scholarly journals Role and Mechanism of circ_0058063/miR-635 Axis in the Malignant Phenotype of Multiple Myeloma RPMI8226 Cells

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Xiaoya Li ◽  
Lingzhi Ding ◽  
Geyu Gu ◽  
Changjun Zheng ◽  
Chenshuai Pan ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Protein Expression ◽  
Migration And Invasion ◽  
Normal Bone Marrow ◽  
Malignant Phenotype ◽  
Ki 67 ◽  
Myeloma Cells ◽  
Bone Marrow Tissue ◽  
Normal Bone

Objective. This study aims to explore circ_0058063 effect on multiple myeloma cells malignant phenotype and its feasible mechanism. Methods. We selected 47 cases of multiple myeloma tissues and 47 cases of normal bone marrow tissues and then used RT-qPCR method to test circ_0058063 and miR-635 expression in the tissues. Myeloma cells RPMI8226 were transfected with si-circ_0058063, miR-635 mimic, and si-circ_0058063 + anti-miR-635, respectively. Then, we adopt CCK-8 method, flow cytometry method, and Transwell and western blot methods to detect the influences of knockdown of circ_0058063 or miR-635 overexpression on RPMI8226 cell proliferation, apoptosis, migration, and invasion and also Ki-67, Bax, Bcl-2, MMP-2, and MMP-9 protein expression. The dual luciferase reporter gene assay experiment proved that it has regulatory relationship between circ_0058063 and miR-635. Results. circ_0058063 expression of multiple myeloma was higher than that in normal bone marrow tissue ( P < 0.05 ), while miR-635 expression was lower than that in normal bone marrow tissue ( P < 0.05 ). Knockdown of circ_0058063 or overexpression of miR-635 could reduce proliferation capacity, migration, invasion cell quantities, and Ki-67, MMP-2, MMP-9, and Bcl-2 protein expression ( P < 0.05 ), while increasing apoptosis rate together with Bax protein expression ( P < 0.05 ). circ_0058063 targets to negatively regulate miR-635, while knocking down miR-635 reverses the influences of knocking down circ_0058063 on RPMI8226 proliferation, apoptosis, migration, and invasion. Conclusion. circ_0058063 expression increased in multiple myeloma tissues. Knocking down its expression may inhibit myeloma proliferation, migration, and invasion by targeting and upregulating miR-635 and also promote cell apoptosis. As for multiple myeloma treatment, circ_0058063/miR-635 may provide new molecular targets.

Prognostication by miRNome-Profiling in Multiple Myeloma

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1822-1822
Author(s):  
Anja Seckinger ◽  
Tobias Meißner ◽  
Vladimir Benes ◽  
Sabine Schmidt ◽  
Jonathan Blake ◽  
...  
Keyword(s):  
Gene Expression ◽  
Multiple Myeloma ◽  
Bone Marrow ◽  
Plasma Cells ◽  
Proliferation Index ◽  
Normal Bone Marrow ◽  
Global Test ◽  
Myeloma Cells ◽  
Normal Bone ◽  
Bone Marrow Plasma

Abstract Abstract 1822 INTRODUCTION. MicroRNAs are an abundant class of small non-protein-coding RNAs that function as negative gene regulators in diverse biological processes including cancer by affecting the stability and translation of mRNAs. METHODS. We determined expression of 559 miRNAs by miChip (Exiqon LNA Array probes V9.2) in CD138-purified myeloma cells from previously untreated patients (n=69), normal bone marrow plasma cells of healthy donors (pooled to n=3), and human myeloma cell lines (n=20). For normalization, an invariant-based method was applied. Gene expression profiling was performed using Affymetrix U133 2.0 DNA-microarrays. RESULTS. We found 29 miRNAs to be significantly up- and 35 down-regulated in myeloma cells vs. normal bone marrow plasma cells, respectively. Expression of 18 miRNAs was simultaneously significantly (P<.01) associated with event-free survival (EFS) and overall survival (OS), and allowed the delineation of prognostic groups. Of these, miRNAs miR-659 (located at 22q12.1) was significantly lower expressed in myeloma cells compared to normal bone marrow plasma cells. For this miRNA, low expression delineates a group with inferior EFS (median 19.7 months vs. not reached, P<.001) and OS (median 52.9 months vs. not reached, P<.001). This group shows a significantly higher gene expression based proliferation index. In contrast, high miR-590 expression (located at 7q11.23) delineates a group with inferior EFS (median 12.4 vs. 36 months, P<0.001) and OS (median 29.4 months vs. not reached, P<.001). By using Goeman's global test, a significant association of the predicted target gene signatures with survival could be found for both, EFS (miR-590-5p, P<.001; miR-659, P<.001) and OS (P=.009; P=.002). CONCLUSION. In conclusion, we demonstrate the prognostic relevance of miRNome profiling in multiple myeloma. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Aberrant Ki-67 expression in normal bone marrow revealed by multiparameter flow cytometric analysis

Cytometry ◽  
1991 ◽  
Vol 12 (1) ◽  
pp. 50-63 ◽  
Author(s):  
Dirk R. Van Bockstaele ◽  
Jar Lan ◽  
Hans-W. Snoeck ◽  
Marcel L. Korthout ◽  
Robrecht F. De Bock ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometric Analysis ◽  
Normal Bone Marrow ◽  
Ki 67 ◽  
Normal Bone ◽  
Flow Cytometric

Role for PP2A Regulatory Subunit B55α as A Regulator of AKT and Other Signaling Pathways In AML.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1668-1668
Author(s):  
Peter P. Ruvolo ◽  
YiHua Qui ◽  
Kevin R Coombes ◽  
Nianxiang Zhang ◽  
Vivian Ruvolo ◽  
...  
Keyword(s):  
Gene Expression ◽  
Bone Marrow ◽  
Protein Expression ◽  
Normal Bone Marrow ◽  
Protein Levels ◽  
B Subunit ◽  
Normal Bone ◽  
Cd34 Cells ◽  
Blast Cells ◽  
B Subunits

Abstract Abstract 1668 Activation of survival kinases such as Protein Kinase B (AKT), Protein Kinase C (PKC), and Extracellular Receptor Activated Kinase (ERK) predict poor clinical outcome for patients with acute myeloid leukemia (AML; Kornblau et al Blood 2006). A better understanding of how the activities of these kinases are regulated by phosphorylation and dephosphorylation will enable the development of targeted therapies directed against this axis. Protein Phosphatase 2A (PP2A) negatively regulates PKC, AKT, and ERK but its role in AML is not clear. In the current study we examined the role of PP2A in regulating AKT in AML. Activation of AKT involves phosphorylation of threonine 308 (T308) and serine 473 (S473). A recent study has indicated that phosphorylation of AKT at T308 but not S473 is a poor prognostic factor for AML patients and that PP2A activity negatively correlated with T308 phosphorylation (Gallay et al Leukemia 2009). PP2A is a family of different isoforms that form hetero-trimers consisting of a catalytic C subunit, a scaffold A subunit, and one of at least 21 different regulatory B subunits. The functionality of each PP2A isoform is determined by the regulatory B subunit. Thus to understand PP2A regulation of AKT in AML, it is essential to study the B subunit that regulates the AKT phosphatase. The PP2A isoform regulating AKT in the AML patients is currently unknown. Evidence suggests that the B55a subunit is responsible for dephosphorylation of AKT at T308. In the current study, we compared B55α gene expression in blast cells derived from AML patients with normal counterpart (i.e. CD34+) cells derived from normal bone marrow donors by real time PCR. Surprisingly, B55α gene expression was higher in the patients. Reverse Phase Protein Analysis (RPPA) is a powerful tool that allows for the analysis of protein expression from patient samples. Protein levels of the PP2A B subunit were analyzed by RPPA in AML blast cells obtained from 511 newly diagnosed AML patients and CD34+ cells obtained from 11 normal bone marrow donors. Levels of B55α protein were significantly lower in the blast cells from the AML patients compared to normal CD34+ cells. While the mechanism for the observed difference in gene versus protein expression in the leukemia cells has yet to be determined, a plausible mechanism is that the B55α protein is being proteolyzed since monomeric PP2A B subunits that are not part of the PP2A hetero-trimer are degraded. Importantly the reduced levels of B55α protein observed would be predicted if AKT were activated in the AML blast cells. We next compared AKT phosphorylation status with B55α protein expression in the AML blast cells using RPPA to answer this question. Analysis of RPPA data revealed that there was no correlation between B55α protein levels and levels of total AKT protein or with levels of AKT phosphorylated at S473 in the AML samples. However, there was a moderate but significant negative correlation between B55α protein levels and levels of AKT phosphorylated at T308. This result suggests that B55α is mediating dephosphorylation of AKT at T308 but not S473 in the AML cells. B55α expression was not associated with FAB classification but was positively correlated with high blast and peripheral blood counts. While the level of expression of the B subunit did not correlate with overall survival, intermediate levels of B55α expression were associated with longer complete remission duration. We predict that higher levels of B55α would reflect low levels of other PP2A B subunits. Consistent with this prediction, B55α expression positively correlated with MYC expression in the AML patients. MYC expression is regulated by a B subunit that competes with B55α (i.e. B56α). These findings suggest that B55α may play an important role in AML as a negative regulator of AKT and perhaps by other as yet unidentified functions. Activation of B55α is a potential therapeutic target for overcoming the AKT activation frequently observed in AML. Disclosures: No relevant conflicts of interest to declare.

scholarly journals IKZF1/3 Protein Expressions Are Associated with a Better Survival in Relapsed/Refractory Multiple Myeloma Patients Treated with Lenalidomide

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4506-4506
Author(s):  
Maryam Pourabdollah ◽  
Mohammad Bahmanyar ◽  
Eshetu G Atenafu ◽  
Donna Reece ◽  
Hong Chang
Keyword(s):  
Risk Factors ◽  
Multiple Myeloma ◽  
Bone Marrow ◽  
Protein Expression ◽  
Research Funding ◽  
Nuclear Staining ◽  
Cytoplasmic Staining ◽  
Myeloma Cells ◽  
Refractory Multiple Myeloma ◽  
Sequence Of Events

Abstract It has been demonstrated that lenalidomide causes selective degradation of IKZF1 (Ikaros) and IKZF3 (Aiolos) which are two essential transcription factors for proliferation of multiple myeloma cells. Consequently, the drug sets up a molecular sequence of events that lead to programmed cell death in the tumoral cells. This anti-proliferative effect is mediated by down-regulation of c-Myc and interferon regulatory factor 4 (IRF4). However, it is not clear whether IKZF1/IKZF3 protein expression in myeloma cells is predictive of clinical outcome. Thus, we evaluated bone marrow samples of 50 relapsed/refractory multiple myeloma (MM) patients regarding IKZF1/3 protein expression before starting lenalidomide. There were 31 males and 19 females with median age of 59 years (range 41-75). They all had received lenalidomide-based therapy after relapse following autologous stem cell transplantation (ASCT), thalidomide, or bortezomib. The median follow-up was 86.4 months. By immunohistochemistry (IHC), CD138 positive myeloma cell aggregates were examined for IKZF1 and IKZF3 protein expression. We used H-score method (range 0-300) based on the intensity and percentage of the stained myeloma cells. Cases were considered positive for IKZF1 or IKZF3 if H-score is equal or over 150 or 200, respectively. IKZF1 showed nuclear staining but IKZF3 showed both nuclear and cytoplasmic staining. IKZF1 and IKZF3 were expressed in 72% and 58% of the bone marrow specimens, respectively. IKZF1 and IKZF3 expressions were strongly correlated (p<0.0001). Both IKZF1 & IKZF3 expressions were associated with longer progression free (p=0.0029 & p<0.0001, respectively) and overall survivals (p=0.0014 & p<0.0001, respectively). IKZF3 (p=0.0025) expression but not IKZF1 (p=0.094) was correlated with the clinical response to lenalidomide. There was no significant association between IKZF1/3 protein expression and other clinical or biological risk factors including age, gender, International staging system (ISS), hemoglobin, calcium, creatinine, bone lytic lesions, β2 micro-globulin, albumin or cytogenetic risk factors such as del (13q), del (17p), t(4;14), and amp (1q21). Our study demonstrates that expressions of the IKZF1/3 proteins detected by IHC are correlated with superior survival outcome in refractory MM patients treated with lenalidomide. IHC is routinely available, robust and inexpensive method. Thus, if confirmed in a larger prospective study, IKZF1/3 immunostaining can be readily adopted in clinical practice for prediction of drug response and clinical outcomes in MM patients receiving lenalidomide therapy. Disclosures Reece: Celgene: Consultancy, Honoraria, Research Funding; Otsuka: Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding; Merck: Research Funding; BMS: Honoraria, Research Funding; Novartis: Honoraria, Research Funding.

The Proliferation Inhibitor CDKN1C (P57KIP2) Is Over Expressed in CD34+ Cells of Patients with MDS and Determines a Worse Prognosis Independently of IPSS Score Factors

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3820-3820
Author(s):  
Sascha Dietrich ◽  
Mindaugas Andrulis ◽  
Andrea Pellagatti ◽  
Aleksandar Radujkovic ◽  
Ulrich Germing ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Overall Survival ◽  
Protein Expression ◽  
Normal Bone Marrow ◽  
Double Staining ◽  
Expression Levels ◽  
Cyclin Dependent Kinases ◽  
Normal Bone ◽  
Cd34 Cells ◽  
Worse Prognosis

Abstract Abstract 3820 Purpose: Progressive cytopenias are the cause of death for the majority of patients with myelodysplastic syndromes (MDS). In order to investigate if the proliferative activity of CD34+ cells in MDS bone marrow correlates with prognosis, we measured expression levels of the proliferation inhibitor CDKN1C (P57KIP2) in two independent study cohorts. Patients and methods: Gene expression profiling data on bone marrow CD34+ cells were obtained from 183 MDS patients (55 with RA, 48 with RARS, 37 with RAEB1 and 43 with RAEB2). mRNA expression levels of CDKN1C (P57KIP2) were correlated with overall survival (OS) and proliferation markers such as PCNA, cyclins and cyclin dependent kinases. In a second independent patient cohort comprising 93 patients (17 had RA, 13 REAB1, 27 RAEB2, 29 AML arising from MDS and 7 CMML), protein expression levels of CDKN1C (P57KIP2) were evaluated by immune histochemistry (IHC) in trephine biopsies. Protein expression levels of CDKN1C (P57KIP2) were correlated with clinical outcome, OS and the proliferation marker KI67 in CD34+ cells (double staining). Results: In purified CD34+ cells, mRNA expression of CDKN1C (P57KIP2) correlated with higher risk and poorer overall survival of patients with MDS (p=0.0006, n=183, Figure1). Furthermore, increased CDKN1C (P57KIP2) expression was significantly associated with loss of CD38 (p=0.002) as well as loss of proliferating cell nuclear antigen (PCNA, p<0.001), cyclins and cyclin-dependent kinases (p<0.001) underlining the role of CDKN1C (P57KIP2) as a proliferation inhibitor. Similarly, protein expression of CDKN1C (P57KIP2) determined in trephine biopsies predicted a poor prognosis of patients with MDS (p=0.0003, HR=2.2, n=93, Figure 1). No expression of CDKN1C (P57KIP2) could be demonstrated in 10 control patients with normal bone marrow. Separate evaluation of WHO risk categories revealed that in patients with less than 10% blasts (RA, RAEB1 and CMML1), CDKN1C (P57KIP2) was not predictive for OS. In contrast, patients with high CDKN1C (P57KIP2) expression and RAEB2 (p=.0.02, HR 2.4) or AML arising from MDS (p=0.002, HR 2.9) had a significantly worse OS than patients with low CDKN1C (P57KIP2) levels. CDKN1C (P57KIP2) expression analysis within the group of allogeneic stem cell recipients showed no impact on survival after transplant. Multivariate cox regression analysis with the confounding co-variates: age, IPSS score factors (cytopenias, cytogenetic risk profile, blast count) and primary versus secondary MDS confirmed the independent impact of CDKN1C (P57KIP2) expression on OS (p=0.0002, HR 2.9). CDKN1C (P57KIP2) protein expression could also be demonstrated in sorted CD34+ cells of MDS patients by western blot analysis and was significantly higher than in CD34- cells (p=0.03, n=7). KI67 expression was evaluated in CD34+ cells by IHC double staining in 34 trephine biopsies and 10 control patients with normal bone marrow. Percentages of CD34+ and KI67 positive cells were higher in control patients and patients with low risk MDS (RA, RAEB1) than in patients with high risk MDS (RAEB2) (p<0.01). Patients with AML (>20% blasts) had significantly higher levels of KI67 (p<0.05). In patients with MDS (<20% blasts) KI67 percentages correlated inversely with CDKN1C (P57KIP2) expression (p=0.04). Conclusions: High CDKN1C (P57KIP2) expression in CD34+ cells of patients with MDS is associated with a reduced fraction of proliferative CD34+ cells and determines a worse prognosis independently of factors used to calculate the IPSS score. Allogeneic stem cell transplantation could overcome the worse prognosis of high CDKN1C (P57KIP2) expression. Further studies are warranted to determine the impact of CDKN1C (P57KIP2) expression to guide clinical decisions. Disclosures: No relevant conflicts of interest to declare.

Proliferating normal bone marrow cells do stain for Ki-67 antigen

1993 ◽  
Vol 85 (4) ◽  
pp. 835-836 ◽  
Author(s):  
Giorgio Cattoretti ◽  
Attilio Orazi ◽  
Johannes Gerdes
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Cells ◽  
Normal Bone Marrow ◽  
Ki 67 ◽  
Normal Bone ◽  
Marrow Cells

scholarly journals High levels of interleukin-6 are associated with low tumor burden and low growth fraction in multiple myeloma

Blood ◽  
1994 ◽  
Vol 83 (7) ◽  
pp. 1903-1908
Author(s):  
OF Ballester ◽  
LC Moscinski ◽  
GH Lyman ◽  
JV Chaney ◽  
HI Saba ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Growth Factor ◽  
Interleukin 6 ◽  
Tumor Burden ◽  
Lower Percentage ◽  
Growth Fraction ◽  
Ki 67 ◽  
Myeloma Cells ◽  
Beta 2 Microglobulin

Interleukin-6 (IL-6) is a multifunctional cytokine postulated to play a central role as a growth factor for multiple myeloma (MM). We evaluated the spontaneous secretion of IL-6 in supernatants of Ficoll-Hypaque-- enriched bone marrow (BM) cultures from 35 patients with MM. The levels of IL-6 were correlated with biological and clinical characteristics of the disease. High levels of IL-6 production defined a subgroup of patients with low tumor burden as determined by lower serum beta 2- microglobulin (B2M) (P = .02) and lower percentage of myeloma cells infiltrating the bone marrow (P = .003), higher synthetic rates of monoclonal protein (P = .006), and low proliferative compartments as measured by the percentage of Ki-67--positive myeloma cells. Patients with high proliferative fractions (Ki-67--positive myeloma cells > 20%) had significantly lower levels of IL-6 when compared with patients with low proliferative fractions (P = .005). Our findings do not support IL- 6 as a major growth factor for MM, but demonstrate an association of high levels of IL-6 secretion with low tumor cell burden and low proliferative fraction.

scholarly journals Restricted Immunoglobulin Joining Chain (IgJ) Protein Expression in B Lymphoblastic Leukemia (B-ALL) Based on B-ALL Subtype

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 7-7
Author(s):  
Catherine K Gestrich ◽  
Kwadwo Asare Oduro
Keyword(s):  
Gene Expression ◽  
Bone Marrow ◽  
Protein Expression ◽  
Hodgkin Lymphoma ◽  
Predictive Value ◽  
Lymphoblastic Leukemia ◽  
B Cell Lymphoma ◽  
Normal Bone Marrow ◽  
Neoplastic Cells ◽  
Normal Bone

Background Philadelphia-like (Ph-like) B Lymphoblastic Leukemia (B-ALL) is a high-risk subtype of B-ALL that lacks the BCR-ABL1 fusion but has a gene expression profile similar to Philadelphia positive (Ph+) B-ALL. Gene expression profiling has previously identified Immunoglobulin Joining chain (IgJ) overexpression at the mRNA level in Ph-like B-ALL. This is surprising since IgJ is normally expressed by mature or maturing B cells. The normal function of IgJ protein is to concatenate monomers of immunoglobulin IgM or IgA into the mature pentameric and dimeric forms of these molecules respectively. IgJ also plays a crucial role in transport of IgA protein across mucosal epithelium to facilitate mucosal humoral immunity. In hematopathology, J chain immunohistochemistry (IHC) has been used to identify neoplastic cells in nodular lymphocyte predominate Hodgkin lymphoma (NLPHL) and can be used in distinguishing this disease from morphologic mimickers. It does not have known diagnostic utility outside of this context. Lymphoblasts do not typically express immunoglobulins at the protein level. Therefore, we sought to determine the protein expression of IgJ in B-ALL and to determine whether IgJ immunohistochemistry may be employed in identifying particular subtypes of B-ALL. Methods We selected a total of 46 B-ALL cases diagnosed from a bone marrow sample at our institution from 2016-2019 with adequate diagnostic material for IHC. This included 5 cases of Ph-like B-ALL, all with a CRLF2 rearrangement and overexpression, 7 de novo Ph+ B-ALL and 34 cases representing the other most commonly recognized WHO subtypes of B-ALL, determined based on cytogenetic studies performed at the time of diagnosis. No cases of B-ALL with t(5;14) and B-ALL with iAMP21 was represented. Our cohort included 23 pediatric cases and 24 adult cases and the patients ranged from 1 to 82 years old at the time of initial diagnosis. A total of 8 normal bone marrow cases (negative staging bone marrow biopsies for diffuse large B cell lymphoma, neuroblastoma or classic Hodgkin lymphoma) were used as controls. IgJ IHC was performed on B-plus fixed paraffin embedded bone marrow biopsy specimens using a commercially available and validated anti-IgJ monoclonal antibody (clone OTI3B3). Staining of bone marrow samples was performed at 2 different dilutions; tonsil secondary follicles and neoplastic cells from NLPHL were used as technical controls. Cellular staining in the lymphoblasts was scored in a blinded manner by a board certified hematopathologist and a pathologist in training as diffusely positive, partially positive, or negative. Results Cellular staining was distinguishable from background staining due to circulating immunoglobulins and there was almost perfect inter-observer concordance in identifying positive and negative cases (agreement of 98%, kappa test 0.94). All normal bone marrow controls cases were negative for IgJ cellular staining. A total of 11/46 (23%) B-ALL cases demonstrated partial or diffuse cellular staining for IgJ in the lymphoblasts. This included 4/5 (80%) Ph-like cases, 5/7 (71%) Ph+ cases, 1/3 MLL rearranged cases and 1/6 ETV6-RUNX1. All TCF3-PBX1 (0/4), hyperdiploid B-ALL (0/10), hypodiploid B-ALL (0/2), and B-ALL, NOS cases (0/9) were negative for IgJ. Diffuse IgJ staining was restricted to Ph-like (2/4) or Ph+ (2/5) B-ALL subtypes; the positive MLL rearranged and ETV6-RUNX1 B-ALL cases only showed weak partial staining. IgJ protein was significantly expressed in Ph+/Ph-like B-ALL (p&lt;0.0001) and in our cohort, detected these cases with a 75% sensitivity, 95% specificity, 82% positive predictive value and 92% negative predictive value. Conclusion We conclude that IgJ protein expression occurs in a subset of B-ALL, predominantly restricted to Ph+ and Ph-like cases. Although, these findings will need to be validated in larger studies, our results suggest that IgJ IHC, in concert with routine standard cytogenetics studies may be a rapid and cost effective method in identifying Ph-like B-ALL. Disclosures No relevant conflicts of interest to declare.

The Parthenolide Derivative LC-1 Is An Effective Single Agent and Is Highly Synergistic with Existing Therapies in Multiple Myeloma.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1708-1708
Author(s):  
Elisabeth J Walsby ◽  
Saman Hewamana ◽  
Alan Burnett ◽  
Steven Knapper ◽  
Chris Fegan ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Cell Lines ◽  
Nuclear Localization ◽  
Plasma Cells ◽  
Bone Marrow Cells ◽  
Binding Capacity ◽  
Single Agent ◽  
Normal Bone Marrow ◽  
Normal Bone

Abstract Multiple myeloma (MM) remains incurable with conventional therapeutic agents and has a median survival of only 3–5 years. Therefore, there is clearly a need for novel treatment strategies that can change the natural pathology of this condition. The nuclear factor κB (NF-κB) family of transcription factors is constitutively activated in MM cell lines and the majority of MM patients. Since NF-κB has known oncogenic activity in a number of human malignancies, targeted inhibition of this family of proteins may be useful in the treatment of MM. We and others have recently shown that the parthenolide derivative LC-1 has activity in acute myeloid leukaemia (AML) and chronic lymphocytic leukaemia (CLL) cells. Unusually, it induces apoptosis via the activation of both the intrinsic and extrinsic pathways and apoptosis is preceded by marked inhibition of NF-κB. Importantly, LC-1 is more potent against primary AML blasts and CLL lymphocytes than normal bone marrow progenitors and normal B-cells and T-cells. In this study we set out to evaluate LC-1 in MM cell lines and plasma cells derived from MM patients. LC-1 was cytotoxic to MM cell lines H929, U266 and JJN3 and induced apoptosis in a dosedependent manner resulting in an overall LD50 of 3.6mM (±1.8) after 48 hours in culture. Primary myeloma plasma cells, identified by CD38 and CD138 positivity, had a mean LD50 for LC-1 of 5.4mM (±1.6) after 48 hours of in vitro culture. Normal bone marrow cells were significantly less sensitive to the effects of LC-1 under the same conditions (P = 0.0007). Treatment of MM cell lines with LC-1 resulted in a decrease in the nuclear localization of NF-κB, as evidenced by a dose-dependent decrease in the DNA binding capacity of the NF-κB subunit RelA after 4 hours of treatment. To demonstrate whether synergy exists between LC-1 and existing MM therapies, the H929 cell line was treated for 48 hours with LC-1 and doxorubicin (32:1), melphalan (1:1) or bortezimib (1:500) and the combination indices (CI) calculated using the median effect method. A combination index of less than 1 denotes synergy. LC-1 did not show synergy with doxorubicin (CI &gt;1) but was synergistic with melphalan and bortezimib (CI values of 0.53 and 0.59 respectively). Taken together our data clearly demonstrate that LC-1 has activity in MM cell lines and primary MM cells. Its ability to inhibit the nuclear localization of NF-κB is important to its cytotoxic effects. Furthermore, it may also provide an explanation for the synergy demonstrated with melphalan and bortezimib. These results provide a rationale for exploring the potential of LC-1 in clinical studies.

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