The Novel JAK2 Inhibitor NVP-BSK805 Has Cytotoxic Activity on Malignant Plasma Cells

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2993-2993
Author(s):  
Renate Burger ◽  
Franziska Rademacher ◽  
Matthias Staudinger ◽  
Matthias Peipp ◽  
Andreas Güunther ◽  
...  
Keyword(s):  
Cytotoxic Activity ◽  
Cell Lines ◽  
Plasma Cell ◽  
Stromal Cells ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Synergistic Activity ◽  
Jak Inhibitors ◽  
Inhibitory Effects ◽  
Jak2 Inhibitor

Abstract Abstract 2993 In multiple myeloma (MM) and plasma cell leukemia, activation of the JAK/STAT pathway is induced by interleukin (IL)-6, which is produced and secreted into the tumor microenvironment primarily by stromal cells. Upon binding of IL-6 to its specific alpha-chain receptor, dimerization of the gp130 signaling subunits leads to activation of associated JAK kinases and STAT transcription factors. In particular, STAT3 has been shown to be essential for myeloma cell growth and survival. NVP-BSK805 (Novartis) is a novel substituted quinoxaline JAK2 inhibitor tool compound which displays more than 20-fold selectivity for JAK2 over the other JAK family members and more than 100-fold selectivity over a panel of additional kinases (Baffert et al., Mol Cancer Ther 9:1945, 2010). The study presented here aims at growth inhibitory effects of NVP-BSK805 in malignant plasma cells. NVP-BSK805 inhibited the growth of six human myeloma cell lines displaying dose-dependent activity with IC50 concentrations between 2.6 μ mol/L and 6.8 μ mol/L. Among the cell lines, IL-6 dependent INA-6 cells were most sensitive to the inhibitory effects of the compound: both IL-6 and bone marrow stromal cell induced proliferation as measured by [3H]-thymidine uptake was completely inhibited at 4 μ mol/L and IC50 concentrations were less than 1 μ mol/L. Viability of the stromal cells was not significantly affected. NVP-BSK805 concentrations as low as 0.5 μ mol/L were sufficient to yield a marked reduction of IL-6 induced STAT3 phosphorylation and complete abrogation at 2 μ mol/L, thereby blocking essential survival signals. Accordingly, treatment of INA-6 cells with NVP-BSK805 for 48 hours led to significant apoptosis starting at 2 μ mol/L with a 30% increase in annexin V-positive cell numbers compared to DMSO controls. Importantly, NVP-BSK805 showed potent cytotoxic activity on plasma cell-enriched primary tumor samples from patients with extramedullary plasma cell disease that are highly responsive to IL-6: in 3 out of 4 tumor samples the IC50 concentrations were between 0.5 μ mol/L and 0.6 μ mol/L. These studies are extended to combinations of NVP-BSK805 with PI3K, mToR, MAPK, HDAC, and IGF-1R inhibitors in order to optimize targeted therapy strategies facing different pathway alterations in individual myeloma patients. In INA-6 cells, synergistic activity was found combining NVP-BSK805 with rapamycin and the MEK1 inhibitor U0126. Preclinical in vivo studies are ongoing. Our results with NVP-BSK805 substantiate the use of JAK inhibitors as a therapeutic strategy for patients with MM. Since results from studies with pan-JAK inhibitors such as pyridone 6 indicate involvement of additional JAK kinases, the choice of the optimal compound will depend on its JAK family selectivity and the biology of JAK signaling in MM. Disclosures: No relevant conflicts of interest to declare.

Inhibition of Interleukin-6 Signaling with CNTO 328 Sensitizes Plasma Cell Dyscrasias to the Effects of Melphalan in Association with Suppression of Signaling through Protein Kinase B/Akt.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1836-1836
Author(s):  
Sally A. Hunsucker ◽  
Valeria Magarotto ◽  
Jairo A. Matthews ◽  
Michael Wang ◽  
Veerabhadran Baladandayuthapani ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Protein Kinase ◽  
Cell Lines ◽  
Plasma Cell ◽  
Stromal Cells ◽  
Plasma Cells ◽  
Protein Kinase B ◽  
Myeloma Cell ◽  
Ortho Biotech ◽  
Plasma Cell Dyscrasias

Abstract Abstract 1836 Poster Board I-862 Background: The neutralizing anti-interleukin (IL)-6 monoclonal antibody (MAb) CNTO 328 acts in an additive to synergistic manner to enhance the activity of bortezomib and dexamethasone against models of multiple myeloma by suppressing several IL-6-induced anti-apoptotic signaling pathways. We therefore sought to evaluate the possibility that blockade of IL-6 signaling could also augment the activity of melphalan, and to determine the potential mechanisms underlying this interaction. Methods: A panel of myeloma cell lines was studied both in suspension and with bone marrow stromal cells to evaluate the activity of CNTO 328 with and without melphalan. The CNTO 328 + melphalan combination was also tested in primary cells from patients with a variety of plasma cell dyscrasias. Results: Treatment of IL-6-dependent KAS-6/1, INA-6, and ANBL-6 myeloma cell lines with CNTO 328 + melphalan reduced plasma cell viability in an additive-to-synergistic manner compared to melphalan with a control MAb. Isobologram analysis demonstrated that the combination was synergistic in KAS-6/1 cells regardless of the sequence of drug treatment (combination indices (CIs) from 0.275-0.607), although the strongest synergy was seen with CNTO 328 pretreatment (CIs from 0.275-0.493). These anti-proliferative effects were accompanied by an enhanced activation of drug-specific apoptosis, and this increased cell death was not rescued by the trophic effects of co-culture of plasma cells with the human-derived stromal cell line HS-5. CNTO 328 increased melphalan-mediated induction of both extrinsic, caspase-8-mediated apoptosis, as well as intrinsic, caspase-9-mediated death, which converged to produce increased levels of caspase-3 activity. Apoptosis was enhanced in part by CNTO 328-stimulated cleavage of Bid to tBid, and alterations in the phosphorylation status of BimEL, as well as increased conversion of Bak and, to a lesser extent, of Bax, to their active forms. Neutralization of IL-6 by CNTO 328 also suppressed signaling through the protein kinase B/Akt pathway, as evidenced by decreased levels of phospho-Akt, and decreased activation of several downstream Akt targets, including p70 S6 kinase and 4E-BP1. Importantly, CNTO 328 + melphalan showed enhanced anti-proliferative effects compared to melphalan and a control MAb against primary CD138+ plasma cells derived from patients with multiple myeloma, monoclonal gammopathy of undetermined significance, and amyloidosis, while demonstrating less toxicity to stromal cells. The enhanced effect of the CNTO 328 + melphalan combination was statistically significant compared to either drug alone (p<0.05) in CD138+ cells isolated from patients who had not received prior melphalan therapy. Conclusions: These studies provide a rationale for translation of CNTO 328 into the clinic in combination with melphalan-based therapies, including either high dose therapy in transplant-eligible patients, or standard dose melphalan-containing induction regimens in transplant-ineligible patients, such as with the combination of bortezomib, melphalan, and prednisone. Disclosures: Voorhees: Millennium Pharmaceuticals: Speakers Bureau; Celgene: Speakers Bureau. Xie:Centocor Ortho Biotech Inc.: Employment. Cornfeld:Centocor Ortho Biotech Inc.: Employment. Nemeth:Centocor Ortho Biotech Inc.: Employment.

scholarly journals Phenotypic analysis of human myeloma cell lines

Blood ◽  
1989 ◽  
Vol 73 (2) ◽  
pp. 566-572
Author(s):  
C Duperray ◽  
B Klein ◽  
BG Durie ◽  
X Zhang ◽  
M Jourdan ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lines ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Light Chains ◽  
Cell Phenotype ◽  
Phenotypic Analysis ◽  
Barr Virus ◽  
Human Myeloma Cell

Multiple myeloma (MM) is a B-cell malignancy characterized by the accumulation, primarily in bone marrow, of a clone of plasma cells. The nature of the stem cells feeding the tumoral compartment is still unknown. To investigate this special point, we have studied the phenotypes of nine well-known human myeloma cell lines (HMCLs) and compared them with those of normal lymphoblastoid cell lines (LCLs). Twenty-four clusters of differentiation involved in B lymphopoiesis were investigated using a panel of 65 monoclonal antibodies (MoAbs). For each cluster, the percentage of positive cells and the antigen density were determined, giving rise to a “quantitative phenotype”. We thus classified the HMCLs into two different groups: those with cytoplasmic mu chains (c mu+) and those without (c mu-). In the first (c mu+) group, comprising seven cell lines, the HMCLs had a phenotype of pre-B/B cells close to that of Burkitt's lymphoma cell lines. They expressed low densities of surface mu chains, without detectable cytoplasmic or surface light chains. Three of them were infected with the Epstein Barr virus (EBV). These c mu+ HMCLs bore most of the B-cell antigens except CD23. They expressed the CALLA antigen (CD10) and lacked the plasma-cell antigen PCA1. In contrast, LCLs expressed surface light chains, high densities of CD23, low densities of PCA1 antigen, and no CD10 antigen. The c mu- HMCLs had a plasma-cell phenotype, lacking most of the B-cell antigens and expressing high densities of PCA1 antigen.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Plasma cells induce apoptosis of pre-B cells by interacting with bone marrow stromal cells

Blood ◽  
1996 ◽  
Vol 87 (8) ◽  
pp. 3375-3383 ◽  
Author(s):  
T Tsujimoto ◽  
IA Lisukov ◽  
N Huang ◽  
MS Mahmoud ◽  
MM Kawano
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Cell Survival ◽  
Cell Lines ◽  
Stromal Cells ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Myeloma Cells ◽  
B Cell Survival

By using two-color phenotypic analysis with fluorescein isothiocyanate- anti-CD38 and phycoerythrin-anti-CD19 antibodies, we found that pre-B cells (CD38+CD19+) signifcantly decreased depending on the number of plasma cells (CD38++CD19+) in the bone marrow (BM) in the cases with BM plasmacytosis, such as myelomas and even polyclonal gammopathy. To clarify how plasma cells suppress survival of pre-B cells, we examined the effect of plasma cells on the survival of pre-B cells with or without BM-derived stromal cells in vitro. Pre-B cells alone rapidly entered apoptosis, but interleukin-7 (IL-7), a BM stromal cell line (KM- 102), or culture supernatants of KM-102 cells could support pre-B cell survival. On the other hand, inhibitory factors such as transforming growth factor-beta1 (TGF-beta1) and macrophage inflammatory protein- 1beta (MIP-1beta) could suppress survival of pre-B cells even in the presence of IL-7. Plasma cells alone could not suppress survival of pre- B cells in the presence of IL-7, but coculture of plasma cells with KM- 102 cells or primary BM stromal cells induced apoptosis of pre-B cells. Supernatants of coculture with KM-102 and myeloma cell lines (KMS-5) also could suppress survival of pre-B cells. Furthermore, we examined the expression of IL-7, TGF-beta1, and MIP-1beta mRNA in KM-102 cells and primary stromal cells cocultured with myeloma cell lines (KMS-5). In these cells, IL-7 mRNA was downregulated, but the expression of TGF- beta1 and MIP-1beta mRNA was augmented. Therefore, these results suggest that BM-derived stromal cells attached to plasma (myeloma) cells were modulated to secrete lesser levels of supporting factor (IL- 7) and higher levels of inhibitory factors (TGF-beta1 and MIP-1beta) for pre-B cell survival, which could explain why the increased number of plasma (myeloma) cells induced suppression of pre-B cells in the BM. This phenomenon may represent a feedback loop between pre-B cells and plasma cells via BM stromal cells in the BM.

The JAK1/2 Inhibitor Ruxolitinib Kills Malignant Plasma Cells and Is Synergistic with PI3K and mToR Inhibitors

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1844-1844
Author(s):  
Renate Burger ◽  
Tim Bugdahn ◽  
Matthias Staudinger ◽  
Anna Lena Michaelis ◽  
Matthias Peipp ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Cell Growth ◽  
Growth Inhibition ◽  
Cytotoxic Activity ◽  
Signaling Pathways ◽  
Cell Lines ◽  
Plasma Cell ◽  
Stromal Cell ◽  
Plasma Cells ◽  
Combination Index

Abstract Abstract 1844 Malignant plasma cell growth and survival in multiple myeloma (MM) is regulated by cytokines produced in the tumor environment. Specifically, IL-6 plays a key role by activating important signaling pathways through its gp130 receptor associated Janus kinases (JAK). Ruxolitinib (INC424/INCB018424; Novartis/Incyte) is the first small molecule JAK inhibitor approved for the treatment of patients with myelofibrosis. It is a potent inhibitor of both JAK1 and JAK2 and has an approx. 6-fold selectivity against Tyk2 and marked selectivity against JAK3 (more than 130-fold) and additional kinases. The aim of our study was to evaluate the effects of ruxolitinib on malignant plasma cells as well as its activity in combination with other pathway inhibitors. Ruxolitinib activity was evaluated in MTS-based colorimetric cell growth assays or by [3]H-thymidine uptake. IC50 concentrations and combination index (CI) were calculated with CalcuSyn (Biosoft). Evaluating seven human plasma cell lines, ruxolitinib showed a strong cytotoxic activity on the only IL-6 dependent line INA-6 (IC50 0.23 μM). Complete growth inhibition was achieved at 1 μM, even in the presence of bone marrow stromal cells, whereas stromal cell viability and IL-6 production, as measured by specific ELISA, were maintained. Consistent with the dose-dependent inhibition of IL-6 induced STAT3 phosphorylation, apoptosis was induced, resulting in 39% and 63% annexin V-positive cells in the presence of 1 μM ruxolitinib after 48 or 72 hours, respectively. Likewise, significant growth inhibition was seen in purified tumor cells from a patient with plasma cell leukemia that were stimulated with IL-6 (IC50 0.16 μM), and in a LIF-responsive tumor subline of INA-6 (IC50 0.12 μM). In contrast, autonomously growing MM cell lines were not directly inhibited by ruxolitinib, pointing to the kinase specificity of the drug. However, IL-6 mediated drug resistance can be reversed as shown in dexamethasone-sensitive MM1.S cells. Simultaneous inhibition of JAKs with additional signaling pathways that may be activated in myeloma cells by mutations and/or cytokines such as insulin-like growth factor-1, is hypothesized to result in increased cytotoxicity. In INA-6 cells, p44/p42 MAPK activation due to mutated N-Ras and phosphorylation of S6 protein, a downstream target of the PI3K/AKT/mToR pathway, were not abrogated by ruxolitinib. Using combinations of ruxolitinib with inhibitors of PI3K (Ly294002, NVP-BKM120) and mToR (rapamycin), synergistic effects were achieved with a combination index (CI) <1 at the effective dose levels ED50, ED75 and ED90. Other combinations are currently under evaluation. In conclusion, ruxolitinib has strong direct cytotoxic activity against malignant plasma cells that are dependent on JAK/STAT pathway activation. The rationale exists to combine it with inhibitors of complementary pathways and other drugs to potentiate its activity and overcome cytokine or stromal cell mediated drug resistance. Thus, ruxolitinib, as a generally well tolerated drug, may offer therapeutic options for patients with MM. Clearly, the identification of molecular markers may be helpful to assess its precise use alone or in combination, and to select for patients who will benefit from such a treatment. Disclosures: No relevant conflicts of interest to declare.

Expression of CD21 antigen on myeloma cells and its involvement in their adhesion to bone marrow stromal cells

Blood ◽  
1995 ◽  
Vol 85 (12) ◽  
pp. 3704-3712 ◽  
Author(s):  
N Huang ◽  
MM Kawano ◽  
MS Mahmoud ◽  
K Mihara ◽  
T Tsujimoto ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Lines ◽  
Stromal Cells ◽  
Plasma Cells ◽  
Cell Types ◽  
Myeloma Cell ◽  
Adhesion Assay ◽  
Myeloma Cells ◽  
Stromal Cell Line

The mature myeloma cells express very late antigen 5 (VLA-5) and MPC-1 antigens on their surface and adhere to bone marrow (BM) stromal cells more tightly than the VLA-5-MPC-1-immature myeloma cells in vitro. The VLA-5 and MPC-1 antigens possibly function as two of the molecules responsible for interaction of mature myeloma cells with BM stromal cells. However, the immature myeloma cells do interact with BM stromal cells, and it is unclear which adhesion molecules mediate their interaction. In this study, we found that both immature and mature myeloma cells expressed CD21, an adhesion molecule known to bind to CD23. CD21 was also detected on normal plasma cells. To evaluate the role of CD21 expression on myeloma cells, two myeloma cell lines, NOP-2 (VLA-5-MPC-1-) and KMS-5 (VLA-5+MPC-1+), were used as representatives of immature and mature myeloma cell types, respectively, and an adhesion assay was performed between the myeloma cell lines and BM stromal cells. Antibody-blocking results showed that adhesion of the mature type KMS-5 to KM102, a human BM-derived stromal cell line, or to short-term cultured BM primary stromal cells was inhibited by monoclonal antibodies (MoAbs) against CD21, VLA-5, and MPC-1, and inhibition of adhesion of the immature type NOP-2 to KM102 by the anti-CD21 MoAb was observed as well. Furthermore, CD23 was detected on KM102. Treatment of KM102 with an anti-CD23 MoAb also inhibited adhesion of either KMS-5 or NOP-2 to KM102. Therefore, we propose that CD21 expressed on myeloma cells likely functions as a molecule responsible for the interaction of immature myeloma cells as well as mature myeloma cells with BM stromal cells, and CD23 may be the ligand on the stromal cells for the CD21-mediated adhesion.

The Immunotoxin HM1.24-ETA′ Is a Potent Inducer of Apoptosis in Malignant Plasma Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3658-3658
Author(s):  
Matthias Staudinger ◽  
Anja Muskulus ◽  
Renate Burger ◽  
Andreas Guenther ◽  
Roland Repp ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Cell Lines ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Annexin V ◽  
Myeloma Cell ◽  
Parp Cleavage ◽  
Single Chain ◽  
Potent Inducer ◽  
Promising Target

Abstract Despite new treatment modalities, the clinical outcome of at least a subgroup of patients with multiple myeloma (MM) still needs improvement. Antibody-based targeted therapies are increasingly used for tumor therapy, and may represent interesting options for MM patients. HM1.24 is a surface molecule that is over expressed on malignant plasma cells and efficiently internalized from the cell surface. It may represent a promising target for the development of myeloma-directed immunoconstructs. Here, the development and characterization of a novel single-chain immunotoxin, HM1.24-ETA′, is described. HM1.24-ETA′ was generated by genetic fusion of an HM1.24-specific single-chain Fv (scFv) antibody and a truncated variant of Pseudomonas aeruginosa exotoxin A (ETA′). The immunotoxin was expressed in E. coli and purified to homogeneity by affinity chromatography. Specific binding to HM1.24 was demonstrated by immunofluorescence staining and flow cytometry using antigen positive and negative cells. HM1.24-ETA′ efficiently inhibited growth of IL-6 dependent and IL-6 independent myeloma cell lines (INA-6, RPMI8226, U266). Half maximal growth inhibition was observed at low nanomolar concentrations. Further analyses demonstrated that target cell killing occurred via induction of apoptosis, as evidenced by Annexin V/propidium iodide staining and detection of PARP cleavage. The cytotoxic effect was completely blocked by adding excess of unconjugated parental antibody, demonstrating that the effect was antigen-specific and not mediated by non-specific uptake of the immunotoxin. Importantly, HM1.24-ETA′ efficiently triggered apoptosis (>80% Annexin V positive cells) of freshly isolated plasma cell leukemia cells within 48h. In conclusion, HM1.24-ETA′ efficiently triggered apoptosis of multiple myeloma cell lines as well as freshly isolated tumor cells. These results indicate that HM1.24 may represent a promising target structure for efficient antigen-specific delivery of cytotoxic compounds to plasma cell tumors.

GPRC5D Is a Cell Surface Plasma Cell Marker Whose Expression Is High In Myeloma Cells and Reduced Following Coculture With Osteoclasts

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3099-3099 ◽  
Author(s):  
Sathisha Upparahalli venkateshaiah ◽  
Rakesh Bam ◽  
Xin Li ◽  
Sharmin Khan ◽  
Wen Ling ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Surface ◽  
Cell Lines ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Nuclear Fraction ◽  
Myeloma Cells ◽  
Cell Markers ◽  
Qrt Pcr

Abstract Identification of unique cell surface markers on myeloma cells is important for development of targeted therapies and detection of residual disease. GPRC5D is an orphan receptor reportedly expressed at high level in bone marrow (BM) aspirates or biopsies from myeloma patients. Other studies detected GPRC5D in skin and brain but not in other tissues. The aims of the study were to assess GPRC5D gene expression and cell surface localization in myeloma cells, the changes in expression of GPRC5D and other typical myeloma cell markers following coculture of primary myeloma cells with osteoclasts and the consequences of GPRC5D gene silencing on myeloma cell growth. Global gene expression profile (GEP), qRT-PCR and immunohistochemistry revealed exclusive high expression of GPRC5D in normal and myeloma plasma cells but not in B cells, MSCs, osteoclasts or BM mononucleated cells. GPRC5D expression was higher in myeloma plasma cells from newly diagnosed patients (n=698) compared to normal plasma cells (n=26, p<0.0001)). Among molecularly classified groups GPRC5D expression is higher in MS, MF and LB subgroups and lowest in CD2 and MM cell lines. Flow cytometry analysis and immunohistochemistry detected cell surface expression of GPRC5D in myeloma plasma cells while nuclear localization was also detected in certain myeloma cell lines (e.g. H929). Western blots analysis confirmed GPRC5D expression in whole cell lysate and nuclear fraction. We and other demonstrated phonotypical plasticity of myeloma plasma cells capable of altered expression of recognizable plasma cell markers (e.g. CD138, CD45) following coculture with stromal cells (Dezorella et al., 2009) or osteoclasts (Yaccoby, 2005). In coculture of primary CD138-selected myeloma cells with osteoclasts (n=8), CD138 (p<0,004), CD38 (p<0.001) and GPRC5D (p<0.007) were commonly and significantly downregulated, CD45 (p<0.02) was upregulated, and IRF4 expression was unaffected in cocultured myeloma cells compared to the control freshly obtained uncultured myeloma cells assessed by GEP and qRT-PCR. We also observed reduced expression of GPRC5D in MM cells purified from focal lesion compared to interstitial marrow in paired clinical samples (n=176, p<1.21E-09), suggesting that high activity of osteoclasts in osteolytic lesions mediates phenotypical alteration in myeloma cells. Stable knockdown of GPRC5D by 70% in CAG myeloma line using lentiviral particles containing shRNA had no effect on short-term growth of these cells assessed by MTT assay. These data indicate that GPRC5D is a novel cell surface marker for myeloma plasma cells and that its expression is reduced in dedifferentiated myeloma cells. Disclosures: No relevant conflicts of interest to declare.

A Novel Role for Histone Deacetylase 11 (HDAC11) in B Cell Lymphopoiesis and Plasma Cell Survival in Multiple Myeloma

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4715-4715
Author(s):  
Jason B. Brayer ◽  
Eva Sahakian ◽  
John Powers ◽  
Mark B Meads ◽  
Susan Deng ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Cell Lines ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Proteasome Inhibitors ◽  
Myeloma Cell ◽  
Resistant Cell ◽  
Rpmi 8226 ◽  
Human Myeloma Cell

Abstract While multiple myeloma (MM) remains incurable presently, expanded therapeutic options over the past decade have improved patient survival markedly. Proteasome inhibitors have redefined the treatment paradigm for myeloma, often serving as the backbone of front-line treatment. Histone deacetylase (HDAC) inhibitors (HDI), although only marginally active as single agent therapy in hematological malignancies, have demonstrated an ability to salvage bortezomib responsiveness in refractory patients, prompting heightened interest in this class of targeted therapeutics in myeloma. HDAC’s represent a family of enzymes, currently with 11 known members in the classical HDAC family, and subdivided into 4 sub-classes. HDAC11 is currently the only member of the sub-class IV and, as the newest member of the HDAC family, its impact on B cell lymphopoiesis and myeloma development is only starting to be unveiled. Intriguingly, we show that mice with germ-line silencing of HDAC11 (HDAC11KO mice) exhibit a 50% decrease in plasma cells in both the bone marrow and peripheral blood plasma cell compartments relative to wild-type mice. Consistent with this, Tg-HDAC11-eGFP mice, a transgenic strain engineered to express GFP under control of the HDAC11 promoter (Heinz, N Nat. Rev. Neuroscience 2001) reveals that HDAC11 expression is increased in the plasma cell population and to a lesser extent B1 B cells, as compared to earlier lineage stages. Similar observations based on measurements of HDAC11 mRNA were seen in normal human plasma cells. Significant increases in HDAC11 mRNA expression were observed in 7 of 11 primary human multiple myeloma samples and 11 of 12 human myeloma cell lines as compared to normal plasma cells, further emphasizing the potential relevance of HDAC11 to the underlying pathologic processes driving myeloma development and/or survival. Targeted silencing of HDAC11 in RPMI-8226 cells lines using siRNA results in a modest decrease in cell viability as measured by Annexin/PI staining and detection of activated caspase-3. Quisinostat, a second generation pan-HDI, has previously demonstrated activity against human myeloma cell lines in vitro (Stuhmer, Brit J Haematol, 2010), and suppressed bone destruction in an in vivo murine myeloma model (Deleu, Cancer Res, 2009). We similarly observe dose-dependent survival impairment in 10 human myeloma cell lines when cultured in the presence of quisinostat, with EC50’s consistently in the 1-10nM range. Importantly, quisinostat acts synergistically with proteasome inhibitiors (bortezomib and carfilzomib) in RPMI-8226 cells; more importantly, the degree of synergism is amplified in the RPMI-6226-B25 bortezomib-resistant cell line. Although a clear mechanism of action remains to be elucidated, preliminary data suggests that RPMI-8226 cells exposed to quisinostat appear to exhibit a decrease nuclear, but not cytosolic HDAC11. Collectively, these data illustrate a previously unknown role for HDAC11 in plasma cell differentiation and survival. Increased HDAC11 expression seen in myeloma patient specimens and primary myeloma cell lines highlights the potential of HDAC11 as a therapeutic target. Furthermore, we show that quisinostat, a pan-HDI with selectivity towards HDAC11 at lower dosing, acts synergistically with proteasome inhibitors in vitro in proteasome inhibitor sensitive and resistant cell lines. Future work will focus on further elucidating the role of HDAC11 in myeloma survival and drug response, with particular emphasis on proteasome inhibitors. Disclosures No relevant conflicts of interest to declare.

scholarly journals Phenotypic analysis of human myeloma cell lines

Blood ◽  
1989 ◽  
Vol 73 (2) ◽  
pp. 566-572 ◽  
Author(s):  
C Duperray ◽  
B Klein ◽  
BG Durie ◽  
X Zhang ◽  
M Jourdan ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Lines ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Light Chains ◽  
Cell Phenotype ◽  
Phenotypic Analysis ◽  
Barr Virus ◽  
Human Myeloma Cell

Abstract Multiple myeloma (MM) is a B-cell malignancy characterized by the accumulation, primarily in bone marrow, of a clone of plasma cells. The nature of the stem cells feeding the tumoral compartment is still unknown. To investigate this special point, we have studied the phenotypes of nine well-known human myeloma cell lines (HMCLs) and compared them with those of normal lymphoblastoid cell lines (LCLs). Twenty-four clusters of differentiation involved in B lymphopoiesis were investigated using a panel of 65 monoclonal antibodies (MoAbs). For each cluster, the percentage of positive cells and the antigen density were determined, giving rise to a “quantitative phenotype”. We thus classified the HMCLs into two different groups: those with cytoplasmic mu chains (c mu+) and those without (c mu-). In the first (c mu+) group, comprising seven cell lines, the HMCLs had a phenotype of pre-B/B cells close to that of Burkitt's lymphoma cell lines. They expressed low densities of surface mu chains, without detectable cytoplasmic or surface light chains. Three of them were infected with the Epstein Barr virus (EBV). These c mu+ HMCLs bore most of the B-cell antigens except CD23. They expressed the CALLA antigen (CD10) and lacked the plasma-cell antigen PCA1. In contrast, LCLs expressed surface light chains, high densities of CD23, low densities of PCA1 antigen, and no CD10 antigen. The c mu- HMCLs had a plasma-cell phenotype, lacking most of the B-cell antigens and expressing high densities of PCA1 antigen.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Interleukin-10 is a proliferation factor but not a differentiation factor for human myeloma cells

Blood ◽  
1995 ◽  
Vol 85 (9) ◽  
pp. 2521-2527 ◽  
Author(s):  
ZY Lu ◽  
XG Zhang ◽  
C Rodriguez ◽  
J Wijdenes ◽  
ZJ Gu ◽  
...  
Keyword(s):  
Cell Lines ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Late Phase ◽  
Interleukin 10 ◽  
Myeloma Cell ◽  
Enzyme Linked Immunosorbent Assay ◽  
Myeloma Cells ◽  
Differentiation Factor

Because interleukin-10 (IL-10) is a potent differentiation factor of human B cells into mature plasma cells, we investigated its effect on human malignant plasma cells. IL-10 did not induce any differentiation and increase in Ig synthesis in four human IL-6-dependent malignant plasma cell lines. However, it stimulated the proliferation of two of four cytokine-dependent cell lines in the absence of IL-6 and IL-10-dependent myeloma cell lines have been obtained. The myeloma cell growth activity of IL-10 was unaffected by anti-IL-6 and anti-IL-6R antibodies. Similarly, IL-10 stimulated (P = .001) the proliferation of freshly-explanted myeloma cells in IL-6-deprived cultures of tumor samples from patients with active multiple myeloma (MM) and produced twice as many myeloma cells in these cultures. Again, this cytokine was unable to induce further differentiation (assessed by rate of Ig production) of fresh myeloma cells. A very sensitive enzyme-linked immunosorbent assay (ELISA; 1 pg/mL) only rarely detected IL-10 in the sera of MM patients (3 of 89). On the contrary, serum IL-10 was detected in 60% of patients with plasma cell leukemia (12 of 20). These data show that IL-10 is an IL-6-unrelated growth factor for malignant plasmablastic cells. This cytokine could be involved in the late phase of MM in vivo.

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