The immunomodulator AS101 induces growth arrest and apoptosis in Multiple Myeloma: Association with the Akt/Survivin pathway

It has been reported that the activation of multiple myeloma (MM) cells by CD40 induces proliferation, growth arrest, and apoptosis. To determine whether the biologic sequelae of CD40 activation in MM cells depends on p53 function, we identified temperature-sensitive p53 mutations in the RPMI 8226 (tsp53E285K) and the HS Sultan (tsp53Y163H) MM cell lines. These cells were then used as a model system of inducible wtp53-like function because wild-type-like p53 is induced at permissive (30°C) but not at restrictive (37°C) temperatures. Using p21-luciferase reporter assays, we confirmed that CD40 induces p53 transactivation in RPMI 8226 and HS Sultan cells cultured under permissive, but not restrictive, conditions. Furthermore, CD40 activation of these MM cells under permissive, but not restrictive, temperatures increased the expression of p53 and p21 mRNA and protein. Importantly, CD40 activation induced the proliferation of RPMI 8226 and HS Sultan cells at restrictive temperatures and growth arrest and increased subG1 phase cells at permissive temperatures. These data confirmed that CD40 activation might have distinct biologic sequelae in MM cells, depending on their p53 status.

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Interleukin-6 promotes multiple myeloma cell growth via phosphorylation of retinoblastoma protein

Blood ◽

10.1182/blood.v88.6.2219.bloodjournal8862219 ◽

1996 ◽

Vol 88 (6) ◽

pp. 2219-2227 ◽

Cited By ~ 67

Author(s):

M Urashima ◽

A Ogata ◽

D Chauhan ◽

MB Vidriales ◽

G Teoh ◽

...

Keyword(s):

Multiple Myeloma ◽

B Cells ◽

Cell Growth ◽

Tumor Cell ◽

Cell Lines ◽

Interleukin 6 ◽

Growth Arrest ◽

Retinoblastoma Protein ◽

Myeloma Cell ◽

Phosphorylated Form

Interleukin-6 (IL-6) mediates autocrine and paracrine growth of multiple myeloma (MM) cells and inhibits tumor cell apoptosis. Abnormalities of retinoblastoma protein (pRB) and mutations of RB gene have been reported in up to 70% of MM patients and 80% of MM-derived cell lines. Because dephosphorylated (activated) pRB blocks transition from G1 to S phase of the cell cycle whereas phosphorylated (inactivated) pRB releases this growth arrest, we characterized the role of pRB in IL-6-mediated MM cell growth. Both phosphorylated and dephosphorylated pRB were expressed in all serum-starved MM patient cells and MM-derived cell lines, but pRB was predominantly in its phosphorylated form. In MM cells that proliferated in response to IL-6, exogenous IL-6 downregulated dephosphorylated pRB and decreased dephosphorylated pRB-E2F complexes. Importantly, culture of MM cells with RB antisense, but not RB sense, oligonucleotide (ODN) triggered IL- 6 secretion and proliferation in MM cells; however, proliferation was only partially inhibited by neutralizing anti-IL-6 monoclonal antibody (MoAb). In contrast to MM cells, normal splenic B cells express dephosphorylated pRB. Although CD40 ligand (CD40L) triggers a shift from dephosphorylated to phosphorylated pRB and proliferation of B cells, the addition of exogenous IL-6 to CD40L-treated B cells does not alter either pRB or proliferation, as observed in MM cells. These results suggest that phosphorylated pRB is constitutively expressed in MM cells and that IL-6 further shifts pRB from its dephosphorylated to its phosphorylated form, thereby promoting MM cell growth via two mechanisms; by decreasing the amount of E2F bound by dephosphorylated pRB due to reduced dephosphorylated pRB, thereby releasing growth arrest; and by upregulating IL-6 secretion by MM cells and related IL-6- mediated autocrine tumor cell growth.

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PU.1 Is Downregulated in a Subset of Multiple Myeloma by the Methylation of Its -17 kb Upstream Regulatory Region and the Promoter.

Blood ◽

10.1182/blood.v108.11.3417.3417 ◽

2006 ◽

Vol 108 (11) ◽

pp. 3417-3417

Author(s):

Yutaka Okuno ◽

Hiro Tatetsu ◽

Shikiko Ueno ◽

Hiroyuki Hata ◽

Yasuhiro Yamada ◽

...

Keyword(s):

Multiple Myeloma ◽

Cell Growth ◽

Cell Lines ◽

Promoter Region ◽

Growth Arrest ◽

Myeloma Cell ◽

Upstream Region ◽

Cell Growth Arrest ◽

Myeloma Cells ◽

Expression Levels

Abstract It has been reported that disruption of transcription factors critical for hematopoiesis, such as C/EBPa and AML1, is involved in leukemogenesis. PU.1 is a transcription factor important for both myeloid and lymphoid development. We reported that mice in which the levels of PU.1 were 20% of that of wild-type developed acute myeloid leukemia, T cell lymphoma, and a CLL-like disease. These findings strongly suggest that PU.1 has tumor suppressive activity in multiple hematopoietic lineages. Last year, we reported that PU.1 is downregulated in a majority of multiple myeloma cell lines and and freshly isolated CD138 positive myeloma cells from certain number of myeloma patients, and that tet-off inducible exogenous expression of PU.1 in PU.1 negative myeloma cell lines induced cell growth arrest and apoptosis. Based on their PU.1 expression levels, we divided the myeloma patients into two groups, namely PU.1 high and PU.1 low-to-negative, (cutoff index of 25th percentile of the PU.1 expression level distribution among all patients). The PU.1 low-to-negative patients had a significantly poorer prognosis than the PU.1 high patients. To elucidate the mechanisms of downregulation of PU.1, we performed sequence and epigenetic analysis of the promoter region and the -17 kb upstream region that is conserved among mammalians and important for proper expression of PU.1. There are no mutations in these regions of all five myeloma cell lines. In contrast, the -17 kb upstream region was highly methylated in 3 of 4 PU.1 negative myeloma cell lines, while the promoter region was also methylated to various levels in all five myeloma cell lines including one PU.1 positive cell line. These data suggested that the downregulation of PU.1 in myeloma cell lines might be dependent on the methylation of both regulatory regions of PU.1 gene, especially the -17 kb upstream region. We also evaluated the mechanisms of cell growth arrest and apoptosis of myeloma cell lines induced by PU.1. Among apoptosis-related genes, we identified that TRAIL was upregulated after PU.1 induction. To evaluate the effect of upregulation of TRAIL, we stably introduced siRNA for TRAIL into myeloma cell lines expressing PU.1, and we found that apoptosis of these cells was partially suppressed by siRNA for TRAIL, suggesting that apoptosis of myeloma cells induced by PU.1 might be at least partially due to TRAIL upregulation. We are currently performing DNA microarray analysis to compare the expression levels of genes between before and after PU.1 induction, in order to further elucidate the mechanisms of cell growth arrest and apoptosis.

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Silica Nanoparticles Sensitize Human Multiple Myeloma Cells to Snake (Walterinnesia aegyptia) Venom-Induced Apoptosis and Growth Arrest

Oxidative Medicine and Cellular Longevity ◽

10.1155/2012/386286 ◽

2012 ◽

Vol 2012 ◽

pp. 1-10 ◽

Cited By ~ 19

Author(s):

Douaa Sayed ◽

Mohamed K. Al-Sadoon ◽

Gamal Badr

Keyword(s):

Cell Cycle ◽

Multiple Myeloma ◽

Silica Nanoparticles ◽

Biological Effects ◽

Growth Arrest ◽

Hematopoietic Stem ◽

Human Multiple Myeloma ◽

Induced Apoptosis ◽

The Impact ◽

Effective Treatments

Background. Multiple myeloma (MM), an almost incurable disease, is the second most common blood cancer. Initial chemotherapeutic treatment could be successful; however, resistance development urges the use of higher toxic doses accompanied by hematopoietic stem cell transplantation. The establishment of more effective treatments that can overcome or circumvent chemoresistance has become a priority. We recently demonstrated that venom extracted fromWalterinnesia aegyptia(WEV) either alone or in combination with silica nanoparticles (WEV+NPs) mediated the growth arrest and apoptosis of prostate cancer cells. In the present study, we evaluated the impact of WEV alone and WEV+NP on proliferation and apoptosis of MM cells.Methods. The impacts of WEV alone and WEV+NP were monitored in MM cells from 70 diagnosed patients. The influences of WEV and WEV+NP were assessed with flow cytometry analysis.Results. WEV alone and WEV+NP decreased the viability of MM cells. Using a CFSE proliferation assay, we found that WEV+NP strongly inhibited MM cell proliferation. Furthermore, analysis of the cell cycle using the propidium iodide (PI) staining method indicated that WEV+NP strongly altered the cell cycle of MM cells and enhanced the induction of apoptosis.Conclusions. Our data reveal the biological effects of WEV and WEV+NP on MM cells that enable these compounds to function as effective treatments for MM.

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2-Methoxyestradiol overcomes drug resistance in multiple myeloma cells

Blood ◽

10.1182/blood-2002-02-0376 ◽

2002 ◽

Vol 100 (6) ◽

pp. 2187-2194 ◽

Cited By ~ 81

Author(s):

Dharminder Chauhan ◽

Laurence Catley ◽

Teru Hideshima ◽

Guilan Li ◽

Richard Leblanc ◽

...

Keyword(s):

Multiple Myeloma ◽

Drug Resistance ◽

Stromal Cells ◽

Growth Arrest ◽

Peripheral Blood Lymphocytes ◽

Leukemic Cells ◽

Vascular Endothelial ◽

Normal Peripheral Blood ◽

Induced Apoptosis ◽

Endothelial Growth Factor

Abstract 2-Methoxyestradiol (2ME2) an estrogen derivative, induces growth arrest and apoptosis in leukemic cells and is also antiangiogenic. In this study, we demonstrate that 2ME2 inhibits growth and induces apoptosis in multiple myeloma (MM) cell lines and patient cells. Significantly, 2ME2 also inhibits growth and induces apoptosis in MM cells resistant to conventional therapies including melphalan (LR-5), doxorubicin (Dox-40 and Dox-6), and dexamethasone (MM.1R). In contrast to its effects on MM cells, 2ME2 does not reduce the survival of normal peripheral blood lymphocytes. Moreover, 2ME2 enhances Dex-induced apoptosis, and its effect is not blocked by interleukin-6 (IL-6). We next examined the effect of 2ME2 on MM cells in the bone marrow (BM) milieu. 2ME2 decreases survival of BM stromal cells (BMSCs), as well as secretion of vascular endothelial growth factor (VEGF), and IL-6 triggered by the adhesion of MM cells to BMSCs. We show that apoptosis induced by 2ME2 is mediated by the release of mitochondrial cytochrome-c (cyto-c) and Smac, followed by the activation of caspases-8, -9, and -3. Finally, 2ME2 inhibits MM cell growth, prolongs survival, and decreases angiogenesis in a murine model. These studies, therefore, demonstrate that 2ME2 mediates anti-MM activity directly on MM cells and in the BM microenvironment. They provide a framework for the use of 2ME2, either alone or in combination with Dex, to overcome drug resistance and to improve outcome in MM.

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The novel non-boronic proteasome inhibitor S-2209 induces apoptosis and growth arrest in multiple myeloma cells

Journal of Clinical Oncology ◽

10.1200/jco.2008.26.15_suppl.8581 ◽

2008 ◽

Vol 26 (15_suppl) ◽

pp. 8581-8581

Author(s):

P. Baumann ◽

K. Müller ◽

S. Mandl-Weber ◽

R. Doblhofer ◽

A. Ammendola ◽

...

Keyword(s):

Multiple Myeloma ◽

Proteasome Inhibitor ◽

Growth Arrest ◽

The Novel ◽

Myeloma Cells

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Autocrine and Paracrine Interactions between Multiple Myeloma Cells and Bone Marrow Stromal Cells by Growth Arrest-specific Gene 6 Cross-talk with Interleukin-6

Journal of Biological Chemistry ◽

10.1074/jbc.m116.733030 ◽

2017 ◽

Vol 292 (10) ◽

pp. 4280-4292 ◽

Cited By ~ 15

Author(s):

Miki Furukawa ◽

Hiroshi Ohkawara ◽

Kazuei Ogawa ◽

Kazuhiko Ikeda ◽

Koki Ueda ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Cross Talk ◽

Stromal Cells ◽

Interleukin 6 ◽

Bone Marrow Stromal Cells ◽

Growth Arrest ◽

Marrow Stromal Cells ◽

Specific Gene ◽

Myeloma Cells

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Myeloma cell growth arrest, apoptosis, and interleukin-6 receptor modulation induced by EB1089, a vitamin D3 derivative, alone or in association with dexamethasone

Blood ◽

10.1182/blood.v88.12.4659.bloodjournal88124659 ◽

1996 ◽

Vol 88 (12) ◽

pp. 4659-4666 ◽

Cited By ~ 30

Author(s):

D Puthier ◽

R Bataille ◽

S Barille ◽

MP Mellerin ◽

JL Harousseau ◽

...

Keyword(s):

Multiple Myeloma ◽

Cell Growth ◽

Vitamin D3 ◽

Interleukin 6 ◽

Plasma Cells ◽

Growth Arrest ◽

Myeloma Cell ◽

Oncostatin M ◽

Receptor Modulation

We have previously shown that malignant plasma cells expressed the specific receptor for 1,25-dihydroxyvitamin D3 and that this derivative could significantly inhibit the proliferation of such malignant cells. More recently, new vitamin D3 derivatives have been generated with extraordinarily potent inhibitory effects on leukemic cell growth in vitro. These new data prompted us to (re)investigate the capacity of such new vitamin D3 derivatives to inhibit myeloma cell growth in comparison with that of dexamethasone, a potent antitumoral agent in multiple myeloma. In the current study, we show that EB1089, a new vitamin D3 derivative, (1) induces G1 growth arrest of human myeloma cells, which is only partially reversed by interleukin-6 (IL-6); (2) induces apoptosis in synergy with dexamethasone, IL-6, leukemia-inhibitory factor, and Oncostatin M, with an agonistic anti-gp130 monoclonal antibody being unable to prevent this apoptosis; (3) downregulates both the gp80 (ie, the alpha chain of the IL-6 receptor [IL-6Ralpha]) expression on malignant plasma cells and the production of soluble IL-6Ralpha, and finally (4) inhibits the deleterious upregulation of gp80 expression induced by dexamethasone while limiting the dexamethasone-induced upregulation of gp130 expression. Considering that these in vitro effects of EB1089 have been observed at doses obtainable in vivo (without hypercalcemic effects), our present data strongly suggest that EB1089 could have a true interest in the treatment of multiple myeloma, especially in association with dexamethasone.

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Interleukin-6 Overcomes p21WAF1 Upregulation and G1 Growth Arrest Induced by Dexamethasone and Interferon-γ in Multiple Myeloma Cells

Blood ◽

10.1182/blood.v90.1.279 ◽

1997 ◽

Vol 90 (1) ◽

pp. 279-289 ◽

Cited By ~ 62

Author(s):

Mitsuyoshi Urashima ◽

Gerrard Teoh ◽

Dharminder Chauhan ◽

Yasutaka Hoshi ◽

Atsushi Ogata ◽

...

Keyword(s):

Cell Cycle ◽

Multiple Myeloma ◽

Protein Expression ◽

Interleukin 6 ◽

Present Report ◽

Growth Arrest ◽

Interferon Γ ◽

Cell Cycle Distribution ◽

P21 Protein ◽

Ifn Γ

Abstract Interleukin-6 (IL-6) is a growth factor for multiple myeloma (MM) cells and can inhibit MM cell apoptosis. Our recent studies show that IL-6 facilitates MM cell growth via phosphorylation of retinoblastoma protein (pRB); however, the effects of IL-6 on those cyclins, cyclin-dependent kinases (CDKs), and CDK inhibitors (CDIs) that are known to regulate phosphorylation of pRB have not been defined in MM cells. In the present report, we cultured MM cell lines and patient cells with IL-6 and/or dexamethasone (Dex) and characterized changes in cell cycle; expression and association of cyclins, CDKs, and CDIs; and phosphorylation of pRB. Dex induced G1 growth arrest in MM cells, whereas IL-6 facilitated G1 to S phase transition; moreover, the effect of Dex was blocked by IL-6. p21WAF1 (p21) protein was constitutively expressed in the majority of MM cells independent of the status of p53. Its expression was upregulated by Dex and downregulated by IL-6; again, IL-6 inhibited the increase in p21 triggered by Dex. These alterations in p21 expression in MM cells were associated with changes in p21 binding to CDK2, CDK4, and CDK6; CDK2, CDK4, and CDK6 kinase activities; and phosphorylation of pRB. In contrast, expression of G1 cell cycle regulatory proteins, including p27KIP1, cyclin D2, and cyclin E, was not altered in MM cells cultured with Dex and/or IL-6. Finally, interferon-γ (IFN-γ) also induced G1 growth arrest and upregulated p21 protein expression; as with Dex, affects of IFN-γ were inhibited by IL-6. Our results therefore show that changes in cell cycle distribution in MM cells triggered by Dex, IL-6, and IFN-γ correlate with changes in p21 protein expression and implicate p21 in the coupling of Dex-, IL-6–, and IFN-γ–related signals to G1 cell cycle regulation in MM cells.

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