scholarly journals Assessment of Minimal/Measurable Residual Disease Testing in Acute Myeloid Leukaemia By Molecular Methods in an Interlaboratory Study

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 3461-3461
Author(s):  
Stuart Scott ◽  
Richard Dillon ◽  
Christian Thiede ◽  
Sadia Sadiq ◽  
Ashley Cartwright ◽  
...  
Keyword(s):  
Cell Line ◽  
Board Of Directors ◽  
Research Funding ◽  
Residual Disease ◽  
Standard Of Care ◽  
Negative Sample ◽  
Research Support ◽  
Advisory Committees ◽  
Log Reduction ◽  
Measurable Residual Disease

Abstract Background Minimal/measurable residual disease (MRD) testing is increasingly utilised and accepted as standard of care to manage a range of different haematological malignancies. It's use as a surrogate outcome in clinical trials of new therapies is being explored, where it has the potential to accelerate drug assessment and approval. The phenotypic and genetic heterogeneity of acute myeloid leukaemia (AML) has limited the use of MRD in this context; however, the European LeukaemiaNet (ELN) MRD working group have recently published consensus guidelines to standardise both flow cytometric and molecular genetic MRD testing. To assess the accuracy of testing and concordance between laboratories, crucial to patient safety, external quality assessment (EQA)/proficiency testing (PT) is required. Aims To determine the performance of molecular methods for measuring of MRD using the t(8;21)(q22:q22) RUNX1-RUNX1T1, inv(16)(p13q22) CBFB-MYH11, t(15;17)(q24.1;q21.2) PML-RARA and NPM1 Type A markers in an international interlaboratory study. Methods A total of 12 batches of lyophilised EQA material were manufactured. These consisted of three batches of samples for each marker all containing 9x10^6 cells: an MRD 'high' sample; an MRD 'low' sample; and an MRD 'negative' sample. The t(8;21)(q22:q22) RUNX1-RUNX1T1 positive samples were manufactured using the KASUMI-1 cell line, the inv(16)(p13q22) CBFB-MYH11 positive samples using the ME-1 cell line; t(15;17)(q24.1;q21.2 PML-RARA positive samples using the NB4 cell line and the NPM1 Type A (NM_002520.6:c.860_863dup) positive samples using the OCI-AML3 cell line. MRD positive samples were diluted with HL60 cells to achieve the desired MRD level. MRD negative samples were manufactured using the HL60 cell line. The samples were shipped at ambient temperature to the 29 laboratories in 12 countries. Participants were asked to test the blinded samples with their in-house assay and report % normalised ratio of the relevant marker alongside additional methodological and technical data. Results For t(8;21) RUNX1-RUNX1T1, all participants who returned results (n=23) classified the MRD 'high' and MRD 'low' samples as positive and the MRD 'negative' sample as negative. The robust mean log reduction between the MRD 'high' and MRD low sample was 2.7 (range 2.5-2.9). For inv(16) CBFB-MYH11, all participants who returned results (n=22) classified the MRD 'high' sample as positive, 21/22 (95.5%) classified the MRD 'low' sample as positive and 21/22 (95.5%) classified the MRD negative sample as negative. The robust mean log reduction between the MRD 'high' and MRD 'low' sample was 3.16 (range 2.8-4.2). For t(15;17) PML-RARA, all participants who returned results (n=22) classified the MRD 'high' sample as positive, 21/22 (95.5%) classified the MRD 'low' sample as positive and 21/22 (95.5%) classified the MRD negative sample as negative. The robust mean log reduction between the MRD 'high' and MRD 'low' sample was 2.1 (range 1.4-2.4). For NPM1, all participants who returned results (n=23) classified the MRD 'high' as positive, 21/23 (91.3%) classified the MRD 'low' sample as positive and, 17/23 (73.4%) classified the MRD negative sample as negative. The robust mean log reduction between the MRD 'high' and MRD 'low' sample was 3.8 (range 3.2-4.2). Summary/Conclusion The majority of participants in this study were able to detect and accurately quantify MRD when assessing the t(8;21)(q22:q22) RUNX1-RUNX1T1, inv(16)(p13q22) CBFB-MYH11, t(15;17)(q24.1;q21.2) PML-RARA and NPM1 markers, at levels that would be expected within a clinical trial or standard of care setting. A high proportion of participants reported false positive results in the NPM1 marker negative sample. This would have significant consequences clinically, with NPM1 marker false-positivity potentially committing patients to unneeded additional chemotherapy and/or transplant with the attendant risk of morbidity and mortality which highlights the need for ongoing EQA in this area. UK NEQAS LI will work with laboratories advocating they undertake a root cause analysis process to identify the source(s) of error contributing to false positive NPM1 marker results and support their subsequent corrective actions; sharing any educational findings with all participants. Figure 1 Figure 1. Disclosures Scott: Novartis: Research Funding; Biorad: Research Funding. Dillon: Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Research Support, Educational Events; Amgen: Other: Research support (paid to institution); Astellas: Consultancy, Other: Educational Events , Speakers Bureau; Menarini: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Other: Session chair (paid to institution), Speakers Bureau; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: educational events; Jazz: Other: Education events; Shattuck Labs: Membership on an entity's Board of Directors or advisory committees. Whitby: Roche: Membership on an entity's Board of Directors or advisory committees; Alexion: Honoraria, Other: Teaching.

scholarly journals NGS-Based Monitoring of Measurable Residual Disease in FLT3-ITD Positive Acute Myeloid Leukemia

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 2778-2778
Author(s):  
Tamara J. Blätte ◽  
Laura K. Schmalbrock ◽  
Sabrina Skambraks ◽  
Sibylle Cocciardi ◽  
Anna Dolnik ◽  
...  
Keyword(s):  
Cell Line ◽  
Board Of Directors ◽  
Myeloid Leukemia ◽  
Research Funding ◽  
Residual Disease ◽  
High Sensitivity ◽  
Analysis Program ◽  
Advisory Committees ◽  
High Coverage ◽  
Measurable Residual Disease

Abstract Background: In acute myeloid leukemia (AML), most patients respond to chemotherapy initially, but the risk of relapse remains high. Continued monitoring of measurable residual disease (MRD) following therapy onset enables early assessment of treatment response and clinical intervention. However, while appropriate assays exist for several genetic markers, some of the most common aberrations in AML, FLT3 internal tandem duplications (FLT3-ITDs), have remained a challenging target due to insertion site and length heterogeneity. The established diagnostic assay based on PCR and fragment analysis (FA), on the other hand, lacks sufficient sensitivity. In consequence, extensive monitoring of FLT3-ITD MRD in AML patients over time and treatment is not yet available. At the same time, with the approval of the FLT3-targeting tyrosine kinase inhibitor (TKI) midostaurin, the need for detailed monitoring of FLT3-ITD clones has increased. Aims: We have developed a new next-generation sequencing (NGS) based assay and analysis program to detect FLT3-ITDs with high sensitivity and specificity. We aimed to demonstrate our method's applicability to MRD monitoring and evaluated the progression of FLT3-ITD MRD in patients treated with midostaurin within the AMLSG 16-10 trial. Methods: FLT3 exons 14 and 15 were PCR-amplified and sequenced on the Illumina MiSeq to generate at least 1 million 250 bp paired-end reads. ITDs were detected using our novel analysis program that performs all steps of the analysis, does not require any manual filtering and reports ITDs extensively annotated to the user. To assess our assay's concordance with established methods, FLT3-ITDs in all diagnosis and relapse samples were characterized by FA and Sanger sequencing (SSeq). Results: Assay specificity was confirmed with 0 ITDs reported in four FLT3-ITD⁻ control samples (healthy volunteers n=2, AML patient n=1, AML cell line HL60 n=1). To assess assay sensitivity, we serially diluted DNA of the ITD⁺ AML cell line MOLM14 in DNA of the ITD⁻ AML cell line HL60. The expected 21 bp ITD was detected in all four dilutions, down to a variant allele frequency (VAF) of 6.7x10⁻⁵ (0.0067 %). We further processed matched diagnosis and relapse samples from 9 ITD⁺ AML patients at low coverage (mean: 430000 paired-end reads) and confirmed concordance of ITD site, length (range 21-198 bp) and VAF with results from FA/SSeq. For 4/9 patients, we independently sequenced a total of 12 matched diagnosis, remission and relapse samples to our target coverage of 1.5 million paired-end reads. In all of these, ITDs detected by FA were confirmed and ITD site, length and VAF were again concordant between FA/SSeq and our NGS-based approach; results from the diagnosis and relapse samples sequenced to both low and high coverage were highly reproducible. In 11/12 samples sequenced to high coverage, our assay identified additional ITD clones that were not identified by FA: The mean number of ITDs detected was 4.8 at diagnosis (1.5 by FA), 2.0 at clinical remission (not measured by FA) and 1.5 at relapse (0.5 by FA). VAFs of these ITDs ranged from 0.007-38 % (mean 5.3 %); ITDs that were solely detected by our NGS-assay were present at lower VAFs than those also identified by FA/SSeq (NGS-assay only: mean 0.21 %, range 0.007-2 %; also detected by FA/SSeq: mean 21.4 %, range 2-38 %). About half of the ITDs detected by NGS at diagnosis (11/19 ITD clones) were not detected at any of the later time points and thus presumably eradicated by treatment. Conversely, 8/19 ITD clones persisted at remission (6/19) or reoccurred at relapse (2/19). In fact, nearly all of the ITD clones present at relapse were already identified at an earlier time point. Conclusion: In sum, we could demonstrate that our NGS-based assay and analysis program achieve high accuracy and precision and enable MRD monitoring in FLT3-ITD positive AML patients. Its high sensitivity enables the detection of additional ITD clones occurring at low allelic frequencies not only at remission but also at diagnosis and relapse. Further evaluation of MRD monitoring in FLT3-ITD positive AML patients treated with midostaurin within the AMLSG 16-10 trial is currently ongoing. Disclosures Bullinger: Bristol-Myers Squibb: Speakers Bureau; Pfizer: Speakers Bureau; Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Bayer Oncology: Research Funding; Amgen: Honoraria, Speakers Bureau; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Janssen: Speakers Bureau; Sanofi: Research Funding, Speakers Bureau.

scholarly journals Venetoclax and Azacitidine for Older Newly Diagnosed Patients with Acute Myeloid Leukemia: A Single-Institution Pilot Study Using Measurable Residual Disease to Guide Therapy

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 2638-2638 ◽  
Author(s):  
Amanda Winters ◽  
Jonathan A Gutman ◽  
Enkhtsetseg Purev ◽  
Brett M. Stevens ◽  
Shanshan Pei ◽  
...  
Keyword(s):  
Board Of Directors ◽  
Myeloid Leukemia ◽  
Research Funding ◽  
Residual Disease ◽  
Refractory Disease ◽  
Advisory Committees ◽  
Ras Pathway ◽  
Measurable Residual Disease ◽  
Count Recovery

Background: Venetoclax (ven) was approved for older untreated acute myeloid leukemia (AML) patients due to high response rates and durable remissions. As a participating site in the dose escalation study, we observed deeper/more durable responses in some who received >400mg ven. We also noted 16/33 discontinued azacitidine (aza) after achieving a response; 9 relapsed and 7 remained in long term remission on ven only. Based on these observations, we designed a study that hypothesized: A)Higher initial doses of ven would allow deeper/more durable responses, and B)Multi modality high sensitivity measurable residual disease (MRD) testing could identify patients able to discontinue aza and remain on maintenance ven. Methods: This is an ongoing phase 2 study (NCT03466294) of 42 untreated AML patients ≥60 who decline/are ineligible for induction. Patients have adequate organ function and white blood cell counts <25x109/L (hydrea permitted). In cycle 1, patients receive aza 75mg/m2 on days (d) 1-7 and ven, escalated from 100 to 200 to 400 to 600mg on d 1-4. Ven continues at 600mg d 5-28 and bone marrow biopsies (BMBXs) are performed on d 8 and 28. Patients who achieve morphologic remission without count recovery have up to 14 days off therapy before subsequent cycles, with growth factor support; "upgraded" responses are recorded if count recovery occurs. Non responders discontinue or receive up to two additional cycles of aza and ven 600mg. Responders who remain MRD+ by multiparameter flow cytometry (MPFC, Hematologics) and/or digital droplet PCR (ddPCR) for as many identifiable diagnostic genes as possible also receive up to 2 additional cycles of aza and ven 600mg. MRD+ responders after 3 cycles continue aza and ven 400mg until toxicity/progression. Patients who experience MRD- responses at any time stop aza and continue ven 400mg daily (Fig 1). Results: 30 patients enrolled between May 2018 and July 2019; median age is 71 (60-88), 10% evolved from MDS and 10% and 73% had intermediate and unfavorable risk disease by ELN, respectively (Table 1). 732 adverse events (AEs) occurred; 46 (6%) were serious, the most common were neutropenic fever (37%) and pneumonia (13%). The most common >grade 2 related AEs were leukopenia (53%), thrombocytopenia (44%) and neutropenia (35%); there were no related grade 5 AEs. The overall response rate was 70% (21/30; CR=19, MLFS=2). Median number of cycles to achieve best response was 1. Significant blast reductions were seen on day 8; of the 28 with interpretable day 8 BMBXs, 10 achieved MLFS on day 8. 4 completed ≥1 cycle and were refractory. An additional 4 did not complete cycle 1: 1 died of disease and 3 elected to come off therapy (all subsequently died of disease). Four (19%) responders relapsed, after a median 180 days (27-279). With median follow up of 214 days, median response duration has not been reached. 10 patients died, after a median 65 days (29-256); 1/30 died within 30 days. Median overall survival has not been reached. Of the 26 who completed ≥1 cycle, 19 were MRD- by MPFC, including 18/19 who achieved CR. Of these 26, 3 were not monitored by ddPCR: for 2 patients this was due to the absence of detectable baseline mutations and for 1 patient it was due to refractory disease. The remaining 23 had ddPCR monitoring; 3 became MRD- by this modality (Fig 2). All 3 were also MRD- by MPFC and per protocol discontinued aza and initiated ven maintenance (Fig 1). MRD negativity by both parameters occurred after cycles 1, 2 and 3, respectively. One MRD- patient relapsed after 216 days; two remain in remission after 301 and 124 days. An additional 4 who achieved MRD+ responses discontinued aza at their insistence (and in violation of the protocol); 1 relapsed after 279 days, and 3 remain in ongoing remission. Univariate predictors of refractory disease were FAB M0/M1 (OR 0.070, p=0.02) and RAS pathway mutations (OR 14.25, p=0.02). Conclusions: Higher initial doses of ven are tolerated in this population. Blast reduction occurs quickly in many patients (day 8), for this low intensity regimen. Response rates are consistent with lower doses of ven. Very deep responses, as measured by highly sensitive MRD methods (MPFC and ddPCR are capable of sensitivity up to 0.02%), are attainable. Longer follow up time will determine if higher ven doses and MRD-driven decisions related to continuation of aza result in more durable responses. Increased maturation of blasts and RAS pathway mutations are predictors for refractory disease. Disclosures Lyle: Pfizer: Membership on an entity's Board of Directors or advisory committees; Daiichi Sankyo Incyte: Membership on an entity's Board of Directors or advisory committees; Agios: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees. Pollyea:Agios: Consultancy, Membership on an entity's Board of Directors or advisory committees; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees; Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celyad: Consultancy, Membership on an entity's Board of Directors or advisory committees; Diachii Sankyo: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees; Forty-Seven: Consultancy, Membership on an entity's Board of Directors or advisory committees.

scholarly journals A Prospective Phase 2 Study of Venetoclax and Low Dose Ara-C (VALDAC) to Target Rising Molecular Measurable Residual Disease and Early Relapse in Acute Myeloid Leukemia

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 1261-1261
Author(s):  
Ing S Tiong ◽  
Sun Loo ◽  
Emad Uddin Abro ◽  
Devendra Hiwase ◽  
Shaun Fleming ◽  
...  
Keyword(s):  
Board Of Directors ◽  
Myeloid Leukemia ◽  
Research Funding ◽  
Residual Disease ◽  
Phase 2 ◽  
Advisory Committees ◽  
Early Relapse ◽  
Phase 2 Study ◽  
Measurable Residual Disease ◽  
Acute Myeloid

Abstract Introduction Rising molecular measurable residual disease (MRD) is an arbiter of clinical relapse in acute myeloid leukemia (AML). Venetoclax (VEN) is active against IDH and NPM1 mutant (mt) AML as monotherapy (Konopleva et al, 2016 and Chua et al, 2020) and can yield MRD negative remission when combined with low dose ara-C (LDAC) in patients unfit for intensive chemotherapy (DiNardo and Tiong et al, 2020). In a retrospective study, we showed that VEN in combination with hypomethylating agents or LDAC could erase rising NPM1mt MRD in 6/7 cases (Tiong et al, 2020). We now present a prospective phase 2 study of VEN and LDAC in patients with molecular MRD failure or oligoblastic AML relapse. Methods This multicenter phase 2 study stratified patients into oligoblastic relapse (marrow blasts 5-15%; Group A), or molecular MRD failure (Group B) as defined by the European LeukemiaNet (ELN) recommendations (failure confirmed by 2 interval samples) (Schuurhuis et al, 2018). Patients received VEN 600 mg (days 1-28) and LDAC 20 mg/m 2 (days 1-10). Primary objectives were morphologic or MRD response (≥1 log reduction) in groups A and B, respectively. Key secondary objectives were allogeneic hematopoietic cell transplantation (allo-HCT) realization and relapse-free (RFS) and overall survival (OS). The study had Alfred Health ethics approval (196/19). NPM1mt and other fusion transcript levels (per 10 5 ABL) from bone marrow were analyzed by RT-qPCR, IDH1 and IDH2 by Bio-Rad TM droplet digital PCR. Results The study enrolled 32 patients, with 29 evaluable (cut-off date 15/7/21). The median age of the study population was 62 years; 79% had intermediate cytogenetic risk, 66% NPM1mt, 11% FLT3-ITD and 37% IDH1/IDH2 mt. Most received prior intensive chemotherapy (93%) and 2 (7%) allo-HCT in first remission. Median interval from AML diagnosis to study entry was 12.6 months (Table 1). After a median follow-up of 7.9 months, patients had received a median of 3 cycles (range 1-14) of VEN-LDAC, with 13 patients ongoing. The main reasons for treatment cessation were allo-HCT (n=10; 34%) or donor lymphocyte infusion (n=2; 7%), treatment failure (n=3) or an adverse event (n=1). Hematologic complete/incomplete response (CR/CRi) among 11 patients with oligoblastic relapse (group A) was 73% and included: CR (n=5, 45%) or CRi (n=3, 27%), with an additional patient with morphologic leukemia-free state and 2 patients with stable disease. Overall, across both groups, median RFS and OS were not reached, estimated at 78% and 91% at 1 year, respectively. Among 18 patients with molecular MRD failure (group B) treated with VEN+LDAC, molecular response (≥1 log reduction) was achieved in 72%, and the RFS and OS were estimated at 83% and 87% at 1 year, respectively. Analysis of a sub-group of patients with NPM1mt (n=18); 6 and 12 from Groups A and B, respectively revealed the median NPM1mt transcript level at study entry to be 8985 copies (IQR 826, 94,431). A molecular response was achieved in 14 (78%) patients, including 9 (50%) with complete molecular remission (CR MRD-), with most responses achieved within 2 cycles of therapy (Figure B). Treatment with VEN-LDAC was generally well tolerated, with 15 serious adverse events reported within the first 2 cycles, including infection (n=6; 19%) and febrile neutropenia (n=3; 9%). Only one subject discontinued treatment due to stroke. Conclusions In this prospective study, in patients with first oligoblastic relapse or MRD failure, VEN in combination with LDAC induced a high rate of molecular MRD remission that was rapidly achieved, resulting in a high rate of survival at 12-months (&gt;90%) and with low toxicity. Follow-up is ongoing to determine the durability of response. Treatment of patients with MRD or early clinical failure may represent an attractive clinical trial setting for investigation of novel, non-intensive AML therapies. This approach will be investigated in a future multi-arm, precision-based platform trial called INTERCEPT (Investigating Novel Therapy to Target Early Relapse and Clonal Evolution as Pre-emptive Therapy in AML). Figure 1 Figure 1. Disclosures Tiong: Servier: Consultancy, Speakers Bureau; Amgen: Speakers Bureau; Pfizer: Consultancy. Hiwase: Novartis: Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees. Fleming: Amgen Inc: Research Funding. Bajel: Amgen: Speakers Bureau; Abbvie, Amgen, Novartis, Pfizer: Honoraria. Fong: Amgen, BMS: Speakers Bureau; Amgen: Research Funding; AbbVie, Amgen, Novartis, Pfizer, Astellas: Honoraria. Wei: Celgene/BMS: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Agios: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Genentech: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Astellas: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Astra Zeneca: Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees; Macrogenics: Membership on an entity's Board of Directors or advisory committees; Servier: Membership on an entity's Board of Directors or advisory committees, Research Funding. OffLabel Disclosure: This presentation will discuss the use of venetoclax in targeting measurable residual disease and early relapse of acute myeloid leukemia.

scholarly journals Comparison of Acute Myeloid Leukemia Measurable Residual Disease Detection By Flow Cytometry in Peripheral Blood and Bone Marrow

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 2729-2729
Author(s):  
Colin D. Godwin ◽  
Yi Zhou ◽  
Megan Othus ◽  
Carole M. Shaw ◽  
Kelda M. Gardner ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Board Of Directors ◽  
Correlation Coefficient ◽  
Myeloid Leukemia ◽  
Research Funding ◽  
Residual Disease ◽  
Spearman Correlation ◽  
Advisory Committees ◽  
Measurable Residual Disease ◽  
Acute Myeloid

BACKGROUND: The current recommendation against the need for bone marrow aspiration (BMA) in routine follow-of persons with acute myeloid leukemia (AML) in remission preceded the recognition that multiparameter flow cytometry (MFC) is a sensitive and specific means to detect imminent morphologic relapse. Given this recognition, we wondered whether BMA is now necessary, or if concordance between MFC results in peripheral blood (PB) and BMA is such as to make BMA unnecessary, at least for evaluation of measurable residual disease (MRD) by MFC. Previous studies have demonstrated a strong correlation between disease detection by MFC in PB and BMA. Here we examined 724 paired PB and BMA samples from 482 patients to further examine the concordance between PB and BMA blast detection by MFC, particularly among patients in morphologic remission. PATIENTS AND METHODS: We included adults in our institutional AML database, covering 2008-2018. Our Hematopathology database was queried to identify PB and BMA MFC sample pairs with samples considered "paired" if measured within one week of each other. If an individual had multiple pairs, all were included unless otherwise specified. Ten-color MFC was performed routinely on BMA aspirates with a panel of three antibody combinations, with the same antibody combinations applied to PB samples. When identified, the abnormal population was quantified as a percentage of the total CD45+ white cell events. Any level of residual disease was considered positive. Complete remission (CR) and relapse were defined according to the European LeukemiaNet 2017 classification. Relationship between PB and BMA blast % was measured using Spearman's Rank-Order Correlation. Relationship between PB and BMA samples identified as positive or negative is illustrated using 2 X 2 tables (Table 1). RESULTS: Considering all 724 sample pairs, the Spearman correlation coefficient between PB and BMA blast percentage was 0.93, and was 0.91 considering only the first sample pair for each individual patient (n= 482). 315 sample pairs were positive by PB, 97% of which were also positive by BMA while 95% of 409 pairs negative by PB were also negative by BMA. Similar results were seen considering only a patient's first pair. Restricting analysis to patients with pairs obtained between the dates of CR and relapse, the Spearman correlation coefficient was 0.82 with 91% of 35 cases positive in PB also positive in BMA; 93% of 114 pairs negative in PB were also negative in marrow. As a complementary means to compare pairs when AML burden was low, we examined only pairs where the BMA MFC showed <5% blasts. Here, the Spearman correlation coefficient between PB and BMA blasts was 0.83. 90% of 70 positive PB cases were also positive by BMA while 95% of 295 negative PB cases were also negative by BMA. Examining pairs taken from patients in morphologic remission immediately prior to undergoing hematopoietic cell transplant yielded a Spearman correlation coefficient of 0.92, with all 9 PB positive cases also being positive in BMA and 96% of PB negative cases being negative in BMA. CONCLUSIONS: This is the largest cohort of AML PB and BMA sample pairs analyzed by MFC to-date. The percentages of blasts measured in PB and BMA are strongly correlated. In the 365 pairs from patients with MRD-level disease, the predictive value of PB MFC positivity for BMA positivity was 90% (63/70) while the predictive value PB MFC negativity for BMA negativity was 95%. Disclosures Othus: Glycomimetics: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees. Gardner:Abbvie: Speakers Bureau. Walter:BioLineRx: Consultancy; BiVictriX: Consultancy; Boehringer Ingelheim: Consultancy; Boston Biomedical: Consultancy; Covagen: Consultancy; Daiichi Sankyo: Consultancy; Kite Pharma: Consultancy; New Link Genetics: Consultancy; Pfizer: Consultancy, Research Funding; Race Oncology: Consultancy; Seattle Genetics: Research Funding; Argenx BVBA: Consultancy; Aptevo Therapeutics: Consultancy, Research Funding; Jazz Pharmaceuticals: Consultancy; Astellas: Consultancy; Agios: Consultancy; Amgen: Consultancy; Amphivena Therapeutics: Consultancy, Equity Ownership.

scholarly journals Clonal Hematopoiesis and Its Implications for Flow Cytometric Assessment of Measurable Residual Disease in Patients with NPM1-mutated Acute Myeloid Leukemia

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 38-39
Author(s):  
Sanam Loghavi ◽  
Courtney D. DiNardo ◽  
Koichi Takahashi ◽  
Rashmi Kanagal-Shamanna ◽  
Tomoyuki Tanaka ◽  
...  
Keyword(s):  
Board Of Directors ◽  
Myeloid Leukemia ◽  
Research Funding ◽  
Residual Disease ◽  
Advisory Committees ◽  
Clonal Hematopoiesis ◽  
Hla Dr ◽  
Measurable Residual Disease ◽  
Acute Myeloid ◽  
Myeloid Progenitors

INTRODUCTION: NPM1 mutations (NPM1mut) occur in ∼30% of acute myeloid leukemia (AML) and frequently co-occur with mutations in other genes including those attributed to clonal hematopoiesis (CH) including DNMT3A and TET2, among others. CH mutations may persist beyond attaining NPM1mut-negative remission. Persistent CH may be associated with immunophenotypic alterations in myeloid progenitors detected by flow cytometry (FC) which poses an interpretive challenge in assessment of measurable residual disease (MRD) by FC. The aim of this study was to characterize the immunophenotypic alterations associated with persistent CH in the setting of NPM1mut clearance and to determine their possible clinical or biologic significance. METHODS: The cohort included 67 consecutive patients (pts) with NPM1mut AML treated at our institution between 01/2017 and 11/2019. FC assessment for MRD was performed an eight-color panel using FACSCanto II instruments (BD Biosciences, San Diego, CA) with a sensitivity of 10-3 to 10-4. Whole bone marrow (BM) DNA was interrogated for mutations with an 81-gene myeloid next-generation sequencing (NGS) panel using an Illumina MiSeq sequencer (Illumina, Inc., San Diego, CA, USA) with a sensitivity: 1% variant allelic frequency (VAF). RESULTS: Pts included 26 men and 41 women with a median age of 64 (range, 19-84) years with newly diagnosed NPM1mut AML. AML blasts had the following immunophenotype at baseline: promyelocytic-like phenotype (CD34-, CD117+, HLA-DR-) in 18 (27%), aberrant myeloid CD34-/CD117+/HLA-DR+ in 15 (22%), aberrant myeloid CD34+ in 13 (19%), myelomonocytic in 11 (16%), and monocytic in 10 (15%) cases. All pts had additional co-mutations at baseline (Fig 1). The most frequently co-mutated genes were DNMT3A (58%) FLT3 (51%), TET2 (27%), IDH2 (24%), PTPN11 (19%), IDH1 (18%), NRAS (16%), and SRSF2 (12%). Pts were treated with intensive (35;52%) and non-intensive induction regimens (32; 48%) (Fig 1); 22 (33%) received an allogeneic hematopoietic stem cell transplant as post-remission consolidation. We compared FC and NGS results in follow-up samples in pts achieving NPM1mut negative remission with adequate data available for comparison (n=50). 13 (26%) pts cleared all mutations whereas 37 (74%) had persistent CH. The most common mutations in the setting of residual CH involved DNMT3A (70%), TET2 (27%), IDH2 (19%) and IDH1 (11%). Among 37 pts with residual CH, 19 (51%) had no phenotypic alterations detected by FC while 17 (49%) had myeloid progenitors with alterations in intensity of antigen expression (increased CD13, CD123, CD117 and/or decreased CD38) or deviation from normal maturation but not diagnostic for AML MRD (herein referred to as pre-leukemic (PL) phenotype); 1 sample was MRD+ by FC. Mutation VAF of ≥5% was significantly more common (p=0.008) in cases with FC PL+ (100%) vs cases with normal FC phenotype (63%). IDH2 and SRSF2 mutation were exclusively observed in PL+CH+ cases with the former being statistically significant when compared with the FC-normal group (p=0.016). PL phenotype by FC did not correlate with intensity of induction therapy (41% treated with intensive regimens vs 59% non-intensive). The CH+/PL+ cohort had more pts ≥60 yrs old (67%) but the difference was not significant. There was no significant association between PL+ and residual mutation count. Presence of PL+ phenotype was not associated with a shorter relapse-free survival (RFS) (median not reached for both groups). CONCLUSIONS: Post-remission clonal hematopoiesis in the setting of NPM1mut clearance is common, and may result in immunophenotypic changes in myeloid progenitors, posing interpretive challenges for MRD assessment by FC. These alterations may be attributable to specific CH characteristics, such as IDH2 and SRSF2 mutations and VAF, but are not associated with a shorter RFS and thus should not be interpreted as residual AML or considered a high-risk attribute. Additional studies in other AML subtypes are warranted to further delineate these changes and their clinical significance. Figure 1 Disclosures DiNardo: Daiichi Sankyo: Consultancy, Honoraria, Research Funding; Notable Labs: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy; Takeda: Honoraria; Jazz: Honoraria; ImmuneOnc: Honoraria; AbbVie: Consultancy, Honoraria, Research Funding; Syros: Honoraria; Agios: Consultancy, Honoraria, Research Funding; Calithera: Research Funding; MedImmune: Honoraria; Celgene: Consultancy, Honoraria, Research Funding. Short:AstraZeneca: Consultancy; Takeda Oncology: Consultancy, Honoraria, Research Funding; Amgen: Honoraria; Astellas: Research Funding. Kadia:JAZZ: Honoraria, Research Funding; Ascentage: Research Funding; Astra Zeneca: Research Funding; Incyte: Research Funding; Celgene: Research Funding; Novartis: Honoraria; Cyclacel: Research Funding; Genentech: Honoraria, Research Funding; Amgen: Research Funding; Abbvie: Honoraria, Research Funding; Cellenkos: Research Funding; Pulmotec: Research Funding; Astellas: Research Funding; BMS: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding. Konopleva:Sanofi: Research Funding; Eli Lilly: Research Funding; AstraZeneca: Research Funding; Rafael Pharmaceutical: Research Funding; Amgen: Consultancy; F. Hoffmann La-Roche: Consultancy, Research Funding; AbbVie: Consultancy, Research Funding; Stemline Therapeutics: Consultancy, Research Funding; Kisoji: Consultancy; Reata Pharmaceutical Inc.;: Patents & Royalties: patents and royalties with patent US 7,795,305 B2 on CDDO-compounds and combination therapies, licensed to Reata Pharmaceutical; Ascentage: Research Funding; Calithera: Research Funding; Forty-Seven: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; Cellectis: Research Funding; Agios: Research Funding; Ablynx: Research Funding. Kantarjian:Adaptive biotechnologies: Honoraria; Aptitute Health: Honoraria; BioAscend: Honoraria; Delta Fly: Honoraria; Janssen: Honoraria; Oxford Biomedical: Honoraria; Ascentage: Research Funding; Daiichi-Sankyo: Honoraria, Research Funding; BMS: Research Funding; Immunogen: Research Funding; Jazz: Research Funding; Novartis: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Sanofi: Research Funding; Actinium: Honoraria, Membership on an entity's Board of Directors or advisory committees; Abbvie: Honoraria, Research Funding; Amgen: Honoraria, Research Funding. Ravandi:Macrogenics: Research Funding; Celgene: Consultancy, Honoraria; AstraZeneca: Consultancy, Honoraria; Xencor: Consultancy, Honoraria, Research Funding; Jazz Pharmaceuticals: Consultancy, Honoraria, Research Funding; Abbvie: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Astellas: Consultancy, Honoraria, Research Funding; BMS: Consultancy, Honoraria, Research Funding; Orsenix: Consultancy, Honoraria, Research Funding.

scholarly journals Establishing a Novel Pipeline That Combines in-Silico Prediction with in-Vitro and Ex-Vivo Validation to Discover Secondary Drug Combinations Against Relapsed and/or Refractory Multiple Myeloma

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 1615-1615
Author(s):  
Sayak Chakravarti ◽  
Suman Mazumder ◽  
Harish Kumar ◽  
Neeraj Sharma ◽  
Ujjal Kumar Mukherjee ◽  
...  
Keyword(s):  
Cell Death ◽  
Cell Line ◽  
Board Of Directors ◽  
In Silico ◽  
Research Funding ◽  
Ex Vivo ◽  
Standard Of Care ◽  
In Silico Prediction ◽  
Advisory Committees

Abstract Multiple myeloma (MM) is the second-most common hematological malignancy in the US. MM is an incurable, age-dependent plasma cell neoplasm with a 5-year survival rate of less than 50%. Extensive inter-individual variation in response to standard-of-care drugs like proteasome inhibitors (PIs) and immunomodulatory drugs (IMiDs), drug resistance, and dose-limiting toxicities are critical problems for the treatment of MM. Clinical success in anti-myeloma treatment, therefore, warrants continuous development of novel combination therapy strategies with the explicit goal to improve the therapeutic efficacy by concomitantly targeting multiple signaling pathways. Previously, we have reported the development of an in-house computational pipeline called secDrug that applies greedy algorithm-based set-covering computational optimization method followed by a regularization technique to predict secondary drugs that can be repurposed as novel synergistic partners of standard-of-care drugs for the management of refractory/ resistant MM. Top among these secondary drugs (secDrugs) were the HSP90 inhibitor 17-AAG. In this study, we used 17-AAG as a proof of principle to establish a pipeline that integrates our in silico predictions with in vitro and ex vivo validation as well as multi-omics technologies to identify, validate, and characterize therapeutic agents that could be used either alone or in combination with standard-of-care drugs for the treatment of R/R MM patients (Figure 1). To screen and validate our in silico prediction results, we performed in vitro cytotoxicity assays using 17-AAG on a panel of human myeloma cell lines (HMCLs; in vitro model systems) that captures a wide range of biological and genetic heterogeneity representing the complexities encountered in clinical settings. These cell lines include HMCLs representing innate sensitive/resistance, &gt;10 pairs of parental and clonally-derived PI- and IMiD-resistant pairs (P vs VR or LenR; representing acquired/emerging resistance/relapse), NRAS mutants which leads to the constitutive activation of oncogenic Ras signaling, and CRISPR-edited HSP90 knockdown cell line. Our results showed that 17-AAG has high synergistic activity in combination with PI in inducing apoptosis even in innate and acquired PI-resistant HMCLs and significantly reduces the effective dose of PI required to achieve IC 50 (Chou-Talalay's Dose Reduction Index or DRI 7±1.4). Moreover, 17-AAG+IMiD showed synergistic cell killing activity in clonally-derived IMiD resistant HMCL. Further, 17-AAG induced cell death was comparable with Hsp90 knockdown as evident from the cytotoxicity assay using PI and 17-AAG in combination in RPMI8226-wild type and RPMI-HSP90AA1 knocked down cell line. Notably, 17-AAG was strikingly effective against the NRAS-mutant cell line indicating an additional niche (NRas mutant myeloma) where 17-AAG could be most effective. Next, we performed RNA sequencing to elucidate the molecular mechanisms behind 17-AAG drug action, drug synergy, 17-AAG-induced cell death. Our gene expression profiling (GEP) followed by Ingenuity Pathway Analysis (IPA) analysis revealed protein ubiquitination, aryl hydrocarbon receptor signalling pathway as the top canonical pathways. 17-AAG induced apoptosis via mitochondrial mediated pathway in myeloma. 17-AAG exerts its cytotoxic effect by activating intrinsic pathway of apoptosis which we further confirmed through the increase in reactive oxygen species generation and decrease in mitochondrial membrane potential. 17-AAG was also effective in reducing the expression of hallmarks of MM such as p65/NF-kB, IRF4, c-Myc. Finally, we performed mass cytometry (CyTOF; Cytometry by time of flight) on primary bone-marrow cells (PMCs) from myeloma patients for further validation of proteomic signatures at the single-cell level. CyTOF analysis confirmed 17-AAG-induced cell death and key changes in MM-specific proteomic markers. 17-AAG treated PMCs showed elevated cleaved caspase levels and down-regulation of IRF4 and phospho-STAT3. GEP and CyTOF results were confirmed using immunoblotting assays. Together, our study demonstrates a unique pipeline for drug repositioning that has the potential to revolutionize clinical decision-making by minimizing the number of drugs required for discovering successful combination chemotherapy regimens against drug-resistant myeloma. Figure 1 Figure 1. Disclosures Kumar: BMS: Consultancy, Research Funding; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Research Funding; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Tenebio: Research Funding; Beigene: Consultancy; Oncopeptides: Consultancy; Antengene: Consultancy, Honoraria; Carsgen: Research Funding; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; KITE: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Merck: Research Funding; Astra-Zeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Research Funding; Roche-Genentech: Consultancy, Research Funding; Bluebird Bio: Consultancy; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Adaptive: Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Research Funding.

scholarly journals Genomic Data Improves Prognostic Stratification in Adult T-Cell Acute Lymphoblastic Leukemia Patients Enrolled in Measurable Residual Disease-Oriented Trials

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 3486-3486
Author(s):  
Celia González-Gil ◽  
Mireia Morgades ◽  
Francisco Fuster-Tormo ◽  
Jesus García-Chica ◽  
Pau Montesinos ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Board Of Directors ◽  
Research Funding ◽  
Residual Disease ◽  
Lymphoblastic Leukemia ◽  
Prognostic Significance ◽  
Gene Signature ◽  
Advisory Committees ◽  
Cell Acute Lymphoblastic Leukemia ◽  
Measurable Residual Disease

Abstract Background: Genetic information has become critical to understand the development of T-cell acute lymphoblastic leukemia (T-ALL) and to elucidate the origin of disease relapse. Several genetic markers, together with measurable residual disease (MRD), are considered strong predictors of patient outcome. However, the prognostic significance of genetic markers can varie according to treatment. Aim: We used targeted deep sequencing to analyze the genetic profile of 125 T-ALL patients enrolled in three consecutive MRD-oriented trials from the Spanish PETHEMA (Programa Español de Tratamientos en Hematología) group. Genomic information was analyzed together with the main clinical and biologic data in a subset of 111 patients with detailed clinical and outcome data to determine the prognostic significance for overall survival (OS) and cumulative incidence of relapse (CIR). Methods: Genetic mutations were detected using a custom gene panel and sequenced on a MiSeq platform. Alignment, variant calling, filtration and annotation of variants were done using standardized pipelines. OS curves were plotted by the Kaplan-Meier method and compared by the log-rank test. CIR was estimated using cumulative incidence functions by competing risks analysis. A Cox proportional hazard regression model was used to identify predictive factors for OS. Statistical significance was set at (two-sided) p-values &lt;0.05. Results: Recurrently mutated genes found in ≥4/125 patients involved transcription factor tumor suppressor genes (PTEN, BCL11B, RUNX1, GATA3, ETV6), epigenetic regulators (PHF6, DNMT3A, EP300, KMT2C, EZH2, TET2), DNA mismatch repair genes (MSH2), ribosomal (RPL5) and RNA splicing (U2AF1) genes, and genes involved in the RAS/MAPK (NRAS), WNT (FAT1, FAT3), IL7R-JAK-STAT (JAK3, JAK1, IL7R) and NOTCH1 signaling pathways, respectively. Mutations in the latest pathway (NOTCH1 & FBXW7) was found in 88/125 (70%) patients. Clinical-genetic correlations revealed that patients with mutations in JAK3, DNMT3A, N/KRAS, IL7R, MSH2 or in U2AF1 were associated with lower OS (vs unmutated patients). None of the mutated genes had impact on CIR. Upon grouping the mutated genes according to their functional role and potential biological impact on T-ALL, two gene signatures were defined. These included the aging gene signature (DNMT3A and U2AF1) characterized by mutations in genes identified in clonal hematopoiesis of indeterminate potential (CHIP); and the treatment resistance gene signature (JAK3, N/KRAS, IL7R and MSH2), defined by mutations in genes involved in resistance to the ALL therapy. Both clusters identified patients with poorer response to therapy (poorer blast clearance on day 14 of induction treatment and lower CR rates). Therefore, we considered together (worse outcome genetics [WOG] signature) for univariate and multivariate analyses. WOG and MRD level (0.1% cut-off) on day 35 after induction therapy (+35d MRD) showed significant prognostic impact in the univariable and multivariable analyses for OS (3y) with a hazard ratio (95% CI) of 2.4 (1.2; 4.8) and 2.7 (1.4; 5.1), respectively (Table 1). OS according to these two variables allowed risk stratification of T-ALL into low, intermediate- and high-risk (HR) patients with significantly different outcomes (p&lt;0.001) (Figure 1). Conclusion: A genetic signature with independent prognostic significance of MRD has been identified in this cohort of patients included in MRD-oriented trials. This gene signature (WOG) together with MRD could help to improve risk-stratification of adult T-ALL patients and would be of interest in the search for new therapies for HR patients Funding: Support from AECC (GC16173697BIGA); ISCIII (PI19/01828 and PI19/01183), co-funded by ERDF/ESF, "A way to make Europe"/"Investing in your future", CERCA/Generalitat de Catalunya SGR 2017 288 (GRC)/ C González-Gil was supported by AGAUR grant (2020 FI_B2 00210). Figure 1 Figure 1. Disclosures Diaz-Beyá: Jazz: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Astellas: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Celgene: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Mercadal: Gilead Sciences, Inc.: Honoraria, Speakers Bureau; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Tormo: Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Jazz Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Barba: Amgen, Celgene, Gilead, Incyte, Jazz Pharmaceuticals, MSD, Novartis, Pfizer and Roche, Jazz Phar,aceuticals: Honoraria; Cqrlos III heqlth Institute, aSOCIACION espanola contra el cancer, PERIS: Research Funding. Maciejewski: Regeneron: Consultancy; Novartis: Consultancy; Bristol Myers Squibb/Celgene: Consultancy; Alexion: Consultancy. Ribera: ARIAD: Consultancy, Research Funding, Speakers Bureau; AMGEN: Consultancy, Research Funding, Speakers Bureau; Pfizer: Consultancy, Research Funding, Speakers Bureau; TAKEDA: Consultancy, Research Funding, Speakers Bureau; NOVARTIS: Consultancy, Speakers Bureau; SHIRE: Consultancy, Speakers Bureau.

scholarly journals Prognostic Role of Multiparameter Flow Cytometry-Based Measurable Residual Disease Assessment in Patients with Acute Myeloid Leukemia Harboring DNMT3A/TET2/ASXL1 Mutation

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 8-9
Author(s):  
Muhned Alhumaid ◽  
Georgina S. Daher-Reyes ◽  
Aaron D Schimmer ◽  
Andre C. Schuh ◽  
Anne Tierens ◽  
...  
Keyword(s):  
Board Of Directors ◽  
Myeloid Leukemia ◽  
Research Funding ◽  
Residual Disease ◽  
Statistical Significance ◽  
Multiparameter Flow Cytometry ◽  
Long Term Outcomes ◽  
Advisory Committees ◽  
Measurable Residual Disease

BACKGROUND: Multiparameter flow cytometry (MFC) has increasingly been used for measurable residual disease (MRD) assessment in patients with acute myeloid leukemia (AML), while next-generation sequencing (NGS)-based MRD monitoring tool is in clinical development for its application. Clonal hematopoiesis (CH), in which leukemia-associated somatic mutations gene are present in individuals with no apparent hematologic disease, adds a challenge in the detection of MRD. In patients with AML, CH could be potentially pre-leukemic, while persistent mutations in DNMT3A, TET2 orASXL1 (DTA) in remission marrow are usually removed from the analysis of residual leukemic cells. However, reports suggest that persistent DTA mutations in remission may be correlated with an increased relapse risk. In the patients with DTA mutations, the use of NGS for MRD monitoring is limited or modified due to the presence of CH clone in the remission marrow. We evaluated whether MFC-MRD can be adjunctive to predict the risk of AML relapse in this population of 221 patients with DTA mutation (DNMT3A (n=123), ASXL1 (n=56) or TET2 (n=100). METHODS: The present study evaluated long-term outcomes in AML patients who achieved first complete remission (CR1) and compared outcomes according to MFC-based MRD status (was defined as negative if patients achieved 0.1 or less) assessed at the time of CR1. A total of 435 patients diagnosed with AML and treated with induction chemotherapy between 2015 and 2018 were included. MFC-MRD was assessed in 336 patients in CR1 (77%). NGS was performed using samples obtained at the time of initial diagnosis and used for mutational subgroup classification. Overall survival (OS) was calculated as the date of CR1 to the date of death and censored on the date of the last follow-up. Relapse-free survival (RFS) was defined as the time from the date of CR1 to the date of relapse or death from any cause. Cumulative incidence of relapse (CIR) and non-relapse mortality (NRM) were calculated considering competing risk. The Kaplan-Meier method using a log-rank test and a multivariate Cox proportional hazard model was used for analyses of time-to-event endpoints. For CIR and NRM, Gray test was performed for the risk factors and the Fine-Gray model was adopted for the multivariate model. RESULTS: According to the MFC-MRD status, i.e., the group with positive MRD (MRDpos; n=118, 35%) vs. those with negative MRD (MRDneg; n=218, 65%), we evaluated OS, RFS, and CIR. The MFC-MRDneg group showed better OS at 2 years 67.0% than the MFC-MRDpos group 40.7% (p&lt;0.001). The MFC-MRDneg group also showed a higher RFS rate at 2 years (58.7%) than the MFC-MRDpos group (40.6%) (p=0.001). The CIR was higher in the MFC-MRDpos group, 26.9%, than in the MFC-MRDneg group 21.1%, but with borderline statistical significance (p=0.083). NRM was slightly higher in the MFC-MRDpos group, 32.5%, than in the MFC-MRDneg group, 20.2%, but with borderline statistical significance (p=0.057). We divided the groups according to the number of induction treatment courses, AML type, cytogenetics risk, and age (&lt;60 vs ≥60), and compared OS, RFS, CIR and NRM between MFC-MRDpos vs MRDneg groups, which showed that MFC-MRD is relevant for risk stratification regardless of above-mentioned clinical variables Tab1. Also, we evaluated MFC-MRD status at CR by mutational profile subgroup. Long-term outcomes such as OS, RFS, CIR or NRM were compared by the mutational subgroup. It consistently showed a trend of superior OS, RFS and lower risk of CIR in patients with MFC-MRDneg compared to MFC-MRDposTab1. Of interest, in the subgroup of patients carrying any DTA mutations (n=221), those with MFC-MRDneg (n=103) showed better OS (HR 1.61 [1.01-2.55%]; p=0.042), RFS (HR 1.66 [1.06-2.61%]; p=0.026) and CIR (HR 1.99[1.03-3.83%]; p=0.04) compared to those MFC-MRDpos (n=64; Fig 1). Multivariate analysis confirmed that the MFC-MRDneg is an independent prognostic factor in patients with DTAmutwith respect to OS: MFC-MRDpos (HR 1.63, p=0.04) and age (≥60; HR 2.04, p=0.008) for OS; for RFS, MFC-MRDpos (HR 1.71, p=0.02) and age (≥60; HR 2.32, p= 0.001); for CIR, MFC-MRDpos (HR 2.31, p=0.01) and HCT (HR 0.14, p=&lt;0.001). Conclusion: These findings suggest that in AML patients with DTAmut, MFC-MRD status at the time of remission assessment can be a tool for MRD assessment when NGS-based MRD assessment is limited. Further study is strongly warranted to reach a clearer conclusion with multiple cohorts. Disclosures Schimmer: Takeda: Honoraria, Research Funding; Novartis: Honoraria; Jazz: Honoraria; Otsuka: Honoraria; Medivir AB: Research Funding; AbbVie Pharmaceuticals: Other: owns stock . Tierens:Amgen: Membership on an entity's Board of Directors or advisory committees; Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Astellas Pharma: Membership on an entity's Board of Directors or advisory committees. McNamara:Novartis: Honoraria. Maze:Pfizer: Consultancy; Novartis: Honoraria; Takeda: Research Funding. Gupta:Pfizer: Consultancy; Bristol MyersSquibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Sierra Oncology: Consultancy, Membership on an entity's Board of Directors or advisory committees; Incyte: Honoraria, Research Funding.

scholarly journals Prognosis Value of Measurable Residual Disease By Multiparameter Flow Cytotometry in Patients with Acute Myeloblastic Leukemia Prior to Allogeneic Hematopoietic Cell Transplantation

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 1821-1821
Author(s):  
Teresa Caballero ◽  
Olga Pérez-López ◽  
Ana Yeguas Bermejo ◽  
Eduardo Rodriguez Arbolí ◽  
Enrique Colado Varela ◽  
...  
Keyword(s):  
Complete Remission ◽  
Board Of Directors ◽  
Cell Transplantation ◽  
Research Funding ◽  
Hematopoietic Cell Transplantation ◽  
Residual Disease ◽  
Hematopoietic Cell ◽  
Advisory Committees ◽  
Measurable Residual Disease ◽  
Reduced Intensity

Abstract Introduction: Acute myeloblastic leukaemia (AML) is an heterogeneous disease with different molecular and prognostic characteristics. According to the comorbidities and the revised 2017 European Leukaemia genetic risk stratification (ELN17), allogeneic hematopoietic cell transplantation (HCT) is the best therapeutic option for many patients with AML (Grimm, Blood Adv 2020). However, relapse remains the main cause of mortality after transplantation. Impact of MRD on the outcome of patients is well recognized and ELN2017 introduced the new response category complete remission (CR) without MRD (Döhner H, Blood 2017). Detection of measurable residual disease (MRD) by multiparameter flow cytometry (MFC) in AML before allogeneic HCT could be a powerful predictor of outcome and decisive when establishing strategies that modify the prognosis of these patients. Methods: Retrospective multicentre analysis of MRD by MFC of patients undergoing transplantation allogeneic in 4 centres during the period from 2012 to 2020. Both Leukaemia Associated Aberrant Immunophenotype (LAIP) and different from normal (DFN) approach were used to analyse the MRD. The MRD was carried out with 8-color panels based on Euroflow protocols. The samples were acquired in 8-color digital cytometers (FACSCanto II) calibrated and compensated according to Euroflow protocols. Results: 295 of 318 patients were evaluated. Table 1 shows the characteristics of the patients. 285 (96.7%) were in complete remission (CR), 207 had negative MRD, in 21 MRD was less than 0.1% (MRD-low) and in 57 greater than or equal to 0.1% (MRD-high). At 2 years, the overall survival (OS) and leukaemia-free survival (LFS) in the whole group were 69% (95% CI 63.18-74.18) and 58.4% (95% CI 52.4-63.9) respectively. In CR patients, MRD levels significantly influenced on outcomes, with OS and LFS of 76.7% and 67.6% for negative MRD, 68.5% and 49.7% MRD-low and 50 % and 36.6% in MRD-high, p &lt;0.001) (Figure 1). Considering only MRD-high as positive, according to ELN17, cumulative incidence of relapse (CIR) at 2 and 5 years were significantly lower among those with positive MRD: 22% (95% CI 17-28.1%) and 27% (95% CI 21%-33.5%) for negative MRD vs 46,5% (95% CI 32.4%-59.5%) and 50% (95% CI 34.8%-63.2%) in positive MRD, p 0.0005. No differences were observed in terms of non relapse mortality (p 0.2).. Likewise, positive MRD also identified different prognostic subgroups within the ELN2017 subgroups: OS and LFS among high-risk ELN2017 patients of 63.6% and 52.3% in negative MRD vs 35.7% and 18.2% in positive MRD patients, p = 0.0085 and p = 0.0094, respectively; for intermediate risk: 77% and 67.6% in negative MRD vs 67% and 50.5% in positive MRD patients, p = 0.23 and p = 0.056; and for favourable: 84% and 77.7% in negative MRD vs 48% and 39.2% in patients with positive MRD, p = 0.0051 and p = 0.0341. Considering the conditioning regimen, patients with MRD negative before transplant had better OS and LFS at 2 years (82% and 71.4% among those received myeloablative conditioning and 65% and 57.6% among those who received reduced intensity, respectively) than those who had positive MRD prior to transplant (56% and 44.4% in myeloablative and 43% and 25.5% in reduced intensity) (p &lt;0.001) (Figure 2). In multivariate time-dependent analysis, age (HR 1.019 p = 0.024-95% CI 1.001-1,038), adverse risk group according to ELN17 (HR 2.13 p = 0.033 CI95 1.54-3.93 ) and MRD before transplant (HR 3.8 p &lt;0.001 95 CI 1.55-3.93) significantly influenced survival. Conclusions: Detection of MRD prior to transplant by MFC identifies a group of patients with a worse prognosis and could be key when selecting the most appropriate therapeutic strategy. Figure 1 Figure 1. Disclosures Caballero: Celgene: Consultancy. Belén Vidriales: Roche: Consultancy; Novartis: Speakers Bureau; Jazz: Consultancy, Speakers Bureau; Astellas: Consultancy, Speakers Bureau. Montesinos: Karyopharm: Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Stemline/Menarini: Consultancy; Tolero Pharmaceutical: Consultancy; Agios: Consultancy; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Incyte: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Janssen: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Forma Therapeutics: Consultancy; Glycomimetics: Consultancy; Teva: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Daiichi Sankyo: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Astellas Pharma, Inc.: Consultancy, Honoraria, Other: Advisory board, Research Funding, Speakers Bureau. Perez-Simon: JANSSEN, TAKEDA, PFIZER, JAZZ, BMS, AMGEN, GILEAD: Other: honorarium or budget for research projects and/or participation in advisory boards and / or learning activities and / or conferences.

scholarly journals A Phase II Single Arm Study of Nivolumab As Maintenance Therapy after Autologous Stem Cell Transplantation in Patients with Hodgkin Lymphoma at Risk of Relapse or Progression

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 2455-2455
Author(s):  
Carlos Bachier ◽  
Henning Schade ◽  
Behyar Zoghi ◽  
Aravind Ramakrishnan ◽  
Nirav N. Shah
Keyword(s):  
Stem Cell ◽  
High Risk ◽  
Maintenance Therapy ◽  
Board Of Directors ◽  
Hodgkin Lymphoma ◽  
Research Funding ◽  
Residual Disease ◽  
Refractory Disease ◽  
Advisory Committees ◽  
Extranodal Disease

Abstract Introduction: Autologous stem cell transplants (ASCT) are standard of care for patients with primary refractory or recurrent Hodgkin lymphoma (HL). While transplant results in cure for some patients, others relapse and succumb from their disease. Studies have found high expression of programmed death ligand 1 (PD-L1) in HL cells. The anti-PD-1 monoclonal antibody, nivolumab, has been safe and efficacious in the treatment of relapsed, refractory HL (Ansell et al. 2015). We evaluated the safety and efficacy of nivolumab maintenance therapy post-ASCT in high risk for relapse Hodgkin disease. Methods: Patients with HL with high risk of residual disease following ASCT ( high risk defined as refractory disease, relapse &lt;12 months, or relapse ≥12 months with extranodal disease after frontline therapy) received nivolumab (240 mg IV every 2 weeks) starting 45-180 days post-transplant for a maximum of 6 months of treatment. Patients were followed for AEs through 100 days after the last dose of drug. PET-CT response assessments were performed 1-3 month, 6 month, and 12 month post-ASCT. The primary objective was to evaluate the safety and tolerability of nivolumab as maintenance therapy early after ASCT. The secondary objective was to evaluate progression-free survival (PFS) at 12 months post-transplant. Results: To date, 37 patients were enrolled; median age 36 years; 25 patients (68%) male. The median number of prior systemic regimens was 2 (range 2-4). 25 patients (68%) had relapsed disease, and 12 patients (32%) had primary refractory disease. 18 patients (49%) had extranodal disease at relapse, 6 patients (16%) had B-symptoms at relapse, and 11 patients (30%) had residual disease after salvage, including 10 patients (27%) of whom had 2-3 prior salvage therapies. 22 patients (60%) had received prior brentuximab, and 3 patients (8%) had received prior nivolumab or pembrolizumab. 36 patients received ASCT and 1 patient received tandem ASCT. At the time of data cutoff, 28 patients (76%) had discontinued nivolumab treatment, 22 patients (60%) because they had completed the 6-month treatment course, 4 patients (11%) due to an adverse event (AE) (1 patient each with pain, pneumonitis, rhabdomyolysis, or hypothyroidism), and 2 patients (5%) due to disease progression. The median duration of treatment was 22.1 weeks. 17 patients (46%) experienced a treatment-related AE (TRAE), of which 5 patients (14%) experienced a ≥Grade 3 TRAE. The most common (≥5%) TRAEs were diarrhea, fatigue, bone pain, neutrophil count decreased, pruritus, rash, and vomiting. 2 patients experienced a treatment-related serious AE (pneumonitis, rhabdomyolysis). There were no treatment-related deaths. With a median follow up of 9.2 months, the median PFS and overall survival (OS) have not been reached. The 6 month PFS is 92.1% and the 12-month OS is 100%. There were no differences in OS when stratified based on prior treatment. Conclusions: The use of nivolumab maintenance early after ASCT is safe and tolerable in this high risk patient population. Early efficacy data is promising, but data need to mature to determine the 12 month PFS. Figure 1 Figure 1. Disclosures Bachier: CRISPR: Membership on an entity's Board of Directors or advisory committees; Autolus: Membership on an entity's Board of Directors or advisory committees; Nkarta: Membership on an entity's Board of Directors or advisory committees; Mana: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; BMS: Membership on an entity's Board of Directors or advisory committees. Shah: Umoja: Consultancy; Incyte: Consultancy; Legend: Consultancy; Kite: Consultancy; Miltenyi Biotec: Consultancy, Honoraria, Research Funding; Lily: Consultancy, Honoraria, Research Funding; Epizyme: Consultancy.

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