Flow Cytometry Based Detection of MRD in Bone Marrow of Patients with Multiple Myeloma: A Comparison Between Fluorescent-Based Cytometry Versus Cytof

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 4195-4195 ◽  
Author(s):  
Arnab Ghosh ◽  
Nicole Carreau ◽  
Alessandra Moscatello ◽  
Adeeb Rahman ◽  
Jingjing Qi ◽  
...  
Keyword(s):  
Heavy Metal ◽  
Multiple Myeloma ◽  
Flow Cytometry ◽  
Metal Ions ◽  
Research Funding ◽  
Heavy Metal Ions ◽  
Residual Disease ◽  
Response To Therapy ◽  
Institutional Research ◽  
Highly Sensitive

Abstract The introduction of novel agents for the treatment of multiple myeloma (MM) has shifted the emphasis towards achieving a molecular complete remission (CR). Traditional methods to detect malignant plasma cells (PC) in minimal residual disease (MRD) utilize multicolor flow cytometry (MFC) to detect aberrant phenotypes. However fluorescent-based MFC assays are limited by the number of markers, difficulty in standardization of assays and overlap of signal between the fluorescent channels. These limitations can be overcome by cyTOF, a flow cytometry assay based on time-of-flight mass spectrometry using antibodies labeled with heavy-metal ions, permitting simultaneous assessment of large panels of markers. We have developed a novel highly sensitive mass cytometry protocol for the detection of MRD. BM from 5 MM patients and 1 non-myeloma control were RBC-lysed and labeled with MM markers for MFC and cyTOF. cyTOF is a flow cytometry based on mass spectometry where antibodies are labeled with heavy-metal ions, permitting utilization of more markers without concern for spillover. The MFC panel included CD38, CD138, CD45, CD56, CD19, CD117. Labeled cells for MFC were acquired by BD LSR Fortessa.. The panel for cyTOF comprised markers for cells across the hematopoietic spectrum and those specific for MM. Labeled cells acquired with cyTOF were analyzed using SPADE. Limits of detection (LOD) and quantification (LOQ) for MFC are based on prior reports. The results were compared with an independent commercial MFC-based assay (Genoptix). Malignant PC could be detected by MFC and cyTOF based assasys (See Figure). We were able to detect MRD in three subjects that had no detectable disease by an independent commercial MFC based assay (Genoptix). This can be attributed to acquiring at least 1x106 for our assays leading to a lower LOD and LOQ (greater sensitivity). Using cyTOF and SPADE, we could detect MRD in 4/5 patients. Overall our panel with cyTOF has a lower LOD than MFC for detection of MRD. Early detection of MRD in MM patients using a highly sensitive flow cytometry like cyTOF will help in risk stratification, predicting relapse and studying response to therapy. Disclosures Chari: Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Millennium/Takeda: Consultancy, Research Funding; Onyx: Consultancy, Research Funding; Array Biopharma: Consultancy, Other: Institutional Research Funding, Research Funding; Biotest: Other: Institutional Research Funding; Novartis: Consultancy, Research Funding. Jagannath:Bristol Myers Squibb: Honoraria; Janssen: Honoraria; Merck: Honoraria; Novartis: Honoraria; Celgene: Honoraria.

Real-World Experience with Minimal Residual Disease Testing with Next Generation Flow Cytometry and Functional Imaging in Multiple Myeloma

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 17-18
Author(s):  
David Böckle ◽  
Paula Tabares Gaviria ◽  
Xiang Zhou ◽  
Janin Messerschmidt ◽  
Lukas Scheller ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Flow Cytometry ◽  
High Risk ◽  
Functional Imaging ◽  
Real World ◽  
Research Funding ◽  
Residual Disease ◽  
Focal Lesions ◽  
High Risk Disease ◽  
Minimal Residual

Background: Minimal residual disease (MRD) diagnostics in multiple myeloma (MM) are gaining increasing importance to determine response depth beyond complete remission (CR) since novel agents have shown to induce high rates of deep clinical responses. Moreover, recent reports indicated combining functional imaging with next generation flow cytometry (NGF) could be beneficial in predicting clinical outcome. This applies in particular to the subset of patients suffering from relapsed/refractory multiple myeloma (RRMM) who tend to show a higher incidence of residual focal lesions despite serological response. Here, we report our institutions experience with implementing both functional imaging and NGF-guided MRD diagnostics in clinical practice. Methods: Our study included patients with newly diagnosed multiple myeloma (NDMM) and RRMM achieving VGPR, CR or sCR. Bone marrow aspirates were obtained for MRD-testing according to IMWG 2016 criteria. Samples were collected between July 2019 and July 2020 and analyzed with NGF (according to EuroFlowTM guidelines) at a sensitivity level of 10-5. Results were compared to functional imaging obtained with positron emission tomography (PET) and diffusion-weighted magnetic resonance imaging (DW-MRI). High-risk disease was defined as presence of deletion 17p, translocation (14;16) or (4;14). Results: We included 66 patients with NDMM (n=39) and RRMM (n=27) who achieved VGPR or better. In patients with RRMM the median number of treatment lines was 2 (range 2-11). Fifteen patients suffered from high-risk disease. Median age at NGF diagnostics was 64 years (range 31-83). Among patients achieving VGPR (n=27), CR (n=10) and sCR (n=29) seventeen (26%) were MRD-negative by NGF testing. CR or better was significantly associated NGF MRD-negativity (p=0.04). Notably, rates of NGF MRD-negativity were similar among patients with NDMM (28%) and RRMM (26%). Even some heavily pretreated patients who underwent ≥ 4 lines of therapy achieved MRD-negativity on NGF (2 of 9). Functional imaging was performed in 46 (70%) patients with DW-MRI (n=22) and PET (n=26). Median time between NGF and imaging assessment was 2 days (range 0-147). Combining results from imaging and NGF, 12 out of 46 (26%) patients were MRD-negative with both methods (neg/neg). Three patients displayed disease activity as measured with both, imaging and NGF (pos/pos). Twenty-nine of the remaining patients were MRD-positive only according to NGF (pos/neg), while two patients were positive on imaging only (neg/pos). More patients demonstrated combined MRD-negativity on NGF and imaging (neg/neg) in the NDMM setting than in RRMM (32% versus 19%). We also observed that 30% of the patients with high-risk genetics showed MRD-negativity on both imaging and NGF. Of note, none of the patients with very advanced disease (≥4 previous lines) was MRD-negative on both techniques. Conclusion In the clinical routine, MRD diagnostics could be used to tailor maintenance and consolidation approaches for patients achieving deep responses by traditional IMWG criteria. Our real-world experience highlights that MRD-negativity can be achieved in patients suffering from high-risk disease and also in late treatment lines, supporting its value as endpoint for clinical trials. However, our data also support MRD diagnostics to be combined with functional imaging at least in the RRMM setting to rule out residual focal lesions. Future studies using MRD for clinical decision-making are highly warranted. Disclosures Einsele: Takeda: Consultancy, Honoraria, Speakers Bureau; Janssen: Consultancy, Honoraria, Research Funding, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; Amgen: Consultancy, Honoraria, Research Funding, Speakers Bureau; Celgene: Consultancy, Honoraria, Research Funding, Speakers Bureau; GlaxoSmithKline: Honoraria, Research Funding, Speakers Bureau; Bristol-Myers Squibb: Consultancy, Honoraria, Research Funding, Speakers Bureau; Sanofi: Consultancy, Honoraria, Research Funding, Speakers Bureau. Rasche:Celgene/BMS: Honoraria; GlaxoSmithKline: Honoraria; Oncopeptides: Honoraria; Skyline Dx: Research Funding; Janssen: Honoraria; Sanofi: Honoraria.

Straw-sheaf-like terbium-based coordination polymer architectures: microwave-assisted synthesis and their application as selective luminescent probes for heavy metal ions

2015 ◽  
Vol 39 (4) ◽  
pp. 2973-2979 ◽  
Author(s):  
Mengmeng Shi ◽  
Chenghui Zeng ◽  
Lei Wang ◽  
Zhiwen Nie ◽  
Yongxia Zhao ◽  
...  
Keyword(s):  
Aqueous Solution ◽  
Heavy Metal ◽  
Coordination Polymer ◽  
Metal Ions ◽  
Heavy Metal Ions ◽  
Microwave Assisted Synthesis ◽  
Luminescent Probes ◽  
Microwave Assisted ◽  
Highly Sensitive ◽  
Polymer Architectures

Terbium-based coordination polymer architectures were successfully synthesized via a microwave heating approach and they showed highly sensitive and selective luminescence quenching to Pb2+ in aqueous solution.

scholarly journals Minimal Residual Disease in Autografts and Bone Marrow of Patients with Multiple Myeloma: 8-Color Multiparameter Flow Cytometry (EuroFlow-NGF) Vs. Next-Generation Sequencing

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 22-23
Author(s):  
Hiroyuki Takamatsu ◽  
Naoki Takezako ◽  
Takeshi Yoroidaka ◽  
Takeshi Yamashita ◽  
Ryoichi Murata ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Flow Cytometry ◽  
Research Funding ◽  
Prognostic Value ◽  
Residual Disease ◽  
Multiparameter Flow Cytometry ◽  
Next Generation ◽  
Generation Sequencing ◽  
Minimal Residual

Background: Autologous stem cell transplantation (ASCT) in conjunction with novel therapeutic drugs can dramatically improve response rates and the prognoses of patients with multiple myeloma (MM). However, most patients with MM ultimately relapse due to minimal residual disease (MRD). Next-generation multiparameter flow cytometry (MFC) (EuroFlow-NGF) and next-generation sequencing (NGS) are currently the standard methods to assess MRD. Aims: To compare the prognostic value of MRD detection in autografts and bone marrow (BM) cells using 8-color MFC (EuroFlow-NGF) and NGS (Adaptive Biotechnologies), and also MRD levels between fresh and cryopreserved autografts using NGF. Methods: The study enrolled 52 newly-diagnosed MM patients who underwent ASCT. The median age ASCT was 61 (range 41-69) years and included 29 males and 23 females at ISS I (n = 17), II (n = 23), and III (n = 12). Of these, 18 patients harbored high-risk chromosomal abnormalities including t(4;14) (n = 15), del17p and t(4;14) (n = 2), and complex (n = 1). Bortezomib-based chemotherapy was used for induction together with melphalan at 140 mg/m2 (n = 1) and 200 mg/m2 (n = 51) for conditioning before ASCT. 39 of 52 (75%) patients received maintenance therapy until progressive disease. The best responses achieved post-ASCT included 30 sCR, 4 CR, 15 VGPR, and 3 PR. Forty autografts, one from each MM patient, were analyzed using NGF and NGS protocols, and BM cells at pre/post-ASCT and autografts derived from 16 patients were analyzed using NGS. The EuroFlow-NGF method uses standard sample preparation; large numbers of cells are evaluated using an optimized 8-color antibody panel that facilitates accurate identification of discrimination between phenotypically aberrant plasma cells (aPCs) and their normal counterparts (Flores-Montero et al., Leukemia 2017). NGS-based MRD assessment was performed using Adaptive's standardized NGS-MRD Assay (Seattle, WA) (Martinez-Lopez et al., Blood 2014). Eight additional autografts were used to assess MRD in both fresh and cryopreserved samples by NGF. Results: MRD was evaluated in 48 of 52 autografts (92%) using NGF and in 44 of 52 autografts (85%) using NGS. We identified aPCs in autografts based on multivariate analysis of individual cell populations (e.g., CD56+, CD19−, CyIgκ+, and CD117+). As the results of NGF revealed a strong correlation with respect to MRD in fresh vs. thawed autografts (r = 0.999, P < 0.0001), MRD was subsequently evaluated in thawed autografts. The sensitivity of NGF was 1 × 10−5-2 × 10−6; the sensitivity of NGS was 1 × 10−6. 28 of 48 (58%) of the autografts were MRD-positive by NGF; 30 of 44 (68%) of the autografts were MRD-positive by NGS. MRD levels in autografts using NGF and NGS correlated with one another (r = 0.69, P < 0.0001; Fig. 1A). MRD negative in autografts by NGF cases (MRDNGF (-)) and MRDNGS (-) tended to show better progression-free survival (PFS) than MRDNGF (+) (P = 0.195) and MRDNGS (+) (P = 0.156), respectively. Furthermore, MRDNGS (-) showed significantly better overall survival (OS) than MRDNGS (+) (P = 0.03) (Fig. 1C) while MRDNGF (-) showed better OS than MRDNGF (+) (P = 0.09) (Fig. 1B). Our data revealed only a minimal correlation between MRD in the autografts (median 1.1 × 10−5,range 0-7.29 × 10−4) and in the BM cells at pre-ASCT (median 5.05 × 10−3,range 6 × 10−6-2.64 × 10−1; r = 0.09, P = 0.7) or at post-ASCT (median 2.11 × 10−4,range 0-9.09 × 10−3; r = 0.14, P = 0.6); MRD detected in the autografts was > 27 times lower than that detected in pre-ASCT BM cells, and MRD detected in the post-ASCT BM cells was > 3 times lower than that detected in pre-ASCT BM cells except for one case in which the ratio was increased by two times. Interestingly, while MRD was detected in all BM cells at pre-ASCT (n = 16), 4 of 16 (25%) of these autografts were MRDNGS-negative. The median of MRD levels of the 4 cases in pre-ASCT and post-ASCT BM cells were 4.14 × 10−4 (range 6-583 × 10−6)and 1.8 × 10−5 (range 0-27 × 10−6), respectively. Conclusion: Although EuroFlow-NGF is a rapid and accurate method for detecting MRD, NGS was more sensitive and provided greater prognostic value than EuroFlow-NGF. Disclosures Takamatsu: Adaptive Biotechnologies: Honoraria; Bristol-Myers Squibb: Honoraria, Research Funding; Janssen Pharmaceutical: Consultancy, Honoraria, Research Funding; Ono pharmaceutical: Honoraria, Research Funding; SRL: Consultancy, Research Funding. Takezako:Bristol-Myers Squibb: Honoraria, Research Funding; Takeda: Honoraria, Research Funding; Janssen: Research Funding; Abbvie: Research Funding. Nakao:Symbio: Consultancy; Kyowa Kirin: Honoraria; Alexion: Research Funding; Novartis: Honoraria.

scholarly journals Retrospective Analysis of Minimal Residual Disease Testing By High Throughput Immunosequencing Versus High Sensitivity Flow Cytometry in Multiple Myeloma

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 1625-1625
Author(s):  
Anwar Khan ◽  
Nagehan Pakasticali ◽  
Omar Fathalla ◽  
Taiga Nishihori ◽  
Mohammad O Hussaini
Keyword(s):  
Multiple Myeloma ◽  
Flow Cytometry ◽  
Minimal Residual Disease ◽  
Correlation Coefficient ◽  
Research Funding ◽  
Residual Disease ◽  
High Sensitivity ◽  
Spearman Correlation ◽  
Color Flow ◽  
Minimal Residual

Abstract Introduction: Detection of minimal residual disease (MRD) is one of the strongest predictors of outcome in multiple myeloma (MM). Until recently, the most commonly available method to detect MRD in clinical practice has been high sensitivity flow cytometry (FC) which can detect MRD with at 10 -5 sensitivity. In recent years, next-generation sequencing (NGS) has become a viable method to assess the MRD in MM patients with a 10 -6 sensitivity. NGS appears to have some advantages over HC-FC by circumventing subjectivity of analysis. However, real-world comparison between these two methodologies in the literature is limited and is important to inform daily hematopathology and oncology ordering practices. Methods: We retrospectively identified all cases of MM with NGS MRD data from bone marrow specimens at the Moffitt Cancer Center and collated corresponding flow MRD data and clinical data (OS, patient demographics) electronically and via chart review. 10-color flow cytometry was performed on a Gallios System and analyzed on Kaluza (Beckman Coulter, IN). Two million events were collected on all cells. Validated lower limit of detection was at least 0.01%. Antibodies included CD28, CD81, CD56, CD138, CD319, CD20, CD19, CD117, CD38, CD45, CD27, CD200 (BD, Biolegend, Beckman Coulter). clonoSEQ ® (Adaptive Biotechnologies, Seattle, WA) testing was performed which uses multiplex polymerase chain reaction (PCR) and NGS to identify, characterize, and monitor clonotypes of immunoglobulin (Ig) IgH (V-J), IgH (D-J), IgK, and IgL receptor gene sequences, and translocated BCL1/IgH (J) and BCL2/IgH (J) sequences Statistical analysis was performed by Spearman correlation coefficient and Kaplan-Meier analysis. Results: 192 samples from 122 unique patients were identified that had both NGS and FC data performed on the same sample. FC+ values ranged from 1x10 -7 to 0.39. NGS+ values ranged from 2.3 x 10 -7 to 0.15. Spearman correlation coefficient showed moderate concordance between NGS and FC at r=0.67 (p<0.001). Six samples were positive by FC (mean tumor burden (MTB)= 0.0007) but missed by NGS; whereas 59 samples were positive by NGS (MTB= 0.002) but missed by flow cytometry. Two cases by FC were equivocal and these were both definitively designated as MRD+ by NGS. Overall survival was worse for MRD+ (by NGS or FC) vs MRD(-) (Figure 1). Conclusion: Our study confirms the importance of MRD detection in MM and shows the robust utility of NGS for MRD detection in routine hematopathology practice. While both FC and NGS are complementary given that each can potentially detect MRD missed by another method, the data supports the increased sensitivity of NGS over FC. Figure 1 Figure 1. Disclosures Nishihori: Novartis: Research Funding; Karyopharm: Research Funding. Hussaini: Stemeline Therapeutics: Honoraria.

scholarly journals Measurable Residual Disease in Extracellular Vesicles from Bone Marrow and Peripheral Blood of Patients with Multiple Myeloma: A Proof-of-Concept Study

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 4712-4712
Author(s):  
Rui Bergantim ◽  
Mélanie A.G. Barbosa ◽  
Sara Peixoto da Silva ◽  
Bárbara Polónia ◽  
Hugo R. Caires ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Flow Cytometry ◽  
Complete Remission ◽  
Research Funding ◽  
Residual Disease ◽  
Disease Diagnosis ◽  
Invasive Monitoring ◽  
Protein Markers ◽  
Measurable Residual Disease

Abstract BACKGROUND: Multiple myeloma (MM) treatment improved substantially in the last years, with unprecedented survival outcomes. However, even when achieving complete remission, patients ultimately relapse. Therefore, monitoring measurable residual disease (MRD) is crucial to assess treatment response and define the depth of patients' remission status. However, this currently still requires invasive bone marrow (BM) aspirates, which severely hinders real-time monitoring of the disease. Therefore, the identification of biomarkers of MRD in the peripheral blood (PB) of patients would allow a more frequent and minimally invasive monitoring of MRD. Extracellular Vesicles (EVs) are small particles (30-1000nm) shed by all cells, which are found in all biofluids including the BM and PB. These particles carry a specific cargo from their cell of origin, including proteins, enclosed by a lipidic layer. Therefore, they have been described as a possible source of cancer biomarkers, with potential to monitor MRD. AIMS: This study aimed to implement a protocol for the isolation of EVs from the BM and PB of MM patients at distinct stages of the disease (diagnosis and remission), in order to detect and compare the levels of known MRD biomarkers in their cargo. METHODS: The study was previously approved by the Ethical Committee of CHSJ and patient's consent was obtained. EVs from BM and PB Platelet-Poor Plasma (PPP) were isolated by size-exclusion chromatography (SEC), and further concentrated by ultrafiltration (UF). Then, the EVs were characterized according to their size and concentration (by Nanoparticle Tracking Analysis), morphology (by Transmission Electron Microscopy), protein concentration (Lowry protein assay) and presence of EV-associated protein markers (Western Blot - WB). In addition, 16 known MRD and MM biomarkers were analyzed by WB in the isolated EVs from PB and BM of seven patients, at two main stages of the disease - diagnosis versus response after autologous stem cell transplant (ASCT). Clinical features regarding cytogenetics and immunophenotypic markers using multi-parameter flow cytometry (MFC) were analyzed and compared. RESULTS: The two-step protocol described allowed the isolation of size-resolved EVs from both PB and BM of MM patients. The EVs isolated (both from PB and BM) presented a size-range from 50 to 500nm and presented EV-associated protein markers, such as CD81 and CD63. Moreover, several MM MRD biomarkers (e.g. CD56, CD45, CD38 and light chain) were detected in the cargo of the EVs from BM and PB at diagnosis and complete remission. The biomarkers of MM and MRD detected in the cargo of PB EVs were mainly the same as the ones detected in the cargo of BM EVs. The complete remission after ASCT was mostly associated with a decrease in the expression of EV-associated MM markers in both the BM and the PB; however, in some patients a few of the markers persisted at this stage when compared to diagnosis. In fact, the expression of CD45 and HLA-DR persisted at the remission stage in 3 and 2, respectively, out of 5 patients presenting these markers at diagnosis. Moreover, an increased expression of CD56 was also detected at remission in 3 out of 7 patients. By correlating these data with patient's routine work-up it was found that patients with persistent CD45 didn't reach 10^-5 MRD negative by flow cytometry. CONCLUSIONS: Taken together, this work suggests that it is possible to detect MM markers in EVs from either BM or PB of MM patients and compare their expression at different stages of the disease (diagnosis and remission after ASCT). Importantly, our results demonstrate the importance and potential of analyzing EVs cargo from PB, suggesting the possibility of using them for minimally invasive monitoring of MRD in MM patients. ACKNOWLEDGEMENTS: The authors acknowledge Celgene/BMS for providing funding to this work (Project Looker - Grant_138800). The authors acknowledge Cytogenetics Laboratory, Department of Clinical Hematology, Centro Hospitalar e Universitário São João and Flow Cytometry Laboratory, Department of Clinical Pathology, Centro Hospitalar e Universitário São João. Disclosures Bergantim: Amgen: Consultancy, Research Funding, Speakers Bureau; Janssen: Consultancy, Speakers Bureau; BMS: Consultancy, Research Funding, Speakers Bureau; Takeda: Consultancy, Speakers Bureau. Barbosa: BMS: Research Funding. Silva: BMS: Research Funding. Polónia: BMS: Research Funding. Caires: BMS: Research Funding. Guimarães: BMS: Research Funding; Amgen: Research Funding. Vasconcelos: BMS: Research Funding; Amgen: Research Funding.

Chemostat-like microfluidic platform for highly sensitive detection of heavy metal ions using microbial biosensors

2015 ◽  
Vol 65 ◽  
pp. 257-264 ◽  
Author(s):  
Minseok Kim ◽  
Ji Won Lim ◽  
Hyun Ju Kim ◽  
Sung Kuk Lee ◽  
Sang Jun Lee ◽  
...  
Keyword(s):  
Heavy Metal ◽  
Metal Ions ◽  
Heavy Metal Ions ◽  
Sensitive Detection ◽  
Microfluidic Platform ◽  
Highly Sensitive ◽  
Highly Sensitive Detection

scholarly journals Synthesis of bismuth nanoparticle-loaded cobalt ferrite for electrochemical detection of heavy metal ions

RSC Advances ◽  
2020 ◽  
Vol 10 (46) ◽  
pp. 27697-27705 ◽  
Author(s):  
Ying He ◽  
Zihan Wang ◽  
Li Ma ◽  
Liya Zhou ◽  
Yanjun Jiang ◽  
...  
Keyword(s):  
Heavy Metal ◽  
Metal Ions ◽  
Electrochemical Detection ◽  
Electrode Material ◽  
Heavy Metal Ions ◽  
Cobalt Ferrite ◽  
Bismuth Nanoparticles ◽  
Highly Sensitive ◽  
Sensitive Sensor ◽  
Photocatalytic Material

As an efficient modified electrode material for the detection of heavy metal ions, bismuth nanoparticles (BiNPs) were loaded on cobalt ferrite (CoFe2O4), a unique magnetic photocatalytic material, to fabricate a highly sensitive sensor.

scholarly journals Allogeneic Transplant Can Abrogate the Relapse Risk in the Patients with Detectable Measureable Residual Disease By Multicolor Flow-Cytometry at the Time of Assessment of Acute Myeloid Leukemia Patients in First Remission

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 36-37
Author(s):  
Muhned Alhumaid ◽  
Georgina S. Daher-Reyes ◽  
Arjun Law ◽  
Auro Viswabandya ◽  
Armin Gerbitz ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Research Funding ◽  
Residual Disease ◽  
Response To Therapy ◽  
Time Dependent ◽  
Time Interval ◽  
Relapse Risk ◽  
Acute Myeloid

BACKGROUND: Acute myeloid leukemia (AML) is a highly heterogeneous group of diseases with variable response to therapy. Several factors have a prognostic impact for an outcome. Despite intensive chemotherapy and hematopoietic stem cell transplant (HCT), a significant proportion of patients eventually relapse, indicating that morphological assessment is not adequate due to limitations in sensitivity, requiring a better tool for assessment of remission. METHODS: A retrospective analysis was performed in AML patients who achieved first complete remission (CR1) and the outcomes compared according to the performance of HCT, and multi-color flow cytometry (MFC)-based measurable residual disease (MRD) status (defined as negative if patients achieved 0.1% or less) assessed at the time of CR1. In order to take account of the time interval from the MFC-MRD assessment to HCT, we applied a Mantel-Byar test for overall (OS) and relapse-free survival (RFS), considering time-to-HCT as a time-dependent covariate, while Simon and Makuch plot was used. Time-dependent Cox proportional hazard models were applied for multivariate analysis. Cumulative incidence of relapse (CIR) and non-relapse mortality (NRM) where evaluated using Fine-Gray model. RESULTS: A total of 435 patients diagnosed with AML and treated with induction chemotherapy between 2015 and 2018, of whom 380 patients (87%) achieved remission, were included. MFC-MRD was assessed in 336 patients in CR1 (77%), out of 380 patients who achieved CR1, and 200 patients (53%) proceeded to HCT. We evaluated OS, RFS, CIR and NRM according to MFC-MRD status in those patients who had negative MRD (MRDneg; n=218, 65%) vs. those with MRD (MRDpos; n=118, 35%). The OS at 2 years was 67.0% vs.40.7% (p≤0.001), RFS at 2 years was 8.7% vs. 40.6% (p≤0.001), CIR 26.9% vs.21.1% but with borderline significance (p=0.08), and NRM 32.5% vs. 20.2% with borderline significance (p=0.057). In patients who achieved CR, we compared OS, CIR, NRM and RFS between the HCT group (n=200) vs. those who did not undergo HCT (no-HCT; n=235). Between the 2 groups, the OS at 2 years was 55.7% vs. 47.2% (p=0.004); CIR 9.7% vs. 34.6% (p≤0.001); NRM 40.9% vs. 12.6% (p≤0.001). There was no difference in RFS: 49.4% vs. 52.8% (p=0.505). There was no difference in the time interval from the MFC-MRD assessment to HCT between the groups (MRDpos vs MRDneg) with a median of 96 days in overall patients who received HCT (p=0.31). In the overall population, when HCT was accounted as a time-dependent covariate, we failed to observe any difference of OS (HR 1.23; p=0.19) or RFS (HR 1.09; p=0.60) between the HCT vs. no-HCT groups. Then, we compared the OS, RFS, CIR, and NRM between the HCT vs no-HCT groups confined to the subgroups of patients with MFC-MRDneg vs MFC-MRDpos, separately. In the MFC-MRDpos subgroup, patients who underwent HCT did better: OS 54.8% in HCT vs. 25.5% in no-HCT (HR 0.52; p≤0.001) and RFS 48.7% vs. 24.1% (HR 0.45; p≤0.001). However, in the MFC-MRDneg subgroup, similar outcomes were noted between the HCT vs no-HCT groups in terms of OS 60.8% vs. 70.7% (HR 1.27, p=0.32), RFS 51.6% vs. 62.4% (HR 1.25; p=0.46) (Fig 1). With respect to the cause of treatment failure according to treatment modality (HCT vs no-HCT) and MFC-MRD status, Fig 2 revealed different patterns of relapse vs NRM between the HCT and the no-HCT groups. In the MFC-MRDneg subgroup, HCT group showed a higher NRM over the no-HCT group (38.0% vs 8.7%; HR 2.08; p≤0.001), while relapse risk was lower in the HCT group (10.4% vs 29.3%; p≤0.001). In the MFC-MRDpos subgroup, relapse incidence was strikingly different in favor of HCT (9.5% vs 50.0%; HR, p≤0.001). Conclusion: These findings suggest that in AML patients HCT could abrogate the relapse risk in patients who are MFC-MRDpos at the time of remission assessment, while the benefit from HCT was minimal in the subgroup that are MFC-MRDneg. Further study is strongly warranted to reach a clearer conclusion with a larger number of cohorts. Disclosures Lipton: Bristol-Myers Squibb: Honoraria; Takeda: Consultancy, Honoraria, Research Funding; BMS: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Pfizer: Consultancy, Honoraria, Research Funding; Ariad: Consultancy, Research Funding.

scholarly journals Combination of Flow Cytometry and Functional Imaging for Monitoring of Residual Disease in Myeloma

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 3185-3185
Author(s):  
Leo Rasche ◽  
Daisy Alapat ◽  
Manoj Kumar ◽  
Grant Gershner ◽  
James E McDonald ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Flow Cytometry ◽  
Functional Imaging ◽  
Late Stage ◽  
Salvage Therapy ◽  
Research Funding ◽  
Residual Disease ◽  
Background Suppression ◽  
Imaging Method ◽  
Medical Sciences

Abstract Introduction The iliac crest is the usual sampling site for minimal residual disease (MRD) monitoring in Multiple Myeloma (MM). However, the disease distribution in the bone marrow (BM) is often heterogeneous. Functional imaging can be used to complement MRD detection at a single site, thereby accounting for asymmetrically distributed disease. Diffusion weighted MRI with background suppression (DWIBS) is a novel functional imaging method that can detect disease in a higher proportion of newly diagnosed MM (NDMM) patients than 18F-fluorodeoxyglucose positron emission tomography (PET), as it is independent of the tumor metabolism. Yet, its performance for monitoring of residual disease has not been described. The aims of this study were 1) to compare DWIBS to PET for the detection of residual disease in patients achieving complete remission (CR), and 2) to test whether DWIBS and PET could complement MRD flow cytometry with a sensitivity of 1x10-5. To address these aims, we investigated 168 NDMM and 33 relapsed patients for whom DWIBS, PET, and MRD were available at the onset of CR during first-line and salvage therapy, respectively. Methods All patients signed written consent in accordance with the Declaration of Helsinki. Residual focal lesions (FLs) were defined as well delineated focal intensities above the surrounding BM background. For DWIBS FLs were considered if restriction could be confirmed on ADC maps. 8-color MRD flow cytometry with a limit of detection of 1x10-5 was available for 83 NDMM and all 33 salvage therapy patients. The Kaplan-Meier method was used for survival analyses. PFS time was measured from onset of CR to relapse or death from any cause or censored at the date of last contact. Paired-end whole exome sequencing of CD138-enriched MM cells was performed on an Illumina HiSeq 2500. Mutations were called from BWA aligned sequencing reads using MuTect. Subclonal reconstruction was done using SciClone. Results Compared to PET, DWIBS detected more CR patients with residual FLs (21% vs. 6%), and the concordance between PET and DWIBS was low. Only 6 of the DWIBS-positive patients also presented with FLs in PET. Yet, 5 patients had PET+/DWIBS- FLs, suggesting that the two techniques are complementary. Both, DWIBS+ and PET+ FLs negatively impacted PFS (p<0.05). For 83 patients MRD data were available. Combining MRD and imaging, residual disease was detectable in 53 patients (64%). The best outcome was seen for 30 double negative (MRD-/Imaging-) patients (3 events with a median follow-up of 3.6 years), the worst outcome was seen for 10 double positive (MRD+/Imaging+) patients (median PFS: 2.1 years). Only 4 of 86 patients were MRD-/Imaging+, indicating that residual FLs are rare in MRD-negative NDMM patients at a sensitivity of 1x10-5. A heterogeneous disease distribution is a common feature of late-stage patients. To test if this increased heterogeneity confounded MRD, we investigated a set of 33 heavily pretreated patients who achieved CR during salvage therapy. Combining MRD and imaging data, we detected residual disease in 25 patients (76%). Of note, the proportion of patients, who were MRD-negative but had residual FLs on functional imaging was significantly higher compared to NDMM (8/16 vs 4/34 patients, p=0.01). At the same time, 10 patients (30%) were MRD+ but Imaging-, supporting the idea that a combined MRD/Imaging approach can improve detection of residual disease and should be used in late-stage patients. To obtain insights in the underlying biology, we performed longitudinal multi-region sequencing of a subset of these CR patients. Our findings support the concept of persistence and progression of multiple spatially separated clones in the BM irrespective of being in an MRD-negative CR. Thereby, focal residual disease could be shown to contribute to relapse. Conclusion DWIBS is a promising tool for detection of residual disease and complements PET. The combination of MRD diagnostics and functional imaging improves prediction of outcome, with double-negativity and double positivity defining groups with excellent and dismal PFS, respectively. Prospective trials using this information to tailor therapy are warranted. From a biological perspective, this study highlights the confounding effects of spatial heterogeneity and limited dissemination of clones within the BM on MRD diagnostics. This may especially be true for patients achieving deep responses during salvage therapies. Disclosures Roy Choudhury: University of Arkansas for Medical Sciences: Employment, Research Funding. Epstein:University of Arkansas for Medical Sciences: Employment. Barlogie:International Workshop on Waldenström's Macroglobulinemia: Other: travel stipend; Millenium: Consultancy, Research Funding; Multiple Myeloma Research Foundation: Other: travel stipend; ComtecMed- World Congress on Controversies in Hematology: Other: travel stipend; Myeloma Health, LLC: Patents & Royalties: : Co-inventor of patents and patent applications related to use of GEP in cancer medicine licensed to Myeloma Health, LLC; European School of Haematology- International Conference on Multiple Myeloma: Other: travel stipend; Celgene: Consultancy, Research Funding; Dana Farber Cancer Institute: Other: travel stipend. Davies:Takeda: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Janssen: Consultancy, Honoraria. Morgan:Celgene: Consultancy, Honoraria, Research Funding; Bristol-Myers Squibb: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Janssen: Research Funding.

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