scholarly journals Long-term reduction of amplified ornithine decarboxylase sequences in human myeloma cells

1995 ◽  
Vol 310 (1) ◽  
pp. 299-303
Author(s):  
J Wahlfors ◽  
S Myöhänen ◽  
V P Korhonen ◽  
L Alhonen ◽  
J Jänne
Keyword(s):  
Enzyme Activity ◽  
Cell Line ◽  
Ornithine Decarboxylase ◽  
Copy Number ◽  
Methylation Status ◽  
Gene Copy Number ◽  
Myeloma Cell ◽  
Gene Copy ◽  
Myeloma Cell Line ◽  
Myeloma Cells

(1) Human myeloma cell line Sultan, resistant to 20 mM difluoro-methylornithine (DFMO) owing to ornithine decarboxylase (ODC) gene amplification, was grown in the absence of DFMO for a period of 10 months. The gene copy number and methylation status of the ODC gene were monitored after withdrawal of DFMO. Moreover, levels of ODC mRNA, immunoreactive ODC protein, ODC activity and polyamine levels were recorded recurrently during the course of the study. (2) The results revealed that ODC gene copy number started to decrease after 4 weeks growth without DFMO, to a final level of less than 30% of the original gene dosage. The methylation status of the ODC gene, however, remained almost unaltered, displaying only a modest increase in methylation after 10 months without DFMO. The amount of ODC message dropped very rapidly to 75% of the original value, then started to decrease in a gene copy-number-dependent manner. The amount of ODC protein closely followed the levels of mRNA during the study, whereas the ODC activity, after a transient increase during the first week, decreased to half of the original level after 4 weeks. Between 6 and 16 weeks ODC activity stabilized to a fifth of the original value and no more changes were detected during the subsequent period of observation. (3) Due to the grossly elevated ODC enzyme activity, levels of putrescine and spermidine first peaked and then stabilized at 6 weeks after DFMO withdrawal. The final spermidine level was comparable with that of the parental Sultan cell line with only one copy of active ODC gene. However, putrescine content was strikingly elevated, being stabilized to a level that was 20 times higher than in parental cells. Spermine concentration was practically unchanged during the study. (4) According to the results obtained in this study, the abnormal level of ODC expression in human myeloma cells is suppressed partially at the level of transcription or post-transcriptionally, but it is not due to increased methylation of the gene. The major regulatory mechanism to compensate for a highly elevated ODC expression was modulation of the enzyme activity. After 10 months without DFMO, the cells still displayed about 20 times higher ODC activity and putrescine concentration than the myeloma cell line with a single copy of the ODC gene. They did not, however, show any signs of growth retardation or other features different from the parental cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Increased Copy Number of Interleukin-6 Receptor Gene Is Associated with Adverse Survival In Multiple Myeloma Patients Treated with Autologous Stem Cell Transplantation

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1895-1895
Author(s):  
Seon Young Kim ◽  
Hyun Jung Min ◽  
Hyun Kyung Park ◽  
Bora Oh ◽  
Tae Young Kim ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Monoclonal Antibody ◽  
Stem Cell ◽  
Cell Line ◽  
Copy Number ◽  
Myeloma Cell ◽  
Newly Diagnosed ◽  
Myeloma Cell Line ◽  
Myeloma Cells

Abstract Abstract 1895 Background: Interleukin-6 (IL-6) is a potent pleiotropic cytokine that regulates plasma cell (PC) growth via IL-6 receptor (IL-6R). We hypothesized that upregulation of IL-6R in myeloma cells might confer the growth privilege to myeloma cells over bone marrow (BM) hematopoietic cells. We investigated the frequency and prognostic implication of increased copy number of IL-6R gene by fluorescence in situ hybridization (FISH) in patients with newly diagnosed multiple myeloma (MM). To confirm the role of IL-6R, the change of proliferative potentials after blocking of IL-6R by monoclonal antibody was observed in a myeloma cell line. Methods: 102 patients with newly diagnosed MM were enrolled. FISH study for IL-6R was performed using home-made probe for the locus. The BAC (bacterial artificial chromosome) clone RP11 350G8 (BACPAC Resources, Oakland, CA) was used as the IL-6R gene-positive clone and the FISH probe was labeled by a nick translation reaction. FISH signals were counted among BM plasma cells sorted by immunoglobulin light chain staining (cIg FISH). U266 myeloma cell line was treated with IL-6R monoclonal antibody (MRA, Chugai, Japan) at concentration of 0.06 to 1.25 μM and then cell viability test were performed. Results: The amplification of IL-6R was detected in 53/102 patients (52.0%). The 5-year overall survival (OS) rate of patients with IL-6R gene amplification was 41.3% versus 44.8% for those with a normal IL-6R (P = 0.425). In 44 patients treated with high-dose chemotherapy and autologous stem cell transplantation (ASCT), patients with 3.1 or more mean IL-6R gene copy number per PC showed worse 5-year OS compared to those who had less than 2.1 mean copies of IL-6R gene (44.4% versus 78.0%, P = 0.024). In multivariate analysis, the increase of IL-6R copy numbers (mean copy/PC ≥ 3.1) could be considered as an independent prognostic factor for MM patients underwent ASCT (hazard ratio, 30.9; 95% CI, 1.74–548.6; P = 0.020). Treatment of IL-6R monoclonal antibody on the U266 cell line induced marked reduction of cell viability compared with control, ranged from 7.1% to 98.7% according to the increase in treated antibody level. Conclusions: The gain of IL-6R gene is frequent in myeloma, showing an association with adverse prognosis in MM patients treated with ASCT. In vitro suppression of cell proliferation by IL-6R blocker suggests the potential role of IL-6R in myeloma cell growth and therapeutic implication of IL-6R blocker in the future. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Levels of ornithine decarboxylase genomic sequences, heterogeneous nuclear RNA and mRNA in human myeloma cells resistant to α-difluoromethylornithine

1991 ◽  
Vol 278 (3) ◽  
pp. 871-874 ◽  
Author(s):  
J M Hyttinen ◽  
M Halmekytö ◽  
L Alhonen ◽  
J Jänne
Keyword(s):  
Ornithine Decarboxylase ◽  
Reverse Transcription ◽  
Copy Number ◽  
Gene Dosage ◽  
Gene Copy Number ◽  
Gene Copy ◽  
Dot Blot ◽  
Semiquantitative Assessment ◽  
Myeloma Cells ◽  
Human Myeloma Cells

We previously isolated and characterized a human myeloma cell line overproducing ornithine decarboxylase (ODC) due to gene amplification [Leinonen, Alhonen-Hongisto, Laine, Jänne & Jänne (1987) Biochem. J. 242, 199-203]. We have now employed the PCR combined with reverse transcription to determine semiquantitatively ODC gene dosage and the amounts of heterogeneous nuclear (hn) RNA and of mature mRNA of the enzyme in parental and alpha-difluoromethylornithine-resistant human myeloma cells. Experiments with dilution series revealed that the ODC gene copy number and the amount of both hnRNA and mRNA were increased to the same extent (about 100-fold) in the resistant cells. Similar dot-blot analyses of ODC-specific genomic DNA and total RNA indicated that the ODC gene copy number was increased by a factor of 380 and the amount of ODC mRNA by a factor of 700. Our results indicate that the PCR combined with reverse transcription is at least as useful as blot analyses to give semiquantitative assessment of the amounts of specific DNA or RNA sequences. In addition, the use of the PCR enables the analysis of minute sample amounts in extremely short time.

scholarly journals Nongenotoxic ABCB1 activator tetraphenylphosphonium can contribute to doxorubicin resistance in MX-1 breast cancer cell line

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Raimonda Kubiliute ◽  
Indre Januskeviciene ◽  
Ruta Urbanaviciute ◽  
Kristina Daniunaite ◽  
Monika Drobniene ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Line ◽  
Protein Level ◽  
Copy Number ◽  
Gene Copy Number ◽  
Gene Copy ◽  
Common Mechanism ◽  
Dna Hypomethylation ◽  
Mesenchymal Transition ◽  
Molecular Features

AbstractHyperactivation of ABC transporter ABCB1 and induction of epithelial–mesenchymal transition (EMT) are the most common mechanism of acquired cancer chemoresistance. This study describes possible mechanisms, that might contribute to upregulation of ABCB1 and synergistically boost the acquisition of doxorubicin (DOX) resistance in breast cancer MX-1 cell line. DOX resistance in MX-1 cell line was induced by a stepwise increase of drug concentration or by pretreatment of cells with an ABCB1 transporter activator tetraphenylphosphonium (TPP+) followed by DOX exposure. Transcriptome analysis of derived cells was performed by human gene expression microarrays and by quantitative PCR. Genetic and epigenetic mechanisms of ABCB1 regulation were evaluated by pyrosequencing and gene copy number variation analysis. Gradual activation of canonical EMT transcription factors with later activation of ABCB1 at the transcript level was observed in DOX-only treated cells, while TPP+ exposure induced considerable activation of ABCB1 at both, mRNA and protein level. The changes in ABCB1 mRNA and protein level were related to the promoter DNA hypomethylation and the increase in gene copy number. ABCB1-active cells were highly resistant to DOX and showed morphological and molecular features of EMT. The study suggests that nongenotoxic ABCB1 inducer can possibly accelerate development of DOX resistance.

scholarly journals Heterogeneous expression of a novel MPC-1 antigen on myeloma cells: possible involvement of MPC-1 antigen in the adhesion of mature myeloma cells to bone marrow stromal cells

Blood ◽  
1993 ◽  
Vol 82 (12) ◽  
pp. 3721-3729 ◽  
Author(s):  
N Huang ◽  
MM Kawano ◽  
H Harada ◽  
Y Harada ◽  
A Sakai ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
Cell Line ◽  
Bone Marrow Stromal Cells ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Myeloma Cell Line ◽  
Myeloma Cells ◽  
Human Myeloma Cell Line ◽  
Heterogeneous Expression

Abstract Recent immunophenotypic analysis has shown that the heterogeneous expression of the adhesion molecule VLA-5 classifies myeloma cells into VLA-5+ mature and VLA-5- immature subpopulations. To further clarify the two myeloma subpopulations, we generated a monoclonal antibody, MPC- 1, by immunizing mice with an adherent human myeloma cell line, KMS-5. The MPC-1 antibody recognized a 48-Kd surface antigen on KMS-5 but not on U-266, a nonadherent human myeloma cell line. Specificity characterization showed that MPC-1 antigen was expressed on mature myeloma cells, normal plasma cells, and mature B cells, whereas pre-B cells and germinal center B cells lacked its expression. Monocytes and a human bone marrow stromal cell line, KM102, also expressed this antigen. Two subclones of MPC-1+ VLA-5+ (KMS-5Ad) and MPC-1-VLA-5+ (KMS- 5NAd) were separated from the KMS-5 cell line. The KMS-5NAd adhered to KM102 more tightly than did the KMS-5NAd, and the U-266 (MPC-1-VLA-5-) displayed almost no adherence to the KM102. The adhesion of the KMS-5Ad was partially inhibited by the MPC-1 antibody. These results, taken together, suggest that the MPC-1 antigen serves as a differentiation marker for B-lineage cells, including plasma cells, and may function as an adhesion molecule involved in the interaction of mature myeloma cells with bone marrow stromal cells.

Enforced CD19 Expression Leads to Growth Inhibition and Reduced Tumorigenicity

Blood ◽  
1999 ◽  
Vol 94 (10) ◽  
pp. 3551-3558 ◽  
Author(s):  
Maged S. Mahmoud ◽  
Ryuichi Fujii ◽  
Hideaki Ishikawa ◽  
Michio M. Kawano
Keyword(s):  
Growth Inhibition ◽  
Cell Line ◽  
Myeloma Cell ◽  
Inhibitory Effect ◽  
Myeloma Cell Line ◽  
Growth Inhibitory Effect ◽  
Myeloma Cells ◽  
Growth Inhibitory ◽  
Human Myeloma Cell Line

In multiple myeloma (MM), the cell surface protein, CD19, is specifically lost while it continues to be expressed on normal plasma cells. To examine the biological significance of loss of CD19 in human myeloma, we have generated CD19 transfectants of a tumorigenic human myeloma cell line (KMS-5). The CD19 transfectants showed slower growth rate in vitro than that of control transfectants. They also showed a lower capability for colony formation as evaluated by anchorage-independent growth in soft agar assay. The CD19 transfectants also had reduced tumorigenicity in vivo when subcutaneously implanted into severe combined immunodeficiency (SCID)-human interleukin-6 (hIL-6) transgenic mice. The growth-inhibitory effect was CD19-specific and probably due to CD19 signaling because this effect was not observed in cells transfected with a truncated form of CD19 that lacks the cytoplasmic signaling domain. The in vitro growth-inhibitory effect was confirmed in a nontumorigenic human myeloma cell line (U-266). However, introduction of the CD19 gene into a human erythroleukemia cell line (K-562) also induced growth inhibition, suggesting that this effect is CD19-specific, but not restricted to myeloma cells. These data suggest that the specific and generalized loss of CD19 in human myeloma cells could be an important factor contributing to the proliferation of the malignant plasma cell clones in this disease.

Expression of TRAIL Receptors on the Multiple Myeloma Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4868-4868
Author(s):  
Juan Li ◽  
Junhe Li ◽  
Shaokai Luo ◽  
Yin Zhao
Keyword(s):  
Multiple Myeloma ◽  
Cell Line ◽  
Mononuclear Cells ◽  
Myeloma Cell ◽  
Myeloma Cell Line ◽  
Killing Effect ◽  
Myeloma Cells ◽  
Decoy Receptors ◽  
Trail Receptors ◽  
Chemotherapy Agents

Abstract Objective To study the different expression of death receptors and decoy receptors on mononuclear cells from patients with multiple myeloma and myeloma cell line KM3 and compare the different expression of TRAIL receptors after chemotherapy or exposure to doxorubicin, to explore the mechanisms by which TRAIL selectively kills tumor cells. Methods Semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and flow cytometry was used to investigate the expression of four receptors on mononuclear cells in 23 multiple myeloma patients and myeloma cell line KM3 and 15 controls, we furthermore compared the changes of expression mode after chemotherapy and incubation of KM3 cell with sub-clinical concentration of Doxorubicin. Results There finds only DR4 and DR5 on KM3 cell line without the expression of DcR1 and DcR2. Expression of DR4 and DR5 on mononuclear cells of MM patients is higher than that of controls (P<0.05), but DcR1 and DcR2 expression was lower than that of controls (P<0.05), after chemotherapy and exposure to Doxorubicin, the expression of DR5 on MM cells was up-regulated (P<0.05) Conclusions The expression of four receptors on myeloma cells and normal controls was significantly different, which might account for the selective killing effect of TRAIL on MM cells. DR5 was up-regulated on KM3 when incubating with Doxorubicin and after chemotherapy which suggests chemotherapy agents might enhance the apopotosis of MM cells through up-regulating of DR5 receptor.

Multiple Myeloma: Interleukin-15 Antagonizes Bone Morphogenetic Protein-Induced Apoptosis and Growth Inhibition.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3444-3444
Author(s):  
Magne Rekvig ◽  
Anne-Tove Brenne ◽  
Torstein Baade Ro ◽  
Anders Waage ◽  
Magne Borset ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Growth Inhibition ◽  
Cell Line ◽  
Caspase 3 ◽  
Myeloma Cell ◽  
Stimulus Figure ◽  
Myeloma Cell Line ◽  
Myeloma Cells ◽  
Induced Apoptosis

Abstract Multiple myeloma has two distinct features: Expansion of malignant plasma cells within the bone marrow accompanied by skeletal destruction. Bone morphogenetic proteins (BMPs) have been shown to induce apoptosis and inhibit growth in myeloma cells. BMPs are members of the TGF-β superfamily of proteins capable of inducing bone formation, and regulate proliferation, differentiation and apoptosis. We have investigated myeloma cell apoptosis and proliferation with BMP-4 and −6 in concert with the myeloma cell growth factors interleukin (IL)-2, IL-6, IL-10, IL-15, IL-21, tumor necrosis factor (TNF)-α and insulin-like growth factor (IGF)-1. Eight samples of highly purified myeloma cells from patients and a human myeloma cell line, IH-1 (Brenne AT et al. Blood. 2002 May 15;99(10):3756–62.), were used in this study. Cytokine concentrations used in the referred experiments were for BMP-4 20ng/ml, BMP-6 250ng/ml, IL-15 20ng/ml and IL-6 0,1ng/ml, respectively. Growth inhibition was measured in a proliferation assay by methyl-[3H]-thymidine incorporation and apoptosis by annexin V- FITC-binding/PI-uptake on flow cytometry. IL-15 antagonized growth inhibition (Figure A) and prevented apoptosis induced by BMP-4 (Figure B) and BMP-6 in the myeloma cell line IH-1. IL-15 also antagonized the growth inhibition induced by BMP-4 and/or BMP-6 in three out of eight patient samples. Neither IL-6, nor any of the other investigated cytokines were able to rescue the myeloma cells from growth inhibition and apoptosis induced by BMP-4 and -6. Among the investigated cytokines, we found that IL-15 has a unique capability to antagonize BMP- induced apoptosis and growth inhibition in myeloma cells. We examined cleavage of the proapoptotic protein caspase-3 and found that BMP-4 activated caspase-3 in the IH-1 cell line. This activation of caspase-3 was blocked by IL-15 but not by IL-6. We have demonstrated a possible mechanism for myeloma cells to escape apoptosis and growth-inhibition within the bone marrow. Intramedullar levels of IL-15 and BMPs may play a role in the pathogenesis of multiple myeloma. Figure A. Proliferation in response to BMP-4 stimulus Figure A. Proliferation in response to BMP-4 stimulus Figure B. Apoptosis in response to BMP-4 stimulus Figure B. Apoptosis in response to BMP-4 stimulus

MYC Responsible for down-Regulation of CD33 Expression in Human Myeloma Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 5116-5116
Author(s):  
Karim Shamsasenjan ◽  
Ken-ichiro Otsuyama ◽  
Mohd S. Iqbal ◽  
Maged S. Mahmoud ◽  
Michio M. Kawano
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Myeloma Cell ◽  
Myeloma Cell Line ◽  
Down Regulation ◽  
Myeloma Cells ◽  
Expression Levels ◽  
Stat3 Phosphorylation ◽  
Serum Crp ◽  
Human Myeloma Cells

Abstract Human myeloma cells from about 10% of cases with multiple myeloma expressed CD33 and have monocytoid morphology with convoluted nuclei, and all these patients had no increase in serum CRP values. In CD33(+) myeloma cells as well as myeloma cell lines, CD33 expression levels were correlated with the increased expression levels of CEBPA (C/EBPα) gene. This correlation was confirmed by the finding that transfection with the CEBPA gene induced CD33 expression in a CD33(−) myeloma cell line. As suggested by the lack of an increase in serum CRP values in CD33(+) myelomas, IL-6 down-regulated the expression of CD33 in CD33(+) myeloma cell lines along with the down-regulation of CEBPA gene expression. Cucurbitacin I (STAT3 inhibitor) but not U0126 (MAPK inhibitor) could abolish the effect of IL-6. Furthermore, IL-6 up-regulated the expression of MYC via STAT3 phosphorylation and MYC bound to the promoter region of CEBPA gene followed by the down-regulation of the CEBPA expression. It was confirmed that introduction of shRNA for MYC into a CD33(+) myeloma cell line blocked the IL6-induced down-regulation of CD33 and CEBPA expression. Therefore, these results indicate that IL-6 can reverse the expression level of CD33 by up-regulating MYC followed by the down-regulation of CEBPA expression.

Panobinostat Induces Upregulation of CD38 and Augments the Anti-Myeloma Efficacy of Daratumumab in Pre-Clinical Models

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4481-4481 ◽  
Author(s):  
Estefania Garcia-Guerrero ◽  
Tea Gogishvili ◽  
Sophia Danhof ◽  
Martin Schreder ◽  
Celine Pallaud ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Flow Cytometry ◽  
Cell Line ◽  
Myeloma Cell ◽  
Target Cells ◽  
Myeloma Cell Line ◽  
Myeloma Cells ◽  
Cd38 Expression ◽  
Clinical Models ◽  
Pre Treatment

Abstract Background: Immunotherapy with monoclonal antibodies (mAbs) has recently entered the clinical arena in multiple myeloma, including Daratumumab that targets CD38 on malignant plasma cells. The efficacy of mAbs depends on antigen density and expression of accessory ligands on target cells to initiate cell- and complement-dependent effector mechanisms. Here, we investigate the use of the histone deacetylase inhibitor (HDACi) Panobinostat to modulate target antigen expression and ligand profile on myeloma in favor of potent mAb-mediated recognition and destruction. We show that Panobinostat augments CD38 expression specifically on myeloma cells and demonstrate powerful synergy with anti-CD38 mAb Daratumumab in pre-clinical models. Methods: The myeloma cell line MM1.S and primary myeloma cells were treated with titrated doses of Panobinostat (0, 10, 25 nM) and expression of CD38 and a panel of additional target molecules including B-cell maturation antigen (BCMA) and SLAMF7, as well as accessory ligands analyzed by flow cytometry at 24, 48 and 72 hours. Antibody-dependent cellular cytotoxicity (ADCC) against Panobinostat treated and untreated myeloma cells was analyzed at 4 and 20 hours after addition of PBMC at an effector to target ratio of 25:1 in the presence of Daratumumab (1, 10, 50 ug/mL) or an isotype control antibody. Results: We first treated the myeloma cell line MM1.S with Panobinostat and analyzed its direct cytotoxic anti-myeloma effect. Consistent with previous work, the percentage of live MM1.S myeloma cells had decreased to 85% and 50% after 48 hours of exposure to 10 and 25 nM respectively. We analyzed expression of CD38 on residual live, i.e. 7-AAD negative MM1.S cells by flow cytometry and observed a 1.5 (10 nM) and 2-fold (25 nM) increase of CD38 expression by mean fluorescence intensity (MFI) compared to baseline levels and untreated control cells. The increase in CD38 expression was already detectable after 24 hours and plateaued between 48 and 72 hours. We confirmed our observation in primary myeloma cells from multiple donors (n=4) and detected an even stronger increase to 2 (10 nM) and 4-fold (25 nM) higher CD38 expression compared to untreated cells at 48 hours. Interestingly, expression of BCMA and SLAMF7 was not increased after Panobinostat treatment at all tested concentrations and time points in both MM1.S and primary myeloma. We confirmed that Panobinostat-induced upregulation of CD38 specifically occurred in myeloma, and neither observed this phenomenon in a panel of leukemia and lymphoma cell lines including Raji (Burkitt) and JeKo-1 (mantle cell), nor on resting/activated primary CD8+ and CD4+ T cells that we isolated from peripheral blood of several donors (n=3). Next, we were interested in determining whether the increase in CD38 expression enabled superior anti-myeloma activity of the anti-CD38 mAb Daratumumab. Panobinostat pre-treatment was done for 48 hours at 10 nM as this is a clinically achievable serum level with currently approved regimens. Indeed, significantly higher ADCC was mediated by Daratumumab at all tested concentrations (1, 10 and 50 ug/mL) against MM1.S that we had exposed to Panobinostat. At 4 hours, ADCC was 45% and 25% in Panobinostat-treated and untreated MM1.S respectively, and at 20 hours, near-complete, >90% ADCC of Panobinostat-pre-treated MM1.S had occurred, whereas only 65% of MM1.S were eliminated by Daratumumab without Panobinostat pre-treatment. These data were confirmed in multiple experiments with MM1.S and PBMC from different donors, and with primary myeloma cells. Experiments to evaluate synergy of Panobinostat and Daratumumab therapy in a xenograft model (NSG/MM1.S) are ongoing. Conclusions: Our data demonstrate that the HDACi Panobinostat induces upregulation of CD38 on myeloma and a subsequent dramatic increase of Daratumumab-mediated ADCC in pre-clinical models. These data suggest that Panobinostat could be used synergistically with Daratumumab in a clinical setting to increase response rates and extend duration of responses to Daratumumab. Panobinostat has a known ability to modulate the transcriptional profile of myeloma cells and our data demonstrate for the first time that this ability can be utilized to augment the therapeutic index of antibody-based immunotherapy in multiple myeloma. Disclosures Pallaud: Novartis: Employment. Lehmann:Novartis: Employment. Hudecek:Novartis: Research Funding.

Sphingolipids as a novel target for the treatment of multiple myeloma.

2013 ◽  
Vol 31 (15_suppl) ◽  
pp. e19534-e19534
Author(s):  
Yubin Kang ◽  
Jagadish Kummetha Venketa
Keyword(s):  
Multiple Myeloma ◽  
Cell Death ◽  
Cell Line ◽  
Cell Lines ◽  
Apoptotic Cell ◽  
Myeloma Cell ◽  
Sphingolipid Metabolism ◽  
Myeloma Cell Line ◽  
Myeloma Cells ◽  
Novel Target

e19534 Background: Multiple myeloma (MM) is the second most common hematological malignancy in the United States and accounts for ~10,600 deaths annually. MM remains an incurable disease and almost all patients will eventually relapse and become refractory to currently available therapeutic agents. There is an unmet need for better understanding the disease’s molecular pathways and for identifying novel therapeutic targets. Sphingolipid metabolism is being increasingly recognized as a key pathway in tumor cell proliferation and in tumor sensitivity to anticancer drugs. We hypothesize that altered sphingolipid metabolism plays an important role in the pathogenesis of MM, thus providing a novel target in the treatment of MM. Methods: We first assayed sphingolipid metabolism including sphingolipid metabolites and sphingolipid metabolizing genes in myeloma cell lines, in freshly isolated human primary CD138+myeloma cells, and in publically available dataset. We then tested the efficacy of the selective SK2 inhibitor (ABC294640) and the SK2 shRNA in killing myeloma cells in vitro. Results: 1) Compared to immortalized B cells, the levels of pro-apoptotic ceramides were decreased whereas the proliferative sphingosine 1-phosphate (S1P) was increased in myeloma cell lines. 2) The expression of several key sphingolipid-metabolizing genes including sphingosine kinase (SK) 1 and 2 was altered in freshly isolated human primary bone marrow myeloma cells and in publically available microarray dataset. 3) The selective SK2 inhibitor (ABC294640) induces apoptotic cell death and inhibits myeloma cell growth with an IC50of ~20 μM in 9 myeloma cell lines. 4) Interestingly, OPM-1 myeloma cell line was extremely sensitive to ABC294640 with an IC50of <5 µM whereas U266 myeloma cell line was resistant to ABC294640. SK2 shRNA induced apoptotic cell death in OPM-1, but not in U266 cells. We are currently investigating the molecular mechanisms underlying the resistance of U266 myeloma cells to ABC294640. Conclusions: Our data demonstrated that sphingolipid metabolism provides an attractive target in the treatment of refractory/relapased multiple myeloma.

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