Simvastatin Induces Death of Multiple Myeloma Cell Lines

2004 ◽  
Vol 52 (5) ◽  
pp. 335-344 ◽  
Author(s):  
Naomi Gronich ◽  
Liat Drucker ◽  
Hava Shapiro ◽  
Judith Radnay ◽  
Shai Yarkoni ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Multiple Myeloma ◽  
Cell Death ◽  
Cell Lines ◽  
Myeloma Cell ◽  
Cytoplasmic Calcium ◽  
Multiple Myeloma Cell ◽  
Cycle Arrest ◽  
Rpmi 8226

BackgroundAccumulating reports indicate that statins widely prescribed for hypercholesteromia have antineoplastic activity. We hypothesized that because statins inhibit farnesylation of Ras that is often mutated in multiple myeloma (MM), as well as the production of interleukin (IL)-6, a key cytokine in MM, they may have antiproliferative and/or proapoptotic effects in this malignancy.MethodsU266, RPMI 8226, and ARH77 were treated with simvastatin (0-30 μM) for 5 days. The following aspects were evaluated: viability (IC50), cell cycle, cell death, cytoplasmic calcium ion levels, supernatant IL-6 levels, and tyrosine kinase activity.ResultsExposure of all cell lines to simvastatin resulted in reduced viability with IC50s of 4.5 μM for ARH77, 8 μM for RPMI 8226, and 13 μM for U266. The decreased viability is attributed to cell-cycle arrest (U266, G1; RPMI 8226, G2M) and cell death. ARH77 underwent apoptosis, whereas U266 and RPMI 8226 displayed a more necrotic form of death. Cytoplasmic calcium levels decreased significantly in all treated cell lines. IL-6 secretion from U266 cells was abrogated on treatment with simvastatin, whereas total tyrosine phosphorylation was unaffected.ConclusionsSimvastatin displays significant antimyeloma activity in vitro. Further research is warranted for elucidation of the modulated molecular pathways and clinical relevance.

Aptamer TY04 Inhibits Multiple Myeloma Cell Growth Via Cell Cycle Arrest

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 5390-5390
Author(s):  
Jing Liu ◽  
Hong-Juan Dai ◽  
Bian-Ying Ma ◽  
Jian-Hui Song ◽  
Hui-yong Chen ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Multiple Myeloma ◽  
Flow Cytometry ◽  
Cell Growth ◽  
Cell Lines ◽  
Myeloma Cell ◽  
Multiple Myeloma Cell ◽  
Cycle Arrest ◽  
M Phase ◽  
Target Molecules

Abstract Multiple myeloma (MM), also known as plasma cell myeloma, is characterized by accumulation of clonal plasma cells in the bone marrow and overproduction of monoclonal immunoglobulin (Ig) in the blood or urine. MM accounts for approximately 10% of all hematologic malignancies. Despite recent advances in the understanding and treatment of this disease, MM remains an incurable disease in the vast majority. With conventional chemotherapy, the 5-year median survival rate for MM patients is approximately 25%. Aptamers are single-stranded RNA or DNA sequences that bind to target molecules with high affinity and specificity. Compared with antibodies, aptamers have unique advantages including easy chemical synthesis and modification, low toxicity, lack of immunogenicity, and rapid tissue penetration, Based on these advantages, aptamers show great potential for therapeutic application. The aptamer TY04 is a single-stranded DNA (ssDNA) generated by a method named cell-based systematic evolution of ligands by exponential enrichment (cell-SELEX), We found TY04 strongly inhibited the growth of multiple myeloma cell lines including MM1.S, NCI-H929, KM3 and OPM2,The concentration of TY04 to inhibit 50% cell growth (IC50) on MM1.S was 3.89 μM. In contrast, TY04 had no effect on the growth of non-tumor cell lines — immortal B lymphoblastoid cell lines. Next, we used MM1.S cell line as the model to study the mechanism of TY04 anti- multiple myeloma. Flow cytometry analysis showed that TY04 with the sequence specifically bind to MM1.S cells when compared with unselected ssDNA library control. To investigate whether the target molecules of TY04 are membrane proteins on cell surface, MM1.S cells were treated with trypsin and proteinase k for 2 or 10 minutes before incubation with TY04. The result revealed that TY04 lost partly recognition ability on treated cells, indicating that the target molecules were most likely membrane proteins. Furthermore, we evaluated the cell cycle distribution of MM1.S after TY04 treatment. We found that TY04 significantly caused cell-cycle arrest in G2/M phase. The percentage of G2/M phase cells increased from 30.1±1.56 to 53.2±6.36. To identify the underlying molecular mechanism, G2/M-related proteins were assayed by flow cytometry. Following TY04 treatment, a concomitant inhibition of ERK1/2, cyclin B, CDK1 and γ-tubulin expression occurred. Meanwhile, human cell cycle PCR array was used to analyze the expression of 84 genes key to cell cycle regulation in TY04-treated MM1.S cells. Our results indicated that aptamer TY04 decreased the genes expression of CCNB1, CCNB2, BIRC5, BRCA1 and CCNH, which were involved in the progress of G2/M phase. All these results are significant in that they provide a framework for further exploring the use of TY04 as a novel anti-multiple myeloma agent. Disclosures: No relevant conflicts of interest to declare.

Tetraspanin Overexpression Induces Multiple Myeloma Cell Death.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 5067-5067
Author(s):  
Tali Tohami ◽  
Liat Drucker ◽  
Judith Radnay ◽  
Hava Shapiro ◽  
Michael Lishner
Keyword(s):  
Cell Cycle ◽  
Multiple Myeloma ◽  
Cell Death ◽  
Cell Lines ◽  
Caspase 3 ◽  
Annexin V ◽  
Myeloma Cell ◽  
Transfected Cells ◽  
Over Expression ◽  
Rpmi 8226

Abstract Background: Medullary and extra-medullary dissemination of multiple myeloma (MM) cells involves cell-cell and cell-extracellular matrix (ECM) interactions. Proteins coordinating these intricate networks regulate the signaling cascades in a spatial and time dependent manner. Tetraspanins facilitate multiprotein complexing in defined membranal microdomains and select family members have been identified as metastasis suppressors. In preliminary studies, we observed that tetraspanins CD82, frequently down regulated or lost at the advanced clinical stages of various cancers, was absent in MM (8 BM samples, 5 cell lines) and CD81, characteristically expressed in leukocytes plasma membranes, was under-expressed (4/8 BM samples, 4/5 cell lines). We aimed to investigate the consequences of CD81 and CD82 over-expression in myeloma cell lines. Methods: CAG and RPMI 8226 were transfected with pEGFP-N1/C1 fusion vectors of CD81 and CD82. Transfected cells were assessed for - cell morphology (light and fluorescent microscope); cell survival (eGFP+/PI- cells); cell death (Annexin V/7AAD, pre-G1, activated caspase-3 (IC), caspase dependence with pan caspase inhibitor z-VAD-fmk); cell cycle (PI staining). Results: CD82 induced cell death was determined by morphologic characteristics in stably transfected CAG cells (50%) compared to their mock-transfected counterparts (8%) (p<0.05). Activated caspase-3 was also detected (40% of the CD82 transfected cells) (p<0.05). In CD82 transiently transfected MM cell lines a reduced fraction of surviving cells was observed compared to mocks (~60%) (p<0.05) yet, no increases in pre-G1 or Annexin V+/7AAD- subgroups were observed. Moreover, CD82 induced cell death could not be inhibited by the use of z-VAD-fmk. CD82 transfection did not affect the cell cycle of CAG and RPMI 8226 lines. CD81 stably transfected cell lines (CAG and RPMI 8226) could not be established. Indeed, in transiently transfected cells we determined a massive rate of CD81 induced cell death. This is demonstrated in a surviving fraction of only 10% CAG cells and 30% RPMI 8226 (compared to mock) (p<0.05). The CD81 transfected cells were negative for PS exposure, pre-G1 sub-population, or inhibition of death with z-VAD-fmk. The death inducing effect of both tetraspanins in the two cell lines was evident with the pEGFP-N1 orientation vector only. Conclusions: CD81 and CD82 over-expression in MM cell lines causes cell death. Based on the restriction of the killing effect to the pEGFP-N1 clone it may be speculated that its implementation is either dependent on the interactions of the N1 tetraspanin terminus or the proteins’ conformation. It is of interest that CD81 though normally expressed in RPMI 8226 still induced cell death when over-expressed, possibly indicative of ’negative signaling’. Tetraspanins’ suppressive effects on adhesion, motility, and metastasis in solid tumors combined with its capacity to induce myeloma cell death underscore the significance of its absence in MM cell lines and patients. We suspect that a better understanding of CD81/82 mediated signaling pathways will promote future treatment of myeloma cell in their microenvironment. Current studies designed to assess the involvement of oxidative stress in CD81/CD82 induced death are underway.

scholarly journals Investigation of the Proliferation, Apoptosis/Necrosis, and Cell Cycle Phases in Several Human Multiple Myeloma Cell Lines. Comparison ofViscum albumQuFrF Extract with Vincristine in anIn VitroModel

2010 ◽  
Vol 10 ◽  
pp. 311-320 ◽  
Author(s):  
Eva Kovacs
Keyword(s):  
Cell Cycle ◽  
Multiple Myeloma ◽  
Cell Lines ◽  
Myeloma Cell ◽  
Cytostatic Effect ◽  
Multiple Myeloma Cell ◽  
Cytocidal Effect ◽  
Human Multiple Myeloma ◽  
Cell Cycle Phases ◽  
Rpmi 8226

Multiple myeloma is a haematological disorder of malignant plasma cells. Interleukin-6 (IL-6) is a potent growth factor for the proliferation of these cells. Vincristine as a chemotherapeutic agent is used mainly in combination with other chemotherapeutic substances in the treatment of different haematological disorders.Viscum albumQuFrF (VAQuFrF) extract is an experimental drug that is not used in the treatment in tumour patients. It contains 2000 ng lectin and 10 µg viscotoxin in 10 mg extract. In this study, the effects of VAQuFrF extract were compared with those of vincristine in six human multiple myeloma cell lines (Molp-8, LP-1, RPMI-8226, OPM-2, Colo-677, and KMS-12-BM) using anin vitromodel. As parameters, the IL-6 production, proliferation, apoptosis/necrosis, and cell cycle phases of the cells were taken. To measure the IL-6 production, apoptosis/necrosis, and cell cycle phases, the substances were tested in dose ranges of 10, 50, and 100 µg/106cells. To measure the proliferation of the cells, the substances were tested in dose ranges of 1, 5, and 10 µg/105cells. The profile of the antitumour effects of the two substances is identical. (1) Neither VAQuFrF extract nor vincristine produced IL-6 in any cell line. (2) Both substances inhibited the proliferation of the cells (cytostatic effect), arrested the cell cycle phases, and increased the number of apoptotic/necrotic cells (cytocidal effect). At a dose of 10 µg/105cells, VAQuFrF more effectively inhibited the proliferation than vincristine (p< 0.01) in the cell lines Molp-8, LP-1, and RPMI-8226. (3) VAQuFrF affected the tumour cells mainly via cytostatic effect. Vincristine had a clear cytocidal effect. These findings indicate that VAQuFrF extract could be a novel drug in the treatment of multiple myeloma.

A Novel Class of Sulfonanilides Entering Clinical Trials for Targeted Treatment of Multiple Myeloma: Dual-Mechanism Compounds Inhibiting HIF1A-Signaling and Inducing Apoptosis

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2987-2987 ◽  
Author(s):  
Dirk Hose ◽  
Anja Seckinger ◽  
Hartmut Goldschmidt ◽  
Tobias Meißner ◽  
Blanka Leber ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Multiple Myeloma ◽  
Bone Marrow ◽  
Cell Cycle Arrest ◽  
Cell Lines ◽  
Mechanism Of Action ◽  
Myeloma Cell ◽  
Cycle Arrest ◽  
Dual Mechanism

Abstract Abstract 2987 Background. We have recently shown HIF1A to be expressed in 95.4% of CD138-purified myeloma cell samples from previously untreated patients (n= 329), with significantly higher [lower] expression in case of presence of t(4,14) [hyperdiploidy] vs. patients without the respective aberration. This makes HIF1A an interesting target in myeloma treatment. Additionally, we have shown about 40% of myeloma cell samples to have a proliferation-index above the median plus three standard-deviations of normal bone-marrow plasma cells, and we and others have proven proliferation to be associated with adverse prognosis in myeloma. Here, we report on 2 members of a novel class of sulfonanilides, their preclinical activity and pharmacology, and their dual mechanism of action, targeting HIF1A-signaling and inducing apoptosis via cell cycle arrest and tubulin depolymerization. Patients and Methods. The effect of the novel sulfonanilides ELR510444 and ELR510552 on the proliferation of 20 human myeloma cell lines and the survival of 5 primary myeloma cell-samples cultured within their microenvironment were tested. The results of efficacy studies in in two murine models (RPMI8226-xenograft-model and 5T33-model) are also presented. The mechanism of action was investigated using a variety of in-vitro assays (see below). Results. Preclinical activity in Myeloma. i) The sulfonanilides ELR510444 and ELR510552 completely inhibit proliferation of 20/20 tested myeloma cell lines at low nM concentrations and ii) induce apoptosis in 5/5 primary myeloma cell-samples at 6.4 – 32 nM concentration, without major effect on the bone marrow microenvironment. iii) They significantly inhibit tumor growth (xenograft; RPMI8226 mouse model, 6 mg p.o. bid for ELR510444, 15 mg p.o. bid for ELR510552) and bone marrow infiltration (5T33-model; ELR510444, 6 mg/kg p.o. bid × 4d, rest 3d (cycle)). Mechanism of action. Apoptosis induction and G2/M-block. i) Both compounds lead to caspase-3/7 activation and subsequent apoptosis with cellular EC50 values of 50–100 nM. ii) The compounds induce an initial cellular arrest in G2/M and a significant tubulin depolymerizing effect, followed by an increase in a sub-G1 (apoptotic) population after 24h. HIF1A-inhibition. i) Both compounds show a potent inhibition of HIF1A signaling in a cell based reporter assay (HRE-bla HCT-116) at EC50s of 1–25nM, whereas ii) at concentrations of 1 μ M, neither of the compounds shows an effect in assay systems monitoring the JAK/STAT, NFκB, PI3K/AKT/FOXO or Wnt/β-catenin-signaling pathways. iii) Kinase inhibition profiling showed no significant inhibition at 1μ M in two assays assessing 100 (Invitrogen) and 442 (Ambit) kinases, respectively. Pre-clinical pharmacology. Single dose exposure of 25 mg p.o. yields a maximum concentration of 1.1 μ M with a half life time of 3.6 hours (ELR510444) and 2.7 μ M and 6.6 h (ELR510552) in mice, respectively. The compounds are well-tolerated at levels that are significantly above the in vitro EC50 in all myeloma cell lines and primary samples tested. Conclusion. ELR510444 and ELR510552 are very active on all tested myeloma cell lines and primary myeloma cells without major impact on the bone marrow microenvironment, and show activity in two different mouse models. The compounds inhibit HIF1A-signaling and induce apoptosis via cell cycle arrest and tubulin depolymerization. Preclinical pharmacology data show favorable in vivo profiles with exposure levels in mice significantly higher than concentrations required for in vitro activity. Therefore, this novel class of compounds represents a promising weapon in the therapeutic arsenal against multiple myeloma entering a phase I/II trial within the next year. Disclosures: Leber: ELARA Pharmaceuticals GmbH: Employment. Janssen:ELARA Pharmaceuticals GmbH: Employment. Lewis:ELARA Pharmaceuticals GmbH: Employment. Schultes:ELARA Pharmaceuticals GmbH: Employment.

Bortezomib (PS-341) Sensitizes Multiple Myeloma Cell Lines to Etoposide-Induced Apoptosis.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 5166-5166
Author(s):  
Ivana Zavrski ◽  
Martin Kaiser ◽  
Marleen Rosche ◽  
Ulrike Heider ◽  
Christian Jakob ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Multiple Myeloma ◽  
Cell Lines ◽  
Synergistic Effects ◽  
Myeloma Cell ◽  
Cellular Level ◽  
Treatment Schedule ◽  
Topoisomerase Iiα ◽  
Multiple Myeloma Cell ◽  
Cycle Arrest

Abstract Proteasome inhibitor bortezomib interacts with the regulation of protein turnover in eukaryotic cells. The results of proteasomal inhibition consist in cell cycle arrest and induction of apoptosis. Etoposid targets topoisomerase IIα and is included in several protocols for myeloma treatment. In this study, we examined the effects of simultaneous and sequential treatments of bortezomib and the topoisomerase IIα inhibitor etoposide in multiple myeloma cell lines, particularly in terms of potential synergistic effects between both drugs. Using the MTT assay, cytotoxicity levels for dosages ranging from 0.01 nM and 100 nM for bortezomib and 0.001 μM and 100 μM for etoposide in multiple myeloma cell lines OPM-2, NCI-H929 and RPMI-S were determined and IC50 values calculated. In each experiment, cells were treated with each drug individually and with fixed ratios of both drugs simultaneously (co-incubation over 48 h) and sequentially (pre-incubation with etoposide for 24 h and co-incubation with bortezomib for additional 24 h). The data were analyzed using the median effect method of Chou and Talalay, whereas the combination indices (CI) were calculated for each level of cytotoxicity. A CI&lt; 1 indicated synergy, a CI= 1 indicated additivity and a CI&gt; 1 indicated antagonism. In the sequential treatment schedule, we found synergistic effects in all three cell lines, even at low single-agent cytotoxicity levels (fractional inhibition &lt;50%). Synergistic action were found in RPMI-S with a CI-range between 0.08 and 0.9, in OPM-2 between 0.05 and 0.07 and in NCI-H929 between 0.65 and 0.86 in different doses of both drugs. Interestingly, when cells were treated simultaneously with both drugs, no synergistic effects were observed. On cellular level, we found cell cycle arrest in the G2-M phase of the cell cycle, when cells were treated with both drugs sequentially. Furthermore, we noticed a correlation between the etoposide-sensitivity and G2-M/S-fraction of the sample, indicating that prolonged drug action in the G2-M/S-phase might contribute to enhanced growth inhibitory effects. On sub-cellular level, the synergy was accompanied by diminished activation of transcriptional factor NF-κB, whereas bortezomib abrogated the etoposide-induced NF-κB up-regulation. This effect was accompanied by down-regulation of Bcl-2, an anti-apoptotic protein and transcriptional product of NF-κB. Furthermore, when cells were treated simultaneously with both drugs, we noticed a translocation of topoisomerase IIα from nucleus into the cytoplasm, where the enzyme remained inactive. In sequential treatment schedule, no translocation of topoisomerase IIα was observed, indicating that the drug target remained in its functional compartment. In conclusion, our data show strong synergistic effects between bortezomib and etoposide in appropriate treatment schedules at low drug concentrations in vitro, indicating that the drug combination might be useful for clinical trials in multiple myeloma patients.

The Novel NFkB Inhibitor V1810 Induces Apoptosis and Cell Cycle Arrest in Multiple Myeloma Cells and Overcomes NFkB Mediated Drug Resistance.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1715-1715 ◽  
Author(s):  
Felix Meinel ◽  
Sonja Mandl-Weber ◽  
Philipp Baumann ◽  
Johann Leban ◽  
Ralf Schmidmaier
Keyword(s):  
Cell Cycle ◽  
Multiple Myeloma ◽  
Drug Resistance ◽  
Cell Death ◽  
Cyclin D1 ◽  
Cell Cycle Arrest ◽  
Cyclin D3 ◽  
Cycle Arrest ◽  
Rpmi 8226

Abstract Multiple Myeloma (MM) is a fatal malignancy characterised by the accumulation and expansion of antibody producing plasma cells in the bone marrow. Evidence is increasing that nuclear factor kappa B (NFκB) is a promising target for new anti-myeloma therapies. In this study, we assessed the in vitro activity of V1810, a novel NFκB inhibitor. V1810 potently induces cell death in all tested MM cell lines (OPM-2, U266, NCI-H929, RPMI-8226) with an IC50 ranging between 5μM to 10μM as well as in primary MM cells from patients. Cell death induced by V1810 clearly shows biological features of apoptosis such as DNA fragmentation and caspase 3 cleavage. In OPM2, U266 and RPMI-8226 cells induction of apoptosis is accompanied by cell cycle arrest. Western blots revealed downregulation of cyclin D1 (U266) or cyclin D2 (OPM2, NCI-H929, RPMI-8226) respectively, but not cyclin D3. Consistent with the downregulation of cyclin D1/2, retinoblastoma protein was found to be hypophosphorylated. Considering that cyclin D1 and D2 are known to be NFκB target genes, this is in line with our finding that V1810 inhibits baseline NFκB activity in MM cells (36% relative reduction). Importantly, V1810 also abrogates NFκB activation induced by genotoxic drugs like melphalan and doxorubicin. Accordingly, V1810 and melphalan synergistically decrease MM cell viability. Taken together, V1810 induces apoptosis and cell cycle arrest in MM cells by inhibition of NFκB and overcomes NFκB mediated drug resistance to melphalan. The maximum tolerable dose (MTD) of V1810 in BalbC mice was 10mg/kg i.v. and plasma concentrations of 9.5μM are achievable in NRMI mice after 5mg/kg V1810 i.v., which corresponds well to the used in vitro concentrations. This study strongly supports the further development of NFkB inhibitors in MM, especially in combination with genotoxic drugs.

scholarly journals In Vitro JAK1/2 Inhibition, in Association with Bortezomib and Lenalidomide, Is Comparable with Bortezomib, Lenalidomide and Dexametasone: An Alternative for Multiple Myeloma Patients with JAK2 overexpression?

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 5663-5663
Author(s):  
Mariana Bleker de Oliveira ◽  
Veruska Lia Fook Alves ◽  
Angela Isabel Eugenio ◽  
Rodrigo Carlini Fernando ◽  
Mihoko Yamamoto ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Multiple Myeloma ◽  
Cell Death ◽  
Cell Lines ◽  
U266 Cell ◽  
Late Apoptosis ◽  
Rpmi 8226 ◽  
Number Of Cells ◽  
Stat Pathway

Abstract Background: Despite the absence of JAK1 and JAK2 mutations in multiple myeloma (MM), high levels of IL-6 constitutively activate the JAK/STAT pathway promoting survival and proliferation of tumor cells. Therefore, pharmacological inhibition of JAK proteins can be a potentially therapeutic strategy for myeloma treatment. Aims: 1) to identify expression of JAK1 and JAK2 in MM cell lines and in recently diagnosed MM patients; 2) to perform functional in vitro studies in MM cell lines treated with JAK/STAT pathway inhibitor (ruxolitinib), associated with drugs currently used in MM first line treatment (bortezomib, lenalidomide and dexamethasone), with and without co-culture with normal stromal cells; 3) to evaluate global gene expression of JAK/STAT pathway in cell lines treated with ruxolitinib to elucidate its mechanism of action in MM. Methods: JAK1 and JAK2 expression were analyzed in four cell lines (RPMI-8226, U266, SKO-007 and SKM-M2) and in bone marrow samples from 30 MM patients and 3 healthy controls by real time PCR. After IC50 calculation, drugs concentrations were: bortezomib (B) 10 nM for both RPMI-8226 and U266 cell lines; ruxolitinib (R) 30 µM for RPMI-8226 and 40 µM for U266 cell lines; lenalidomide (L) 10 µM for both cell lines; and dexamethasone (D) 1 µM for both cell lines. Apoptosis and cell cycle were evaluated by flow cytometry. PCR array for 92 JAK/STAT pathway related genes (Taqman® Array Human JAK/STAT Pathway, Applied Biosystems, Foster City, CA, USA) was performed in RPMI-8226 and U266 wild type and B+R treated cell lines, in duplicates. Results: Among the four cell lines, U266 presented the highest expression of JAK1 and JAK2 genes. JAK1 was overexpressed in 27% and JAK2 in 57% of 30 MM patients (considering at least 2-fold increase). After B+R treatment, RPMI-8226 showed increased number of cells in SubG0 phase (p<0.001) with reduction of cells in S (p<0.01) and G2/M (p<0.001) phases. In U266 cell line, there is a slight increase of cells in SubG0 phase (p<0.05). Also, after B+R treatment, both RPMI-8226 and U266 presented 50% of cells in late apoptosis, which was accompanied by reduction of expression levels of BCL-2 and BCL-XL anti-apoptotic genes. The expression profile of JAK/STAT pathway after B+R treatment showed that many JAK/STAT, Ras/Raf/MAPK and PI3K/Akt/mTOR pathways genes lost their expression, mainly in RPMI-8226, with insignificant changes in U266 expression pattern. Co-culture of RPMI-8226 with normal stromal cell line HS5 protected tumor cells from apoptosis, as the number of cells in late apoptosis decreased from 50% to 32% (p<0.001). The addition of immunomodulatory drug lenalidomide to the schedule (B+R+L) increased tumor cell death from 32% to 73% in co-culture (p<0.001). Despite the impressive results, B+R+L schedule was equivalent to currently used treatment for standard risk MM patients B+L+D (67% of cell death, p>0.05), in co-culture. Conclusion: B+R combination induced cell cycle arrest and apoptosis in U266 and RPMI-8226. The new drug combination B+R+L has in vitro results comparable with B+L+D and presents an alternative for MM treatment of almost 60% of cases bearing JAK2 overexpression. Our results support future studies using JAK inhibitors as an alternative for MM treatment in a Precision Medicine approach. Financial support: FAPESP 2010/17668-6 and CNPq. Disclosures No relevant conflicts of interest to declare.

Stew in its Own Juice: Protein Homeostasis Machinery Inhibition Reduces Cell Viability in Multiple Myeloma Cell Lines

2019 ◽  
Vol 19 (2) ◽  
pp. 112-119 ◽  
Author(s):  
Mariana B. de Oliveira ◽  
Luiz F.G. Sanson ◽  
Angela I.P. Eugenio ◽  
Rebecca S.S. Barbosa-Dantas ◽  
Gisele W.B. Colleoni
Keyword(s):  
Multiple Myeloma ◽  
Cell Death ◽  
Cell Viability ◽  
Cell Lines ◽  
Treatment Options ◽  
Compensatory Mechanism ◽  
Protein Homeostasis ◽  
Protein Response ◽  
Rpmi 8226

Introduction:Multiple myeloma (MM) cells accumulate in the bone marrow and produce enormous quantities of immunoglobulins, causing endoplasmatic reticulum stress and activation of protein handling machinery, such as heat shock protein response, autophagy and unfolded protein response (UPR).Methods:We evaluated cell lines viability after treatment with bortezomib (B) in combination with HSP70 (VER-15508) and autophagy (SBI-0206965) or UPR (STF- 083010) inhibitors.Results:For RPMI-8226, after 72 hours of treatment with B+VER+STF or B+VER+SBI, we observed 15% of viable cells, but treatment with B alone was better (90% of cell death). For U266, treatment with B+VER+STF or with B+VER+SBI for 72 hours resulted in 20% of cell viability and both treatments were better than treatment with B alone (40% of cell death). After both triplet combinations, RPMI-8226 and U266 presented the overexpression of XBP-1 UPR protein, suggesting that it is acting as a compensatory mechanism, in an attempt of the cell to handle the otherwise lethal large amount of immunoglobulin overload.Conclusion:Our in vitro results provide additional evidence that combinations of protein homeostasis inhibitors might be explored as treatment options for MM.

CD40 activation mediates p53-dependent cell cycle regulation in human multiple myeloma cell lines

Blood ◽  
2000 ◽  
Vol 95 (3) ◽  
pp. 1039-1046 ◽  
Author(s):  
G. Teoh ◽  
Y.-T. Tai ◽  
M. Urashima ◽  
S. Shirahama ◽  
M. Matsuzaki ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Cell Lines ◽  
Growth Arrest ◽  
Myeloma Cell ◽  
Luciferase Reporter ◽  
Temperature Sensitive ◽  
Multiple Myeloma Cell ◽  
Cd40 Activation ◽  
Human Multiple Myeloma ◽  
Rpmi 8226

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.

scholarly journals Induction of Apoptosis, Autophagy and Ferroptosis by Thymus vulgaris and Arctium lappa Extract in Leukemia and Multiple Myeloma Cell Lines

Molecules ◽  
2020 ◽  
Vol 25 (21) ◽  
pp. 5016
Author(s):  
Aveen N. Adham ◽  
Mohamed Elamir F. Hegazy ◽  
Alaadin M. Naqishbandi ◽  
Thomas Efferth
Keyword(s):  
Cell Cycle ◽  
Multiple Myeloma ◽  
Cell Death ◽  
Ethyl Acetate ◽  
Cell Cycle Arrest ◽  
Cell Lines ◽  
Thymus Vulgaris ◽  
Arctium Lappa ◽  
Cycle Arrest ◽  
Induction Of Apoptosis

Thymus vulgaris and Arctium lappa have been used as a folk remedy in the Iraqi Kurdistan region to deal with different health problems. The aim of the current study is to investigate the cytotoxicity of T. vulgaris and A. lappa in leukemia and multiple myeloma (MM) cell lines and determine the mode of cell death triggered by the most potent cytotoxic fractions of both plants in MM. Resazurin assay was used to evaluate cytotoxic and ferroptosis activity, apoptosis, and modulation in the cell cycle phase were investigated via Annexin V-FITC/PI dual stain and cell-cycle arrest assays. Furthermore, we used western blotting assay for the determination of autophagy cell death. n-Hexane, chloroform, ethyl acetate, and butanol fractions of T. vulgaris and A. lappa exhibited cytotoxicity in CCRF-CEM and CEM/ADR 5000 cell lines at concentration range 0.001–100 μg/mL with potential activity revealed by chloroform and ethyl acetate fractions. NCI-H929 displayed pronounced sensitivity towards T. vulgaris (TCF) and A. lappa (ACF) chloroform fractions with IC50 values of 6.49 ± 1.48 and 21.9 ± 0.69 μg/mL, respectively. TCF induced apoptosis in NCI-H929 cells with a higher ratio (71%), compared to ACF (50%) at 4 × IC50. ACF demonstrated more potent autophagy activity than TCF. TCF and ACF induced cell cycle arrest and ferroptosis. Apigenin and nobiletin were identified in TCF, while nobiletin, ursolic acid, and lupeol were the main compounds identified in ACF. T. vulgaris and A. lappa could be considered as potential herbal drug candidates, which arrest cancer cell proliferation by induction of apoptosis, autophagic, and ferroptosis.

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