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.

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.

Mcl-1 Is Overexpressed in Multiple Myeloma and Correlates with Disease Severity.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1419-1419
Author(s):  
Soraya Wuilleme-Toumi ◽  
Nelly Robillard ◽  
Patricia Gomez-Bougie ◽  
Philippe Moreau ◽  
Steven Le Gouill ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Disease Severity ◽  
Cell Lines ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Lymphocytic Leukemia ◽  
Myeloma Cells

Abstract Multiple Myeloma (MM) is a fatal malignancy of B-cell origin characterized by the accumulation of plasma cells within the bone marrow. The expression of the pro-survival members of the Bcl-2 family has been shown to be a key process in the survival of myeloma cells. More particularly, Mcl-1 expression turned out to be critical for their survival. Indeed, knockdown of Mcl-1 by antisenses induces apoptosis in myeloma cells. Finally, Mcl-1 was found to be the only anti-apoptotic Bcl-2 family member which level of expression was modified by cytokine treatment of myeloma cells. For these reasons, we have evaluated the expression of Mcl-1 in vivo in normal, reactive and malignant plasma cells (PC) i.e., myeloma cells from 55 patients with MM and 20 human myeloma cell lines using flow cytometry. We show that Mcl-1 is overexpressed in MM in comparison with normal bone marrow PC. Forty-seven percent of patients with MM at diagnosis (p=.017) and 80% at relapse (p=.014 for comparison with diagnosis) overexpress Mcl-1. Of note, only myeloma cell lines but not reactive plasmocytoses have abnormal Mcl-1 expression, although both plasmocyte expansion entities share similar high proliferation rates (>20%). Of interest, Bcl-2 as opposed to Mcl-1, does not discriminate malignant from normal PC. This shows that the overexpression of Mcl-1 is clearly related to malignancy rather than to proliferation. It will be important to know whether the overexpression of Mcl-1 is related to an abnormal response to cytokines like Interleukin-6 or to mutations of the promoter of the Mcl-1 gene as already described in B chronic lymphocytic leukemia. Finally, level of Mcl-1 expression is related to disease severity, the highest values being correlated with the shortest event-free survival (p=.01). In conclusion, Mcl-1 which has been shown to be essential for the survival of human myeloma cells in vitro is overexpressed in vivo in MM and correlates with disease severity. Mcl-1 represents a major therapeutical target in MM.

Dual Src/Abl Kinase Targeted Inhibition in Myeloma Microenvironment Promotes Myeloma Cell Apoptosis Both in Vitro and In Vivo.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2813-2813
Author(s):  
Karthik Ramasamy ◽  
Lee Macpherson ◽  
Ghulam J Mufti ◽  
Stephen Schey ◽  
Yolanda Calle
Keyword(s):  
Bone Marrow ◽  
Cell Proliferation ◽  
Stromal Cells ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Myeloma Cells ◽  
Abl Kinase ◽  
Osteoclast Function

Abstract Abstract 2813 Poster Board II-789 Osteoclast, in addition to eroding the bone resulting in lytic lesions, enhances plasma cell proliferation and survival via direct cell to cell contact. Src family protein tyrosine kinases (SFKs) and c-Abl kinase play important role downstream of integrin adhesion receptors, and regulate the cytoskeletal organisation, cell motility and gene expression in response to cell adhesion. We hypothesised targeting SFKs and Abl kinase with the small molecule tyrosine kinase inhibitor Dasatinib has potential to reduce adhesion of plasma cells to ECM proteins in the bone marrow and modify the microenvironment by inhibiting osteoclast function, specifically bone resorption. As a result, myeloma cells could be sensitised to drugs with cytotoxic properties such as dexamethasone. Osteoclasts were generated from primary bone marrow mononuclear cells of myeloma and MGUS patients (n=10). Using Immunofluorescence, we found that Dasatinib 100nM but not dexamethasone inhibited osteoclastogenesis and disrupted the actin cytoskeletal organisation with actin clusters formed in the periphery of the cell. There was absence of actin ring formation at sealing zones which is essential for bone resorption. This effect consistently led to impaired osteoclast function, evidenced by fewer resorption pits formed on rabbit dentine slices on toluidine blue staining. Experiments were repeated ≥ 3 times. In plasma cells, the combination of dexamethasone and Dasatinib synergistically (Calcusyn software) inhibited cell proliferation at clinically relevant concentrations and induced apoptosis of human and murine myeloma cell lines alone and in cocultures with human stromal cells ( p<.001). Dasatinib alone at 200 nM concentration does not inhibit plasma cell proliferation with maximal serum concentration achieved in Phase I CML trials being 180nM. Additionally, Dasatinib and Dexamethasone in combination inhibited secretion of IL-6 but not MIL -1 alpha in stromal cell cocultures. Dasatinib but not dexamethasone significantly inhibited adhesion of myeloma cell lines on Fibronectin despite integrin activation with Magnesium EGTA. This effect was mediated through down regulation of both Src and Abl phosphorylation. Both Dasatinib and Dexamethasone inhibited adhesion of PC on stromal cells and osteoclasts. Taken together, our in vitro results suggest that Dasatinib and dexamethasone could be an effective therapeutic combination with Dasatinib impairing adhesion of plasma cells to the bone marrow microenvironment as well as osteoclast function and resultant bone disease thereby sensitising myeloma cells to the cytotoxic effect of dexamethasone. We have also established that the combination of Dasatinib 75mg/kg and dexamethasone 1mg/kg is not toxic to C57BL/KaLwRij mice. The anti-myeloma efficacy of these drugs alone and in combination is being currently studied. The combination of Dasatinib 100 mg OD days 1-28 and Dexamethasone 20mg OD on Day 1-4, 15-18 has resulted in a partial response (EBMT criteria) in 2 multiply relapsed and steroid refractory myeloma patients without significant toxicity. Serum calcium levels fell commensurate with disease response and we are currently performing experiments to analyse the effect of the drug combination on osteoclast function in vivo. These findings warrant exploring this drug combination in steroid resistant myeloma and patients with extensive skeletal disease prospectively in a phase I/II trial. Disclosures: Off Label Use: Dasatinib is not licensed for Myeloma.

Identification of Vacuolar H+- ATPase as a Target for Myeloma Therapeutics.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 257-257
Author(s):  
Yuan Xiao Zhu ◽  
Chung Xin ◽  
Sheng Ben Liang Lian ◽  
Wee-Joo Chng ◽  
Suzanne Trudel ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Viability ◽  
Cell Lines ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Normal Donor ◽  
Bafilomycin A1 ◽  
Myeloma Cells ◽  
Treatment Conditions

Abstract From a high throughput RNAi screen of the human druggable genome targeting the KMS11 cell line, we identified the suppression of the vacuolar H+- ATPase (V-ATPase) family as cytotoxic to myeloma cells. In the screen, two oligos against each gene for the V-ATPase subunits ATP6V1A and ATP6V1B1 resulted in suppression of cell growth (50% and 60% inhibition of cell viability respectively). We further confirmed this result using both lentiviral shRNA knockdown and two small molecule inhibitors specific for V-ATPase. Silencing of ATP6V1A by lentiviral shRNA knock-down in KMS11 and in OPM1 myeloma cell lines caused 75–80% reduction of cell viability at 5 days post infection (measured by MTT assay). Consistent with this result, the V-ATPase specific inhibitors, bafilomycin A1 and REATA 203, both inhibited the growth of a genetically heterogeneous and standardized panel of 14 human myeloma cell lines in vitro with an IC50 ranging from 2.2 – 8.9 nM (mean 5.25 nM) for bafilomycin A1 and 46–1594 nM (mean 542.5 nM)) for REATA 203. We further demonstrated that patient samples (n=10) were sensitive to 20nM bafilomycin A1 which induced a mean of 58% of MM cells to undergo apoptosis (range 10% to 93%) after 24 hours of treatment. Similar to bafilomycin A1, treatment of primary patient-derived MM cells with 500 nM REATA 203 for 72 hours resulted in a mean 69% apoptosis (range 24% to 97%). In contrast, non-myeloma cells (the CD138- fractions of the bone marrow samples) were less sensitive - mean 9% apoptosis (range from 0% to 34%) under the same treatment conditions. Of high interest, however, unlike most drugs we have studied in pre-clinical myeloma models, the cytotoxicity induced by bafilomycin A1 in MM cell lines is abrogated by co-culture with patient bone marrow stromal cells but is not affected by IL-6 or IGF-1 treatment. Dexamethasone- or melphalan-resistant MM cell lines were also highly sensitive to both bafilomycin A1 and REATA 203. In a xenographic JJN3 mouse model, bafilomycin A1 suppresses and delays growth of tumor in a dose-dependent fashion. Gene expression analysis of normal-donor bone marrow plasma cells (n=19), primary tumor samples from MM patients (n=107) and normal somatic tissues demonstrates ubiquitous expression of most subunits of V-ATPase, however, some subunits are preferentially expressed in myeloma cells compared with normal plasma cells, including ATP6V1F (84% vs. 11%), ATP6V1E1 (29% vs. 5%), ATP6V1G2 (17% vs. 0%) and ATP6V0E 2 (36% vs. 16%). In conclusion, our data indicate that vacuolar H+-ATPase inhibitors are of interest as potential therapeutics for MM.

Bone Morphogenic Protein 6: A Prognostic Factor Expressed by Normal Plasma Cells and Multiple Myeloma Cells Inhibiting Their Proliferation and Angiogenesis Induction

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2701-2701
Author(s):  
Anja Seckinger ◽  
Tobias Meißner ◽  
Jérôme Moreaux ◽  
Hartmut Goldschmidt ◽  
Axel Benner ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Overall Survival ◽  
Cell Lines ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
High Dose Chemotherapy ◽  
High Dose ◽  
Myeloma Cells

Abstract BACKGROUND: Pathogenesis of multiple myeloma is partly attributed to an aberrant expression of proliferation-, pro-angiogenic and bone-metabolism modifying factors by malignant plasma-cells. AIM. Given the long and variable time-span from first diagnosis of early-stage plasma-cell dyscrasias to overt myeloma and the low proliferation rate of malignant plasma-cells, we hypothesize these to concomitantly express a novel class of anti-proliferative factors of potential prognostic relevance. Here, bone morphogenic proteins (BMPs) represent possible candidates, as they inhibit proliferation, stimulate bone formation, and have an impact on the survival of cancer patients. PATIENTS AND METHODS. We assessed expression of BMPs and its receptors by Affymetrix DNA-microarrays (n=434) including CD138-purified primary myeloma-cell-samples, normal bone-marrow plasma-cell-samples, polyclonal plasmoblasts-samples, human myeloma-cell-lines (HMCL), and whole bone-marrow. Presence and differential gene expression was determined by PANP-algorithm and empirical Bayes statistics. Event-free (EFS) and overall survival (OAS) were investigated for the 168 patients undergoing high-dose chemotherapy (HM-group) using Cox’s proportional hazard model. Findings were validated using the same strategy on an independent group of 345 patients from the Arkansas-group. For validation, quantitative real-time PCR and flow cytometry were performed. In vitro induction of angiogenesis was assessed using the AngioKit-assay. Effect of BMP6 on proliferation of HMCL was assessed by 3H-thymidine uptake. RESULTS. BMP6 is the only BMP expressed by normal- (13/14 samples) and malignant plasma-cells (228/233 samples). It is significantly lower expressed in proliferating non-malignant plasmablastic cells and human myeloma cell-lines. In vitro, BMP6 significantly inhibits proliferation of myeloma-cell-lines with an IC50 ranged from 0.08–2.15μg/ml, survival of primary myeloma-cells, and in vitro tubule formation down to the level of the negative control. High BMP6-expression in malignant plasma cells delineates significantly superior overall-survival for patients undergoing high-dose chemotherapy in both independent series of patients (n=168, P=.02 and n=345, P=.03, respectively, see below). CONCLUSION. With BMP6 we report for the first time the autocrine expression of a prognostically relevant anti-angiogenic and anti-proliferative factor and its receptors by normal and malignant plasma-cells. Figure Figure

A Method for Measurement of Drug Sensitivity of Myeloma Cells Co-Cultured with Bone Marrow Stromal Cells

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1373-1373
Author(s):  
Kristine Misund ◽  
Katarzyna Anna Baranowska ◽  
Toril Holien ◽  
Christoph Rampa ◽  
Dionne Klein ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Lines ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Stromal Cell ◽  
Drug Sensitivity ◽  
Myeloma Cell ◽  
Marrow Stromal Cells ◽  
Myeloma Cells

Abstract Abstract 1373 The aim of this work was to establish a robust and simple method for the measurement of drug sensitivity in myeloma cells under conditions mimicking aspects of the bone marrow microenvironment. In particular we wanted to measure drug sensitivity in myeloma cells cultivated in the presence of stromal cells. The tumor microenvironment can profoundly affect tumor cell survival as well as alter antitumor drug activity, and it is generally believed that growth and survival of myeloma cells is critically dependent on the bone marrow microenvironment. Bone marrow stromal cells (BMSC) have been shown to protect myeloma cells from common cytostatic or cytotoxic drugs in vitro. Common in vitro assays used for high-throughput drug screening cannot easily discriminate between stromal and tumor cell responses in co-cultures. Although a few recent studies have overcome this problem (Ramasamy K. et al., 157(5):564–79,2012, McMillin D. et al., 16(4):483–9, 2010), the application of stable transfection for labeling of cells limits the practical application of these co-culture studies to cell lines, excluding primary myeloma cells that inherently may be hard to transduce even by retroviral vectors. Here, we analyzed survival of myeloma cells co-cultured with BMSC using an automated fluorescence microscope, ScanR. ScanR is a microscope based screening station. By staining the cell nuclei with DRAQ5, we were able to discriminate between BMSC and myeloma cells, based on their staining intensity and nuclear shape. Using the apoptotic marker YO-PRO-1, the effects of drug treatment on the viability of the myeloma cells in the presence of stromal cells could be measured. The main advantages of this method are the non-necessity of cell manipulation before co-culture and the low number of myeloma cells (5000 primary cells) that are needed per measurement, which makes the method ideal for experiments with primary myeloma cells. In fact, the analysis was easier and more robust when using slowly growing cells, i.e. by using primary myeloma cells compared to more rapidly proliferating myeloma cell lines. This method should be well-suited for high throughput analysis, as the cells are stained in situ with no washing, centrifugation, or fixation steps before analysis. The method was compared to a conventional method for detecting cell viability; flow cytometry where annexin V labeling was used to detect apoptotic cells. As shown in figure 1, the dose-response curves obtained for ANBL-6 cells treated with different doses of melphalan were similar and showed the same trends for both methods. However, the effects of melphalan treatment were more evident analyzed by the ScanR system than by flow cytometry (EC50 YO-PRO-1 = 11μM versus EC50Annexin V= 15μM). The stromal cell population applied in this study was able to support IL-6 dependent myeloma cell lines without addition of IL-6. This as IL-6 dependent INA-6 cells cultivated in the presence of BMSC survived in the absence of added IL-6. This study shows the importance of stromal cell support for primary myeloma cell survival in vitro, as half of the cell samples had a marked increase in their viability when cultured in the presence of BMSC. Stromal cell-induced protection against common myeloma drugs was also observed with this method. For instance, experiments with primary myeloma cells from patient MM7, showed that in the presence of BMSC, the EC50 for the common myeloma drug cyclophosphamide was increased from 5 μM to approximately 10 μM (figure 2). Figure 1 Figure 1. Figure 2 Figure 2. Disclosures: No relevant conflicts of interest to declare.

Plasma Levels of SDF-1 and Expression of SDF-1 Receptor on Multiply Myeloma Cells of Human Multiply Myeloma.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4874-4874
Author(s):  
Caixia Li ◽  
De Pei Wu ◽  
Junjie Cao ◽  
Xiaojin Wu ◽  
Xiao Ma ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Lines ◽  
Plasma Levels ◽  
Myeloma Cell ◽  
Migration Ability ◽  
Myeloma Cells ◽  
And Migration ◽  
Cxcr4 Axis ◽  
Healthy Persons

Abstract Multiple myeloma(MM) is a monoclonal expansion of malignant cells with a plasmablast-plasma cell morphology that is almost exclusively localized to the bone marrow, except at the final stages of disease, when they proliferate in the extramedullary area. The mechanisms of the selective homing of MM cells to the bone marrow compartment are poorly understood. The chemokine stromal cell-derived factor-1 (SDF-1) and its receptor CXCR4 contribute to stem cell homing and play a role in trafficking of leukemic cells. In this study we have investigated expression and biological behavior of SDF-1/CXCR4 in MM-derived cell lines and primary MM cells. FACS and RT-PCR analysis was used to study the expression of CXCR4 and ICAM-1(CD54) on the surface of MM cells from 4 IL-6 dependant cell lines (XG1,XG2,XG6 and XG7) and 25 freshly isolated tumor samples from patients with diagnosed MM. Mononuclear cells were purified by positive selection of magnetical and FACS sorting. Chemotaxis assay through transwell bore polycaronate and ELISA assay were employed to monitor the SDF-1, IL-6, and sICAM-1 levels. We found that[circ1]Fresh MM cells and MM cell lines expressed various levels of functional CXCR4 ranging from 23.1% to 77.7%,which was correlated with the in vitro migration ability of MM cells[(23.2±1.08)%, P<0.01]; [circ2]SDF-1 levels in the bone marrow(BM) of MM patients were significantly higher than the those of healthy persons (3489.23±651.63)pg/ml, (2818.57±597.79)pg/ml, P<0.05; but plasma levels of SDF-1 in peripheral blood of MM patients were lower than those of healthy persons[(1973±133)pg/ml, (2334.857±574.92), P=0.062]; [circ3]Plasma levels of PCL(4097.14±680.71) were significantly higher than those of healthy persons, P<0.01. The results firstly demonstrated abnormal expression of SDF-1 and its receptor CXCR4 on Human MM cells, which is closely correlated with the migration of MM cells. Furthermore, we discovered that SDF-1 could up-regulate the expression of ICAM-1 on MM cells; the plasma level of soluble ICAM-1 was correlated with the expression of CXCR4 on MM cells. These findings suggested that SDF-1/CXCR4 axis play a key role on the trafficking of MM cells via mediating the effect of adhesion molecules. Moreover, we observed higher plasma levels of IL-6 in PB of 60% MM patients compared with those of healthy individuals. Finally, the levels of IL-6 were closely correlated with SDF-1 levels (γ=0.8, P<0.01), These data indicated that in the IL-6-dependent myeloma cell lines or fresh myeloma samples and myeloma cell growth triggered by SDF-1 maybe due to up-regulation of autocrine and paracrine IL-6 by myeloma cells and stromal cells in BM. The results suggested that the expression of CXCR4 have an essential role in the proliferation and migration of myeloma cells in patients with multiple myeloma.In conclusion, MM cells expressed various levels of functional CXCR4, which were correlated with the migration ability of MM cells in vitro; SDF-1/CXCR4 axis plays a key role in the trafficking of MM cells via mediating the effect of adhesion molecules; The plasma levels of IL-6 closely correlated with SDF-1 plasma levels, myeloma cell growth triggered by SDF-1 may be due to up-regulation of autocrine and paracrine IL-6 by myeloma cells and stromal cells in BM. All these suggested that the expression of CXCR4 play an essential role in the proliferation and migration of myeloma cells in patients with multiple myeloma.

SDF-1 Is Responsible for the Constitutively High NF-kB Activity in Human Myeloma Cells.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4737-4737
Author(s):  
Abul Islam ◽  
Ken-ichiro Otsuyama ◽  
Jakia Amin ◽  
Saeid Abroun ◽  
Karim Shamsasenjan ◽  
...  
Keyword(s):  
Cell Lines ◽  
Target Genes ◽  
Plasma Cells ◽  
Plasma Concentrations ◽  
Myeloma Cell ◽  
Binding Activity ◽  
Myeloma Cells ◽  
Expression Levels ◽  
Human Myeloma Cells

Abstract The chemokine, stromal cell-derived factor 1 (SDF-1; CXCL12) and its receptor, CXCR4 are considered to be essentially required for plasma cell homing to the bone marrow (BM). It is well known that plasma cells in the BM (long-lived plasma cells) survive for a long time and have the constitutively high NF-kB activity. Since human myeloma cells are considered to be derived from these committed long-lived plasma cells, we investigated the role of SDF-1 on the survival of primary myeloma cells from myeloma patients and the possible relationship with NF-kB activity. First, we confirmed that all primary myeloma cells expressed CXCR4 but not CCR9 or CCR10 receptors on their surface and the levels of CXCR4 expression apparently correlated with maturity of BM plasma cells; mature myeloma cells (MPC-1+) as well as polyclonal plasma cells expressed higher levels of CXCR4 than those on immature myeloma cells (MPC-1-). The production of SDF-1 was found strongly in BM stromal cells but not in primary myeloma cells as well as myeloma cell lines. On the other hand, high DNA binding activity of NF-kB was constitutively detected in primary myeloma cells as well as myeloma cell lines, and these NF-kB activities significantly correlated with the expression levels of CD54 on their surface, for CD54 gene is one of the strict NF-kB target genes. Based on the expression levels of CD54 protein, interestingly, primary myeloma cells showed weaker NF-kB activities than those in monoclonal plasma cells from MGUS and polyclonal plasma cells from polyclonal gammopathy. Plasma concentrations of SDF-1 were also significantly correlated to the expression levels of CD54 on primary myeloma cells significantly (P<0.01). Furthermore, it was confirmed that addition of SDF-1 significantly increased the expression levels of CD54 in the in vitro culture of primary myeloma cells. Therefore, these results indicate that SDF-1 is responsible for high expression levels of CD54 and possibly the constitutively high NF-kB activity in primary myeloma cells.

MDX-1097 Binds Specifically to Kappa Myeloma Cells and Anti-Tumour Activity Is Mediated by Multiple Effector Cells.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1846-1846
Author(s):  
Mae Wong ◽  
Parisa Asvadi ◽  
Rosanne Dunn ◽  
Darren Jones ◽  
Douglas Campbell ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Lines ◽  
Human Tissue ◽  
Plasma Cells ◽  
Specific Binding ◽  
Myeloma Cell ◽  
Myeloma Cells ◽  
Effector Cells ◽  
Anti Tumour Activity ◽  
Tumour Activity

Abstract Abstract 1846 Poster Board I-872 Previous studies have described a murine monoclonal antibody, mKap, that specifically recognizes a cell surface antigen expressed on kappa myeloma cells and not on normal lymphoid cells. This antigen has been identified and designated kappa myeloma antigen (KMA). KMA consists of free kappa light chains (kFLC) not associated with heavy chain and is present on plasma cells isolated from kappa myeloma (MMk) patient bone marrow aspirates, kappa myeloma cell lines and kappa macroglobulinemia. In vitro data demonstrated that mKap was able to inhibit cell growth and induce apoptosis in myeloma cell lines. In addition, pre-clinical studies demonstrated that mKap was well tolerated and showed significant efficacy in a SCID xenograft model of MM. MDX-1097 is a chimeric version of mKap that is currently in development for the treatment of kappa restricted multiple myeloma. The antibody retains the binding affinity and specificity of mKap. Specific binding of MDX-1097 to malignant plasma cells isolated from MMk patient bone marrow aspirates has recently been demonstrated by flow cytometry. In addition a human tissue cross-reactivity study was performed using immunohistochemistry to assess the potential binding of MDX-1097-FITC to cryosections taken from a human tissue panel of three normal donors. The results demonstrated that MDX-1097 bound to bone marrow plasma cells from two patients with kappa cell dyscrasia but did not bind to normal human tissue samples or to plasma cells from a patient with lambda plasmacytoma. The ability of serum kFLC to inhibit MDX-1097 binding to the myeloma cell line, JJN3, was assessed by flow cytometry using serum derived from 32 MMk patients. The results indicated that MDX-1097 at a concentration of 100μg/mL (equivalent to an estimated serum concentration of 5mg/kg dose) is capable of binding to myeloma cells in the presence of 0–250μg/mL of serum kFLC. In vitro functional studies have demonstrated that MDX-1097 engages Fc receptor bearing effector cells and induces antibody dependent cellular cytotoxicity (ADCC) in kappa myeloma cell lines in the presence of healthy donor peripheral blood mononuclear cells. Further investigations have verified that purified natural killer cells (NK) play a major role in MDX-1097 anti-tumour activity. Importantly, recent studies have demonstrated that antibody dependent cellular phagocytosis by macrophages contributes to the anti-tumour activity of several therapeutic monoclonal antibodies. Preliminary data indicates that MDX-1097 may be capable of inducing enhanced uptake by macrophages. In conclusion MDX-1097 showed specific binding to KMA on myeloma cells isolated from patient's bone marrow samples and antibody binding is observed in the presence of kFLC in patient serum. In addition MDX-1097 anti-tumour activity is probably mediated by multiple Fc receptor bearing effector cells. Disclosures: Wong: Immune System Therapeutics: Employment. Asvadi:Immune System Therapeutics: Employment. Dunn:Immune System Therapeutics: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Jones:Immune System Therapeutics: Employment. Campbell:Immune System Therapeutics: Employment.

scholarly journals A Role for Syntenin-1 in Multiple Myeloma Cell Survival

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 1008-1008
Author(s):  
Tyler Moser-Katz ◽  
Catherine M. Gavile ◽  
Benjamin G Barwick ◽  
Sagar Lonial ◽  
Lawrence H. Boise
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Cell Death ◽  
Cell Survival ◽  
Cell Lines ◽  
Plasma Cells ◽  
Surface Expression ◽  
Myeloma Cell ◽  
Myeloma Cells ◽  
Rpmi 8226

Abstract Multiple myeloma is the second most common hematological malignancy in the U.S. with an estimated 30,700 new diagnoses in 2018. It is a clonal disease of plasma cells that, despite recent therapeutic advances, remains incurable. Myeloma cells retain numerous characteristics of normal plasma cells including reliance on survival signals in the bone marrow for long term viability. However, malignant transformation of plasma cells imparts the ability to proliferate, causing harmful bone lesions in patients, and in advanced stages independence of the bone-marrow microenvironment. Therefore, we are investigating the molecular mechanisms of myeloma cell survival that allow them to become extramedullary. We identified syntenin-1 (SDCBP) as a protein involved in myeloma cell survival and a potential therapeutic target. Syntenin-1 is an adapter protein that has been shown to regulate surface expression of several transmembrane proteins by binding with membrane phospholipids and mediating vesicular trafficking of proteins throughout the cell. Syntenin-1 regulates the surface expression of CD138, a plasma/myeloma cell marker. Syntenin-1 has been shown to regulate apoptosis in numerous cancer cell lines including breast cancer, glioma, and pancreatic cancer but its role in multiple myeloma survival has not been studied. To determine if syntenin-1 expression has an effect on myeloma cell survival, we utilized the CoMMpass dataset (IA12), a longitudinal study of myeloma patients that includes transcriptomic analysis throughout treatment. We found that patients with the highest expression of syntenin-1 mRNA (top quartile) had significantly worse overall survival, progression-free survival, and a shorter response duration than those in the bottom quartile of expression. To determine if syntenin-1 has a role in myeloma cell survival, we used short hairpin RNA to knock down syntenin-1 (shsyn) in RPMI 8226 and MM1.s myeloma cell lines. We then determined the amount of cell death using Annexin-V staining flow cytometry four days following lentiviral infection. We found increased cell death in syntenin-1-silenced cells compared to our empty vector control in both RPMI 8226 (control=42.17%, shsyn=71.53%, p=0.04) and MM1.s cell lines (control=8.57%, shsyn=29.9%, p=0.04) suggesting that syntenin-1 is important for myeloma cell survival. Syntenin-1 contains two PDZ domains that allow it to bind to receptor proteins via their corresponding PDZ-binding motifs. We therefore wanted to look at correlation of syntenin-1 expression with CD138 and CD86, two PDZ-binding domain containing proteins expressed on the surface of myeloma cells. Using the CoMMpass dataset, we found patients with high expression of syntenin-1 had a median expression of CD86 that was twice as high as the total population (P<0.0001) while syntenin-1-low patients expressed CD86 at levels that were half as much as the population (P<0.0001). In contrast, there was no clear relationship between syntenin-1 and CD138 mRNA expression. Indeed if one takes into account all patients, there is a positive correlation between CD86 and syntenin-1 expression (r=0.228, P<0.0001) while there is a negative correlation between CD138 and syntenin-1 (r=-0.1923, P<0.0001). The correlation with CD86 but not CD138 suggests a previously undescribed role for syntenin-1 in myeloma cells. Our lab has previously shown that expression of CD86 is necessary for myeloma cell survival, and signals via its cytoplasmic domain to confer drug resistance. Silencing syntenin-1 results in a decrease in CD86 surface expression. However, there is no change in CD86 transcript or total cellular CD86 protein levels in our shsyn treated cells. Moreover, knockdown of CD86 resulted in increased protein expression and transcript levels of syntenin-1. Taken together, these data suggest that syntenin-1 may regulate CD86 expression on the cell surface. Our data supports a novel role for syntenin-1 in myeloma cell viability and as a potential regulator of CD86 surface expression. The role of syntenin-1 has not previously been explored in multiple myeloma and determining its molecular function is warranted as it may be an attractive target for therapeutic treatment of the disease. Disclosures Lonial: Amgen: Research Funding. Boise:AstraZeneca: Honoraria; Abbvie: Consultancy.

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