells

Abstract Multiple myeloma (MM) is a B-cell neoplasm that is characterized by the accumulation of malignant plasma cells in the bone marrow. The transcription factor PRDM1 is a master regulator of plasma cell development and is considered to be an oncosuppressor in several lymphoid neoplasms. The PRDM1β isoform is an alternative promoter of the PRDM1 gene that may interfere with the normal role of the PRDM1α isoform. To explain the induction of the PRDM1β isoform in MM and to offer potential therapeutic strategies to modulate its expression, we characterized the cis regulatory elements and epigenetic status of its promoter. We observed unexpected patterns of hypermethylation and hypomethylation at the PRDM1α and PRDM1β promoters, respectively, and prominent H3K4me1 and H3K9me2 enrichment at the PRDM1β promoter in non-expressing cell lines compared to PRDM1β-expressing cell lines. After treatment with drugs that inhibit DNA methylation, we were able to modify the activity of the PRDM1β promoter but not that of the PRDM1α promoter. Epigenetic drugs may offer the ability to control the expression of the PRDM1α/PRDM1β promoters as components of novel therapeutic approaches.

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Dysregulated Apurinic/Apyrimidinic Endonucleases (Ape1 and Ape2) Lead to Genetic Instability in Multiple Myeloma.

Blood ◽

10.1182/blood.v104.11.1418.1418 ◽

2004 ◽

Vol 104 (11) ◽

pp. 1418-1418

Author(s):

Masood A. Shammas ◽

Hemant Koley ◽

Sima Shah ◽

Ramesh B. Batchu ◽

Pierfrancesco Tassone ◽

...

Keyword(s):

Multiple Myeloma ◽

Homologous Recombination ◽

Genomic Instability ◽

Cell Lines ◽

Plasma Cells ◽

Dna Recombination ◽

Endonuclease Activity ◽

Single Nucleotide ◽

Genetic Changes

Abstract Multiple myeloma (MM) is associated with significant genomic instability. Homologous recombination (HR), which is elevated in MM, is considered to be responsible for this instability. As endonucleases play an important role in mediating HR, here we have evaluated the role of endonuclease in biology and progression of MM. Gene expression profile using Affymetrix U133 array showed > 2 fold elevation of Ape1 or Ape2 or both in 5 of 6 MM cell lines and 12 of 15 patient samples. Immunocytochemistry confirmed upregulation of Ape1 protein in MM cell lines. A Plasmid degradation assay confirmed significantly elevated endonuclease activity in MM cells compared to normal plasma cells. To identify the pre-dominating endonuclease activity, the degradation assay was carried out in the presence of specific endonuclease inhibitors. Harmane and methoxyamine (MA), specific inhibitors of apurinic/apyrimidinic endonucleases effectively inhibited significant endonuclease activity, while other endonuclease inhibitors ACPD and FK506 had minimal effects, confirming predominant role of apurinic/apyrimidinic endonucleases (APE) in mediating increased endonuclease activity in MM. We investigated the role of elevated APE endonuclease activity on DNA recombination and subsequent genomic re-arrangements. Using a plasmid-based assay we have previously demonstrated significantly elevated homologous recombination (HR) in MM. Inhibition of endonuclease by methoxyamine suppressed HR activity by 85 ± 2% in MM cells. Next, we evaluated whether inhibition of HR by methoxyamine can affect the frequency of acquisition of new genetic changes in MM cells using single nucleotide polymorphism (SNP) arrays (Affymetrix) as indicator of genomic instability. In three independent experiments, methoxyamine reduced the acquisition of new loss of heterozygocity (LOH) loci by an average of 71%. These data suggest that the dysregulated APE endonucleases contribute significantly to the genomic instability, acquisition of new mutations and progression of MM and provides the rationale for targeting endonuclease activity to prevent disease progression including development of drug resistance.

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Microrna Expression Profile Identifies Distinct Clinically Relevant Sub-Groups in Multiple Myeloma: Novel Prognostic Markers and Potential Targets for Therapy

Blood ◽

10.1182/blood.v112.11.96.96 ◽

2008 ◽

Vol 112 (11) ◽

pp. 96-96 ◽

Cited By ~ 2

Author(s):

Sophia Adamia ◽

Herve AvetLoiseau ◽

Samirkumar B Amin ◽

Yu-Tzu Tai ◽

Steven P. Treon ◽

...

Keyword(s):

Multiple Myeloma ◽

Cell Differentiation ◽

Clinical Outcome ◽

Signaling Pathways ◽

Cell Lines ◽

Plasma Cells ◽

Normal Plasma ◽

Group A ◽

Group B

Abstract MicroRNA, an abundant class of small endogenous RNAs, regulate target genes through inducing translational inhibition and cleavage of targeted transcripts. To date, microRNAs have been implicated in normal biological processes, including development, cell differentiation, apoptosis and proliferation as well as in malignant transformation. However, their role in multiple myeloma (MM) remains unknown. Here we investigated role of microRNAs in myelomagenesis, and their influence on prognosis and clinical outcome. We evaluated profiles of 384 microRNAs in bone marrow derived CD138+ plasma cells (PC) from 79 uniformly treated MM patients, 11 MM cell lines and 9 healthy donors using qRT-PCR based microRNA array. The relative expression was calculated using comparative Ct method, and data was normalized using endogenous controls and analyzed using SDS, RQ manager, R and dChip softwares. MicroRNA expression profiles detected in MM patients were correlated with clinical outcome measures. We observed significant modulate expression of 61 microRNAs in myeloma cells compared to normal plasma cells. When more stringent criteria were used, we identified 24 differentially expressed microRNAs in patient samples. Further, unsupervised hierarchical clustering of filtered microRNAs, based on their DCt values, identified two major groups within the MM population (groups A and group B). Samples of Group A clusters with MM cell lines, indicating more proliferative nature of MM patient cells. Within B group, a second degree node group B2, clusters with normal plasma cells indicating more indolent course, while patients in an additional node B1 represented an assorted pattern. The unsupervised clustering of all MM samples showed consistent changes in miR-30b, -30c, -30d, -142-5p, -24, -191, -181d, -374, -146b, -140, -145, -125a, -151, -223, -155, let7b, indicative of a role of these microRNA in myelomagenesis; while supervised analysis of samples within groups A and B identified modulated expression of different sets of miRNAs. In group A miR-585 and let-7f were upregulated 8–12 fold, while miRs -125a, -126, -155, -223, -146a, -374 -19a, -20a, -26a, -30a -5p, -30b, and -30d were significantly downregulated; in group B, all differentially expressed microRNAs were downregulated (p<0.001) compared to normal plasma cells. These modulated miRNAs target critical signaling pathways including apoptosis, hematopoietic cell differentiation and proliferation, survival and angiogenesis by upregulating function of HOX9, c-myc, VCAM-1, Bcl-2, E2F1, SHP1, SHP2, VEGF, and DUSp6 molecules. We further analyzed the effect of microRNA on clinical outcome. We have observed significantly superior event free and overall survival of patients in group B2 compared to patients in group A (2 yr estimated EFS 79% versus 54% respectively; p=0.05; and 2 yr estimated OS 94% versus 70% respectively; p =0.017). Taken together this data identifies critical microRNAs as modulators of gene expression and signaling pathways and provides potential novel microRNA and gene targets in MM to both understand biological behavior and for therapeutic application.

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DNA Methyltransferase Inhibitors Increase Expression of CD38 and Enhance Daratumumab Efficacy in Multiple Myeloma

Blood ◽

10.1182/blood-2018-99-114206 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 5618-5618 ◽

Cited By ~ 1

Author(s):

Priya Choudhry ◽

Margarette C. Mariano ◽

Arun P Wiita

Keyword(s):

Multiple Myeloma ◽

Flow Cytometry ◽

Cell Lines ◽

Research Funding ◽

Plasma Cells ◽

Cpg Island ◽

Ex Vivo ◽

Single Agent ◽

Cd38 Expression

Abstract Introduction: The anti-CD38 monoclonal antibody Daratumumab is highly effective against multiple myeloma, is well tolerated, and has high single agent activity as well as combination effects with lenalidomide-dexamethasone as well as bortezomib-dexamethasone. Patient response to daratumumab monotherapy is highly correlated with pretreatment levels of CD38 expression on MM plasma cells (Nijhof et al, Leukemia (2015) 29:2039) and CD38 loss is correlated with daratumumab resistance (Nijhof et al, Blood (2016) 128:959). As a result, there is significant interest in elucidating the regulation and role of CD38 in MM. Recently, All Trans Retinoic Acid (ATRA), a known small molecule inducer of CD38 in myeloid cells, as well as the FDA-approved histone deacetylase inhibitor panobinostat, were both demonstrated to induce CD38 in MM plasma cells leading to increased lysis by daratumumab. Examining ENCODE data, we found the presence of a CpG island at the first exon of CD38. We hypothesized that removing methylation sites from this CpG island may de-repress CD38 transcription and lead to increased CD38 protein at the cell surface in MM plasma cells. Therefore, here we studied the role of DNA methyl-transferase inhibitors (DNMTis), currently FDA-approved for treatment of myelodysplastic syndrome, as agents to potentiate daratumumab therapy. Methods: We treated MM cell lines (RPMI-8226, MM.1S, XG-1, KMS12-PE) with two different DNMTis, 5-Azacytidine and decitabine, and assessed CD38 cell surface expression by flow cytometry. Similarly, we treated MM patient bone marrow aspirates ex vivo and assessed induction of CD38 expression in the CD138 positive population by flow cytometry. We analyzed CD38 mRNA levels and total CD38 protein levels by qRT-PCR and western blotting respectively. ATRA was used as a positive control in all experiments. We further tested the functional effect of DNMTi treatment on MM cell lines using an Antibody Dependent Cell Cytotoxicity (ADCC) assay. Briefly, live treated cells were incubated overnight with daratumumab and NK92-CD16 transgenic cells at and E:T ratio of 20:1, and lysis was measured using CytoTox-Glo (Promega). Results: Flow analysis revealed that DNMTi treatment induces a 1.2-2 fold increase in CD38 surface protein expression in a dose-dependent manner across MM cell lines. DNMTi treatment consistently yielded similar or higher increases in CD38 expression than that seen in ATRA- or panobinostat-treated cells. Despite significantly lower single-agent cytotoxicity, we found that decitabine led to similar surface CD38 induction as 5-Azacytidine. By RT-qPCR, 5-Azacytidine increased CD38 mRNA expression ~3 fold versus DMSO control, compared to ~2 fold mRNA increase with ATRA. In functional ADCC assays, DNMTi treatment also led to greater lysis than ATRA. Furthermore, the combination of both DNMTi and ATRA was additive, leading to the greatest lysis by NK cells. In contrast, in ex vivo-treated patient samples, ATRA induced greater CD38 expression than 5-Azacytidine on malignant plasma cells. However, this result is expected since MM plasma cells from patients typically do not proliferate in standard ex vivo culture, and active DNA replication is a requirement for successful DNMT inhibition based on known mechanism of action. In patients, however, we anticipate that continual plasma cell proliferation will lead to effective increases in CD38 after DNMTi treatment, as found in MM cell lines here. Summary and Conclusions: Our results here demonstrate that CD38 expression in MM cells is regulated by DNA methylation and targeting DNMTs with small molecule inhibitors leads to increased vulnerability to Daratumumab treatment. We propose that combination treatment with DNMTi and Daratumumab can lead to higher efficacy of daratumumab in daratumumab-naïve MM, as well as reversal of daratumumab-resistance. These combinations should be tested in clinical trials. Disclosures Wiita: Sutro Biopharma: Research Funding; TeneoBio: Research Funding.

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Identification of EpHA3 As a New Potential Molecular Target in Multiple Myeloma.

Blood ◽

10.1182/blood.v120.21.2926.2926 ◽

2012 ◽

Vol 120 (21) ◽

pp. 2926-2926

Author(s):

Antonella Caivano ◽

Francesco La Rocca ◽

Alessandra Favole ◽

Sonia Carturan ◽

Enrico Bracco ◽

...

Keyword(s):

Multiple Myeloma ◽

Monoclonal Antibody ◽

Cell Lines ◽

Plasma Cells ◽

Molecular Target ◽

Over Expression ◽

In Vitro Angiogenesis ◽

Novel Therapeutic

Abstract Abstract 2926 Introduction Angiogenesis plays a central role in the progression of both solid and hematological tumors. In particular, in multiple myeloma (MM) the critical role of bone marrow (BM) microenvironment and angiogenesis has been well documented. The past decade has witnessed a dramatic improvement in the therapeutic options in MM. However, the disease remains incurable, underscoring the need for continued efforts towards understanding MM biology and exploitation of novel therapeutic approaches. In this setting, monoclonal antibodies against myeloma-specific cell surface antigens represent a promising therapeutic approach, which is however hampered by a lack of appropriate target structures expressed across all pathogenic myeloma cells. The Eph receptors, a large family of receptor tyrosine kinases (RTKs) activated by ephrins binding, have been implicated in many processes involved in malignancy, including alteration of the tumor microenvironment and in angiogenesis, in both of which EpHA3 likely plays an active role. Aberrant expression of EpHA3 is seen in many types of hematolologic malignancies (some leukemic cell lines, T-cell lymphoma, acute lymphoblastic leukemia, myeloproliferative neoplasms) although it is not expressed ubiquitously. Finally, the over-expression of Eph is believed to be sufficient to confer tumorigenic potential although probably further mechanisms can occur to abnormally activate the receptor. Basing on the role of EpHA3 in haematological malignancies, a first-in-class engineered IgG1 antibody targeting the EpHA (KB004) was developed and it is now under phase I clinical trials in USA and Australia for the treatment of EpHA3 overexpressing hematological myeloid malignancies refractory to conventional treatment. We investigated the EpHA3 role and its preferential membrane–bound by GPI linker ligand EFNA5, in MM patients in order to define EpHA3 as new molecular target for a novel therapeutic approach with a specific anti EpHA3 monoclonal antibody. The EpHA3 expression has been studied through a comparative proteomic analysis between BM endothelial cells (ECs) of patients with MM (MMECs) or with monoclonal gammopathy of undetermined significance (MGECs), of control subjects (normal ECs) and in MM cell lines. Methods After written informed consent, BM aspirates have been collected from 20 MM and 4 MGUS patients. Normal ECs were derived from 3 BM aspirates of subjects with anemia due to iron or vitamin B12 deficiency. We analyzed the expression levels of EpHA3 in normal ECs, MGECs and MMECs and MM cell lines evaluating the mRNA and protein levels by RT-qPCR and by WB coupled to ImmunoFluorescence analysis. The biological effects of EpHA3 targeting in MMECs have been studied silencing the EpHA3 mRNA in MMECs and testing them at 72h after silencing in series of functinal assays including viability assay by trypan blue exclusion staining and by in vitro angiogenesis assay followed by measurement of mesh areas and vessel length. Moreover, we studied EFNA5 mRNA expression levels in Normal ECs, MGECs and MMECs and in MM cell lines by PCR. Results Our data showed that EpHA3 mRNA levels are progressively increased from ECs to MGECs reaching the highest values in MMECs. Subsequent analysis by WB and immunofluorescence confirmed EpHA3 protein upregulation among the different EC types. The MMECs in which EpHA3 has been silenced revealed a protein level reduction of approximately 60% when compared to the control. We could not detect major viability defects. Furthermore, in vitro angiogenesis inhibition was marginal when compared to the not silenced counterpart. To know whether EpHA3 may impact not only MM angiogenesis but also plasma cells, three MM cell lines were studied for the EpHA3 expression. We found the plasma cell lines gave constant over expression of EpHA3. Finally, the preliminary data regarding EFNA5 mRNA expression level showed it is expressed in either MMECs and MM plasma cell lines. The evaluation of KB004 effect on MMECs in term of apoptosis induction and in vitro tube formation inhibition, as well as the analysis of EpHA3 levels in primary MM plasma cells are in progress. Conclusions From this study we expect to characterize the role of the EpHA3in MM patients and to provide experimental evidences supporting the possibility of using EpHA3 as a new molecular target for MM by proving the in vitro efficacy of a monoclonal antibody to target the angiogenesis of MM. Disclosures: No relevant conflicts of interest to declare.

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The Somatostatin Analog Octreotide Inhibits Growth of Interleukin-6 (IL-6)–Dependent and IL-6–Independent Human Multiple Myeloma Cell Lines

Blood ◽

10.1182/blood.v93.5.1724 ◽

1999 ◽

Vol 93 (5) ◽

pp. 1724-1731 ◽

Cited By ~ 28

Author(s):

Patrik Georgii-Hemming ◽

Thomas Strömberg ◽

Eva Tiensuu Janson ◽

Mats Stridsberg ◽

Helena Jernberg Wiklund ◽

...

Keyword(s):

Multiple Myeloma ◽

Growth Inhibition ◽

Cell Lines ◽

Interleukin 6 ◽

Plasma Cells ◽

Somatostatin Receptor ◽

Somatostatin Receptors ◽

Somatostatin Analog ◽

Igf I ◽

Human Multiple Myeloma

Abstract Somatostatin and its analogs can inhibit growth in several cell types, in part by interfering with insulin-like growth factor-I (IGF-I) signaling. Our previous studies point to the importance of paracrine and autocrine IGF-I in the support of growth and survival of human multiple myeloma (MM) cell lines. In this report, we have investigated the potential role of a somatostatin analog, octreotide, in regulating growth and/or survival in MM. The results show that all MM cell lines express functional somatostatin receptors (sst). The MM cell lines express the subtypes sst2, sst3, and predominantly sst5 as determined by reverse-transcriptase polymerase chain reaction and fluorescence-activated cell sorter analysis. Octreotide inhibited the growth of both the interleukin-6 (IL-6)–dependent and the IL-6–independent MM cell lines. The effect is mainly cytostatic, resulting in 25% to 45% growth inhibition, and in three of eight of the MM cell lines a weak induction of apoptosis was recorded. Our results also show that octreotide may act as an inducer of apoptosis in primary B-B4+ plasma cells isolated from bone marrow of MM patients. In conclusion, the results show a novel pathway for growth inhibition of MM cells: the activation of somatostatin receptor signaling.

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Role of N6-Methyladenosine (m6A) RNA Modification in Multiple Myeloma

Blood ◽

10.1182/blood-2018-99-115820 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 5601-5601

Author(s):

Christian Bach ◽

Magdalena Leffler ◽

Cindy Flamann ◽

Jan Kronke ◽

Dimitrios Mougiakakos ◽

...

Keyword(s):

Multiple Myeloma ◽

Cell Lines ◽

Messenger Rna ◽

Plasma Cells ◽

Conflicts Of Interest ◽

Stop Codon ◽

Mrna Levels ◽

Molecular Abnormalities ◽

Gene Expression Control

Abstract Multiple myeloma (MM) is considered a chronic and incurable disease due to its highly complex and heterogeneous molecular abnormalities. In recent years, integrating proteasome inhibitors and immunomodulatory drugs into MM frontline therapy has significantly improved treatment efficacy with a median overall survival (OS) being prolonged from 3-4 to 7 years. Despite this progress, patients refractory to the aforementioned agent classes display a median OS of only 9 months. Thus, the clinical necessity for developing novel therapeutic alternative approaches is self-evident. Methylation of N6-adenosine (m6A) is known to be important for diverse biological processes including gene expression control, translation of protein, and messenger RNA (mRNA) splicing. m6A regulatory enzymes consist of "writers" METTL3 and METTL14, "readers" YTHDF1 and YTHDF2, and "erasers" FTO and ALKBH5. However, the functions of m6A mRNA modification and the specific role of these enzymes in MM remain unknown. Here we report that METTL3, a key component of the m6A methyltransferase complex, is highly expressed in MM cell lines and in isolated patient's MM cells. In contrast, we found no significant differences in the expression of the m6A demethylases FTO and ALKBH5. Accordingly, compared to plasma cells from healthy donors, global PolyA+ RNA showed a significant increase in m6A content in patient's MM plasma cells. In MM cell lines, global m6A profiling by methylated RNA-immunoprecipitation sequencing revealed m6A peaks near the stop codon in mRNAs of multiple oncogenes including MAF and CCND1. Cross-linking immunoprecipitation showed that METTL3 bound to the m6A peak within MAF and CCND1 mRNA. Depletion of METTL3 by shRNA had little effect on global mRNA levels, but specifically reduced protein levels of c-Maf and Cyclin D1. Moreover, downregulation of METTL3 in several MM cell lines results in cell cycle arrest and apoptosis. Together, these results describe a role for METTL3 in promoting translation of a subset of oncogenes in MM and identify this enzyme as a potential therapeutic target for multiple myeloma. Disclosures No relevant conflicts of interest to declare.

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The Role of PHF19 As a Promoter of Tumorigenicity and Therapeutic Target in Multiple Myeloma

Blood ◽

10.1182/blood-2019-130595 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 508-508

Author(s):

Carolina D. Schinke ◽

Pingping Qu ◽

Shmuel Yaccoby ◽

Valeriy V Lyzogubov ◽

Veronica MacLeod ◽

...

Keyword(s):

Cell Cycle ◽

Multiple Myeloma ◽

Candidate Gene ◽

Cell Lines ◽

Crucial Role ◽

Plasma Cells ◽

Total Therapy

Introduction - Multiple Myeloma (MM) is a hematologic malignancy characterized by clonal growth of differentiated plasma cells (PCs). Despite improvement in MM therapy, the disease remains mostly incurable and is characterized by recurrent relapses with development of resistant clones that eventually lead to patient death. The pathways that lead to resistant and aggressive MM are not fully understood highlighting the need to improve our understanding of MM biology to identify potential new pathways and therapeutical targets. PHD Finger Protein 19 (PHF19) is a regulator of Polycomb Repressive Complex 2 (PRC2), the sole methyltransferase complex capable of catalyzing H3K27me3 to induce and enforce gene repression. PRC2 employs enhancer of zeste homolog 1 and 2 (EZH1/EZH2) as enzymatic subunits to hypermethylate H3K27. While overexpression and gain of function mutations of EZH1/2 have been observed in many cancers the role of this particular pathway in MM remains poorly understood. In the present study, we report on PHF19 as a new candidate gene to play a potential crucial role in MM oncogenesis. Methods- Gene expression profiling (GEP; Affymetrix U133 Plus 2.0) was performed on 739 MM patients (from total therapy trials [TT] 3-5; low risk MM n=636, high risk MM n=103), 42 patients with monoclonal gammopathy of undetermined significance (MGUS), 73 smoldering MM patients, 42 patients with primary plasma cell leukemia and 34 healthy donors. Myeloma risk was determined by the GEP 70 signature as previously defined. To test the implications of functional PHF19 knock down (KD) we used TRIPZ inducible PHF19 shRNA vs. scrambled control (Dharmacon) in two MM cell lines (JJN3 and ARP1). Real time PCR as well as western blotting was used to confirm PHF19 KD as well as to elucidate the effect on H3K27me3 (Cell Signaling). Functional in vitro studies included proliferation (Promega), clonogenic assays (StemCell), cell cycle and apoptosis assays (both Invitrogen). In vivo studies were performed using SCID mice that were subjected to tail vain injection with PHF19 KD JJN3 cells (n=10) or scrambled shRNA control (n=10). Weekly ELISA (Bethyl) and in vivo imaging (Xenogen) were performed and survival was recorded. Results- GEP of the previously mentioned patient populations and healthy controls identified PHF19 (chr9q33.2) as a candidate gene that was consistently dysregulated in MM patients. Mean expression levels at different MM stages correlated with disease aggressiveness (ANOVA, p<0.0001), Figure 1. High expression of PHF19 (log2>10.46) at diagnosis correlated significantly with adverse clinical parameters, including ISS III, anemia and elevated LDH, as well as worse overall survival (5 yr OS = 29% for patients with high PHF19 expression vs 77% for patients with low PHF19 expression [log2<10.46], p< 0.0001). These results led us to test the implications of functional PHF19 KD using TRIPZ inducible PHF19 shRNA vs. scrambled control in the JJN3 and ARP1 MM cell lines. PHF19 KD led to a drastic reduction of H3K27me3 thereby resulting in significantly reduced proliferation via cell cycle arrest, while apoptosis was not substantially altered. Clonogenic assays showed a significant reduction in colony numbers and size of MM cells with PHF19 KD compared to the control (>75% reduction in both cell lines, p<0.05). Xenograft studies showed consistently less tumor burden in the mice injected with PHF19 KD cells compared to scrambled control, evident through ELISA testing for IgG Kappa (Median =180 mg/ml for scrambled control vs 80 mg/ml for PHF19 KD at week 8, p=0.07) and bioimaging (Median bioilumisence 2.1x108 p/s for scrambled control vs. 0.8x108 p/s for PHF19 KD at week 8, non-significant). Median OS in mice injected with PHF19 KD cell was substantially longer (66 days) compared to mice subjected to scrambled control cells (54 days), p=0.052. Conclusion- In summary we show that PHF19 is upregulated in malignant plasma cells of MM patients and that PHF19 expression levels increase with advanced MM stages. High PHF19 expression was a marker of adverse prognosis in our total therapy (TT 3-5) cohort. Most importantly, in-vitro and in-vivo functional studies showed that PHF19 has important biological functions in MM. These results suggest that epigenetic regulation through histone methylation, in particular, H3K27 trimethylation, plays a crucial role in MM and the affected downstream pathways should be further elucidated. Disclosures Boyle: Janssen: Honoraria, Other: Travel; Abbvie: Honoraria; Amgen: Honoraria, Other: travel; Takeda: Honoraria, Other: travel; Celgene Corporation: Honoraria, Other: Travel. van Rhee:Kite Pharma: Consultancy; Adicet Bio: Consultancy; Karyopharm Therapeutics: Consultancy; Takeda: Consultancy; Sanofi Genzyme: Consultancy; Castleman Disease Collaborative Network: Consultancy; EUSA: Consultancy. Walker:Celgene: Research Funding.

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The Somatostatin Analog Octreotide Inhibits Growth of Interleukin-6 (IL-6)–Dependent and IL-6–Independent Human Multiple Myeloma Cell Lines

Blood ◽

10.1182/blood.v93.5.1724.405k21_1724_1731 ◽

1999 ◽

Vol 93 (5) ◽

pp. 1724-1731

Author(s):

Patrik Georgii-Hemming ◽

Thomas Strömberg ◽

Eva Tiensuu Janson ◽

Mats Stridsberg ◽

Helena Jernberg Wiklund ◽

...

Keyword(s):

Multiple Myeloma ◽

Growth Inhibition ◽

Cell Lines ◽

Interleukin 6 ◽

Plasma Cells ◽

Somatostatin Receptor ◽

Somatostatin Receptors ◽

Somatostatin Analog ◽

Igf I ◽

Human Multiple Myeloma

Somatostatin and its analogs can inhibit growth in several cell types, in part by interfering with insulin-like growth factor-I (IGF-I) signaling. Our previous studies point to the importance of paracrine and autocrine IGF-I in the support of growth and survival of human multiple myeloma (MM) cell lines. In this report, we have investigated the potential role of a somatostatin analog, octreotide, in regulating growth and/or survival in MM. The results show that all MM cell lines express functional somatostatin receptors (sst). The MM cell lines express the subtypes sst2, sst3, and predominantly sst5 as determined by reverse-transcriptase polymerase chain reaction and fluorescence-activated cell sorter analysis. Octreotide inhibited the growth of both the interleukin-6 (IL-6)–dependent and the IL-6–independent MM cell lines. The effect is mainly cytostatic, resulting in 25% to 45% growth inhibition, and in three of eight of the MM cell lines a weak induction of apoptosis was recorded. Our results also show that octreotide may act as an inducer of apoptosis in primary B-B4+ plasma cells isolated from bone marrow of MM patients. In conclusion, the results show a novel pathway for growth inhibition of MM cells: the activation of somatostatin receptor signaling.

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Role of 1q21 in Multiple Myeloma: From Pathogenesis to Possible Therapeutic Targets

Cells ◽

10.3390/cells10061360 ◽

2021 ◽

Vol 10 (6) ◽

pp. 1360

Author(s):

Jessica Burroughs Garcìa ◽

Rosa Alba Eufemiese ◽

Paola Storti ◽

Gabriella Sammarelli ◽

Luisa Craviotto ◽

...

Keyword(s):

Multiple Myeloma ◽

Plasma Cells ◽

De Novo ◽

Current Knowledge ◽

Therapeutic Targets ◽

Poor Response ◽

Signal Pathway ◽

Pathological Features ◽

Therapeutic Approaches

Multiple myeloma (MM) is characterized by an accumulation of malignant plasma cells (PCs) in the bone marrow (BM). The amplification of 1q21 is one of the most common cytogenetic abnormalities occurring in around 40% of de novo patients and 70% of relapsed/refractory MM. Patients with this unfavorable cytogenetic abnormality are considered to be high risk with a poor response to standard therapies. The gene(s) driving amplification of the 1q21 amplicon has not been fully studied. A number of clear candidates are under investigation, and some of them (IL6R, ILF2, MCL-1, CKS1B and BCL9) have been recently proposed to be potential drivers of this region. However, much remains to be learned about the biology of the genes driving the disease progression in MM patients with 1q21 amp. Understanding the mechanisms of these genes is important for the development of effective targeted therapeutic approaches to treat these patients for whom effective therapies are currently lacking. In this paper, we review the current knowledge about the pathological features, the mechanism of 1q21 amplification, and the signal pathway of the most relevant candidate genes that have been suggested as possible therapeutic targets for the 1q21 amplicon.

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Lower expression of the RASSF10 gene induces myeloma cell proliferation due to hypermethylation of the gene promoter in multiple myeloma

10.21203/rs.3.rs-23701/v1 ◽

2020 ◽

Author(s):

Jin Chen ◽

Hui Liu ◽

Zhaoyun Liu ◽

Qing Shao ◽

Fengjuan Jiang ◽

...

Keyword(s):

Multiple Myeloma ◽

Overall Survival ◽

Cell Lines ◽

Plasma Cells ◽

Malignant Neoplasm ◽

Gene Promoter ◽

Myeloma Cell ◽

Suppressor Gene ◽

Induced Apoptosis

Abstract Background Multiple myeloma (MM) is an incurable malignant neoplasm of plasma cells, in which genetic defects, epigenetic aberrations and bone marrow microenvironment are involved in the pathogenesis. RASSF10 acts as a tumor suppressor gene by methylation in glioma and several other cancers, but its role in MM remains unknown. Methods In order to explore the role of RASSF10, mRNA expression was detected in MM patients and analyzed with overall survival. Results Expression of the RASSF10 gene significantly decreased in newly diagnosed MM patients, and was positively correlated with overall survival. RPMI-8226 and OPM-2 cell lines with lower RASSF10 expression were selected for further study. Overexpression of RASSF10 in these two cell lines inhibited proliferation and induced apoptosis. The RASSF10 gene promoter in MM cell lines was hypermethylated, and downregulated after decitabine treatment. Meanwhile, expression of the RASSF10 gene was upregulated. MM cells with overexpression of RASSF10 were injected into nude mice and exerted anti-MM activity in vivo. Conclusions Low expression of RASSF10 contributed to the proliferation of myeloma cells by hypermethylation of its promoter.

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