Dynamics Of Minimal Residual Disease Research Articles

Abstract PO2-20-01: The STOP-HER2 Trial: A Phase 2 Study of Stopping Trastuzumab - Outcomes in Patients with HER2+ Metastatic Breast Cancer (TBCRC 062)

Abstract BACKGROUND: Anti-HER2 therapies have transformed the trajectory for patients with HER2-positive (HER2+) MBC. A subset of patients with HER2+ MBC do exceptionally well and remain on maintenance anti-HER2 therapy for many years. However, current therapy is non-curative and patients are treated indefinitely. While these therapies often are well-tolerated, they may cause significant toxicities, are expensive, and typically require frequent infusion and provider visits. Case reports and retrospective series demonstrate that some patients may do well after stopping anti-HER2 treatment. We currently lack prospective data on the likelihood of clinical impact of successful treatment discontinuation, outcomes among patients who progress and restart HER2-directed therapy, and tools to predict who may safely stop therapy. In this study, we will enroll pts with an exceptional response to anti-HER2 therapy. Pts may elect to either continue or stop anti-HER2 therapy and will be followed clinically and radiographically on a protocol-specified schedule. Considering the central role of patients in choosing the study intervention arm, and the unique nature of stopping therapy as a treatment intervention, patient advocates have been deeply involved in the study design to ensure patient concerns are being heard and addressed. We will retrospectively assess minimal residual disease (MRD), measured in plasma, as a biomarker of ongoing response, in a step toward shifting the treatment paradigm for pts with HER2+ MBC. METHODS: STOP-HER2 is a prospective, non-randomized, phase II, multicenter study of either continuation (cohort 1) or cessation (cohort 2) of anti-HER2 therapy in exceptional responders with HER2+ MBC. Eligible patients must have biopsy-proven HER2+ MBC by ASCO/CAP guidelines and must have been receiving first-line anti-HER2 therapy for MBC for at least 3 years without evidence of progressive disease (PD) as determined by the treating investigator. Patients with prior brain-only PD or oligo-PD outside the brain are eligible at least 2 years after local treatment to the central nervous system (CNS) and in absence of subsequent PD. Up to fifty-two participants will be enrolled into cohort 2, and an additional thirty participants enrolled into the cohort 1, at 10 US sites within the Translational Breast Cancer Research Consortium. Co-primary endpoints are the 1-year progression-free survival (PFS) in cohorts 2 and 1, separately. Secondary endpoints include the clinical benefit rate upon re-initiation of anti-HER2 therapy for patients in cohort 2 experiencing PD after treatment discontinuation; and 3-year overall survival in both cohorts. Correlative endpoints include MRD status at baseline and the correlation between MRD dynamics and outcomes. Patient-reported outcomes will be collected and analyzed, including quality of life, illness intrusiveness, financial toxicity, anxiety, and decision regret. The study will use a two-stage design to assess the primary objective of estimating the rate of participants who are free of progression at one year after discontinuing anti-HER2 therapy. If 50% or less are free of progression at one year, stopping HER2 therapy would not be considered a viable treatment plan (null hypothesis). The study is designed to have 80% power to reject the null hypothesis at the 10% type I error rate when 68% of exceptional responders are free of progression at one year after stopping HER2 therapy. STOP-HER2 began enrollment in Q2 2023 (NCT05721248). Citation Format: Heather Parsons, Kathryn Ruddy, Stefania Morganti, Karen Smith, Victoria Attaya, Eliza Kallfelz, Michelle DeMeo, Jamie LaScala, Shirley Mertz, Teri Pollastro, Patricia Spears, Adam Brufsky, Chau Dang, Susan Dent, Ahmed Elkhanany, WIlliam Gwin, Ciara O'Sullivan, Kathy Miller, Sara Nunnery, Elaine Walsh, Anna Maria Storniolo, Sara Tolaney, Nabihah Tayob, Antonio Wolff, Eric Winer, Mothaffar Rimawi, Ian Krop, Nancy Lin. The STOP-HER2 Trial: A Phase 2 Study of Stopping Trastuzumab - Outcomes in Patients with HER2+ Metastatic Breast Cancer (TBCRC 062) [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-20-01.

Droplet digital PCR: An effective method for monitoring and prognostic evaluation of minimal residual disease in JMML.

Juvenile myelomonocytic leukaemia (JMML) is a rare myeloproliferative neoplasm requiring haematopoietic stem cell transplantation (HSCT) for potential cure. Relapse poses a significant obstacle to JMML HSCT treatment, as the lack of effective minimal residual disease (MRD)-monitoring methods leads to delayed interventions. This retrospective study utilized the droplet digital PCR (ddPCR) technique, a highly sensitive nucleic acid detection and quantification technique, to monitor MRD in 32 JMML patients. The results demonstrated that ddPCR detected relapse manifestations earlier than traditional methods and uncovered molecular insights into JMML MRD dynamics. The findings emphasized a critical 1- to 3-month window post-HSCT for detecting molecular relapse, with 66.7% (8/12) of relapses occurring within this period. Slow MRD clearance post-HSCT was observed, as 65% (13/20) of non-relapse patients took over 6 months to achieve ddPCR-MRD negativity. Furthermore, bone marrow ddPCR-MRD levels at 1-month post-HSCT proved to be prognostically significant. Relapsed patients exhibited significantly elevated ddPCR-MRD levels at this time point (p = 0.026), with a cut-off of 0.465% effectively stratifying overall survival (p = 0.007), event-free survival (p = 0.035) and cumulative incidence of relapse (p = 0.035). In conclusion, this study underscored ddPCR's superiority in JMML MRD monitoring post-HSCT. It provided valuable insights into JMML MRD dynamics, offering guidance for the effective management of JMML.

Analysis of Peripheral Blood and Bone Marrow Residual Disease Dynamics during Intensive Treatment and Maintenance in Patients with Multiple Myeloma Included in the GEM12MENOS65 and GEM14MAIN Clinical Trials

Introduction: In patients (pts) with multiple myeloma (MM), analysis of minimal residual disease (MRD) dynamics in bone marrow by Next Generation Flow Cytometry (NGF) has shown that conversion of minimal residual disease (MRD) status modulates the risk of progression. Mass spectrometry measuring low-level monoclonal immunoglobulins has shown potential for Peripheral-blood based Residual Disease (PRD) assessment. In this study, we have analyzed the clinical impact of PRD and MRD dynamics in MM pts receiving intensive treatment as per the GEM2012MENOS65 trial and during maintenance as per the GEM2014MAIN trial. Patients and Methods: In GEM2012MENOS65 trial, pts received six cycles of VRD-GEM induction, autologous stem cell transplantation conditioned with melphalan or busulfan plus melphalan and consolidation with two cycles of VRD-GEM. Patients achieving at least minimal response were offered to be enrolled in the GEM2014MAIN and randomized to maintenance with lenalidomide and low-dose dexamethasone (Rd) or Rd plus ixazomib for two years; if not reaching MRD negativity at this point, pts received three more years of Rd. PRD dynamics were analyzed separately in each trial by Quantitative Immunoprecipitation Mass Spectrometry with anti IgG/A/M, total k and total l beads using the EXENT ® Solution (The Binding Site, part of Thermo Fisher Scientific). MRD was analyzed following the recommendations of the IMWG and according to the Euroflow guidelines. Only those pts with available samples at all the time points analyzed (GEM2012: post-induction, post-ASCT and after-consolidation; GEM014: post-consolidation and after 1 and 2 years of maintenance) were included. Results: 134 out of the 458 pts enrolled in the GEM2012MENOS65 trial and receiving intensive treatment as per protocol were analyzed. At treatment completion (post-consolidation), PRD and MRD status were associated with almost identical prognostic value: median progression-free survival (mPFS) in PRD - not reached (n.r.) vs 3.98 years in PRD + cases (p=0.0006) and in MRD - n.r. vs 3.99 years in MRD + cases (p=0.0001). When dynamics were analyzed, sustained PRD and MRD negativitywas observed in 58 (43.3%) and 44 (33.8%) pts and sustained positivity in 42 (31.2%) and 57 (42.5%) pts, respectively. In 35 (26.1%) and 33 (24.6%) pts, PRD and MRD converted from positive to negative, respectively. Sustained PRD or MRD positivity at the 3 time points analyzed was associated with a significantly shorter PFS (mPFS 4.04 years, p=0.0042 and 3.9 years, p=0.0008, respectively), compared to pts remaining negative or converting from positive to negative, in whom mPFS was not reached (Fig.1A). 109 pts out of the 332 pts enrolled in the GEM2014MAIN trial and receiving maintenance treatment as per protocol were analyzed. At treatment completion (after two years of maintenance), PRD or MRD status were associated with comparable prognostic value: mPFS in PRD - and PRD + cases n.r. (p=0.0039) and in MRD - cases n.r. vs MRD + 2.87 years (p<0.0001). When dynamics were analyzed, sustained PRDand MRD negativity was observed in 68 pts (62.4%) and 61 (55.9%) pts and sustained positivity in 19 (17%) and 19 (17%) cases, respectively. In 17 (15.6%) pts PRD and MRD converted from positive to negative and in 5 (5.6%) and 10 (0.91%) from negative to positive, respectively. Sustained PRD positivity at the 3 time points analyzed was associated with a shorter PFS (p=0.0185), compared to pts who remained PRD - or converted from PRD + to PRD -, in whom mPFS was not reached. The mPFS in pts remaining MRD - or converting from MRD + to MRD - was similar and has not been reached yet; the mPFS in pts with sustained MRD positivity at the 3 time points analyzed or converting from MRD - to MRD + was also similar (2.99 years and 2.88 years, respectively). Interestingly, pts converting from PRD - to PRD + had a poorer PFS (mPFS of 0.86 years) as compared to patients converting from MRD - to MRD + (mPFS of 2.28 years) (Fig.1B). Conclusions: In conclusion, assessment of PRD and MRD dynamics during intensive treatment and maintenance allows to define more precisely the patients' clinical outcome as compared to the analysis of isolated time points. Reaching MRD or PRD negativity post-consolidation or after 2 years of maintenance is associated with a similar prognostic value, thus highlighting the clinical utility of mass spectrometry as an alternative, non-invasive technique for disease evaluation.

Parallel Monitoring of Measurable Residual Disease and Chimerism By NGS in Myelodysplastic Syndrome after Allogenic STEM Cell Transplant

Background: Minimal Residual Disease (MRD) and chimerism monitoring in patients with Myelodysplastic Syndrome (MDS) is essential to track and quantify the proportion of donor and recipient cells to assess engraftment and treatment efficacy. Next Generation Sequencing (NGS) is rapidly evolving, can identify several somatic mutations simultaneously, quantifies allele burden, and is a suitable technology for this task. Traditionally its main limitations have been high single base error rates, which limited the sensibility to 0.1-1% Variant Allele Frequency (VAF). However, sensitivities up to 10 -4 VAF can be achieved by applying novel ultrasensitive personalized sequencing techniques. Aims: This study explores the applicability of PhasedSNP, a deep sequencing approach that quantifies chimerism in parallel to MRD in MDS patients after Allogenic Stem Cell Transplantation (ASCT). Methods: We included 24 patients older than 18 years who underwent ASCT between 2015 and 2022 at four third-level hospitals. We excluded patients with missing follow-up due to procedure-related mortality or relapse before day 100 and patients with unavailable samples. Fifty-three post-transplant samples were sequenced (six at day +30; two at +60; 21 at +100; 21 at +180, and three at day +360). Seven additional pre-transplant samples were included to confirm the donor and receptor signal switch. The genomic DNA of the donor and receptor was sequenced with an in-house PhasedSNP panel that covers 58 regions with paired polymorphisms in less than 15bp. Receptor DNA was also sequenced with a 42-gene panel (Ion Torrent TM) to identify somatic mutations used as biomarkers in follow-up samples to quantify MRD as previously described in ( Onecha E. et al. Haematologica 2019). Briefly, an amplicon-multiplexed mini-panel was defined for every patient to detect the selected MRD biomarkers and donor/receptor phased SNPs identified at diagnosis. The mini-panel included molecular-tagged primer pairs to amplify every selected marker (PhasedSNPs and somatic mutations) in three biological replicates. Final libraries were sequenced on the Ion S5 System platform with a median coverage of 211,190 reads per amplicon. A median of 558 ng of genomic DNA (range 73.44 - 858.6 ng) was sequenced in follow-up cases. Results: After excluding potential CHIP and Germline mutations, the most frequently mutated genes in the cohort were STAG2 (21%), SETBP1TP53RUNX1 andETV6 (16% each one). The baseline genotyping permitted to identify at least one somatic MRD biomarker in 79% of the patients (19/24), and 50% (12/24) presented more than one somatic mutation. MRD could be calculated for 42 of 53 samples. Thirteen patients presented sample at day +180 and clinical information. Four of them progressed, being three of them MRD positive. At this time-point the sensitivity of the test was 75% and the negative predictive value of 86%. The baseline PhasedSNP panel identified chimerism biomarkers (donor and receptor) for all patients. The PhasedSNP chimerism test presented a dynamic detection range below 10 -5 in serial dilutions. The test was positive for 48/53 samples using a threshold of 10 -4, with an average signal of 7·10 -3 (range 2·10 -5 to 4·10 -2). The correlation between PhasedSNP chimerism and MRD was high (R=0.921, p<0.001). However, the chimerism signal was consistently above the MRD signal, most likely due to the detection of healthy receptor cells with the PhasedSNP test. This can be seen in the two examples presented in the figure, with one patient ( MDS7, Figure, top) with an equivalent signal of chimerism at days +30, +100, and +180, but with MRD negativity at day +420 and two logarithms reduction in chimerism compared the highest value (day 100). Regarding the other patient, MDS13 ( Figure, bottom), chimerism and MRD dynamics evolve parallel but with more than two logarithmic differences between both values. This patient was MRD negative at day +100, but positive at day +180, and relapses one year later. Summary/Conclusion: NGS-based parallel detection of chimerism and MRD in MDS is feasible and has high applicability and sensitivity, with several advantages compared to traditional methods. Nevertheless, further research and a larger cohort is necessary to draw meaningful conclusions.

Pursuing the Clonal Dynamics of Minimal Residual Disease (MRD) in Relapsed and Refractory Acute Myeloid Leukemia

Acute Myeloid Leukemia (AML) is a myelogenous malignancy that frequently relapses despite complete remission (CR) and elicits non-responsiveness (refractory) to treatment. Recent studies have emphasized that acquiring resistance to treatment and/or residual subclones bearing drug persistency such as Cancer stem cells (CSCs) or Minimal residual disease (MRD) clones can lead to relapse and treatment-refractory in AML. However, due to the AML's cytogenetic clonal heterogeneity, drug persistent MRD having leukemogenic properties and its clinical relevance have not yet been fully explained. In this study, we investigate the clonal dynamics of AML MRD by pursuing their genotypic/phenotypic changes in the individual AML patient's Bone Marrow (BM) niche. To investigate, we collected longitudinal BM samples at diagnosis (Dx), remission (CR), relapse (Rel), and treatment-refractory (Per) from two patients of relapsed/refractory AML. By bulk Whole Exome Sequencing (WES), somatic variants were analyzed, and selected four oncogenic variants (Patient #01: FLT3 D835E, WT1 R375Pfs; Patient #02: KIT D816Y, ASXL1 A722L) showing a significant change according to the treatment process. Seven blood cell fractions (4 HSPCs, Hematopoietic Stem and Progenitors: HSC/MPP, LMPP, CMP/MEP, GMP, LSC-like populations; 3 Mature leukocytes: Myeloid cells, T/B Lymphocytes) from AML patient's bone marrow samples were sorted by cell surfaces antigens. Selected AML-associated variants were detected by droplet digital PCR (ddPCR) in DNA of each sorted seven single cell fractions. In the AML patient BM, we found that the clones with an oncogenic (or likely oncogenic) variant were distributed in not only primitive cell populations with the HSPC (LSC-like) phenotype, but also mature (differentiated) cell populations such as leukocytes. These clones were not completely disappeared even after intense AML chemotherapy, and remained in minimal quantities. In our sequential analysis, some residual clones distributed in LSC-like populations exhibited the traditionally well-known MRD's character, such as repopulating LSC proportions. Furthermore, some other residual clones only in mature populations also seemed to be played a role in functional MRD resulting in the repopulation of LSCs. However, whether these residual clones in LSC-like populations or mature populations eventually induced repopulating of LSC-like fractions during AML relapse, and particularly in the relapse before an endpoint, these clones also tended to be highly enriched in LSC-like populations by 30% (based on the VAF). While the variant clones in mature populations may be the temporal salvages of AML-MRD clones (functional MRD) to survive against treatment, the variant clones in LSC-like populations might be the AML-MRDs practically contributing to AML recurrence and treatment-refractory. This approach suggests the methodology for specifying AML-MRD and pursuing its clonal evolution according to the treatment progress. This will help to improve AML risk stratification and the diagnostic methodology, enabling the preemptive detection of AML MRD at the initial diagnosis/treatment. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Dynamics of Minimal Residual Disease Defines a Novel Risk-Classification and the Role of Allo-HSCT in Adult Ph-Negative B Cell Acute Lymphoblastic Leukemia

Dynamics of Minimal Residual Disease Defines a Novel Risk-Classification and the Role of Allo-HSCT in Adult Ph-Negative B Cell Acute Lymphoblastic Leukemia

Dynamics of minimal residual disease defines a novel risk-classification and the role of allo-HSCT in adult Ph-negative B-cell acute lymphoblastic leukemia

The prognosis of minimal residual disease (MRD) in acute lymphoblastic leukemia (ALL) patients is well established. However, the implementation of dynamic MRD for risk classification and decision-making for allogeneic hematopoietic stem cell transplantation (allo-HSCT) remains vague. In this study, we collected multiparameter flow cytometry (MFC)-MRD data of Ph-negative B-ALL patients (n = 134) from the Precision-Classification-Directed-Target-Total-Therapy-ALL-2016 (PDT-ALL-2016) cohort and stratified it into high-(HR), medium-(MR), and standard-risk (SR) groups. With a median of 3.65 years follow-up (95% CI: 3.037–4.263), 3-year OS rate was 51.8 ± 8.3% in HR, compared with MR 61.5 ± 10.8% (p = 0.472), and SR 73.3 ± 5.9% (p = 0.006). Multivariate analysis shows that integrated dynamic MRD is an independent factor for overall survival. Compared to pediatric-inspired chemotherapy, allo-HSCT significantly improves the survival of the HR cohort (p < 0.001), but not in MR and SR. Finally, our study suggests that integrated dynamic MRD defines a novel risk-classification criteria and highlights the benefits of allo-HSCT in adult patients with Ph-negative ALL.

Longitudinal minimal residual disease assessment in multiple myeloma patients in complete remission \u2013 results from the NMSG flow-MRD substudy within the EMN02/HO95 MM trial

BackgroundMultiple myeloma remains an incurable disease with multiple relapses due to residual myeloma cells in the bone marrow of patients after therapy. Presence of small number of cancer cells in the body after cancer treatment, called minimal residual disease, has been shown to be prognostic for progression-free and overall survival. However, for multiple myeloma, it is unclear whether patients attaining minimal residual disease negativity may be candidates for treatment discontinuation. We investigated, if longitudinal flow cytometry-based monitoring of minimal residual disease (flow-MRD) may predict disease progression earlier and with higher sensitivity compared to biochemical assessments.MethodsPatients from the Nordic countries with newly diagnosed multiple myeloma enrolled in the European-Myeloma-Network-02/Hovon-95 (EMN02/HO95) trial and undergoing bone marrow aspiration confirmation of complete response, were eligible for this Nordic Myeloma Study Group (NMSG) substudy. Longitdudinal flow-MRD assessment of bone marrow samples was performed to identify and enumerate residual malignant plasma cells until observed clinical progression.ResultsMinimal residual disease dynamics were compared to biochemically assessed changes in serum free light chain and M-component. Among 20 patients, reaching complete response or stringent complete response during the observation period, and with ≥3 sequential flow-MRD assessments analysed over time, increasing levels of minimal residual disease in the bone marrow were observed in six cases, preceding biochemically assessed disease and clinical progression by 5.5 months and 12.6 months (mean values), respectively. Mean malignant plasma cells doubling time for the six patients was 1.8 months (95% CI, 1.4–2.3 months). Minimal malignant plasma cells detection limit was 4 × 10–5.ConclusionsFlow-MRD is a sensitive method for longitudinal monitoring of minimal residual disease dynamics in multiple myeloma patients in complete response. Increasing minimal residual disease levels precedes biochemically assessed changes and is an early indicator of subsequent clinical progression.Trial registrationNCT01208766

Normalization of the Immune Microenvironment during Lenalidomide Maintenance Is Associated with Sustained MRD Negativity in Patients with Multiple Myeloma

Normalization of the Immune Microenvironment during Lenalidomide Maintenance Is Associated with Sustained MRD Negativity in Patients with Multiple Myeloma

Minimal Residual Disease Dynamics after Venetoclax-Obinutuzumab Treatment: Extended Off-Treatment Follow-up From the Randomized CLL14 Study.

The CLL14 study has established one-year fixed-duration treatment of venetoclax and obinutuzumab (Ven-Obi) for patients with previously untreated chronic lymphocytic leukemia. With all patients off treatment for at least three years, we report a detailed analysis of minimal residual disease (MRD) kinetics and long-term outcome of patients treated in the CLL14 study. Patients were randomly assigned to receive six cycles of obinutuzumab with 12 cycles of venetoclax or 12 cycles of chlorambucil (Clb-Obi). Progression-free survival (PFS) was the primary end point. Key secondary end points included rates of undetectable MRD and overall survival. To analyze MRD kinetics, a population-based growth model with nonlinear mixed effects approach was developed. Of 432 patients, 216 were assigned to Ven-Obi and 216 to Clb-Obi. Three months after treatment completion, 40% of patients in the Ven-Obi arm (7% in the Clb-Obi arm) had undetectable MRD levels < 10-6 by next-generation sequencing in peripheral blood. Median MRD doubling time was longer after Ven-Obi than Clb-Obi therapy (median 80 v 69 days). At a median follow-up of 52.4 months, a sustained significant PFS improvement was observed in the Ven-Obi arm compared with Clb-Obi (median not reached v 36.4 months; hazard ratio 0.33; 95% CI, 0.25 to 0.45; P < .0001). The estimated 4-year PFS rate was 74.0% in the Ven-Obi and 35.4% in the Clb-Obi arm. No difference in overall survival was observed (hazard ratio 0.85; 95% CI, 0.54 to 1.35; P = .49). No new safety signals occurred. Appearance of MRD after Ven-Obi is significantly slower than that after Clb-Obi with more effective MRD reduction. These findings translate into a superior long-term efficacy with the majority of Ven-Obi-treated patients remaining in remission.

BTK, NUTM2A, and PRPF19 Are Novel KMT2A Partner Genes in Childhood Acute Leukemia.

Chromosomal rearrangements of the human KMT2A/MLL gene are associated with acute leukemias, especially in infants. KMT2A is rearranged with a big variety of partner genes and in multiple breakpoint locations. Detection of all types of KMT2A rearrangements is an essential part of acute leukemia initial diagnostics and follow-up, as it has a strong impact on the patients’ outcome. Due to their high heterogeneity, KMT2A rearrangements are most effectively uncovered by next-generation sequencing (NGS), which, however, requires a thorough prescreening by cytogenetics. Here, we aimed to characterize uncommon KMT2A rearrangements in childhood acute leukemia by conventional karyotyping, FISH, and targeted NGS on both DNA and RNA level with subsequent validation. As a result of this comprehensive approach, three novel KMT2A rearrangements were discovered: ins(X;11)(q26;q13q25)/KMT2A-BTK, t(10;11)(q22;q23.3)/KMT2A-NUTM2A, and inv(11)(q12.2q23.3)/KMT2A-PRPF19. These novel KMT2A-chimeric genes expand our knowledge of the mechanisms of KMT2A-associated leukemogenesis and allow tracing the dynamics of minimal residual disease in the given patients.

Improving Outcomes for Patients with High-Risk Myeloma Via Prospective Trial Evidence: The Myeloma UK Nine Optimum Trial

Abstract INTRODUCTION Outcomes for patients with molecular high-risk (HR) multiple myeloma (MM) are poor with traditional approaches. Recent trials have demonstrated that novel combination therapies such as daratumumab/lenalidomide/dexamethasone improve outcome for HR disease at relapse, but access to these novel combination therapies remains limited in many public health care systems. Innovative clinical trials with robust and inclusive molecular testing strategies are needed to generate evidence for stratified therapy for HRMM in standard care. We reported that genetic double-hit, i.e. presence of two or more HR genetic lesions, predicts poor outcome in MM (Shah V, Leukemia 2017). Gene expression profiling (GEP) with the EMC92/SKY92 signature (SkylineDx) identifies patients with short survival (Kuiper R, Leukemia 2012). We present here updated data on an integrated genetic and GEP molecular risk stratification strategy, which will be used in the Myeloma UK (MUK) nine OPTIMUM trial to generate evidence for improved treatment of HRMM. METHODS AND RESULTS We present updated survival analyses (data cut-off July 2016) for 336 representative patients from the transplant-eligible cohort of the UK NCRI Myeloma XI trial (median follow-up 42 months). For all patients a complete dataset for adverse genetic lesions del(1p32), gain(1q21), del(17p), adverse translocations was generated using MLPA and qRT-PCR (Shah V, Leukemia 2017). All tumors were GEP risk profiled using the SKY92 MMprofiler on an Affymetrix DX2 platform. 32% of MM tumors carried at least one HR genetic lesion, and 22% carried a double-hit. Double-hit was associated with a hazard ratio of 2.1 for progression-free survival (PFS) (P The phase IIb MUKnine OPTIMUM trial investigates innovative combination therapy with Daratumumab, Cyclophosphamide, Velcade, Lenalidomide, Dexamethasone (DCVRd) induction followed by high dose melphalan + ASCT augmented by Velcade, intensified consolidation (DVR) and maintenance (DR) in patients with molecular high risk status as above. Its design follows recommendations for trials in rare cancers (Bogaerts J, Eur J Canc 2015), using a Bayesian strategy to compare outcomes with DCVRd against near-concurrent control data from the Myeloma XI trial, as reported above. The primary endpoint is PFS, with interim futility analyses of PFS and Minimal Residual Disease (MRD) negativity in OPTIMUM patients vs. Myeloma XI HR patients defined by the same molecular criteria. OPTIMUM is designed to detect an increase in PFS of 25% compared to Myeloma XI control data and, based on Bayesian modelling, up to 620 newly diagnosed patients will be molecularly screened for HR disease and up to 105 patients with HR status will be treated on the OPTIMUM trial. All other patients will be treated according to UK standard and data will be collected as for the HR group. Secondary endpoints include response, PFS2 and OS. Exploratory research accompanying the trial is planned to include longitudinal MRD dynamics and diffusion-weighted MRI imaging. CONCLUSION The UK Myeloma Research Alliance (MRA) MUKNine OPTIMUM study is one of the first trials in MM designed to provide comparative evidence for stratified treatment of patients with molecular high-risk MM. The trial has regulatory approval and is scheduled to start Q3 2017 in 22 hospitals of the Myeloma UK Clinical Trial Network. Disclosures Owen: Celgene: Consultancy, Honoraria, Research Funding; Takeda: Honoraria, Other: travel support; Janssen: Consultancy, Other: travel support. Drayson: Abingdon Health: Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Gregory: Celgene: Consultancy, Honoraria; Janssen: Honoraria. Jackson: Chugai: Honoraria; JJ Amgen: Honoraria; Takeda: Honoraria; Celgene: Honoraria. Cook: Glycomimetcs: Consultancy, Honoraria; Bristol-Myers Squibb: Consultancy, Honoraria; Takeda: Consultancy, Honoraria, Research Funding, Speakers Bureau; Amgen: Consultancy, Honoraria, Research Funding, Speakers Bureau; Celgene: Consultancy, Honoraria, Research Funding, Speakers Bureau; Janssen: Consultancy, Honoraria, Research Funding, Speakers Bureau; Sanofi: Consultancy, Honoraria, Speakers Bureau. Jenner: Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Support, Research Funding, Speakers Bureau; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel Support , Research Funding, Speakers Bureau; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Chugai: Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Kaiser: Janssen: Honoraria; Amgen: Consultancy, Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Takeda: Consultancy; BMS: Consultancy, Other: Travel expenses; Chugai: Consultancy.

Dynamics of minimal residual disease in patients with multiple myeloma on continuous lenalidomide maintenance: a single-arm, single-centre, phase 2 trial

In this single-centre, single-arm, phase 2 study, we enrolled patients aged 18 years and older from the Memorial Sloan Kettering Cancer Center (New York, NY, USA) who had newly diagnosed multiple myeloma following unrestricted frontline therapy and an Eastern Cooperative Oncology Group Performance Status of 2 or lower, including patients who started maintenance before study enrolment. All participants received lenalidomide maintenance at 10 mg for 21 days of 28-day cycles until progression or unacceptable toxic effects for up to 5 years on protocol. The primary endpoint was progression-free survival at 60 months per protocol and key secondary endpoints were MRD rates after completion of the 12th, 24th, and 36th cycle of maintenance and the association between progression-free survival and annual measurement of MRD status. MRD was assessed from first-pull bone marrow aspirates at baseline and annually by flow cytometry per International Myeloma Working Group criteria, (limit of detection of at least 1 × 10-5) up to a maximum of 5 years. Patients who completed at least four cycles of treatment were included in the analysis of the primary endpoint, and patients who had completed at least one dose of treatment on protocol were assessable for secondary endpoints. The study was registered at ClinicalTrials.gov, NCT02538198, and is now closed to accrual. Between Sept 8, 2015, and Jan 25, 2019, 108 patients (100 evaluable for the primary endpoint) were enrolled. Median follow-up was 40·7 months (95% CI 38·7-45·0). At 60 months, progression-free survival was 64% (95% CI 52-79). Median progression-free survival was unreached (95% CI unreached-unreached). MRD dynamics were assessed using 340 MRD assessments done over 5 years for 103 evaluable patients. Patients who sustained MRD negativity for 2 years (n=34) had no recorded disease progression at median 19·8 months (95% CI 15·8-22·3) past the 2-year maintenance landmark. By contrast, patients who lost their MRD-negative responses (n=10) were more likely to progress than those with sustained MRD negativity (HR infinite; p<0·0001) and those with persistent MRD positivity (HR 5·88, 95% CI 1·18-33·33; p=0·015) at the 2-year landmark. Haematological and non-haematological serious adverse events occurred in 19 patients (18%). The most common adverse events of grade 3 or worse were decreased lymphocyte count in 48 (44%) patients and decreased neutrophil count in 47 (44%) patients. One death occurred on study due to sepsis and heart failure and was considered unrelated to the study drug. Serial measurements of MRD allow for dynamic assessment of risk for disease progression. Early intervention should be investigated for patients with loss of MRD negativity. Sustained MRD positivity is not categorically an unfavourable outcome and might portend prolonged stability of low-level disease. Memorial Sloan Kettering and Celgene.

Long-Term Sustained Minimal Residual Disease (MRD) Negativity in Patients with Multiple Myeloma Treated with Continuous Lenalidomide Maintenance Therapy: A Clinical and Correlative Phase 2 Study

Long-Term Sustained Minimal Residual Disease (MRD) Negativity in Patients with Multiple Myeloma Treated with Continuous Lenalidomide Maintenance Therapy: A Clinical and Correlative Phase 2 Study

Monitoring for Minimal Residual Disease before and after Allogeneic Hematopoietic Stem Cell Transplantation in Newly Diagnosed Acute Leukemia

Objective:To assess the relevance between minimal residual disease (MRD) levels and disease recurrence and prognosis in patients with acute leukemia before and after hematopoietic stem cell transplantation (HSCT), and further to explore potential benefit based on pre-transplant MRD stratification. Methods: A total of 113 patients with newly diagnosed acute leukemia in Fujian Union hospital from April 2013 to April 2019 were retrospectively analyzed. 94 patients were complete remission (CR)( 50 cases of AML and 44 cases of ALL). 19 patients didn't achieve CR( 10 patients with AML and 9 patients with ALL). The median age was 26(1-56) years. 67 patients were male and 46 patients were female. The sources of hematopoietic stem cell include bone marrow, cord blood and peripheral blood stem cell. All patients underwent myeloablative conditioning before transplantation. Standard graft-versus-host disease (GVHD) prophylaxis was applied in all patients. MRD was measured by flow cytometry and real-time quantitative polymerase chain reaction(PCR). MRD values &gt;10-4 or BCR-ABL were considered positive for minimal residual disease and compared with bone marrow morphological findings. Median follow time was 15.2 (2.2 to 70.5) months. Results: Among patients in CR, achieving pre-MRD negativity was associated with longer 2-year relapse-free survival (2-year RFS; 81.4% vs 49.1%; P=0.003) and 2-year overall survival (2-year OS; 84% vs 50.6%; P=0.012). Compared to pre-MRDneg patients, pre-MRDpos patients had a higher incidence of relapse (31.2% vs. 5.1%, P=0.001). There was no significant difference between OS and MRD level after transplantation (OS; post-MRDneg 82.9% vs. post-MRDpos 66.7%, P =0.468). In the second set of analyses, CR patients were classified into the MRDpos /MRDpos group, the MRDpos /MRDneg group, the MRDneg /MRDpos group, and the MRDneg /MRDneg group according to MRD dynamics. Compared to the other three groups, patients from the MRDpos /MRDpos group had higher cumulative incidences of relapse (MRDpos /MRDpos, 48.3%; MRDpos /MRDneg, 22.2%, MRDneg /MRDpos,40.0%; MRDneg /MRDneg, 2.7%; P&lt;0.000) and worse 2-year RFS(MRDpos /MRDpos, 34.3%; MRDpos /MRDneg, 60.0%, MRDneg /MRDpos,60.0%; MRDneg /MRDneg, 76.6%; P=0.012). 1-year RFS and 1-year OS were similar in patients who didn't achieve CR, regardless of the post-MRD response(1-year RFS; 50.0% vs. 14.3%, P=0.063 and 1-year OS; 50.0% vs. 14.3%, P=0.184).In this group, 7 patients with MRD-positive after transplantation did not survive, but 12 post-HSCT MRD-negative patients had not relapsed. Conclusion: MRD can be used as a sensitive indicator to evaluate disease prognosis. Patients who are in morphologic remission and have no evidence of MRD before the HSCT come to encouraging leukemia-free survival. Continuous regular dynamic observation is important for guiding disease recurrence and prognosis. Disclosures No relevant conflicts of interest to declare.

Dynamics of Minimal Residual Disease in Neuroblastoma Patients.

Neuroblastoma is a common extracranial solid tumor of neural crest (NC) origin that accounts for up to 15% of all pediatric cancer deaths. The disease arises from a transient population of NC cells that undergo an epithelial-mesenchymal transition (EMT) and generate diverse cell-types and tissues. Patients with neuroblastoma are characterized by their extreme heterogeneity ranging from spontaneous regression to malignant progression. More than half of newly diagnosed patients present highly metastatic tumors and are stratified into a high-risk group with dismal outcome. As many as 20% of high-risk patients have residual disease that is refractory or progressive during induction chemotherapy. Although a majority of high-risk patients achieve remission, larger part of those patients has minimal residual disease (MRD) that causes relapse even after additional consolidation therapy. MRD is composed of drug-resistant tumor cells and dynamically presented as cancer stem cells (CSCs) in residual tumors, circulating tumor cells (CTCs) in peripheral blood (PB), and disseminated tumor cells (DTCs) in bone marrow (BM) and other metastatic sites. EMT appears to be a key mechanism for cancer cells to acquire MRD phenotypes and malignant aggressiveness. Due to the restricted availability of residual tumors, PB and BM have been used to isolate and analyze CTCs and DTCs to evaluate MRD in cancer patients. In addition, recent technical advances make it possible to use circulating tumor DNA (ctDNA) shed from tumor cells into PB for MRD evaluation. Because MRD can be detected by tumor-specific antigens, genetic or epigenetic changes, and mRNAs, numerous assays using different methods and samples have been reported to detect MRD in cancer patients. In contrast to the tumor-specific gene-rearrangement-positive acute lymphoblastic leukemia (ALL) and the oncogenic fusion-gene-positive chronic myelogenous leukemia (CML) and several solid tumors, the clinical significance of MRD remains to be established in neuroblastoma. Given the extreme heterogeneity of neuroblastoma, dynamics of MRD in neuroblastoma patients will hold a key to the clinical validation. In this review, we summarize the biology and detection methods of cancer MRD in general and evaluate the available assays and clinical significance of neuroblastoma MRD to clarify its dynamics in neuroblastoma patients.

PB2188 ANALYSIS OF MINIMAL RESIDUAL DISEASE IN PATIENTS WITH MULTIPLE MYELOMA RECEIVING DARATUMUMAB

Background:Over the past two decades, with the advent of new drugs, the tactics of treatment of patients with multiple myeloma (MM) has changed significantly; however, the general problem of the effectiveness of MM therapy remains unresolved to date. This disease is characterized by a progressive course and development of resistance to both proteasome inhibitors (bortezomib) and immunomodulatory drugs (lenalidomide), i.e. double refractoriness. Anti‐CD38 monoclonal antibodies are suggested for the treatment of heavily‐pretreated patients, including patients with double refractoriness.Aims:To study the dynamics of minimal residual disease (MRD) in patients with MM with double refractoriness receiving daratumumab.MethodsSeven patients with MM refractory to prior bortezomib and lenalidomide treatment were included in the study from 2017 to 2019. The number of lines of previous therapy was 5 (median = 3). The average age of patients–61 years (48–76 years). Daratumumab was administered to all patients as an intravenous infusion at a dose of 16 mg/kg: weekly at Weeks 1‐8, biweekly at Weeks 9‐24 and from Week 25 till progression. MRD assessment in bone marrow aspirate using multicolor flow cytometry (with a sensitivity of 10−5) was performed every 3 months from the start of treatment. To assess the effectiveness of treatment, generally accepted criteria were used.Results:Overall response was 86%. Fifty‐seven % of patients achieved a very good partial response, 29% had a partial response and 14% developed disease progression during therapy. After 3 months of therapy, 6 (86%) patients were MRD‐positive (median 0.072%, range 0.017–0.15%), of these, in 5 patients the monitoring showed a decrease in the number of myeloma cells, while in one patient MRD‐negativity was achieved. One patient (14%) with a very good partial response, after 3 months from the start of therapy, achieved an indicator of MRD‐negativity &lt;0.01% (median ‐ 0.005%), however, after 6 months the MRD status changed to positive.Summary/Conclusion:Daratumumab treatment regimen demonstrated a high level of general response. The MRD score reflects the clearance of residual tumor cells and is an important independent prognostic criterion for determining the depth of remission, for stratification by risk groups and disease prognosis.

PF630 MINIMAL RESIDUAL DISEASE MONITORING AND DEPTH OF RESPONSE IN MULTIPLE MYELOMA

Background:Minimal residual disease (MRD) assessment is a known surrogate marker for survival in multiple myeloma (MM). The majority of data comes from retrospective or subset analyses of patients enrolled in clinical trials.Aims:We present here a single institution's experience assessing MRD in MM patients receiving frontline therapy as well as those receiving therapy for relapsed disease. We describe the impact of depth of response, duration of response, and direction of response, on prognosis.Methods:181 multiple myeloma patients treated at The University of California, San Francisco (UCSF) from 2008 to 2016 were included. 126 were newly diagnosed patients versus 55 in second line or later. Patients received anti‐MM therapy per provider preference with the aim of obtaining maximal response by International Myeloma Working Group (IMWG) criteria and MRD was assessed in those achieving VGPR or better. MRD assessment was performed by commercially available next generation sequencing (NGS) of immunoglobulin genes (Adaptive[SN1] biotechnologies, Seattle, WA, USA). The sensitivity of this technique ranged from 10–4‐to‐10–6) and was 10–6in most cases. Progression Free Survival (PFS) curves were plotted by the Kaplan‐Meier method, and the log‐rank test was used to estimate the statistical significance of differences observed between curves.Results:A total of 398 MRD samples were analyzed at various time points during the disease course. MRD data was available at 33 time points for 59 patients, 2 time points for 36 patients and 1 time point for 86. Median follow up was 26 months. Overall, 66 of 181 total patients (36%)achieved MRD negativity (&lt;10–6) on one or multiple assessments.In the newly diagnosed group, 43 of 126 (34%), patients achieved undetectable MRD on at least one occasion. These patients had a prolonged PFS in comparison with patients who were persistently MRD positive (NR vs 49m[jm1] [SN2], p 0.006) Figure 1.A.Of the 55 patients who received therapy for relapsed disease, 21 of them achieved MRD negativity (38%) and PFS was also prolonged versus patients who never achieved MRD negative status (53m vs 23m, p = 0.03). Figure 1.BAlso, when we analyzed the effects of depth of response on survival, patients who were MRD negative or who were MRD positive at a very low level (between 10–5and 10–6), had a better prognosis than those with higher disease burdens (&gt;10–5)(p 0.001) Figure 1.C.Finally, we analyzed the effect of repeated MRD monitoring on PFS. Three categories were identified in newly diagnosed patients: (A) patients with ≥3 MRD‐ samples, (B) patients with continuously declining detectable clones, and (C) patients with a stable number of clones. Groups A and B had a more prolonged PFS than group C (NR vs 31m, p &lt; 0.0001). Figure 2.Summary/Conclusion:Conclusion. This study shows that MRD assessment in a real world setting likely has the same predictive power as that seen in clinical trials. MRD dynamics can accurately predict disease evolution and ultimately could drive clinical decision‐ making. MRD is an important predictor of PFS in newly diagnosed and relapsed patients. This study supports the concept of MRD‐driven decision‐making and validates the relevance of MRD in a real‐world MM practice.

The significance of peri-transplantation minimal residual disease assessed by multiparameter flow cytometry on outcomes for adult AML patients receiving haploidentical allografts.

A retrospective study (n = 460) was performed to assess the relationship between minimal residual disease (MRD) and transplant outcomes in a haplo-stem cell transplantation (SCT) setting. Patients from the pre-MRDneg group and the pre-MRDpos group had comparable outcomes. Compared to post-MRDneg patients, post-MRDpos patients had a higher incidence of relapse (100.0% vs. 8.3%, p < 0.001), lower incidences of overall survival (OS) (16.9% vs. 78.2%, p < 0.001) and leukemia-free survival (LFS) (0% vs. 76.5%, p < 0.001), and comparable probability of NRM (13.4% vs. 16.9%, p = 0.560). In a second set of analyses, all adult AML patients undergoing haplo-SCT were classified into the MRDneg/MRDneg group, the MRD decreasing group, and the MRD increasing group according to MRD dynamics by flow cytometry peri-SCT. Compared to the other two groups, patients from the MRD increasing group had higher cumulative incidences of relapse (MRD increasing, 100.0%; MRDneg/MRDneg, 9.6%; MRD decreasing, 19.2%; p < 0.001) and worse probabilities of OS (MRD increasing, 28.5%; MRDneg/MRDneg, 76.3%; MRD decreasing, 76.0%; p < 0.001) and LFS (MRD increasing, 0.0%; MRDneg/MRDneg, 73.9%; MRD decreasing, 74.0%; p < 0.001). The results indicated that haploidentical allografts might have a beneficial anti-leukemia effect in eradicating pretransplantation MRD, and MRD assessment peri-SCT is useful for risk stratification from a practical perspective.

Molecular Minimal Residual Disease Detection in Acute Myeloid Leukemia

Minimal residual disease (MRD) detection based on the standardized molecular monitoring of the t(9;22)-related BCR-ABL1 fusion transcript is well established for patients with chronic myeloid leukemia (CML). The levels of BCR-ABL1 serve as a guide to tailor treatment of the CML patient. In acute myeloid leukemia (AML) MRD detection based on polymerase chain reaction (PCR) approaches targeted towards the acquired molecular abnormalities is less well established. MRD measurement of the CBFB-MYH11 and RUNX1-RUNX1T1 fusion transcripts after induction therapy has been shown to be of some clinical importance. However, these transcripts can persist during long term complete remission, without having an effect on treatment outcome. In contrast, sequential MRD monitoring of the PML-RARA fusion transcript in acute promyelocytic leukemia (APL) is a strong predictor of relapse. Initial molecular MRD studies were limited to these favorable AML subtypes. Due to the discovery of novel recurrent abnormalities in AML the potential of molecular MRD detection has increased substantially. Although, certain acquired mutations, such as those in NPM1, are known for a number of years, only recently the application of these molecular abnormalities for MRD detection has been investigated in larger clinical trials. By NPM1 mutant MRD detection we can now recognize patients with higher risk of relapse. Highly sensitive targeted detection of the hotspot mutations in AML subsets is feasible by means of real-time PCR, but detection of patient specific mutations with this technology is still challenging. Next generation sequencing (NGS) revealed that AML is an extremely heterogeneous disease, as illustrated by the multitude of acquired mutations, but this technology has also opened possibilities for detection of MRD in virtually every patient. With NGS there is no need for patient specific assays since practically all mutations are detected. These molecular abnormalities, as single marker or in combination, will most certainly improve MRD monitoring of AML. However, it remains yet to be determined how MRD levels are assessed and which combination of markers in a MRD detection result in clinically relevant information, requiring extensive validation in large clinical AML trials. Smaller studies already demonstrated the variable dynamics of MRD during treatment and associations between somatic mutations persistence and risk of relapse. However, clonal hematopoiesis of undetermined potential, i.e., preleukemic mutations that may persist after treatment, provides an extra layer of complexity to the applicability of MRD detection. For example, the clinical applicability of MRD detection in the setting of mutant DNMT3A and IDH mutations is likely less effective due to the persistent DNMT3A and IDH mutant preleukemic cells following treatment. However, should all mutations be cleared after treatment or can preleukemic mutations in otherwise normal hematopoiesis persist without resulting in relapse? Taken together, there is need for molecular approaches to understand the dynamics of residual disease in AML during treatment. DisclosuresNo relevant conflicts of interest to declare.