scholarly journals Bivalent Histone Modifications By Reiibp Leads to Transcriptional Reprogramming and TLR7-BTK Driven IL-6 Pro-Inflammatory Response in t(4;14) Myelomagenesis

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 2202-2202
Author(s):  
Phyllis SY Chong ◽  
Julia Lim ◽  
Jing Yuan Chooi ◽  
Wee-Joo Chng
Keyword(s):  
Cell Line ◽  
Research Funding ◽  
Plasma Cells ◽  
Cancer Cell Line ◽  
Transcriptome Profiling ◽  
Myeloma Cell ◽  
Stable Cell Line ◽  
Dependent Manner ◽  
Myeloma Cells ◽  
Chip Sequencing

Abstract Multiple Myeloma (MM) is characterized by the uncontrolled proliferation of malignant plasma cells that are incapable of producing functional antibodies. Current treatment regime involves novel drug classes such as proteasome inhibitors, immunomodulatory drugs and monoclonal antibodies, which have significantly improved survival outcomes in patients. Patients with t(4;14) translocation represents ~15% of MM cases, and displays a dysregulation of the MMSET locus. It has one of the worst prognosis when compared to other subgroups, but represents an intermediate-risk group given its response towards bortezomib. REIIBP is a t(4;14)-deregulated isoform arising from alternative promoter usage within the MMSET locus. Despite sharing identical sequence with the C-terminus of MMSET II, we found that REIIBP displayed mutually exclusive expression with the full-length MMSET II arising from MB4-1 breakpoint. Additionally, the expression of REIIBP can be regulated through microRNAs by another histone methyltransferase, EZH2 in a Dicer-dependent manner. We generated a stable cell line that overexpresses REIIBP in the t(4;14)-negative human myeloma cells, RPMI8226, and revealed REIIBP as an epigenetic regulator of repressive H3K27me3 and active H3K4me3 modifications. Transcriptome profiling and ChIP-sequencing identified an upregulation of Toll-like receptor 7 (TLR7) by REIIBP through the enrichment of H3K4me3 on its promoter coupled with decrease in intragenic H3K27me3. This led to a BCR-independent activation of Bruton's tyrosine kinase (BTK) and NF-ĸB signaling in t(4;14) myeloma cells. Using Cancer Cell Line Encyclopedia and DepMap portal (Broad Institute) to compare the expression of BTK and BTK dependency scores across lineages revealed that BTK is an important target in MM, and REIIBP expression correlated with BTK in the CoMMpass dataset. Activation of TLR7-BTK by REIIBP conferred bortezomib resistance through the dysregulation of pro-inflammatory cytokine expression such as IL-6. Importantly, cells with REIIBP overexpression displayed enhanced lethality towards BTK inhibitor Ibrutinib, and combination with Bortezomib potentiated inhibition in myeloma cell lines and mice engrafted with RPMI8226-REIIBP tumors. Altogether, our results indicated that blockade of REIIBP in t(4;14) cells through combining proteasome and BTK inhibitors is a therapeutic strategy in the clinic for further evaluation. Disclosures Chng: Novartis: Honoraria, Research Funding; Antengene: Honoraria; Pfizer: Honoraria; Sanofi: Honoraria; AbbVie: Honoraria; Amgen: Honoraria; BMS/Celgene: Honoraria, Research Funding; Johnson & Johnson: Honoraria, Research Funding; Takeda: Honoraria; Aslan: Research Funding.

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.

CD28 and CD86 Are Necessary for Myeloma Cell Survival.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2946-2946
Author(s):  
Catherine M Gavile ◽  
Jayakumar R Nair ◽  
Kelvin P Lee ◽  
Sagar Lonial ◽  
Lawrence H. Boise
Keyword(s):  
T Cells ◽  
Growth Factor ◽  
Cell Survival ◽  
Cell Line ◽  
Cell Lines ◽  
Plasma Cells ◽  
Surface Expression ◽  
Myeloma Cell ◽  
Myeloma Cells ◽  
Growth And Survival

Abstract Abstract 2946 Multiple myeloma (MM) is a hematologic malignancy characterized by the aberrant proliferation of plasma cells. Myeloma cells retain most of the physiological characteristics of their normal counterpart – the long-lived plasma cell. Myeloma cells secrete immunoglobulin and reside in the bone marrow, where they rely heavily on interactions with the stroma for survival signals. While recent advances in therapeutics have led to an increase in median survival post-diagnosis, the disease remains incurable. Understanding the pathways which mediate growth and survival of these cells will help in identifying new targets that can potentially further improve patient outcomes. CD28 is a receptor better known for its role in T-cell signaling through interaction with its ligands, CD80 or CD86. Interaction between CD28 on T-cells and CD80/86 on antigen-presenting cells leads to survival and proliferation of T-cells. Recent work has shown that the CD80/86-CD28 pathway also plays an important role in normal plasma cell generation and survival. Interestingly, high expression of CD28 and CD86 are poor prognostic markers for myeloma patients. Previous work has shown that CD28 activation provides survival signals for myeloma cells in growth-factor deficient conditions. It has also been shown that CD28 on the myeloma cell interacts with CD80/86 on the dendritic cell, which induces secretion of IL-6 (by the DC), an important myeloma growth factor. However, it is not known if CD28 or CD86 play a role in steady state growth and survival of myeloma cells. In order to determine the role of each of these 2 molecules in myeloma physiology, we knocked-down either CD28 or CD86 on the myeloma cell via lentivirus-mediated shRNAs. We found that knockdown of CD86 leads to apoptosis in 3 myeloma cell lines (RPMI8226, MM1.s, and KMS18). Four days after infection with the lentivirus containing shCD86, 45.7±4.9 and 60.3±4.6 percent control apoptosis was observed in RPMI8226 and MM1.s respectively, while less death was observed in KMS18 (17.6±1.6). CD28-knockdown resulted in apoptosis as well (24.9±4.3 for RPMI8226, 26.8±4.1 for MM1s, 21.8±3.8 for KMS18, percent control apoptosis). Consistent with these findings, we were unable to establish a myeloma cell line with stable knockdown of either CD28 or CD86. Additionally, RPMI8226 cells stably transfected to over-express either Bcl-2, Bcl-xL, or Mcl-1 are protected from cell death induced by CD86 or CD28 silencing. These data suggest that CD28 and CD86 are essential to prevent apoptosis of myeloma cells in vitro. To confirm these findings we determined the effects of CTLA4-Ig on myeloma survival. CTLA4-Ig inhibits CD86-CD28 signaling by binding to CD86, blocking its interaction with CD28. We found that treatment of RPMI8226 and MM1.s cells with CTLA4-Ig caused apoptosis in the myeloma cells after 2 days (23.9±3.9 for RPMI8226 and 20.4±6.2 for MM1.s, percent control apoptosis). Thus like normal plasma cells, CD28 and CD86 are required for the survival of myeloma cells. To determine why silencing of CD86 has a more potent effect than CD28 silencing on myeloma cell survival in 2 out of 3 cell lines, we investigated the effects of silencing on cell surface expression of each of these proteins. CD28 and CD86 mRNA and protein levels were silenced to similar levels by their cognate hairpins. However, in MM.1s and RPMI8226 we found that silencing of CD28 resulted in an increase in CD86 surface expression. This increase was also observed at the mRNA level and in the cells over-expressing Bcl-2 family members, indicating that this is not simply due to the selection of the highest expressing cells. These data suggest a feedback loop exists to regulate CD28-CD86 signaling in myeloma cells. Surprisingly, in the KMS18 cell line, we observe the converse effect, where silencing of CD86 resulted in upregulation of CD28. This provides a likely explanation for why these cells are less susceptible to CD86 silencing than the other two lines. Interestingly, blocking CD86 with CTLA4-Ig treatment also resulted in a modest upregulation in CD28 surface expression of MM.1s and RPMI8226, which suggests that silencing CD86 and binding of CD86 with a soluble receptor are not equivalent, and that multiple signaling feedback pathways exist to regulate the expression of this receptor-ligand pair that is necessary for myeloma cell survival. Disclosures: No relevant conflicts of interest to declare.

Semaphorin 4D to suppress bone formation in multiple myeloma.

2017 ◽  
Vol 35 (15_suppl) ◽  
pp. 8039-8039 ◽  
Author(s):  
Konstantinos Lontos ◽  
Juraj Adamik ◽  
Peng Zhang ◽  
Quanhong Sun ◽  
David Roodman ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Formation ◽  
Cell Line ◽  
Cancer Cell Line ◽  
Myeloma Cell ◽  
Conditioned Media ◽  
Myeloma Cells ◽  
Myeloma Bone Disease ◽  
Semaphorin 4D ◽  
Pre Treatment

8039 Background: Myeloma bone disease is characterized by osteoclast activation and long-term osteoblast suppression. We investigated if Semaphorin 4D (Sema4D; CD100) plays a role in these processes. Sema4D has been shown to be a potent osteoblast inhibitor (Negishi-Koga T et al, Nat Med. 2011). A study recently identified that the breast cancer cell line MDA-MB-231 utilizes Sema4D to create osteolysis (Yang Y et al, PLOS One 2016). There have been previous data that Sema4D is increased in the serum of myeloma patients (Terpos et al, Blood 2012) and that co-culturing myeloma cell lines with osteocytes increases the expression of Sema4D mRNA in both (Suvannasankha et al, Blood 2016). We sought to investigate if myeloma cells are using Sema4D to suppress bone formation and if they affect the levels of Sema4D produced by osteoclasts. Methods: We used lentivirus carrying shRNA for Sema4D or control (Scr) to knock down the level of the protein in the 5TGM1 murine myeloma cell line. Knockdown was verified by qPCR and Western Blot. We subsequently co-cultured the 5TGM1 cells with the MC3T3-subclone M4 (MC4) murine stromal cell line for 2 days, removed the myeloma cells and then differentiated the MC4 cells using ascorbic acid and β-glycerolphosphate. At day 5, we analyzed the cells for Runx2 (a critical gene for the differentiation of stromal cells into osteoblasts) expression utilizing qPCR. Also, we performed qPCR in primary osteoclast (OCL) mouse cells differentiating into OCL with RANKL with or without pre-treatment with myeloma-conditioned media for 3 days before the addition of RANKL. Results: When 5TGM1-Scr were co-cultured with MC4 cells the expression of Runx2 on day 5 was decreased (p=0.02). Strikingly, the 5TGM1-shSema4D cells when co-cultured with MC4s did not have the same effect and allowed the upregulation of Runx2 expression on day 5 (p=0.01). Myeloma-conditioned media increased Sema4D expression by OCL throughout the 5 days of differentiation 2 to 3-fold (p=0.01 for day 5). Conclusions: The myeloma cells seem to be utilizing Sema4D both directly and indirectly to inhibit bone formation. Targeted therapy against Sema4D may improve outcomes and fracture-free survival for multiple myeloma patients.

Efficacy Of ABT-199 In Multiple Myeloma

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4453-4453 ◽  
Author(s):  
Shannon M. Matulis ◽  
Cathy Sharp ◽  
Ajay K. Nooka ◽  
Jonathan L. Kaufman ◽  
Sagar Lonial ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Cell Line ◽  
Cell Lines ◽  
Research Funding ◽  
Plasma Cells ◽  
Survival Rates ◽  
Specific Inhibitor ◽  
Small Subset ◽  
Potent Inducer ◽  
Myeloma Cells

Great strides have been made in the last 15 years in the treatment of multiple myeloma, with FDA approval of both IMIDs and proteasome inhibitors, 10-year survival rates are now achievable in around 25% of patients. However, for the remaining 75%, disease relapse due to drug resistance remains a significant clinical concern. Therefore novel therapeutic approaches to be used in combination with the current active agents are required. The Bcl-2 family of proteins regulates apoptosis and presents an exciting target for therapy. We have demonstrated that myeloma cell lines can be dependent on the anti-apoptotic protein Mcl-1 or can be co-dependent on Mcl-1 and Bcl-2/xL for survival. The distinction between Mcl-1 dependence and co-dependence is the distribution of the pro-apoptotic BH3-only protein Bim. In Mcl-1-dependent lines, Bim is primarily associated with Mcl-1. In contrast in the co-dependent lines, Bim is either predominately associated with Bcl-2/xL or when it is released from Bcl-2/xL it cannot bind to Mcl-1 because of the presence of the Mcl-1 inhibitor, Noxa. This renders these cells sensitive to the Bcl-2/xL inhibitor ABT-737. We have confirmed these findings in freshly isolated patient samples and demonstrated that among the 5 patient samples tested, Bim was associated with both Mcl-1 and Bcl-xL and the cells were sensitive to ABT-737. This suggests that co-dependence on Mcl-1 and Bcl-2/xL may be a common phenomena in myeloma. However, given the adverse effects seen with Bcl-xL inhibition, ABT-737 or the related compound Navitoclax may be difficult to use for the treatment of multiple myeloma. However, ABT-199, a Bcl-2-specific inhibitor, has been developed and we report on preclinical testing in multiple myeloma. Freshly isolated plasma cells from 3 myeloma patients were treated with either ABT-737 or ABT-199 for 24 h and IC50s for each drug were compared. In each patient sample, the plasma cells were less sensitive to ABT-199 than to ABT-737 (MM49: 199 IC50 1.4 μM vs. 737 IC50 0.9 μM; MM51: 199 IC50 0.34 μM vs. 737 IC50 0.07 μM; MM52: 199 IC50 1.3 μM vs. 737 IC50 0.25 μM), and only MM51 was truly sensitive to ABT-199. CoIP studies in MM51 revealed very little Bim bound to Mcl-1 and none bound to Bcl-xL, suggesting the majority of the Bim present in the cell must be bound to Bcl-2. These data suggest that myeloma cells are more likely to be co-dependent on Mcl-1 and Bcl-xL for survival than Mcl-1 and Bcl-2. However Bcl-2 dependence can exist in myeloma. We also performed dose curves in 4 multiple myeloma cells lines, representing 4 different Bim binding patterns and sensitivity to ABT-737. 8226, MM.1s, and KMS18 are all co-dependent cell lines (sensitive to ABT-737), while KMS11 is Mcl-1 dependent. Of these cell lines, KMS18 is the only one with the majority of Bim bound to Bcl-2, therefore should be sensitive to ABT-199. Surprisingly this was not the case. 8226 was the only cell line that was sensitive to ABT-199 (IC50 2.2 μM), while MM1.s, KMS18, and KMS11 were insensitive with IC50s of 6.2 μM, 6.8 μM, and 24.7 μM respectively. In order to gain a mechanistic understanding of these data, we performed CoIP studies to determine the pattern of Bim binding to Mcl-1, Bcl-xL, and Bcl-2 following treatment with ABT-199. We found in KMS18 cells, upon treatment, Bim is released from Bcl-2 and bound by Mcl-1 thereby preventing apoptosis. We have already demonstrated that very little Bim is bound to Bcl-2 in KMS11 and MM.1s, which is consistent with the lack of sensitivity in these cell lines. However in 8226 cells, the high expression of Noxa prevents the Bim released from Bcl-2 from binding to Mcl-1, thereby promoting apoptosis. Silencing of Noxa in this cell line raises the IC50 of ABT-199 2-fold. Next we investigated the effectiveness of combining ABT-199 with the proteasome inhibitor carfilzomib, which has been shown to be a potent inducer of Noxa. A synergistic response to this drug combination was seen in KMS18 cells. CoIP studies revealed that in the presence of Cz, the Bim released from Bcl-2 by ABT-199 could no longer bound to Mcl-1, and was free to activate Bak and Bax. Only additive responses were seen in 8226, MM.1s and KMS11 cell lines. Taken together these data suggest ABT-199 alone may only be an effective treatment for multiple myeloma in a small subset of patients. However, combining it with either Noxa inducers or Mcl-1 inhibitors could be a promising approach for the treatment of this disease. Disclosures: Kaufman: Jansenn: Consultancy; Millennium Pharmaceuticals: Consultancy; Celgene: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Onyx: Consultancy; Merck: Research Funding. Lonial:Millennium: Consultancy; Celgene: Consultancy; Novartis: Consultancy; BMS: Consultancy; Sanofi: Consultancy; Onyx: Consultancy. Boise:Onyx Pharmaceuticals: Consultancy.

IL-6 and IGF-1 Act in Synergy with HGF in Myeloma Cells by Modulating the Ras-MAPK Pathway.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4793-4793
Author(s):  
Håkon Hov ◽  
Erming Tian ◽  
Anders Waage ◽  
Magne Børset ◽  
Anders Sundan
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Growth Factors ◽  
Cell Line ◽  
Plasma Cells ◽  
Kinase Inhibitor ◽  
Mapk Pathway ◽  
Myeloma Cell ◽  
Myeloma Cell Line ◽  
Myeloma Cells

Abstract Multiple myeloma is an incurable malignancy of plasma cells homing to the bone marrow. Myeloma cells are dependent on factors in their microenvironment for survival and expansion. HGF may be produced both by myeloma cells and the bone marrow microenvironment and serum levels of HGF are known to be a prognostic factor in multiple myeloma. Both IL-6 and IGF-1 are known growth factors for myeloma cells. In the human myeloma cell line (HMCL) INA-6, HGF alone had low effect, but together with IL-6 and IGF-1 it became a potent growth factor increasing thymidin incorporation two- to three-fold above the levels obtained with IL-6 or IGF-1 alone. Similar results were obtained for the myeloma cell line OH-2. The ANBL-6 cell line harbours an autocrine growth promoting HGF-loop. When inhibiting this autocrine HGF loop with a specific c-Met receptor tyrosine kinase inhibitor (PHA-665752), IL-6- and IGF-1-induced proliferation was reduced by 80% and 50% respectively. Thus, in the prescence of HGF, both IL-6 and IGF-1 are dependent on the HGF-receptor c-Met for full effect on cell proliferation. There seems to be two interconnected explanations for the synergy between HGF- and either IL-6- or IGF-1-signalling in myeloma cells. IL-6 and IGF-1 treatment increased the expression of c-Met in INA-6 cells Secondly, we found that HGF was unique among the three growth factors in activating both Ras and p44/42 MAPK in INA-6 cells. Also in ANBL-6 cells, IGF and IL-6 was dependent on functional c-Met signalling to fully activate this pathway. Thus, the reason for synergy between HGF and IL-6 or IGF-1 seemed to be that full activation of the Ras-Mapk pathway through Gab1 and SHP-2 by these cytokines was dependent on operating c-Met signalling. Taken together, HGF and c-Met signalling would be attractive targets for therapy of multiple myeloma.

scholarly journals Altered expression of Pax-5 gene in human myeloma cells

Blood ◽  
1996 ◽  
Vol 87 (10) ◽  
pp. 4311-4315 ◽  
Author(s):  
MS Mahmoud ◽  
N Huang ◽  
M Nobuyoshi ◽  
IA Lisukov ◽  
H Tanaka ◽  
...  
Keyword(s):  
Gene Expression ◽  
B Cell ◽  
Cell Line ◽  
Cell Lines ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Altered Expression ◽  
Myeloma Cells ◽  
Normal Plasma ◽  
Human Myeloma Cells

Recent phenotypic analysis of plasma cells showed that normal plasma cells do express the B-cell lineage-specific molecule CD19, but their malignant counterpart (myeloma cells) are CD19-. To clarify the meaning of loss of CD19 antigen on myeloma cells, we first compared the expression of CD19 and Pax-5 genes among B cells, normal plasma cells, myeloma cell lines, and primary myeloma cells, because the Pax-5 gene was reported to encode the transcriptional factor, B-cell-specific activating protein (BSAP), necessary for CD19 gene expression. Neither CD19 nor Pax-5 mRNA could be detected in those primary myeloma cells and cell lines, whereas normal plasma cells did express both CD19 and Pax-5 mRNA. Furthermore, we could confirm that BSAP-binding activity was not detected in the nuclear extract from CD19- myeloma cell line (KMS-5) but was detected in CD19+ B-cell line (Raji) by gel-shift assay. We further examined the expression of E2A and Id genes, because E2A and Id are considered to be positive and negative regulators in the expression of Pax-5 gene, respectively. However, no significant differences in the expression of these E2A and Id-2 genes could be observed between myeloma cells and normal plasma cells. Therefore, these data suggest that the altered expression of Pax-5, but not E2A or Id, is responsible for the loss of CD19 expression in human myeloma cells, although the underlying mechanism of the altered Pax-5 gene expression remains to be clarified.

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

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.

Regulation of Multiple Myeloma Survival and Progression by CD1d

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2720-2720
Author(s):  
Emmanouil Spanoudakis ◽  
Ming Hu ◽  
Kikkeri Naresh ◽  
Evangelos Terpos ◽  
Valeria Melo ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Cell Death ◽  
Cell Survival ◽  
Cell Lines ◽  
Plasma Cells ◽  
Hla Class I ◽  
Myeloma Cell ◽  
Dependent Manner ◽  
Myeloma Cells ◽  
Therapeutic Implications

Abstract Downregulation of conventional HLA molecules from the surface of tumour cells is an important mechanism for tumour immune evasion, survival and progression. Whether CD1d, a non-conventional, glycolipid-presenting HLA class I-like molecule can affect tumour cell survival is not known. To test this we studied expression of surface CD1d on plasma cells from different stages of multiple myeloma (MM) using flow-cytometry. Expressing results as the ratio of the Geo MFI CD1d/isotype IgG1 we found that while CD1d expression was comparable between MGUS (n=8) and newly diagnosed MM patients (n=14; Geo MFI MGUS: 8.61±4.3 vs new MM: 7.1±4.72, p>0.05), in relapsed/advanced disease CD1d was significantly lower (Geo MFI:1.92±0.9, p<0.003 vs MGUS and new MM) and completely lost in 4 out of 5 myeloma cell lines at protein and RNA level. Further, 4 out of 8 paired, same-patient trephine biopsies stained with anti- CD1d showed drastic loss of CD1d expression in advanced compared to early disease. These results confirmed loss of CD1d expression during disease progression and suggested that CD1d impacts negatively on myeloma cell survival. Consistent with this, we found that engagement of CD1d by 2 different anti-CD1d mAbs and as compared to isotypic IgG or media control, induces cell death (i.e., Annexin+) of the CD1d-expressing B lymphoblastoid cell line C1R-CD1d, of myeloma cell lines with retrovirally restored expression of CD1d and purified, CD1d-expressing primary myeloma cells in a dose- and time-dependent manner, coincident with loss of mitochondrial membrane potential (MMP) as assessed by DioC3 staining. Biochemical analysis of relevant cell death pathways showed that MMP loss is associated with overexpression of the pro-apoptotic protein Bax but as demonstrated by immunoblotting and pharmacological inhibition it is caspase- independent. By introducing appropriate CD1d retroviral constructs into CD1d- myeloma cell lines we showed that anti-CD1d-induced cell death requires the cytoplasmic tail but not a Tyr residue critical for lysosomal sorting of CD1d. Finally, we found that anti-CD1d co-operates with anti-myeloma agents in the killing of myeloma cells. Thus, these findings provide evidence linking a novel function of CD1d in the regulation of cell death with tumour survival and progression and might have pathogenetic and therapeutic implications for other CD1d-expressing hematopoietic malignancies as well as myeloma.

Ruxolitinib Regulates the Autophagy Machinery in Multiple Myeloma Cells

2020 ◽  
Vol 20 (18) ◽  
pp. 2316-2323 ◽  
Author(s):  
Alican Kusoglu ◽  
Bakiye G. Bagca ◽  
Neslihan P.O. Ay ◽  
Guray Saydam ◽  
Cigir B. Avci
Keyword(s):  
Multiple Myeloma ◽  
Cell Line ◽  
B Lymphocyte ◽  
Plasma Cells ◽  
Annexin V ◽  
Myeloma Cell ◽  
Control Group ◽  
Multiple Myeloma Cell ◽  
Ic50 Values ◽  
Stat Pathway

Background: Ruxolitinib is a selective JAK1/2 inhibitor approved by the FDA for myelofibrosis in 2014 and nowadays, comprehensive investigations on the potential of the agent as a targeted therapy for haematological malignancies are on the rise. In multiple myeloma which is a cancer of plasma cells, the Interleukin- 6/JAK/STAT pathway is emerging as a therapeutic target since the overactivation of the pathway is associated with poor prognosis. Objective: In this study, our purpose was to discover the potential anticancer effects of ruxolitinib in ARH-77 multiple myeloma cell line compared to NCI-BL 2171 human healthy B lymphocyte cell line. Methods: Cytotoxic effects of ruxolitinib in ARH-77 and NCI-BL 2171 cells were determined via WST-1 assay. The autophagy mechanism induced by ruxolitinib measured by detecting autophagosome formation was investigated. Apoptotic effects of ruxolitinib were analyzed with Annexin V-FITC Detection Kit and flow cytometry. We performed RT-qPCR to demonstrate the expression changes of the genes in the IL-6/JAK/STAT pathway in ARH-77 and NCI-BL 2171 cells treated with ruxolitinib. Results: We identified the IC50 values of ruxolitinib for ARH-77 and NCI-BL 2171 as 20.03 and 33.9μM at the 72nd hour, respectively. We showed that ruxolitinib induced autophagosome accumulation by 3.45 and 1.70 folds in ARH-77 and NCI-BL 2171 cells compared to the control group, respectively. Treatment with ruxolitinib decreased the expressions of IL-6, IL-18, JAK2, TYK2, and AKT genes, which play significant roles in MM pathogenesis. Conclusion: All in all, ruxolitinib is a promising agent for the regulation of the IL-6/JAK/STAT pathway and interferes with the autophagy mechanism in MM.

Design, Synthesis and Cytotoxicity Evaluation of New 3, 5-Disubstituted-2-Thioxoimidazolidinones

2018 ◽  
Vol 18 (4) ◽  
pp. 573-582 ◽  
Author(s):  
Khaled R.A. Abdellatif ◽  
Mostafa M. Elbadawi ◽  
Mohammed T. Elsaady ◽  
Amer A. Abd El-Hafeez ◽  
Takashi Fujimura ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Line ◽  
Cell Lines ◽  
Cancer Cell ◽  
Topoisomerase I ◽  
Cancer Cell Line ◽  
Cytotoxic Agents ◽  
Dependent Manner ◽  
Human Lung Fibroblast ◽  
Mcf 7

Background: Some 2-thioxoimidazolidinones have been reported as anti-prostate and anti-breast cancer agents through their inhibitory activity on topoisomerase I that is considered as a potential chemotherapeutic target. Objective: A new series of 3,5-disubstituted-2-thioxoimidazolidinone derivatives 10a-f and their S-methyl analogs 11a-f were designed, synthesized and evaluated for cytotoxicity against human prostate cancer cell line (PC-3), human breast cancer cell line (MCF-7) and non-cancerous human lung fibroblast cell line (WI-38). </P><P> Results and Method: While compounds 10a-f showed a broad range of activities against PC-3 and MCF-7 cell lines (IC50 = 34.0 – 186.9 and 24.6 – 147.5 µM respectively), the S-methyl analogs 11a-f showed (IC50 = 22.7 – 198.5 and 16.9 – 188.2 µM respectively) in comparison with 5-fluorouracil (IC50 = 60.7 and 40.7 µM respectively). 11c (IC50 = 22.7 and 29.2 µM) and 11f (IC50 = 28.7 and 16.9 µM) were the most potent among all compounds against both PC-3 and MCF-7 respectively with no cytotoxicity against WI-38. Conclusion: The newly synthesized compounds showed good activity against PC-3 and MCF-7 cell lines in comparison with 5-fluorouracil. Compounds 11c and 11f bound with human topoisomerase I similar to its known inhibitors and significantly inhibited its DNA relaxation activity in a dose dependent manner which may rationalize their molecular mechanism as cytotoxic agents.

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