Expression and Clinical Significance of Notch1 On the Membrane of Bone Marrow CD38+CD138+Plasma Cells in the Patients with Multiple Myeloma

Abstract Abstract 4981 Objective: To investigate the expression of bone marrow CD38+CD138+, CD38+CD138-plasma cells and the expression of Notch1 on the membrane of them in the patients with multiple myeloma(MM), and explore the importance of Notch signaling pathway in the formation and progression of MM further. Methods: Thirty-three MM patients and 15 healthy controls were enrolled in this study. The expression of bone marrow CD38+CD138+, CD38+CD138-plasma cells and the expression of Notch1 on the membrane of them were analyzed by flow cytometry. The expression of Notch1 mRNA of bone marrow mononuclear cells were analyzed by RT-PCR. Results: The ratio of CD38+CD138+ plasma cells from 24 newly diagnosed MM patients was (51. 50%±12. 48%) which was significantly higher than CD38+CD138- plasma cells of MM patients (42. 88%±11. 41%)(P=0. 016)and controls 20. 13%±5. 8(P=0. 000). The expression of CD38+CD138+ plasma cells from 24 newly diagnosed MM patients was correlated to the level of malignant plasma cells in there bone marrow(r=0. 546, p=0. 006), serum level of lactate dehydrogenase(LDH)(r=0. 567, p=0. 004), and β2-MG(r=0. 431, p=0. 035). The ratio of Notch1 on the membrane of CD38+CD138+ plasma cells of MM patients was (60. 21%±25. 06%) which was significantly higher than those of CD38+CD138- plasma cells of MM patients 39. 84%±18. 94%(P=0. 000)and controls (38. 34%±19. 39%)(P=0. 004). There was no statistical difference between the two latter groups(P>0. 05). The expression of Notch1 on CD38+CD138+ plasma cells from 24 newly diagnosed MM patients was correlated to the level of malignant plasma cells in there brone marrow(r=0. 914, p=0. 000), serum level of lactate dehydrogenase(LDH) (r=0. 604, p=0. 002), and β2-MG(r=0. 455, p=0. 026). The ratio of Notch1 on the membrane of CD38+CD138+ plasma cells of MM patients who had renal dysfunction was correlated to their abnormal serum creatinine levels. The expression of Notch1 on CD38+CD138+ plasma cells from 17 MM patients who received VD chemotherapy was correlated to the ratio of plasma cell reduction after the first VD chemotherapy(r=0. 842, p=0. 000). The expression of Notch1 mRNA of bone marrow mononuclear from 10 MM patients was (0. 8252±0. 4079) which was significantly higher than those of controls (0. 3759±0. 0813)(p=0. 032). Conclusion: Notch1 over expressed on CD38+CD138+ plasma cells with relation to the effects of early VD therapy and long term outcome of MM. Disclosures: No relevant conflicts of interest to declare.

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The Frequency of Genetic Anomalies According to Clonal Plasma Cells' Phenotype in Patients with Newly Diagnosed (ND) Multiple Myeloma (MM)

Blood ◽

10.1182/blood.v126.23.5316.5316 ◽

2015 ◽

Vol 126 (23) ◽

pp. 5316-5316

Author(s):

Andrei Garifullin ◽

Irina Martynkevich ◽

Sergei Voloshin ◽

Alexei Kuvshinov ◽

Ludmila Martynenko ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Plasma Cells ◽

Conflicts Of Interest ◽

Risk Group ◽

Risk Groups ◽

Fish Analysis ◽

Newly Diagnosed ◽

Definition Of ◽

Standard Risk

Abstract Background. Genetic anomalies (GA) are primary link of pathogenesis in MM. GA lead to formation of clonal plasma cells, which has different phenotype. Aim. To estimate the incidence of GA and their correlation with clonal plasma cells' phenotype in patients with ND MM. Methods. We analysed 22 patients with ND MM (median age 57 years, range 38-80; male/female - 1:1.75). Cytogenetic analysis was performed on bone marrow samples using standard GTG-method. Metaphase FISH analysis was performed according to the manufacturer's protocol using DNA probes: LSI 13(RB1)13q14, IGH/CCND1, IGH/FGFR3, LSI TP53 (17q13.1). 8-color immunophenotypic by flow cytometry using antibody to CD45, CD38, CD138, CD56, CD19, CD20, CD27 and CD117 antigenes. Results. Translocation t(11;14) was detected in 3/14 (21.4%) patients, del(13q) - 2/14 (14.3%), t(11;14) - 3/14 (21.4%), hypodyploidy - 1/20 (5%), del(17р) - 0% patients. Clonal plasma cells' phenotype CD38+CD138+CD45- was detected in 100%. Expression CD56+ was revealed in 11/22 (50%) patients, CD19+ in 9/22 (40.9%), CD117+ in 5/22 (22.7%), CD20+ in 1/22 (4.5%), CD27+ in 1/22 (4.5%). The frequency of GA didn't depend on clonal plasma cells' phenotype and was 27.3%(3/11) in CD56+ phenotype, 23.8%(5/21) - CD20-, 23.8%(5/21) - CD27-, 23.5%(4/17) - CD117-, 23%(3/13) - CD19-, 22.2%(2/9) - CD19+, 20%(1/5) - CD117+, 18.2%(2/11) - CD56-, 0%(0/1) - CD20+, 0%(0/1) - in CD27+ phenotype. Patients of standard risk group according to mSMART 2.0 with GA had CD19-negative plasma cells' phenotype vs. CD19-positive phenotype in patients of intermediate and high-risk groups (p<0.05). 3-years overall survival in standard risk group with CD19- phenotype was 92,3%, CD19+ - 77,7% (p>0.05). Conclusion . Identification of GA, which has adverse forecast, correlates with CD19+ plasma cells phenotype. The combined definition of plasma cells phenotype and GA can improve the system of risk stratification in MM. Disclosures No relevant conflicts of interest to declare.

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Multiple Myeloma Course with Renal Insufficiency in Young Patient: Case Report

Blood ◽

10.1182/blood-2020-143302 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 26-26

Author(s):

Hugo Henrique de Freitas Ferreira ◽

Alessandra Suelen Jardim Silva ◽

Lenilton Silva DA Silva Júnior ◽

Gustavo Henrique de Medeiros Oliveira ◽

Maria das Graças Pereira Araujo ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Young Patient ◽

Mononuclear Cells ◽

Plasma Cells ◽

Serum Proteins ◽

Conflicts Of Interest ◽

Young Patients ◽

Treatment Strategies ◽

Malignant Neoplasm

Introduction: Multiple myeloma (MM) is a malignant neoplasm characterized by the clonal proliferation of abnormal plasma cells in the bone marrow (OM). The average age of patients diagnosed with MM is approximately 70 years, being relatively uncommon in younger individuals. Objective: To report a case of a young patient with multiple myeloma. Case Description: A 42-year-old male patient presented with continuous and progressive low back pain for 3 months, associated with adynamia, weight loss (10 kg), episodes of constipation and bleeding in the oral cavity in this period. Examinations at the first appointment revealed moderate anemia (Hb 7.4 g / dL), leukocytosis, thrombocytopenia, hypercalcemia, and altered renal function (Cr 5.9 and Ur 178), chest tomography indicating vertebral fracture in T6, T11, L2 and L4. Referred for specialized follow-up, he performed electrophoresis of serum proteins with the presence of a monoclonal peak in the gamma globulin fraction. The immunofixation test confirmed monoclonality for IgA isotype and Kappa light chain (IgA / Kappa). The myelogram showed plasmacytosis of more than 50% of mononuclear cells in the bone marrow. He developed renal failure (with dosage of creatinine of 10.1 mg/ dL. and urea of 208 mg/dL) and hypercalcemia requiring dialysis therapy on the third day of hospitalization, having undergone chemotherapy with Bortezomib, cyclophosphamide and dexamethasone. During this period, infection by the multisensitive S. aureus in catheter occurred and, despite being treated with specific antibiotic therapy, it evolved with clinical worsening and hemodynamic instability and was referred to the Intensive Care Unit, going to death after 2 days. Conclusion: Young patients with MM may study with more aggressive characteristics. Despite the use of new therapeutic agents, more effective treatment strategies need to be studied more for patients in this age group. Disclosures No relevant conflicts of interest to declare.

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A Fully Automated Method for Isolating Highly Enriched Population of Multiple Myeloma Plasma Cell From Whole Bone Marrow for Enhanced Sensitivity in Downstream Assays

Blood ◽

10.1182/blood.v118.21.5064.5064 ◽

2011 ◽

Vol 118 (21) ◽

pp. 5064-5064

Author(s):

Hossein Mossafa ◽

Sabine Defasque ◽

Hamid Belaouni ◽

Adrian Arechiga

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Dna Extraction ◽

Mononuclear Cells ◽

Plasma Cells ◽

Conflicts Of Interest ◽

Snp Array ◽

Fish Analysis ◽

Automated Method

Abstract Abstract 5064 Introduction, Multiple myeloma (MM) is characterized by a huge clinical heterogeneity despite the homogenous morphologic appearance of malignant plasma cells (PCs). The advent of interphase fluorescence in situ hybridization (FISH) or MicroArrays (MA) allows an increased rate of aberration detection and identification of some recurrent cryptic changes, which have been increasingly implemented as additional diagnostic and prognostic factors. To heighten sensitivity of Single Nucleotide Polymorphism (SNP) arrays, or FISH it is necessary to have a purified population of cells as starting material. Screening must be performed systematically on the purified CD138+ PCs. After testing different systems for cell purification, we encountered some challenges. We didn't obtain enough PCs for FISH and SNP array studies. This was due to excess M-protein accumulating in the blood stream, increasing hyper viscosity and also due to the morphology and size variations of PCs at various stages of differentiation. Additionally, downstream DNA extraction can be a challenge since EDTA found in most buffers is an inhibitor for chemical PCR reaction for some MA chips. Given the challenges, CERBA laboratory and Miltenyi Biotec GmbH have developed a fully automated process (FAP) for purification for CD138+ PCs. In a study of 100 BM patient samples, we compared the specificity, efficiency, performance, purity, ease of use, technologists' time and the quality of DNA after CD138+ PCs purification. Two methods were compared. In the first method, cells were directly purified from bone marrow samples by FAP using Automated Magnetic Cell Sorter (AMCS). In the second method, mononuclear cells from fresh whole bone marrow (WBM) were enriched by Ficoll, followed by cell selection procedure with anti-CD138+ MicroBeads using the AutoMACS®. Before separation and following the separation, the percentage of PCs was determined by Flow cytometry (FC) on WBM by multiparameter FC (MFC) for CD138/CD38 expression. Additionally, DNA quality on separated cells was assessed by Nanodrop. A fraction of the CD138+ PCs were used after hypotonic shock and Carnoy fixation, applied to glass slides for FISH application and another fraction for DNA extraction for MA (SNP.6 Affymetrix®) FISH was performed with the recommended unbalanced alterations & reciprocal rearrangements: del(13) (q14)(D13S25), del(17)(p13)(TP53),+3(D3Z), +9(D9Z1), +15(D15Z14), t(4;14)(p16;q32)/IGH-FGFR3. Results, the specificity and purity were the same for both process but the efficiency and performance were considerably better for FAP than mononuclear cells enriched by Ficoll (MCEFicoll) process. With FAP, in 95% of the MM cases we obtained enough PCs for performance of the recommended panel of FISH and for 50% of them we could extract DNA for SNP array. For the MCEFicoll, we observed inferior performance, with very few plasma cells after isolation. Having enough PSc for only 65% of the cases and we could only extract DNA for 28% of them. The quality of DNA was the same for both process and the technologists' time was longer by 30' /patient for MCEFicoll process than for FAP. Currently in CERBA lab, we realize more than 20 plasma cells isolation per week for patients with MM and from October 2007 to July 2011 we have separated more than 5.000 specimens using CD138 Whole Blood MicroBeads (CD 138 WBMB) from Miltenyi Biotec, in combination with the AMCS. This has allowed isolation directly from WBM without any sample preparation required, such as density gradient centrifugation (ficoll) or erythrocyte lysis. The detection rate of chromosomal abnormalities and the number of abnormalities per case in MM and PCs dyscrasia significantly improves when there are enough CD138+PCs for analysis. Conclusion, in this report we describe the benefits of fully automated isolations of CD138+ cells from WBM. We have developed an SOP for an automated reliable and standardized method which allows the processing of multiple samples in a single day, while maintaining sample integrity and increasing sensitivity of FISH analysis and WG arrays for a diagnosis lab. Disclosures: No relevant conflicts of interest to declare.

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Prognostic Implications of Metabolic Total Volume and Total Lesion Glycolysis Assessed By PET/CT Combined with High Levels of Bone Marrow Plasma Cells Percentages As a Potential Risk Model in Patients with Newly Diagnosed Multiple Myeloma

Blood ◽

10.1182/blood-2019-126784 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 3193-3193

Author(s):

Toshiki Terao ◽

Yoichi Machida ◽

Takafumi Tsushima ◽

Akihiro Kitadate ◽

Daisuke Miura ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Risk Stratification ◽

Tumor Volume ◽

Mononuclear Cells ◽

Total Lesion Glycolysis ◽

Whole Body ◽

Prognostic Impact ◽

Newly Diagnosed ◽

Pet Ct

Introduction: Multiple myeloma (MM) is a heterogeneous malignant plasma cell (PC) disorder and the survival ranges from several months to > 10-years. Several risk stratification systems such as the Revised International Staging System (R-ISS) have been developed. PET/CT allows the direct assessment of metabolic tumor burden in various malignancies. Therefore, metabolic tumor volume (MTV) and total lesion glycolysis (TLG), which are volumetric parameters applicable to PET/CT, are emerging tools for MM prognostication. This study was aimed to determine the value of MTV and TLG using PET/CT in the prognostication and in combination with various hematologic parameters such as bone marrow PC (BMPC) percentages and circulating tumorous PCs (CPCs) to identify the patients with high-risk features. Methods: A total of 196 consecutive patients with newly diagnosed MM (NDMM) who underwent baseline whole-body PET/CT between January 2009 and June 2019 at Kameda Medical Center, Kamogawa-shi, Japan, were retrospectively analyzed. PET/CT was performed using dedicated PET/CT scanners (Discovery ST Elite Performance; GE Healthcare, Milwaukee, USA). The standard uptake value (SUV) was normalized according to the injected dose and lean body mass. The baseline SUVmax of all lesions was recorded, and the highest value was considered as the SUVmax of the patient. MTV was defined as the myeloma lesions volume visualized on PET/CT scans with SUV greater than or equal to the fixed absolute threshold of SUV = 2.5. TLG was calculated as the sum of the product of average SUV (SUVmean) and MTV of all lesions. Computer‐aided analysis of PET-CT images for MTV and TLG calculations was performed using an open-source software application of Metavol (Hokkaido University, Sapporo, Japan). The CPCs were measured using an 8-color flowcytometry and reported as the percentage per total mononuclear cells using the monoclonal antibodies of CD19, 38, 45, 56, 117, 200, κ, λ, and CD138. The BMPC was calculated by counting the percentages of CD138-stained PCs among the all nucleated cells on bone marrow biopsy samples. Eleven patients (13.8%) were excluded because the MTV data could not be retrieved. Ultimately, 185 patients were included in our analysis. Written informed consent was obtained from all patients. Results: Among the 185 patients, 28 patients (15.1%) were negative for avid lesion on PET/CT. Whole-body MTV and TLG ranged from 0 to 2440.7 mL, with a median of 34.2 mL and from 0 to 12582.4 g, with a median of 97.0 g, respectively. The best cut-off values of MTV and TLG that discriminate the survival using a receiver-operating-characteristic curve analysis were 56.4 mL and 166.4 g, respectively. The overall survival (OS) and progression-free survival (PFS) of patients with a lower cut-off value of MTV (≤56.4 mL) had better survival with not reached (NR) and 37.3 months as compared to those with a higher cut-off value (>56.4 mL) that reached 52.9 and 23.8 months, respectively (p=0.003 and 0.019). Similarly, the OS and PFS of patients with a lower cut-off value of TLG (≤166.4 g) showed better survivals with NR and 37.3 months as compared to those with a higher cut-off value (>166.4 g) that reached 54.3 and 28.8 months, respectively (p=0.0047 and 0.012). Next, we explored the prognostic impact of the clinical variables including MTV or TLG, CPCs, and BMPC. High levels of CPCs and BMPCs levels were defined as ≥0.018% of the total mononuclear cells and BMPCs of ≥57%, respectively. Univariate analysis showed that age≥70, serum creatinine≥2.0 mg/dL, R-ISS stage 3, higher cut-off value of MTV, and higher cut-off value of TLG were the associated with shorter OS. To measure the tumor volume with accuracy, we combined BMPC or CPCs and MTV or TLG. On multivariate analysis, age≥70 and the combination of higher cut-off value of MTV or TLG and high level of BMPC percentage were significantly associated with shorter OS [Hazard Ratio (HR) 2.12, p=0.038, HR 2.66, p=0.027 and HR 2.57, p=0.029, respectively] and PFS (Not assessed, HR 2.52, p=0.018 and HR 2.7, p= 0.011, respectively) (Figure 1). Conclusion: Our findings demonstrated that MTV and TLG calculated from pretreatment PET/CT were useful for risk stratification in patients with NDMM when combined with BMPC. The prognostic performance of the combined high-burden of TLG or MTV and high levels of BMPC were independent of the established risk factors. Disclosures Matsue: Novartis Pharma K.K: Honoraria; Janssen Pharmaceutical K.K.: Honoraria; Celgene: Honoraria; Takeda Pharmaceutical Company Limited: Honoraria; Ono Pharmaceutical: Honoraria.

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CS1: A Potential New Therapeutic Target for the Treatment of Multiple Myeloma.

Blood ◽

10.1182/blood.v108.11.3457.3457 ◽

2006 ◽

Vol 108 (11) ◽

pp. 3457-3457 ◽

Cited By ~ 3

Author(s):

Eric D. Hsi ◽

Roxanne Steinle ◽

Balaji Balasa ◽

Aparna Draksharapu ◽

Benny Shum ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Flow Cytometry ◽

B Cell ◽

Mononuclear Cells ◽

Plasma Cells ◽

Target Cells ◽

Dependent Manner ◽

Myeloma Cells ◽

Effector Cells

Abstract Background: To identify genes upregulated in human memory B and plasma cells, naïve B cell cDNA was subtracted from plasma cell and memory B cell cDNA. One gene that was highly expressed in plasma cells encodes CS1 (CD2 subset 1, CRACC, SLAMF7), a cell surface glycoprotein of the CD2 family. CS1 was originally identified as a natural killer (NK) cell marker. Monoclonal antibodies (mAbs) specific for CS1 were used to validate CS1 as a potential target for the treatment of multiple myeloma (MM). Methods: Anti-CS1 mAbs were generated by immunizing mice with a protein comprising of the extracellular domain of CS1. Two clones, MuLuc63 and MuLuc90, were selected to characterize CS1 protein expression in normal and diseased tissues and blood. Fresh frozen tissue analysis was performed by immunohistochemistry (IHC). Blood and bone marrow analysis was performed using flow cytometry with directly conjugated antibodies. HuLuc63, a novel humanized anti-CS1 mAb (derived from MuLuc63) was used for functional characterization in non-isotopic LDH-based antibody-dependent cellular cytotoxicity (ADCC) assays. Results: IHC analysis showed that anti-CS1 staining occurred only on mononuclear cells within tissues. The majority of the mononuclear cells were identified as tissue plasma cells by co-staining with anti-CD138 antibodies. No anti-CS1 staining was detected on the epithelia, smooth muscle cells or vessels of any normal tissues tested. Strong anti-CS1 staining was also observed on myeloma cells in 9 of 9 plasmacytomas tested. Flow cytometry analysis of whole blood from both normal healthy donors and MM patients showed specific anti-CS1 staining in a subset of leukocytes, consisting primarily of CD3−CD(16+56)+ NK cells, CD3+CD(16+56)+ NKT cells, and CD3+CD8+ T cells. Flow cytometry of MM bone marrow showed a similar leukocyte subset staining pattern, except that strong staining was also observed on the majority of CD138+CD45−/dim to + myeloma cells. No anti-CS1 binding was detected to hematopoietic CD34+CD45+ stem cells. To test if antibodies towards CS1 may have anti-tumor cell activity in vitro, ADCC studies using effector cells (peripheral blood mononuclear cells) from 23 MM patients and L363 MM target cells were performed. The results showed that HuLuc63, a humanized form of MuLuc63, induced significant ADCC in a dose dependent manner. Conclusions: Our study identifies CS1 as an antigen that is uniformly expressed on normal and neoplastic plasma cells at high levels. The novel humanized anti-CS1 mAb, HuLuc63, exhibits significant ADCC using MM patient effector cells. These results demonstrate that HuLuc63 could be a potential new treatment for multiple myeloma. HuLuc63 will be entering a phase I clinical study for multiple myeloma.

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TIMP-1 and TIMP-2 Levels Are Directly Correlated with Neoangiogenesis and Are Prognostic Factors in Multiple Myeloma Patients.

Blood ◽

10.1182/blood.v114.22.4866.4866 ◽

2009 ◽

Vol 114 (22) ◽

pp. 4866-4866

Author(s):

Luciana Correa Oliveira de Oliveira ◽

Juliana Alves Uzuelli ◽

Ana Paula Alencar de Lima Lange ◽

Barbara Amelia Aparecida Santana-Lemos ◽

Marcia Sueli Baggio ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Overall Survival ◽

Prognostic Factors ◽

Bone Disease ◽

Cox Regression ◽

Conflicts Of Interest ◽

Newly Diagnosed ◽

Significant Difference ◽

The Impact

Abstract Abstract 4866 Background Multiple myeloma (MM) is an incurable malignant disease, characterized by increased angiogenesis in the bone marrow (BM) microenvironment and aberrant BM metabolism. Matrix metalloproteinases (MMP) are a family of zinc-dependent endopeptidases implicated in tumour progression, invasion, metastasis and angiogenesis, via proteolytic degradation of extracellular matrix. MMPs are inhibited by tissue inhibitors of metalloproteinase (TIMP). Although recent studies have implicated MMP 9 in MM bone disease, little is known about the role of the TIMPs. Objectives a) to compare levels of sRANKL, OPG, MMP-2, MMP-9, TIMP-1, TIMP-2, VEGF, bFGF, microvessel density (MVD) between newly diagnosed MM patients and healthy controls; b) to determine the association of these molecules with disease progression, bone disease and neoangiogenesis and c) to evaluate the impact of these variables on survival. Patients and Methods As of July 2009 38 newly diagnosed and untreated multiple myeloma patients were enrolled in the study. The median age was 61years-old (range 39-91) with 24 (63%) males. Patients were diagnosed and categorized according The International Myeloma Working Group criteria and ISS, respectively. Bone involvement was graded according to standard X-ray: patients with no lesions, or with one/ two bones involved or diffuse osteoporosis were classified as low score, whereas patients with lesions in more than two bones or presence of bone fracture were classified as high score. MMP-2 and MMP-9 were determined by PAGE gelatin zymography from plasma as previously described. MMP-9, TIMP-1 and TIMP-2, OPG and sRANKL concentrations were measured by ELISA. The levels of VEGF, bFGF were obtained using cytometric bead array. Ten healthy volunteers were used as controls. Bone marrow MVD measured in hotspots was evaluated in 26 out of 38 patients at diagnosis and 15 patients with Hodgkin Lymphoma stage IA and IIA (used as controls) by staining immunohistochemically for CD34. Comparisons among groups were analyzed by ANOVA and the correlation by the Spearman's correlation coefficient. Cox regression were performed for overall survival (OS) analysis. Results Patients with MM had elevated TIMP-1, TIMP-2 and OPG values compared with controls. No significant difference was found between plasma sRANKL, pro-MMP2, pro-MMP9 and MMP-9 levels. We found that plasma TIMP-1 levels correlated positively with bFGF, VEGF, MVD, beta-2 microglobulin (B2M) and OPG (r: 0.514, p=0,001, r: 0.350, p=0,031; r: 0.610, p<0.0001; r: 0.760, p<0.0001 and r: 0.701, p<0.0001, respectively) and TIMP-2 levels with bFGF, DMV, B2M and OPG (r: 0.512, p=0.002; r: 0.595, p<0.0001; r: 0.587, p<0.0001 and r: 0.552, p<0.0001, respectively). TIMP-1 and TIMP-2 levels correlated with the ISS stage (p<0.0001, p=0.006, respectively). The only variables that correlated with clinical bone disease staging were hemoglobin, B2M and albumin levels, whereas TIMP-1, TIMP-2, bFGF, VEGF and OPG correlated with DMV. On the univariate analyses, age, gender, proMMP2, TIMP-1, TIMP-2, creatinine, B2M and MVD were significantly associated with overall survival. In Cox regression model, TIMP-1, TIMP-2 and B2M levels remained to be significantly associated with OS. In conclusion, our results suggest that TIMP-1 and TIMP-2 levels are strongly associated with neoangiogenesis and are independent prognostic factors in MM. Disclosures No relevant conflicts of interest to declare.

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Increased Expression of Cyclin-D1 on Trephine Bone Marrow Biopsies Independently Predicts for Shorter Overall Survival In Patients with Multiple Myeloma Treated with Novel Agents

Blood ◽

10.1182/blood.v116.21.2965.2965 ◽

2010 ◽

Vol 116 (21) ◽

pp. 2965-2965

Author(s):

Evangelos Terpos ◽

Maria Roussou ◽

Anna Tasidou ◽

Magdalini Migkou ◽

Maria Gavriatopoulou ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Overall Survival ◽

Cyclin D1 ◽

Plasma Cells ◽

Univariate Analysis ◽

Novel Agents ◽

Positive Staining ◽

Newly Diagnosed ◽

Immunohistochemical Expression

Abstract Abstract 2965 The cyclin-D1 proto-oncogene is an important cell regulator of G1 to S phase progression. The overexpression of cyclin-D1 has been linked to the development and progression of several malignancies. The aim of our study was to evaluate the impact of the immunohistochemical expression of cyclin-D1on the plasma cells of trephine biopsies on survival of newly-diagnosed patients with multiple myeloma (MM) who were treated with novel agents. We evaluated formalin-fixed, paraffin-embedded, bone marrow sections of 130 consecutive patients with newly-diagnosed MM (67M/63F; median age 68 years) before any kind of therapy administration. One hundred and fifteen patients had symptomatic disease that required therapy: 29 (25%) received bortezomib-based regimens and 31 (26%) received thalidomide-based regimens as first line therapy, while all patients received regimens containing bortezomib or an IMiD at some point during the course of their disease. Immunohistochemistry was performed in all trephine biopsies using monoclonal antibodies against cyclin-D1 (Cell Marque Corp., Rocklin, CA, USA), but also against CD56 (Cell Marque Corp., Rocklin, CA, USA), CD27 (Novocastra, Newcastle upon Tyne, UK), CD117 and MUM-1 (DAKO A/S, Glostrup, Denmark), as recommended by the manufacturers. A case was considered positive if there was unequivocal positive staining of at least 20% of the plasma cells for cyclin-D1, CD56 and MUM-1 and a positive staining of at least 10% of the plasma cells for CD117 and CD27. Among patients with symptomatic myeloma (N=115), positive staining for cyclin-D1 was found in 35 (30%) patients, for CD56 in 45 (39%), for CD117 in 94 (81%) and for CD27 in 72 (62%) patients. In patients with asymptomatic myeloma, positive staining for Cyclin-D1 was found only in 1 (7%) patient, for CD56 in 9 (64%), and for CD117 in 6 (43%) (p<0.01 for all comparisons compared to symptomatic patients). There were significant positive correlations between positivity for CD27 and CD56 (p<0.001), between positivity for cyclin-D1 and CD117 (p=0.045) and a negative correlation between positivity for CD117 and CD56 (p=0.001). We also observed significant correlations between CD56 positivity and ISS-1 or ISS-2 (p=0.01) and between CD117 positivity and ISS-3 disease (p=0.002). The median overall survival (OS) for patients with symptomatic MM was 57 months (range 22–120 months). In the univariate analysis, positivity for cyclin-D1 (41 vs. 62 months, p=0.03) and for CD117 (50 vs. 75 months p=0.018) were associated with inferior survival, while positivity for CD56 (47 vs. 62 months, p=0.286), MUM-1 (52.7 vs. 63.8 months, p=0.528) and CD27 (57 vs. 50 months, p=0.445) were not. Other factors associated with inferior OS, in the univariate analysis, included ISS-3 (median OS 37 months, vs. 57 months for ISS-2 and 73 months for ISS-1, p=0.005), Hb <10 g/dl (56 vs. 73 months, p=0.044), corrected serum calcium >11.5 g/dl (29 vs. 62 months, p=0.02), serum LDH above upper normal limit (31 vs. 61 months, p=0.05), serum creatinine >2 mg/dl (26 vs. 64 months, p=0.007), low platelet counts (<100,000/ml) (22 vs. 62 months, p=0.031) and age >65 years (45 months vs. not reached for younger patients, p=0.002). In the multivariate analysis, positivity for cyclin-D1 (HR: 2.6; p=0.001), ISS stage (HR: 1.8; p=0.001) and age >65 (HR 2.7, p=0.003) were independently associated with inferior survival. Immunohistochemistry for cyclin-D1 identified subgroups of patients in ISS-2 and in ISS-3 who had extremely poor outcome. Patients with cyclin-D1 positivity had a median survival of 22 months in ISS-2 (vs. 64 months for the rest of ISS-2 patients, p=0.01) and of 13 months in ISS-3 (vs. 47 months for the rest of ISS-3, p=0.012). Our findings underline that the immunohistochemical expression of cyclin-D1 in the bone marrow trephine biopsies has independent prognostic value in MM patients, even in the era of novel agents. This marker can easily be assessed in patients who undergo a trephine biopsy as part of their initial evaluation and offers significant prognostic information. Furthermore, novel agents targeting cyclin-D1 may be of therapeutic value in MM. Disclosures: No relevant conflicts of interest to declare.

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Bortezomib Induction Followed by ASCT in Patients with Multiple Myeloma: Achievement of Response After Induction and Achieving CR Post-ASCT Are Both Important Prognostic Markers

Blood ◽

10.1182/blood.v118.21.1866.1866 ◽

2011 ◽

Vol 118 (21) ◽

pp. 1866-1866

Author(s):

Min Kyoung Kim ◽

Chang-Ki Min ◽

Myung Soo Hyun ◽

Kihyun Kim ◽

Sung-Soo Yoon ◽

...

Keyword(s):

Multiple Myeloma ◽

Maintenance Therapy ◽

Conflicts Of Interest ◽

Induction Treatment ◽

Term Outcome ◽

Long Term Outcome ◽

Free Survival

Abstract Abstract 1866 Background: In multiple myeloma (MM), the association between the response to induction before autologous stem cell transplantation (ASCT) and long-term outcome is less clear but the situation may change with the introduction of novel agents. We therefore assessed the clinical relevance of response of bortezomib induction treatment or post-ASCT response for patients who received bortezomib-combined induction chemotherapy followed by ASCT. Methods: We retrospectively assessed 183 MM patients who received bortezomib-containing induction therapy (BTZ-IT) followed by ASCT in 24 institutions throughout Korea between 2003 and 2010. Records of these patients were reviewed using the Korean Myeloma Registry database (www.myeloma.or.kr). Each institution was requested to reconfirm the data using additional case report forms. Patients who had overt MM based on International Myeloma Working Group diagnostic criteria were selected. Results: One-hundred seventy eight patients were eligible. Their median age was 56 years (range, 28–69 years) and 96 (53.9%) were male. Forty nine (27.5%) received bortezomib as front-line therapy and 129 (72.5%) as second-line treatment. All patients underwent ASCT and 22 (12.4%) were treated with tandem ASCT. Ninety-seven (54.5%) patients were treated with maintenance therapy after ASCT. After BTZ-IT, the response rates in this selected series of patients were 37.6% CR, 12.4% VGPR, 41.0% PR, 7.3% SD and 1.7% PD (Figure 1A, 1B, 1C); the corresponding post-ASCT rates were 69.2% CR, 14.0% VGPR, 11.0% PR, 2.9% SD and 2.9% PD. At a median follow-up of 46.6 months, the 3-year overall survival (OS) and event-free survival (EFS) rates were 70.0% and 31.9%, respectively. Multivariate analysis showed that factors independently predictive of OS and EFS included achievement of BTZ-IT response °Ã PR (P=0.025 and P=0.014, respectively) and the treatment with maintenance therapy (P=0.048 and P=0.001, respectively). Post-ASCT CR vs. °Â VGPR was also an independent prognostic factor for OS and EFS (P=0.0001 and P=0.002, respectively). Conclusion: At least PR to BTZ-IT and CR after ASCT were predictive of survival. These findings suggest that patients who responded to BTZ-IT may benefit from ASCT due to an enhanced quality of response. Maintenance therapy can also affect patient outcomes. Disclosures: No relevant conflicts of interest to declare.

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Metabolomics Identifies Potential Biomarkers of Multiple Myeloma Development and Progression

Blood ◽

10.1182/blood.v120.21.3985.3985 ◽

2012 ◽

Vol 120 (21) ◽

pp. 3985-3985

Author(s):

Francesca Fontana ◽

Josè Manuel garcia Manteiga ◽

Magda Marcatti ◽

Francesca Lorentino ◽

Giovanni Tonon ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Plasma Cells ◽

Conflicts Of Interest ◽

In Vitro Tests ◽

Incurable Cancer ◽

Disease Evolution ◽

Cell Counts ◽

Bone Marrow Plasma ◽

Active Disease

Abstract Abstract 3985 Multiple myeloma is a malignancy of plasma cells, which grows at multiple foci in the bone marrow, secretes monoclonal immunoglobulins, and typically induces skeletal destruction, hypercalcemia, anemia, and renal failure. Although it remains an incurable cancer, novel therapeutic regimens have improved overall survival in the last decade. Multiple myeloma originates from post germinal center, terminally differentiated B lymphocytes through a multi-step process involving early and late genetic changes. Multiple myeloma is preceded by monoclonal gammopathy of undetermined significance (MGUS), a frequent age-progressive premalignant expansion of bone marrow plasma cells that behave benignly despite the presence of most myeloma-specific genetic abnormalities. Indeed, development and progression of multiple myeloma are believed to rely on vicious interactions with the bone marrow environment, offering a paradigm to investigate the bone-cancer relationship. In particular, bone and stromal cells are known to be diverted by cancer cells through altered cytokine circuitry. The resulting enhanced osteoclastogenesis and neoangiogenesis, and reduced osteoblast differentiation and activity sustain cancer cell survival, proliferation, migration and chemoresistance. Such crucial interactions, however, have only partially been elucidated in their complexity, dynamics and exact role in disease evolution. A better knowledge of this interplay, still elusive, could help identify prognostic markers, pathomechanisms, and therapeutic targets for future validation. Aiming to achieve an unbiased, comprehensive assessment of the extracellular milieu during multiple myeloma genesis and progression, we performed a metabolomic analysis of patient-derived peripheral and bone marrow plasma by ultra high performance liquid and gas chromatography followed by mass spectrometry. By feature transformation-based multivariate analyses, metabolic profiling of both peripheral and bone marrow plasma successfully discriminated active disease from control conditions (health, MGUS or remission). Moreover, both central and peripheral metabolic scores significantly correlated with bone marrow plasma cell counts. Significant changes in the peripheral metabolome were found to be associated with abnormal renal function in the subset of myeloma patients. Noteworthy, however, renal dysfunction-associated features failed to independently predict disease load, while non-overlapping disease vs. control analyses consistently identified a number of metabolites associated with disease. Among these, increased levels of the C3f-derived peptide, HWESASLL, and loss of circulating lysophosphocholines emerged as hallmarks of active disease. In vitro tests on myeloma cell lines and primary patient-derived cells revealed a previously unsuspected direct trophic role exerted by lysophosphocholines on malignant plasma cells. Altogether, our data demonstrate that metabolomics is a powerful approach suitable for studying the complex interactions of multiple myeloma with the bone marrow environment and general metabolism. This novel strategy holds potential to identify unanticipated markers and pathways involved in development and progression of multiple myeloma. Disclosures: No relevant conflicts of interest to declare.

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Multiple Myeloma Associated GI Polyposis

Blood ◽

10.1182/blood.v120.21.5009.5009 ◽

2012 ◽

Vol 120 (21) ◽

pp. 5009-5009

Author(s):

Nassim Nabbout ◽

Mohamad El Hawari ◽

Thomas K. Schulz

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Plasma Cells ◽

Conflicts Of Interest ◽

Case Reports ◽

Bone Marrow Failure ◽

Bone Lesions ◽

Rare Manifestation ◽

History Of ◽

Central Ulceration

Abstract Abstract 5009 Multiple myeloma is a neoplastic proliferation of monoclonal plasma cells that can result in osteolytic bone lesions, hypercalcemia, renal impairment, bone marrow failure, and the production of monoclonal gammopathy. The gastrointestinal tract is rarely involved in myeloma. GI polyposis is a rare manifestation of extra-medullary disease in multiple myeloma. Such cases usually present as gastrointestinal hemorrhage or intestinal obstruction. A 53-year-old African American male recently diagnosed with multiple myeloma presented with three-day history of rectal bleed and fatigue. EGD showed multiple raised, polypoid, rounded lesions with a superficial central ulceration in the stomach. Colonoscopy showed similar lesions in the ascending and transverse areas of the colon that ranged in size from 5 to 16 mm in diameter. Biopsies showed that these polyps were made of plasma cells. A bone marrow biopsy showed diffuse involvement (greater than 90%) of bone marrow with multiple myeloma with anaplastic features. The patient was started on bortezomib at diagnosis, however, he passed away a few weeks later. This type of metastatic disease has been described in isolated case reports in the literature, while solitary GI plasmacytoma has been reported more frequently. In rare cases, multiple myeloma can involve the GI tract which may lead to bleed or obstruction. This involvement is likely a marker of aggressivity. This example of extra-medullary disease in myeloma is an uncommon variant with features of poor prognosis and dedifferentiation. Disclosures: No relevant conflicts of interest to declare.

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