Quantitative Monitoring of NPM1 Mutation A Minimal Residual Disease Identifies Patients At High Risk for Relapse within the ELN Favorable Risk Group

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1404-1404
Author(s):  
Max Hubmann ◽  
Marion Subklewe ◽  
Thomas Köhnke ◽  
Stephanie Schneider ◽  
Annika Dufour ◽  
...  
Keyword(s):  
High Risk ◽  
Induction Therapy ◽  
Residual Disease ◽  
Risk Group ◽  
Disease Recurrence ◽  
Consolidation Therapy ◽  
Rt Pcr ◽  
Npm1 Mutation ◽  
Minimal Residual

Abstract Abstract 1404 Introduction: Molecular analyses of leukemia-specific markers has led to an improvement of the prognosis evaluation in patients (pts) with acute myeloid leukemia (AML). The European Leukemia Net (ELN) has published a classification which separates different subgroups by cytogenetic and molecular genetic analyses. Nevertheless, there are still pts suffering from disease recurrence within the ELN favorable risk group. To identify these pts at high risk for relapse the monitoring of minimal residual disease (MRD) of leukemia-specific markers could become an important diagnostic tool. In this study the potential of MRD monitoring by quantitative real-time PCR (RT-PCR) of NPM1 A mutation (NPM1 A) at different checkpoints within the ELN favorable risk group of pts with NPM1 A and without FLT3-ITD was investigated. Methods: Pts participating in the AMLCG99, AMLCG2004, and AMLCG2008 trial were prospectively or retrospectively screened for NPM1 mutation and FLT3-ITD by melting curve analyses. 334 pts were screened positive for NPM1 mutation and 262 pts showed a NPM1 A, 78.4 % of all NPM1 mutations. For MRD monitoring a relative RT-PCR was performed in 538 samples of 178 NPM1 A positive pts with a sensitivity of 10-6. MRD was monitored at diagnosis, in aplasia, after induction therapy, after consolidation therapy, and during the follow-up. MRD levels were normalized to the housekeeping gene ABL1 and expressed as a ratio to an internal control of known concentration. Results: In the analysis of the NPM1 A positive and FLT3-ITD negative pts (ELN favorable risk group) 82.5% (n=85) achieved complete remission (CR) after induction therapy. With a median follow-up of 26 (range 1–118) months, 36 (42.9%) pts relapsed within this subgroup. In aplasia, and after induction therapy, pts with a long-lasting remission showed significantly lower NPM1 A ratios in contrast to pts who relapsed during the follow-up. Via Receiver-Operating Curves (ROC) we analyzed the diagnostic power to identify pts at high risk for relapse and determined clinical useable cut-offs at the different checkpoints. ROC were significantly associated with disease recurrence at the checkpoints in aplasia and after induction therapy, but not after consolidation therapy. After induction therapy, a cut-off with a ratio of 0.01 was determined. This cut-off separates the patient cohort into two prognostic groups. NPM1 A MRD levels above the cut-offs result in an increased risk of relapse compared to pts with MRD level below this cut-off. This is reflected in a significantly lower 2-year relapse free survival (RFS) of 18% versus 72% (Figure 1). In 25 pts of this favorable risk group follow-up samples in CR were available for analysis of an upcoming relapse within 100 days of sampling. Only 2 of these pts developed relapse within of the next 100 days, but both pts showed increasing MRD levels prior to relapse. 18 relapse samples were available in this subgroup and interestingly, one patient (5.5%) was NPM1 A negative at relapse. When we further enrolled the FLT3-ITD positive pts into our analyses, not surprisingly we found a negative impact on the RFS of MRD positive and MRD negative pts. Conclusions: Our results confirm the observations of other studies that showed the prognostic impact of NPM1 MRD monitoring by RT-PCR. With the MRD monitoring we could identify pts at high risk for relapse within the ELN favorable risk group. Particularly high MRD levels after the induction therapy were strongly associated with a worse RFS. This and previously published data of others demonstrate that in addition to pre-therapeutic factors, the individual MRD course should be used as prognostic factor for the guidance of treatment and pts with high or increasing levels of MRD should undergo allogeneic stem cell transplantation, if eligible. Disclosures: No relevant conflicts of interest to declare.

Minimal Residual Disease Monitored With Fluorescence In Situ Hybridization (FISH) In CD34+CD38- Population Predicts Clinical Outcome In Core Binding Factor Acute Myeloid Leukemia (CBF-AML)

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 1356-1356
Author(s):  
Xiaoxia Hu ◽  
Libing Wang ◽  
Lei Gao ◽  
Sheng Xu ◽  
Shenglan Gong ◽  
...  
Keyword(s):  
Minimal Residual Disease ◽  
Myeloid Leukemia ◽  
De Novo ◽  
Residual Disease ◽  
Consolidation Therapy ◽  
Rt Pcr ◽  
Binding Factor ◽  
Minimal Residual

Abstract Acute myeloid leukemia (AML) is generally regarded as a stem cell disease, known as leukemic initiating cells (LIC), which initiate the disease and contribute to relapses. Although the phenotype of these cells remains unclear in most patients, they are enriched within CD34+CD38- compartment. In core binding factor (CBF) AML, the cytogenetic abnormablities are also existed in LIC. The aim of this study was to determine the prognostic power of minimal residual disease measured by fluorescence in situ hybridization (FISH) in flow sorted CD34+CD38- cells (FISH+CD34+CD38- population) at different period during the therapy. Thirty-six patients under 65 years of age with de novo CBF AML and treated with CHAML 2010 protocol were retrospectively included in this study. FISH efficiently identified the LICs (FISH+CD34+CD38-) in the CD34+CD38- population. The last follow-up was March 31, 2013, and the median follow-up was 336 days (range: 74-814 days). 33 patients with complete remission (CR) were eligible for the study, and 23 patients (23/33, 69.7%) with t (8;21) or AML1/ETO, and the remaining (10/33, 30.3%) with inv(16)/t(16;16) or CBFβ/MYH11. Flow-cytometry based FISH (F-FISH) procedure was performed at diagnosis, before every cycle of consolidation therapy, and every 3 months during follow-up. The FISH+ percentage at diagnosis constituting an average of 2.1% (range: 0.01%-27.5%) of the blast cells and 64.6% (range: 14%-87.8%) of the CD34+CD38- cells. Before the consolidation, FISH+CD34+CD38- population was detected in 13/33 (39.4%) patients. At this checkpoint, we have found the existence of FISH+CD34+CD38- population had prognostic value for the end points relapse free survival (RFS, 12% versus 68%, P=.008), and retained prognostic significance for RFS in multivariate analysis. Furthermore, the detection of FISH+CD34+CD38- before consolidation was found to be significantly associated with decreased OS. (11% versus 75%, P=.0005) Minimal residual disease (MRD) detected with F-FISH had a prognostic value at an earlier checkpoint when compared with flow cytometry and RT-PCR. Meanwhile, the concordance of flow cytomety, RT-PCR and F-FISH was investigated in the same patient cohort. 14 (70%) of 20 samples with detectable fusion transcripts by PCR did not have detectable leukemic cells by F-FISH. Therefore, the concordance for PCR and F-FISH was 63.7%. The concordance of FC and F-FISH was 64.3%: in 40 samples MRD was detected by both methods and in 61 samples MRD was ruled out by a negative result with the tests. With further analysis, the discrepancies among MRD detected with different MRD monitoring approaches before consolidation and after the first consolidation therapy contribute to 84% of the disconcordance. In summary, the detection of FISH+CD34+CD38- cells before consolidation therapy was significantly correlated with long-term survival in de novo CBF AML patients. F-FISH might be easily adopted as MRD monitor approach in clinical practice to identify patients at risk of treatment failure from the early stage during therapy. Disclosures: No relevant conflicts of interest to declare.

Next Generation Sequencing for Minimal Residual Disease Monitoring in AML Patients with FLT3-ITD,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3548-3548
Author(s):  
Felicitas Thol ◽  
Frederik Damm ◽  
Katharina Wagner ◽  
Katarina Reinhardt ◽  
Jan-Henning Klusmann ◽  
...  
Keyword(s):  
Minimal Residual Disease ◽  
Induction Therapy ◽  
Residual Disease ◽  
Rt Pcr ◽  
Fragment Analysis ◽  
Npm1 Mutation ◽  
Allelic Ratio ◽  
The Mean ◽  
Generation Sequencing ◽  
Minimal Residual

Abstract Abstract 3548 Minimal Residual Disease (MRD) monitoring has become an important tool for risk and treatment stratification in hematological malignancies. MRD monitoring in FLT3 mutated patients has been difficult in the past as FLT3-ITDs vary from patient to patient and individual primer/probe sets would be required to assess MRD over time. In the present study we evaluated next-generation sequencing (NGS) as a new tool for MRD monitoring in patients with FLT3-ITD and NPM1 mutations. Five pediatric and 5 adult AML patients with FLT3-ITD and 10 adult patients with NPM1 mutations were analyzed by NGS with a target coverage of 10,000 reads per amplicon. Pediatric samples were collected at diagnosis, day 22 and after consolidation chemotherapy while adult samples were collected at different time points (average 4 timepoints per patient). Samples were sequenced unidirectionally on eight-lane PicoTiterPlates on a GS FLX sequencing system. In total, 2,563,550 sequencing reads were generated, corresponding to a total of 1,176,171 high-quality sequencing reads. NPM1 mutations were analyzed by quantitative RT-PCR using the MutaQuant kit from Ipsogen (Ispogen, Marseille, France). Allelic ratios of FLT3-ITDs were determined by fragment analysis on a DNA sequencer using GeneMapper software 4.0. First, the sensitivity of NGS to detect mutated alleles was evaluated by sequencing serial dilutions of a patient sample that had 46.3 percent mutated FLT3-ITD alleles at diagnosis. With a target coverage of 10,000 sequences and an allelic ratio of 46.3 percent the theoretical detection sensitivity was at most 1 in 4630 sequences. In fact, the allelic ratio in the sequenced samples linearly decreased in the tested dilutions down to the 5×10-4 dilution (Pearson correlation R2=.996). Samples from healthy volunteers were tested negative for both FLT3-ITD and NPM1 mutations (n=3). Allelic ratios from three diagnostic specimens of FLT3-ITD mutated patients were highly reproducible when determined in two independent NGS runs. As proof of principle we analyzed NPM1 mutated patients by NGS and quantitative RT-PCR in parallel. The mean allelic ratio of NPM1 mutants at diagnosis was 0.37 (range 0.29–0.46). An allelic ratio of 0.37 and 0.4 was measured in peripheral blood of two patients, and thus was similar to ratios in bone marrow. Concordant results between NGS and qRT-PCR were found in 38 samples (95%), whereas in two samples one method did not detect the mutation while the other did (NGS and RT-PCR were negative once each). We analyzed relapse samples in four patients. The NPM1 mutation was detected consistently by both methods in three patients at allelic ratios of 0.013, 0.19, and 0.32, while one patient had lost the mutation at relapse. One patient had an atypical NPM1 mutation for which no RT-PCR kit was available. NGS allowed quantification of the allelic ratio in this patient, which was 0.37 at diagnosis, 0.06 after one cycle of induction therapy, and 0 after the second cycle of induction therapy. In FLT3-ITD mutated patients we could determine insertion site, insertion length, number of individual clones, and allelic ratio from NGS data. The mean allelic ratio in diagnostic samples was 0.27 as measured by NGS and 0.4 as measured by fragment analysis. Three follow up samples were negative by fragment analysis, while a small clone could still be detected with NGS in these samples (allelic ratio 0.0004 to 0.001). All other samples were concordant between fragment analysis and NGS. NGS was used to determine MRD status in 5 patients with childhood AML harboring mutated FLT3. A reduction of 2–3 orders of magnitude was achieved during induction chemotherapy. During consolidation a further decrease or disappearance of mutated alleles was achieved in 3 patients, who remained in remission. However, allelic burden increased in 2 patients after first consolidation treatment (HAM) by 9- and 735-fold compared to the allelic ratio after induction therapy, and they relapsed 74 and 303 days later. Thus, accurate determination of the FLT3-ITD allelic ratio by NGS may become useful to identify patients before overt relapse. In summary, we show that NGS can be used for minimal residual disease assessment in FLT3-ITD mutated AML patients. The sensitivity of the method is scalable depending on the read depth, however, an adequate sensitivity level for efficient MRD detection still needs to be determined. Disclosures: No relevant conflicts of interest to declare.

Minimal Residual Disease by Flow Cytometry in Children with Acute Lymphoblastic Leukemia. Single-Center Experience

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4238-4238
Author(s):  
Marcela Eugenia Soria ◽  
Marcela Ema Gutierrez ◽  
Maria Isabel Gaillard ◽  
Maria Cores ◽  
Gimena Gil ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
High Risk ◽  
Residual Disease ◽  
Risk Group ◽  
High Risk Group ◽  
Event Free Survival ◽  
Free Survival ◽  
Wbc Count ◽  
Minimal Residual

Abstract Abstract 4238 Introduction: Minimal residual disease (MRD) allows detection of blasts in patients in morphologically complete remission. Detection of MRD in patients with ALL is an independent risk factor associated with lower event-free survival (EFS). Objectives: 1) Evaluate MRD in bone marrowaspirateby flow cytometry at days 15 and 33 of induction in children with ALL. 2) Determine MRD association with hematologic features. 3) To correlate MRD association with overall survival (OS) and event-free survival (EFS). Design and Statistical Analysis: Descriptive, retrospective study. Univariate analysis was performed by T- test for continuous varibles and byχ2or Fisher tests for categorical variables. Survival was estimated with Kaplan Meier method. Regression multiple was used for multivariate analysis. p values < 0.05 were deemed as statistically significant. SPSS 15.0 program was used for the analyses. Patients: From January 2006 to December 2009, 84 patients with newly diagnosed ALL, aged 1 –18 years, were admitted at our institution. They were treated according to ALLIC / BFM / GATLA 2002 protocol. Immunophenotypic studies were performed by standard Four-color flow cytometry (FC) using FACSCalibur BD cytometer, MDR follow up panels were tailored depending on aberrant finding at diagnosis. 300.000–500.000 events were acquired using the CellQuest Pro and Paint -a-gate software for data analysis. MDR status has been defined as positive if at least 30–50 clustered events displaying leukemia –associated immunophenotypic characteristics (0,01%). Results: The mean age at diagnosis was 7.7 years (r: 2.1–18). Mean follow-up 33.9 months (r: 1–66). 38% were female. Immunophenotype: Pro B 5%, common B 87%, Pre B 2%, and T 6%. Good response to prednisone was achieved by 92% of patients. With regard to risk groups, the distribution was: 28% standard, 55% intermediate and 17% high. MRD at day 15 was evaluable in 66 patients (78%), being positive in 35 of them (45%). MRD at day 33 was evaluable in 75 patients (89%), being positive in 10 (12%). Conclusion: Positive MRD at day 15 was associated with: increased WBC count at diagnosis, high-risk group, higher relapse rate, lower EFS and OS. Otherwise, positive MRD at day 33 was associated: increased WBC count at diagnosis, T immunophenotype, poor response to prednisone, high risk group, lower EFS and OS. Our data indicate that positive MRD at days 15 and 33 result an independent variable of poor prognosis in children with ALL. Disclosures: No relevant conflicts of interest to declare.

Identification of Clinically Relevant Predictive MRD Checkpoints in AML Patients with NPM1 Mutations: A Study of the AML Study Group (AMLSG).

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1586-1586
Author(s):  
Jan Krönke ◽  
Richard Schlenk ◽  
Kai-Ole Jensen ◽  
Karina Eiwen ◽  
Marianne Habdank ◽  
...  
Keyword(s):  
Induction Therapy ◽  
Prognostic Value ◽  
Residual Disease ◽  
High Dose ◽  
Prognostic Impact ◽  
Consolidation Therapy ◽  
Npm1 Mutation ◽  
Cell Counts ◽  
Induction Cycle

Abstract Abstract 1586 Poster Board I-612 Background Mutations in the nucleophosmin 1 (NPM1) gene represent the most frequent gene mutations in acute myeloid leukemia (AML), with highest frequency (50-60%) in cytogenetically normal (CN)-AML. Several studies have shown the applicability and prognostic value of an NPM1 mutation (NPM1mut)-based assay for detection of minimal residual disease (MRD). So far, there are no studies evaluating the prognostic value of NPM1mut MRD levels in a large controlled cohort of AML patients (pts) enrolled on prospective clinical trials. Aims To evaluate the prognostic value of NPM1mut MRD levels in younger (16 to 60 years) AML pts harbouring NPM1 mutations type A, B or D, and to assess the influence of concurrent FLT3 internal tandem duplications (ITD). Methods All pts were enrolled in the prospective AMLSG 07-04 and AML HD98A treatment trials. Treatment comprised double induction therapy with ICE (idarubicin, cytarabine, etoposide) followed by high-dose cytarabine-based consolidation, autologous or allogeneic stem cell transplantation. Levels of NPM1mut expression ratios, defined as NPM1mut copies per 104ABL copies, were determined by RQ-PCR using TaqMan technology. Dilution series showed a maximum sensitivity of 10-6 and high specificity as no wildtype NPM1 could be detected. Results A total of 1079 samples, [bone marrow (BM), n=1062; peripheral blood, n= 17) from 212 pts were analyzed at diagnosis, after each treatment cycle, during follow-up and at relapse (median number of samples per pt, n=4; range, 1-16). NPM1mut expression ratios at diagnosis varied between 1.1×104 and 10.4×106 (median, 6.9×105). Pretreatment transcript levels were not associated with clinical characteristics (e.g., age, white cell counts, BM blasts) and did not impact on relapse-free (RFS) and overall survival (OS). Following the first induction cycle, the median decrease of the MRD level ratio normalized to pretreatment levels was 4.21×10-3, independent of the presence of concurrent FLT3-ITD (p=0.39). After the 2nd induction cycle, the median reduction of MRD levels was significantly stronger in the FLT3-ITDneg group (6.75×10-5) compared with the FLT3-ITDpos group (4.19×10-4) (p=0.003) and this differential effect was observed throughout consolidation therapy. For evaluation of the prognostic impact of NPM1mut MRD levels, we compared patients achieving PCR-negativity with those with positive values at different checkpoints. The first reliable checkpoint was after double-induction therapy: the cumulative incidence of relapse (CIR) at 4 years of PCR-negative patients (n=27) was 0% compared with 48% (SE, 4.4%, p<0.00001) for PCR-positive patients (n=105). This translated into a significant better OS (p=0.0005). The second checkpoint was after completion of consolidation therapy (first measurement during follow-up period). Again, 4-year CIR was significantly (p<.00001) lower in the PCR-negative group with 11% (SE, 6.5%) compared with 51% (SE, 4.8%) in PCR-positive patients, again translating in a significantly better OS (p<.00001). In addition, the level of NPM1mut expression ratio at any time point examined after completion of therapy correlated with the risk of relapse, since 20 of 22 pts with a value above 1000 NPM1mut/104ABL copies relapsed after a median interval of 90 days (range, 11-709 days). The remaining 2 pts had increasing levels at last follow-up but are still in continuous complete remission (CR). In a few cases relapse prediction appeared to be limited due to inadequate increase of NPM1mut expression levels or to loss of NPM1 mutation at the time of relapse (n=5). On the other hand, we observed a number of pts (n=17) in continuous CR who had intermittent low (<1000 NPM1mut/104ABL copies) NPM1mut expression ratios. Conclusions The levels of NPM1mut expression at two distinct checkpoints, after double induction therapy and after completion of consolidation therapy, can be used as a prognostic factor in NPM1mut AML pts. The adverse outcome of pts carrying a concurrent FLT3-ITD is reflected by a significant lower reduction of tumor burden. Disclosures Göhring: Celgene Corp.:. Schlegelberger:Celgene Corp.:.

Characterization of Minimal Residual Disease

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. SCI-52-SCI-52
Author(s):  
Alberto Orfao ◽  
Bruno Paiva ◽  
Juan Flores-Montero ◽  
Noemi Puig ◽  
Laura Gutierrez ◽  
...  
Keyword(s):  
High Risk ◽  
Induction Therapy ◽  
Residual Disease ◽  
Residual Tumor ◽  
Disease Recurrence ◽  
Response To Therapy ◽  
Genetic Profiles ◽  
Minimal Residual

Abstract Minimal residual disease (MRD) defines persistence of minimal numbers (<10-2-10-6) of residual tumor cells after treatment. In recent years, evaluation of MRD has become more frequently used as a mean to assess the quality of response to therapy in multiple myeloma (MM), particularly among those patients who have reached complete remission (CR). At the same time, it has become one of the most relevant prognostic factors in MM, both among patients with standard-risk and those with high-risk cytogenetics. In parallel, the introduction of novel therapies has led to significantly higher CR rates, with also lower rates of MRD-positivity and lower MRD levels. Such improvement in response to therapy of MM has fostered the development of progressively more sensitive approaches that allow deeper evaluation of the quality of the response achieved. However, it is well-known that while most cases that show persistence of MRD after therapy will eventually relapse, some of these patients show persistence of MRD in the absence of disease recurrence. In turn, a significant fraction of MM patients with high-risk cytogenetics, despite reaching deep responses to therapy, show early relapse. Altogether, these findings point out the potential relevance of the biological features of MRD cells, in addition to the MRD levels, in determining long-term MRD control vs. disease recurrence. Therefore, understanding the biologic signature of MRD cells may provide important insight into the mechanisms involved in chemoresistance and the discovery of novel potential therapeutic targets. At present, information about the phenotypic and genetic/genomic features of the chemoresistant myeloma plasma cell (PC) clones remains limited; this is mainly due the minimal levels of residual tumor cells, particularly among the MRD+ patients identified at advanced stages of therapy. Characterization of the phenotypic and genetic profiles of MRD+ myeloma PC which are resistant to induction therapy vs. paired diagnostic myeloma PC from elderly patients treated with novel drugs in the GEM2010MAS65 clinical trial, unravel that therapy-induced clonal selection can be already identified at the MRD stage, after induction therapy. In these settings, chemoresistant myeloma PC showed a specific phenotypic signature that may result from the persistence of clones with unique cytogenetic alterations. Thus, MRD myeloma PC which persisted after induction therapy showed increased expression levels of integrins and adhesion molecules (e.g. CD11c, CD29, CD44, CD49d, CD49e, CD54 and CD138, suggesting that among the initial tumor bulk, the few chemoresistant cells are likely to be those with stronger adhesion properties. Such cells also showed overall different gene expression profiles, with de-regulated genes/pathways related to proteasome-inhibition chemoresistance (e.g.: genes encoding for proteasome subunits or endoplasmic reticulum proteins), and that may influence survival of MM patients. Comparison of both iFISH and copy number variation profiles between patient-paired diagnostic vs. MRD PC revealed different genetic profiles in a substantial percentage of cases, which may potentially be due to the acquisition of new alterations during therapy that render the cell more chemoresistant, and/or the emergence of ultra-chemoresistant MRD cells that represented a subclone of all PC present at diagnosis. Disclosures Paiva: Celgene: Consultancy; Binding Site: Consultancy; Janssen: Consultancy; BD Bioscience: Consultancy; Onyx: Consultancy; EngMab AG: Research Funding; Millenium: Consultancy; Sanofi: Consultancy. Puig:The Binding Site: Consultancy; Janssen: Consultancy. San Miguel:Millennium: Honoraria; Onyx: Honoraria; Bristol-Myers Squibb: Honoraria; Novartis: Honoraria; Celgene: Honoraria; Sanofi-Aventis: Honoraria; Janssen-Cilag: Honoraria.

Depth of response and minimal residual disease status in ultra high-risk multiple myeloma and plasma cell leukemia treated with daratumumab, bortezomib, lenalidomide, cyclophosphamide and dexamethasone (Dara-CVRd): Results of the UK optimum/MUKnine trial.

2021 ◽  
Vol 39 (15_suppl) ◽  
pp. 8001-8001
Author(s):  
Martin F. Kaiser ◽  
Andrew Hall ◽  
Katrina Walker ◽  
Ruth De Tute ◽  
Sadie Roberts ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
High Risk ◽  
Minimal Residual Disease ◽  
Plasma Cell ◽  
Residual Disease ◽  
Cell Leukemia ◽  
Ultra High Risk ◽  
Grade 3 ◽  
Minimal Residual

8001 Background: Patients with ultra high-risk (UHiR) newly diagnosed multiple myeloma (NDMM) and patients with plasma cell leukemia (PCL) continue to have dismal outcomes and are underrepresented in clinical trials. Recently, improved responses with anti-CD38 monoclonal antibody combination therapy have been reported for NDMM patients. We report here outcomes for NDMM UHiR and PCL patients treated in the OPTIMUM/MUKnine (NCT03188172) trial with daratumumab, cyclophosphamide, bortezomib, lenalidomide, dexamethasone (Dara-CVRd) induction, augmented high-dose melphalan (HDMEL) and ASCT. With final analysis follow-up surpassed in Feb 2021, we report here early protocol defined endpoints from induction to day 100 post ASCT. Methods: Between Sep 2017 and Jul 2019, 107 patients with UHiR NDMM by central trial genetic (≥2 high risk lesions: t(4;14), t(14;16), t(14;20), gain(1q), del(1p), del(17p)) or gene expression SKY92 (SkylineDx) profiling, or with PCL (circulating plasmablasts > 20%) were included in OPTIMUM across 39 UK hospitals. Patients received up to 6 cycles of Dara-CVRd induction, HDMEL and ASCT augmented with bortezomib, followed by Dara-VR(d) consolidation for 18 cycles and Dara-R maintenance. Primary trial endpoints are minimal residual disease (MRD) status post ASCT and progression-free survival. Secondary endpoints include response, safety and quality of life. Data is complete but subject to further data cleaning prior to conference. Results: Median follow-up for the 107 patients in the safety population was 22.2 months (95% CI: 20.6 – 23.9). Two patients died during induction due to infection. Bone marrow aspirates suitable for MRD assessment by flow cytometry (10-5 sensitivity) were available for 81% of patients at end of induction and 78% at D100 post ASCT. Responses in the intention to treat population at end of induction were 94% ORR with 22% CR, 58% VGPR, 15% PR, 1% PD, 5% timepoint not reached (TNR; withdrew, became ineligible or died) and at D100 post ASCT 83% ORR with 47% CR, 32% VGPR, 5% PR, 7% PD, 10% TNR. MRD status was 41% MRDneg, 40% MRDpos and 19% not evaluable post induction and 64% MRDneg, 14% MRDpos and 22% not evaluable at D100 post ASCT. Responses at D100 post ASCT were lower in PCL with 22% CR, 22% VGPR, 22% PR, 22% PD, 12% TNR. Most frequent grade 3/4 AEs during induction were neutropenia (21%), thrombocytopenia (12%) and infection (12%). Grade 3 neuropathy rate was 3.7%. Conclusions: This is to our knowledge the first report on a trial for UHiR NDMM and PCL investigating Dara-CVRd induction and augmented ASCT. Response rates were high in this difficult-to-treat patient population, with toxicity comparable to other induction regimens. However, some early progressions highlight the need for innovative approaches to UHiR NDMM. Clinical trial information: NCT03188172.

Minimal Residual Disease (MRD) Analysis in Acute Myeloid Leukemia (AML) with Favorable Cytogenetics [t(8;21) and inv(16)] by Real Time PCR(RT-PCR) and Flow Cytometry (FC).

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2989-2989
Author(s):  
Granada Perea ◽  
Adriana Lasa ◽  
Anna Aventin ◽  
Alicia Domingo ◽  
Neus Villamor ◽  
...  
Keyword(s):  
Myeloid Leukemia ◽  
Residual Disease ◽  
Fusion Transcript ◽  
Good Prognosis ◽  
Rt Pcr ◽  
Free Survival ◽  
End Of Treatment ◽  
The Mean ◽  
Minimal Residual

Abstract Objectives: To analyze MRD in 65 patients (pts) with good prognosis AML: 30 t(8;21) and 35 inv(16), using both FC and RT-PCR, and to investigate the prognostic value of MRD in the pts outcome. Methods: MRD was monitored in CR pts (n=55) by FC in 101 follow-up samples obtained after various cycles of treatment, as follows: 40 post-induction (ind), 30 post-intensification (int) and 31 at the end of treatment (ttm), and by RT-PCR in 76 samples: 31, 23 and 22, respectively. In 35 pts the two techniques were applied at the same time of the ttm. MRD by FC was assessed using fixed combinations of three monoclonal antibodies. AML1/ETO and CBFb/MYH11 were analyzed following the BIOMED protocol. Results: Twenty-seven percent (n=15) of CR pts relapsed: 6 with t(8;21) and 9 with inv(16). The mean MRD by FC was 1.1% after ind, 0.2% after int and 0.1% at the end of ttm. At the end of ttm, the MRD detected by FC in relapsed and not relapsed pts were significativaly different: 0.3% vs 0.08% (p=0.002). By RT-PCR, the mean of fusion transcript copies/ablx104 differed between relapsed and nonrelapsed pts: 2385 vs 122 (p=0.001) after ind, 56 vs 7.6 after int (p=0.0001) and 75 vs 3.3 (p=0.0001) at the end of ttm. Relapses were more commonly observed in those pts with FC MRD level >0.1% at the end of ttm than in pts with ≤0.1%: 50% vs 12% (p=ns); likewise, using RT-PCR, a cutoff level of >10 copies at the end of ttm correlated with high risk of relapse: 80% of pts with RT-PCR >10 relapsed compared to 12% of pts with levels <10 (p=0.009). The overall survival (OS) probability was 86% for pts with CF MRD ≤0.1 at the end of ttm and 0% for pts with MRD >0.1 (p=0.1) and the leukemia free survival (LFS) was 78% and 44%, respectively (p=0.05). For pts with RT-PCR ≤10 at the end of ttm, the OS was 100% and for pts with RT-PCR >10 it was 30% (p=0.007) and the LFS was 87% and 20%, respectively (p=0.001). MRD was identified after ind in 55% of relapsed pts and at the end of ttm in 83% of relapsed pts. Only 1 pt (1/13) with FC MRD <0.1 and RT-PCR <10 at the end of ttm relapsed. For patients in complete remission, the mean copy level of chimeric transcript was higher for pts with t(8;21) than for those with inv(16): 30.2 vs 17.4 (p=0.0001). Comments: In tandem analysis of MRD by FC and RT-PCR could improve MRD detection in AML pts.

Evaluation of Minimal Residual Disease Based on NPM1 Mutations in AML with Intermediate Risk Cytogenetics: A Prospective Study of 36 Patients.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2847-2847
Author(s):  
Aline Renneville ◽  
Florence Pasquier ◽  
Selim Corm ◽  
Nathalie Philippe ◽  
Charikleia Kelaidi ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Peripheral Blood ◽  
Genomic Dna ◽  
Residual Disease ◽  
Normal Karyotype ◽  
Intermediate Risk ◽  
Npm1 Mutation ◽  
Log Reduction ◽  
Minimal Residual

Abstract Mutations in exon 12 of the nucleophosmin (NPM1) gene occur in approximately 50% of adult acute myeloid leukemia (AML) with normal karyotype. More than 40 mutant variants have been identified. Most of these mutations consist of a 4-bp insertion, which can be used as a target for minimal residual disease (MRD) monitoring. We previously checked the stability of NPM1 mutations at relapse in 21 NPM1-mutated patients at initial diagnosis. In this prospective study, we evaluated MRD by real-time quantitative PCR (RQ-PCR) in 36 NPM1-mutated AML patients corresponding to 33 adult and 3 pediatric cases, treated according to the French ALFA9801 or ALFA9802 and ELAM02 protocols, respectively. Out of these patients, 31/34 (91%) had normal karyotype, 13/33 (39%) had a high initial white blood cell count, and 10/36 (28%) were FLT3-Intern Tandem Duplication (FLT3-ITD) positive. 28 (78%) patients carry NPM mutation A, 3 (8%) mutation B and 5 (14%) other rare variants. RQ-PCR assays using a mutation-specific primer were performed on cDNA for mutation A and B and on genomic DNA for other NPM1 mutants. In our experiments, the maximal reproductible sensitivity of NPM1-based MRD detection is about 10−4 on genomic DNA and 10−5 to 10−6 on cDNA. The median follow-up was 260 days [40–791]. 2 to 9 follow-up samples from bone marrow and/or peripheral blood were analysed per patient. No correlation was found between leukocytosis at diagnosis and initial expression ratio of NPM1 mutation. The study of MRD log reduction after induction therapy shows that molecular responses are very heterogeneous (from 4.10−2 to more than 1.10−5), but 50% of patients reach at least a 4 log reduction in NPM1 levels. Patients with FLT3-ITD tend to have lower log reduction after induction than patients without FLT3-ITD, although not statistically significant (P=0.07). The analysis of NPM1-MRD in bone marrow and in peripheral blood at the same follow-up time-points shows that NPM1 levels can be until 1 log higher in bone marrow. This indicates that the evaluation of NPM1-MRD in bone marrow is more informative than in peripheral blood. We found all relapses had NPM1-MRD levels comparable to those observed at diagnosis. Among the 5 patients who relapsed so far, 2 were predictable by increasing MRD levels 1 to 4 months before hematological relapse. In 29 out of 36 cases, we could monitor MRD by both NPM1 mutation and WT1 gene expression. The comparison of the MRD profiles obtained by these two approaches reveals some discordant results, which can be, at least in part, explained by difference in the sensitivity of the RQ-PCR techniques, since the sensitivity of WT1 expression as MRD target is generally not higher than 10−3. In conclusion, NPM1 mutations are very specific and sensitive markers for MRD monitoring in AML. Further studies are required to determine if NPM1-MRD provides an independent prognostic factor and may be useful for therapeutic stratification in AML patients with intermediate risk cytogenetics.

Monitoring of Minimal Residual Disease in NPM1 Mutated Acute Myeloid Leukemia (AML) – a Single Center Experience in 27 Patients.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4672-4672
Author(s):  
Dana Dvorakova ◽  
Zdenek Racil ◽  
Ivo Palasek ◽  
Marketa Protivankova ◽  
Ivana Jeziskova ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Minimal Residual Disease ◽  
Peripheral Blood ◽  
Conflicts Of Interest ◽  
Residual Disease ◽  
Quantitative Polymerase Chain Reaction ◽  
Npm1 Mutation ◽  
Mgb Probe ◽  
Minimal Residual

Abstract Abstract 4672 Background Mutations within NPM1 gene occurs in about 60% of adult cytogenetic normal AML (CN-AML) and represent the single most frequent molecular aberration in this subgroups of patients. These mutations usually occur at exon 12 and induce most frequently a net insertion of four base pairs. Aims To examine the applicability and sensitivity of DNA-based real-time quantitative polymerase chain reaction (RQ-PCR) with mutation-specific reverse primers and common minor groove binding (MGB) probe and to evaluate whether minimal residual disease levels are of prognostic relevance in CN-AML patients with NPM1 mutations. Methods Patients were treated within different AML trials and follow-up samples of peripheral blood or bone marrow were referred to perform an RQ-PCR. Samples were analysed at diagnosis, during, and after therapy. The NPM1 mutations were A (17 pts), B (1 pt), D (2 pts) and 7 patients with individual rare types. For all cases, levels of minimal residual disease were determined by DNA-based RQ-PCR with mutation-specific reverse primer, one common forward primer and one common MGB probe. The NPM1 mutation value was normalized on the number of albumin gene copies and expressed as the number of NPM1 mutations every 106 genomic equivalents. This assay is highly specific as no wildtype NPM1 could be detected. Maximal reproducible sensitivity was 10 plasmide molecules per reaction. Results A total of 950 samples of bone marrow and/or peripheral blood from 27 patients have been analyzed. Twenty of 27 patients (74%) achieved molecular remission (MR), twenty-six of 27 patients (96%) achieved hematological remission (HR). 6 of 27 (22%) patients achieved HR without MR and one patient failed therapy. 8 of 20 patients (40%) with MR after treatment relapsed at molecular level and except one in all these patients hematological relaps occured (one patient is still in HR with bone marrow blast present, but < 5%). Considering relapsed patients, time from molecular to hematological relapse was 1 to 5 months (median: 3 months). Considering all 14 patients with HR without MR (6 pts) or with molecular relapse (8 pts), in 11 of them hematological relaps occured (79%) and molecular positivity anticipating hematological relaps with median of 3,5 month (1-7 months). 3 of these 14 patients are still in HR. Conclusions Mutations within NPM1 gene are a sensitive marker for monitoring minimal residual disease in CN-AML patients. RQ-PCR using a MGB probe is an efficient approach to long-term follow-up of residual leukemia cells and frequent quantitative monitoring is useful for reliably predicting hematological relapse. Achievement of negativity appears to predict favorable clinical outcome. This work was partially supported by research grant No. MSM0021622430 Disclosures: No relevant conflicts of interest to declare.

Sign in / Sign up

Export Citation Format

Share Document