scholarly journals Novel Single Panel Method for Minimal Residual Disease Detection for Plasma Cells By Flow Cytometry

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 6-7
Author(s):  
David Kimmel ◽  
Mohammed A Aljama ◽  
Stephen Ronan Foley ◽  
Hira S Mian ◽  
Catherine A Ross
Keyword(s):  
Bone Marrow ◽  
Minimal Residual Disease ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Residual Disease ◽  
Large Data ◽  
Final Analysis ◽  
Distinct Advantage ◽  
Limit Of Quantitation ◽  
Minimal Residual

Introduction: Advancement in myeloma therapy has significantly improved outcomes including minimal residual disease (MRD) negativity that may be a surrogate for overall survival. We describe an assay using a single 10-colour panel to detect minimal residual disease (MRD) in plasma cell neoplasms to a level < 10-5. Methodology: Bone marrow aspirate specimens for this MRD assay must be from the first pull of 1.0 to 1.5 mL of bone marrow. A 1 in 10 dilution of the bone marrow is prepared and run on the instrument using a WBC count from the following calculation to determine the volume of specimen to process: For the MRD method, a corrective factor of 4 has been empirically determined to provide a sufficient number of cells to achieve the goal of 10-13 million viable cells during analysis. The required volume of bone marrow is added directly to Versalyse (Beckman Coulter) with 10% bovine serum albumin (BSA) in a bulk RBC lysis step. The cells are incubated for 15 minutes at room temperature while rocking. The cells are centrifuged and the following antibodies are added (Table 1): The cells and surface antibodies are incubated for a total of 20 minutes, specimen is gently vortexed at 10 minutes. Intracellular staining is achieved using the IntraPrep Kit (Beckman Coulter) using our standardized laboratory process. Cells are suspended in approximately 2mL of RPMI 1640 with 10% FCS. The specimen is loaded on to the Navios EX flow cytometer (Beckman Coulter)and data is acquired at approximately 5000 to 10000 events per second. The Navios EX is not capable of collecting more 1 700 000 events at acquisition when all 10 fluorescent detectors are in use plus light scatter detectors, so the specimen is repeatedly reloaded a total of 7 or 8 times. All data files are opened in Kaluza (Beckman Coulter) and merged in to a single file. This large data file is then imported in to the analysis template and analyzed for plasma cells. Analysis: A pilot of 20 specimens from patients with varying plasma cell disorders have been analyzed. Half of these specimens contained populations of monoclonal plasma cells. Ranked in order of smallest to largest (Figure 1): The smallest clone detected at 10x10E-4 is comprised of 1311 events. If a theoretical lower limit of quantitation of 50 events or 5x10E-6 in 10 000 000 total cells analyzed is required, this method will meet this criteria .Notably, all bone marrow specimens of adequate quality (not clotted, non-hemodilute) required less than 1.5mL of bone marrow to achieve > 10 000 000 nucleated cells in the final analysis. Analysis is complex using several dozen plots. Plasma cells are identified using CD38 and CD138. Gated plasma cells are analyzed for the immunophenotype of CD56, CD117, CD27, CD45, CD81 and cytoplasmic light chains simultaneously using n-dimensional radar plots (Figures 2, 3a, 3b): Qualitative results can be calculated from adjusted gates (Table 2): Conclusion: This rapid, high-sensitivity assay for immunophenotypically abnormal and clonal plasma cells requires low volumes of bone marrow. Results are ready in approximately 4 hours which is a distinct advantage and sensitivity can be shown to reach 5 X10-6. Disclosures Foley: Amgen,CelgeneJanssen: Honoraria. Mian:Takeda: Consultancy, Honoraria; Sanofi: Consultancy; Amgen: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Celgene: Consultancy.

Evaluation of a 5-Color Flow Cytometric Technique for Immunophenotyping and Quantitation of Plasma Cell Myeloma in Post-Therapy Bone Marrows.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 5036-5036
Author(s):  
Tove Isaacson ◽  
Andrzej Jakubowiak ◽  
Lloyd Stoolman ◽  
Usha Kota ◽  
William Finn ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Minimal Residual Disease ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Residual Disease ◽  
Plasma Cell Myeloma ◽  
Color Flow ◽  
Flow Cytometric ◽  
Minimal Residual

Abstract Multiparameter flow cytometry is a useful tool for comprehensive immunophenotyping of plasma cell myeloma, and has been proposed as a sensitive method for the evaluation of minimal residual disease in patients following treatment. This study aimed to assess the value of flow cytometry in quantitation of residual disease, in comparison to routine morphologic examination of first-pull bone marrow aspirate smears, in myeloma patients post-therapy. Heparinized bone marrow aspirates were obtained from 27 treated patients with plasma cell myeloma. Cells were prepared for 5-color flow cytometric analysis within 24-hours of specimen draw. Surface membrane staining with anti-CD19, CD20, CD38, CD45, CD56, and CD138 was followed by ammonium chloride lysis of red cells. Fixed and permeabilized cells were analyzed for cytoplasmic light chains to confirm clonality. Data were acquired using an FC500 flow cytometer (Beckman-Coulter), analyzed with CXP software with plasma cells isolated based on bright CD38+ or CD138+ expression. A median of 97,639 cellular events (range 14,279 to 262,508) were collected per analysis. Flow cytometric enumeration of plasma cells was compared to 500-cell differential counts of Wright-Giemsa-stained first-pull aspirate smears from the same cases. The median plasma cell count as determined by flow cytometry was 0.5% (range 0–7.9%). The median plasma cell count estimated by morphologic review was 8.0% (range 0–84.4%). Flow cytometry underestimated the plasma cell content in all but one case. Clonal plasma cells expressed CD38 and CD138 in all cases; 87.5% (21/24) coexpressed CD56, 25% (6/24) coexpressed CD45, and 4.2% (1/24) coexpressed CD19. None was positive for CD20. Although detection of minimal residual disease after therapy for acute leukemia is routinely achieved by flow cytometric analysis, successful quantitation of minimal residual disease in treated myeloma patients using flow cytometry remains limited as it usually underestimates the plasma cell content of bone marrow samples compared to routine morphology of first-pull aspirates. We have observed that this holds true for both pre-treatment and post-treatment specimens. Causes for the discrepancy may include hemodilution of second-pull aspirates used for flow cytometry, fragility and loss of plasma cells during preparation for flow cytometry, and incomplete disaggregation of plasma cells from bone marrow spicules. With improved outcome of treatments, better and more reliable methods of detection of minimal residual disease are needed for optimal prognostic stratification. We are currently validating alternative methods, which may offer more sensitivity while at the same time allow more objectivity, for assessing the amount of minimal residual disease in myeloma patients.

scholarly journals The Use of Next Generation Gene Sequencing to Measure Minimal Residual Disease in Patients with AL Amyloidosis and Low Plasma Cell Burden: A Feasibility Study

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 4353-4353 ◽  
Author(s):  
Shayna Sarosiek ◽  
Vaishali Sanchorawala ◽  
Mariateresa Fulcinti ◽  
Allison P. Jacob ◽  
Nikhil C. Munshi ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Minimal Residual Disease ◽  
Plasma Cell ◽  
Peripheral Blood ◽  
Research Funding ◽  
Plasma Cells ◽  
Residual Disease ◽  
Al Amyloidosis ◽  
Next Generation ◽  
Minimal Residual

Background: AL amyloidosis is a bone marrow disorder in which clonal plasma cells produce light chains that misfold and deposit in vital organs, such as the kidneys and heart, leading to organ failure and eventual death. Treatment is directed towards the clonal plasma cell population in an effort to halt the production of toxic light chains and recuperate organ function. Pallidini et al. demonstrated that almost 50% of patients with AL amyloidosis who achieved a complete hematologic response to prior therapy had minimal residual disease (MRD) detectable in their bone marrow by multiparametric flow cytometry (MPF).1. Next generation gene sequencing (NGS) has been a successful tool in measuring MRD among patients with multiple myeloma2 though the data regarding its use in AL amyloidosis are limited. AL amyloidosis is a disease with a much smaller plasma cell burden at baseline (typically 5-10%), making the task of isolating an initial clonal sequence even more challenging. We sought to evaluate NGS as a method of isolating a clonal population of plasma cells among patients with systemic AL amyloidosis in a first-ever feasibility study. Methods: Patients were eligible if they had systemic AL amyloidosis and no clinical evidence of concurrent active multiple myeloma. In this study, feasibility was deemed successful if discovery of a clone could be achieved in 3 out of 10 of patients. Approximately five cc's of peripheral blood and bone marrow aspirate were collected from each patient and processed for CD138 selection and DNA isolation/purification. De-identified samples were sent to Adaptive Biotech Inc. (Seattle, WA) for initial clonal identification using the ClonoSEQ immunoglobulin heavy chain (IGH) assay. Genomic DNA was amplified by implementing consensus primers targeting the IGH complete (IGH-VDJH) locus, IGH incomplete (IGH-DJH) locus, immunoglobulin κ locus (IGK) and immunoglobulin l locus (IGL). The amplified product was sequenced and a clone identified based on frequency. After proof of feasibility in the first 10 patients an additional 27 patients had initial clonal identification via the same process mentioned above. Results: In total, 37 patient samples underwent NGS via the ClonoSEQ IGH assay method. The median patient age was 66 years old (range: 44 to 83), 24% of which were female. All 37 patients had measurable disease based on serum electrophoresis and immunofixation and/or serum free light chain assay (Table 1). Four patients had no monoclonal protein detected on SIFE or UIFE and 13 patients had a normal sFLC ratio. Of the 33 patients with monoclonal disease on immunofixation, 12 patients had only a free lambda monoclonal protein and the remaining 21 patients had a clonal heavy chain with an associated light chain. Bone marrow biopsies demonstrated clonal plasmacytosis of 40% or lower. ClonoSEQ IGH assay identified trackable clones in 31 of 37 patients (84%) (see Table 1). Four patients had at least one trackable sequence (range: 1 to 5 sequences) in the peripheral blood and 29 patients had at least one trackable sequence in the bone marrow aspirate (range: 1 to 7 sequences). No correlation was seen between the detection of a clone and standard measures of plasma cell tumor burden (SIFE, SPEP, UIFE, UPEP, and sFLCs). Conclusion: NGS was successful in identifying an initial clone in 29 of 37 patients with systemic AL amyloidosis, four of which were detectable in the peripheral blood. Due to the low clonal burden in patients with AL amyloidosis, it is often difficult to assess disease status, especially post-treatment. These encouraging results may enhance disease monitoring and improve patient care in this rare disease. We are currently tracking MRD in the patients with identifiable clones as they receive systemic treatment, the results of which will be available for presentation in December 2019. REFERENCES 1. Palladini G, Massa M, Basset M, Russo F, Milani P, Foli A, et al. Persistence of Minimal Residual Disease By Multiparameter Flow Cytometry Can Hinder Recovery of Organ Damage in Patients with AL Amyloidosis Otherwise in Complete Response. Abstr 3261. 2016; 2. Ladetto M, Brüggemann M, Monitillo L, Ferrero S, Pepin F, Drandi D, et al. Next-generation sequencing and real-time quantitative PCR for minimal residual disease detection in B-cell disorders. Leukemia. 2014;28:1299-307. Table 1 Disclosures Sarosiek: Acrotech: Research Funding. Sanchorawala:Proclara: Consultancy, Honoraria; Takeda: Research Funding; Caelum: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Research Funding; Prothena: Research Funding; Celgene: Research Funding. Jacob:Adaptive Biotechnologies: Employment, Other: shareholder. Munshi:Amgen: Consultancy; Adaptive: Consultancy; Celgene: Consultancy; Celgene: Consultancy; Janssen: Consultancy; Janssen: Consultancy; Takeda: Consultancy; Takeda: Consultancy; Oncopep: Consultancy; Oncopep: Consultancy; Amgen: Consultancy; Abbvie: Consultancy; Abbvie: Consultancy; Adaptive: Consultancy.

scholarly journals The Prognostic Significance of Minimal Residual Disease Detection after Induction and ASCT to the Patients with Multiple Myeloma

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 4956-4956
Author(s):  
Weiqin Yao ◽  
Zhu Mingqing ◽  
Yao Feirong ◽  
Lingzhi Yan ◽  
Song Jin ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Minimal Residual Disease ◽  
Induction Therapy ◽  
Plasma Cells ◽  
Residual Disease ◽  
Prognostic Significance ◽  
Depth Of Response ◽  
Minimal Residual

Abstract Objective: In the last decade the outcome in multiple myeloma in CHINA has greatly improved due to the new, effective therapies including PIs and Imids. But responses to treatment and survival remains heterogeneous because of patient characteristic, disease biology and mechanisms of drug resistance. More and more studies have established the link between depth of response and improved PFS and OS. multiparameter-flow cytometry (MFC) is a main method to detect minimal residual disease(MRD) in myeloma. Sensitivity will be at least at 10-4 to 10-5 by 10-color MFC. Imaging techniques such as PET-CT are important for EMD and bone MRD detection. whole body DWI-MRI is a new imaging technique by mean of the apparent diffusion coefficient(ADC) which can qualify the depth of response to antineoplastic treatment. This study was designed to evaluate the prognostic significance of MRD by 10-color MFC and imaging to the MM patients after induction.Methods: 102 patients with newly diagnosed MM were enrolled at the First Affiliated Hospital of Soochow University from July 2015 to July 2017. All patients were diagnosed and the response were assessed by IMWG criteria. The median of age was 58 (31-75).There were 46 patients with IgG type , 24 IgA , 14 light chain, 18 others. 34 Patients in ISS stageⅠ,34 in stage Ⅱ, 30 in stage Ⅲ. All patients received 4-6 cycles of triplet bortezomib based or lenalidomide based induction therapy. Transplantation available patients received APBSCT with BUCY condition followed by 4-6 cycles of bortezomib based or lenalidomide based consolidation which were given to transplantation unavailable patients too. Lenalidomide and thalidomide were used for over 2y of maintenance therapy. Bone marrow aspirates for MRD imaging MRD assessment were obtained at the end of induction and 1year after ASCT.The median of follow-up was 13 (2-29) months.Results: According to MRD by MFC and imaging after induction therapy and 1 year after ASCT, the patients were divided into different groups. MFC negativity was 33%(29/88) after induction therapy compared with 63%(32/51) after ASCT (X2=11.636,P=0.001). After induction therapy, the median PFS was 22 months for MRD positive group compared with not reached with MRD negative group by MFC (P=0.042) in patients with very good partial remission(VGPR) and above. The 2 years PFS was 100% for those with MRD negative compared with 60% for MRD positive by imaging. The 2 years PFS was 80% for those have multiclonal normal plasma cells compared with 52.6% for those without. The median PFS was not reached for MFC MRD negative patients 1 year after ASCT compared with 20 months for positive patients. (P=0.002). Multivariate analysis including high risk cytogenetics(17p-, t(4;14), t(14;16)), sex, age, ISS, chemotherapy, ASCT, CR/VGPR, normal PCs showed that the MFC MRD and ASCT were independent prognostic factor.Conclusions: Patients with MFC MRD negative after induction therapy or ASCT is a better prognostic marker than CR or even the best marker. Imaging MRD negativity and the appearance of normal plasma cells in the bone marrow suggests a better prognosis.We will have a try to do more research on overall survival(OS),include longer follow-up and a larger number of patients enrolled. Figure. Figure. Disclosures No relevant conflicts of interest to declare.

scholarly journals Role of Imaging in the Evaluation of Minimal Residual Disease in Multiple Myeloma Patients

2020 ◽  
Vol 9 (11) ◽  
pp. 3519
Author(s):  
Elena Zamagni ◽  
Paola Tacchetti ◽  
Simona Barbato ◽  
Michele Cavo
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Minimal Residual Disease ◽  
Plasma Cells ◽  
Residual Disease ◽  
Imaging Techniques ◽  
Response Assessment ◽  
Response To Therapy ◽  
Whole Body ◽  
Minimal Residual

The International Myeloma Working Group (IMWG) recently introduced the evaluation of minimal residual disease (MRD) within the multiple myeloma (MM) response criteria, and MRD negativity assessed inside and outside the bone marrow is currently considered the most powerful predictor of favorable long-term outcomes. However, MRD evaluation has thus far relied on flow-cytometry or molecular-based methods, despite the limitations associated with the patchy infiltration of bone marrow (BM) plasma cells and the presence of extra-medullary (EMD). On the contrary, imaging-based sensitive response assessment through the use of functional rather than morphological whole-body (WB) imaging techniques, such as positron emission tomography with computed tomography (PET/CT) and magnetic resonance imaging (MRI), likely is a promising strategy to overcome these limitations in evaluating response to therapy and in the assessment of the MRD status in MM patients. However, despite the significant advances in the development and availability of novel functional imaging techniques for MRD evaluation, a worldwide standardization of imaging criteria for acquisition, interpretation, and reporting is yet to be determined and will be object of future investigations.

scholarly journals Evaluation of Hematopoietic Stem Cell Product As a New Site for Minimal Residual Disease By Next Generation Flow in Multiple Myeloma

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 4927-4927
Author(s):  
Herbert Henrique de Melo Santos ◽  
Glaciano Ribeiro ◽  
Allan de souza Santos ◽  
Marcos Chaves ◽  
Joanna Leal ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Stem Cell ◽  
Minimal Residual Disease ◽  
Hematopoietic Stem Cell ◽  
Plasma Cell ◽  
Residual Disease ◽  
Next Generation ◽  
Hematopoietic Stem ◽  
Minimal Residual

Abstract Introduction- Next generation flow (NGF) is one of the approaches for testing multiple myeloma (MM) minimal residual disease (MRD) over conventional response assessments. Actually, bone marrow (BM) is the preference site of evaluation because of its sensitivity. Because of its invasively technic, other possible sites for MRD evaluation outside the BM have been studied. In the present study we analyzed the MRD between the BM and the hematopoietic stem cell collected product (HSC product), once the concentration of plasma cell in the HSC product could be higher than peripheric blood sample. Aims- To compare MRD quantification of plasma cell between BM and HSC product after induction from Newly Diagnosed MM(NDMM) Transplant Eligible (TE) patients (pts) exposed to daratumumab, cyclophosphamide, thalidomide and dexamethasone (Dara-CTD) protocol. Methods- The SC product and BM samples were collected after four 28 days cycles of induction therapy from pts treated with Dara-CTd protocol described before by (Crusoe E. et al. Blood 2020; 136 (supplement 1): 17-18). MRD was evaluated by next-generation flow (NGF) based in the EuroFlow® protocol. EuroFlow standards was used to identify clonality and aberrant PC immune phenotype, consisting by EuroFlow 8-color 2-tube method (MM MRD kit, Cytognos, Salamanca), with the acquisition of 5 million events each tube and then merged into a single analysis tube on approximately 10 million events. Plasma cells were identified by CD38 multiepitope and CD138. Other markers were used to detect abnormal phenotypes. For comparison of MRD results, Bland-Altman plot comparing BM-MRD and HSC product-MRD was performed. Results- The first pts was enrolled in November 2018. A total of 24 pts were included, the median age was 60 (range 37- 67 years), 23 (92%) were non-white, 5 (21%) had an R-ISS = 1, 12 (54%) had an R-ISS = 2 and 4 (16%), an R-ISS = 3. Six (25%) pts had high-risk chromosomal abnormalities [del17p, t(4;14) or t(14;16)]. To date, all pts have completed induction and 20 have received transplant. Regarding response rates, after the end of induction (cycle 4), 19 (90%) of the pts obtained > PR and 8 (38%) obtained >VGPR, including three MRD negativity by NGF. 19 pts were analyzed for MRD. Negative MRD in sensitivity <10 -5, >=10 -5 and <10 -4, >=10 -4 evaluated in bone marrow was 4/19(21%), 4/19(21%), 11/19(58%) respectively. Negative MRD in sensitivity <10 -5, >=10 -5 and <10 -4, >=10 -4 evaluated in the HSC product was 13/19(68%), 3/19(16%), 3/19(16%) respectively. Median bone marrow sensitivity 10 -4 lower quartile 10 -5 upper quartile 10 -3. Normal distribution of the differences between BM and SC product MRD was first assessed (Kolmogorov-Smirnov's p < 0.001, n = 19). Discussion-Conclusions- The use of HSC product could enhance the plasma cell concentration and may be an alternative and attractive method for MRD detection that diminished the invasiveness of repetitive bone marrow aspirations and tackling the heterogeneity distribution of MM cells. In this preliminary data the sample size did not allow to show a direct correlation between BM and HCS product. A larger sample would be needed to confirm the hypothesis. Figure 1 Figure 1. Disclosures Hungria: Amgen, BMS, Celgene, Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Support for attending meetings/travel ; Abbvie: Honoraria; Sanofi: Honoraria, Other: Support for attending meetings/travel ; Takeda: Honoraria. De Queiroz Crusoe: Janssen: Research Funding.

scholarly journals The Prognostic Significance of Minimal Residual Disease in Multiple Myeloma in the Molecular Genetic Groups Msmart 3.0

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 42-42
Author(s):  
Andrey Garifullin ◽  
Sergei Voloshin ◽  
Sergey Linnikov ◽  
Irina Martynkevich ◽  
Alexey Kuvshinov ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Minimal Residual Disease ◽  
Plasma Cells ◽  
Residual Disease ◽  
Molecular Genetic ◽  
Prognostic Significance ◽  
Risk Groups ◽  
High Dose ◽  
Genetic Abnormalities ◽  
Minimal Residual

Assessment of the role of genetic abnormalities and minimal residual disease (MRD) is an active developing area in hematology. The use of genetic methods makes it possible to predict the course of the disease and apply an individualized approach to antimyeloma therapy. At the same time, the identification of MRD after therapy determines possibility of relapse. Aim. To identify the prognostic potential of MRD in patients in the standard and high molecular risk groups. Materials and methods. We analyzed 72 patients with MM (median age was 59 years (range 37-80), male/female - 1.3:1). All patients received initial therapy with proteasome inhibitors and / or immunomodulators. High dose therapy (MEL200) and autologous stem cell transplantation (ASCT) was carried out 50 (69%) patients. Standard cytogenetic and FISH methods were used to stratify patients in risk groups of mSMART 3.0. The standard risk (SR) was established in 52 (72%) patients, the high risk (HR) - in 20 (28%) patients. The MFC MRD status of bone marrow was evaluated after 4-6 cycles of induction therapy or after ASCT with use of 5-colors flow cytometry. MRD-negative status (MRD-) was based on level of clonal plasma cells <10-4 in bone marrow sample. Results. The MRD- was reached in 36% (26/72) patients. The median of OS in MRD+ group was 104 months, in MRD- was 146 months (p=.01). The median of PFS in MRD+ group was 26 months, in MRD- was 70 months (p=.00021). 2-years PFS in MRD+ group was 56%, in MRD- group was 100% (p=.00021). We divided patients into the following groups for evaluation the effect of MRD on survival in risk groups: SR МRD+ 34/72 (47%), SR МRD- 18/72 (25%), HR МRD+ 12/72 (17%) and HR MFC МRD- 8/72 (11%). The median of OS in HR MRD+ group was 72 months, in SR MRD+ - 104 months, in HR MRD- - 146 months, in SR MRD- was not achieved (p=.02). The median of PFS in HR MRD+ group was 24 months, in SR MRD+ - 26 months, in HR MRD- - 68 months, in SR MRD- - 70 months (p=.003). The 2-years PFS in HR MRD+ group was 44%, in SR MRD+ group was 50% and in SR MRD- and HR MRD- groups were 100% (p=.003). Conclusion. The absence of MRD is the most important prognostic factor. The leveling of negative effect of genetic abnormalities become possible when the MRD-negative response status is achieved. Presumably, this is due to the elimination of clonal plasma cells owing to the use of optimal antimyeloma therapy which is based on the risk stratification. Disclosures Martynkevich: Pfizer: Honoraria, Speakers Bureau; BMS: Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau. Shuvaev:Novartis: Honoraria, Speakers Bureau; BMS: Honoraria, Speakers Bureau; Pfizer: Honoraria, Speakers Bureau.

Flow Cytometric Immunophenotyping for Minimal Residual Disease Analysis in Multiple Myeloma

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2711-2711
Author(s):  
Ritu Gupta ◽  
Archana Bhaskar ◽  
Paresh Jain ◽  
Atul Sharma ◽  
Lalit Kumar
Keyword(s):  
Bone Marrow ◽  
Minimal Residual Disease ◽  
Bone Marrow Aspirate ◽  
Plasma Cells ◽  
Bone Marrow Cells ◽  
Residual Disease ◽  
Protein Electrophoresis ◽  
Tumor Load ◽  
Total Bone Marrow ◽  
Minimal Residual

Abstract Recent studies using multi-parametric flow cytometry (MFC) have shown frequent immunophenotypic aberrations of plasma cells (PCs) and their utility in minimal residual disease (MRD) detection in multiple myeloma (MM). Presence of normal PCs and hemodilution of the bone marrow aspirate are important considerations in MRD assessment by MFC. The purpose of present study was to characterize PCs in MM to determine the incidence of aberrant antigen expression on myeloma PCs and its role in estimation of minimal residual disease. A total of 108 patients of MM were evaluated to study the immunophenotype of PCs and 37 patients for estimation of MRD using pre-titrated volumes of the following monoclonal antibodies: CD19 FITC, CD20 FITC, CD45 FITC, CD117 PE, CD56 PE, CD38 PE-Cy5.5, CD138 APC (BD Biosciences, San Jose, CA, USA), CD52 PE, Kappa (κ) FITC and Lambda (λ) PE (Serotec). CD38 and CD138 were added to all the tubes for specific identification of PCs. 2ml of bone marrow aspirate was collected from all the subjects as per the guidelines of the institute ethics committee, processed for immunophenotyping studies using standard whole blood lysis technique and analyzed with 4-color flow cytometry. At least two antigens were aberrantly expressed in all and three in 92.6% of MM with CD19 being most frequent followed by CD56, CD45, CD52, CD117 and CD20. Using aberrant immunophenotype for identification of neoplastic PCs, MRD by MFC was detectable in all the patients of MM with ≤5% PC on bone marrow smears. The neoplastic PCs as percentage of total bone marrow cells could not differentiate protein electrophoresis (PE) + from PE− samples (Figure 1A). Assuming that in a hemodiluted bone marrow aspirate both the neoplastic and normal PCs would be proportionately reduced and normal PCs would outnumber the neoplastic clone after successful therapy; when we analyzed the tumor load i.e. the neoplastic PC as percentage of total PCs, a cut-off of 50% neoplastic PCs of total PCs could differentiate PE + samples from PE− ones (Figure 1B). To conclude, MRD detection by aberrant antigen expression is useful and evaluation of tumor load i.e. neoplastic PCs as percentage of total PC may help in better assessment of response to therapy and circumvent the problem of hemodilution in MFC based MRD assays in MM. Figure 1: Tumor load i.e. neoplastic plasma cells (NPCs) as % of total PCs (A) & of total bone marrow cells (B) in samples evaluated for minimal residual disease. The NPCs constituted ≤50% of the total PCs in protein electrophoresis (PE) & immunofixation negative samples and >60% of the total PCs in PE+ samples (B). Arrow indicates two cases which were PE- but positive on immunofixation and had relatively low numbers of neoplastic PCs (B) <> Figure 1:. Tumor load i.e. neoplastic plasma cells (NPCs) as % of total PCs (A) & of total bone marrow cells (B) in samples evaluated for minimal residual disease. The NPCs constituted ≤50% of the total PCs in protein electrophoresis (PE) & immunofixation negative samples and >60% of the total PCs in PE+ samples (B). Arrow indicates two cases which were PE- but positive on immunofixation and had relatively low numbers of neoplastic PCs (B) <>

scholarly journals Standardized assay for assessment of minimal residual disease in blood, bone marrow and apheresis from patients with plasma cell myeloma

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Agnieszka Blum ◽  
Katy Haussmann ◽  
Mathias Streitz ◽  
Stephan Schlickeiser ◽  
Carola Tietze-Buerger ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Minimal Residual Disease ◽  
Plasma Cell ◽  
Residual Disease ◽  
Plasma Cell Myeloma ◽  
Minimal Residual
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