Measurable residual disease assessment prior to allogeneic hematopoietic stem cell transplantation in acute myeloid leukemia and myelodysplastic syndromes: a 20-year monocentric study.

Patients with acute myeloid leukemia (AML) or myelodysplastic syndromes (MDS) who undergo allogeneic hematopoietic stem-cell transplantation (alloHSCT) can have divergent survival outcomes while all in morphological complete remission (CR). Techniques of measurable residual disease (MRD) have allowed us to refine their prognosis in two categories: MRD-positive and MRD-negative patients. We conducted a monocentric retrospective study (01/2000-12/2020) to assess the prognosis of pretransplant MRD status measured by multiparametric flow cytometry (MFC) and molecular biology assessed by PCR. 192 patients were included. The median follow-up period was 77 months. Among patients undergoing alloHSCT in CR, overall survival (median-OS: 130.6 vs. 16.0 months, P < 0.001), disease-free survival (median-DFS: 109.6 vs. 7.1 months, P < 0.001) and cumulative incidence of relapse (12-month CIR: 7.3% vs. 33.7%, P < 0.0001) were significantly different between MRD-negative and MRD-positive patients. Patients with discordant intermethod results had intermediate DFS. MRD-negative patients according to molecular PCR-based techniques, WT1 overexpression and MFC had longer median-DFS, compared to MRD-positive patients (P = 0.001, P < 0.001, P < 0.001, respectively). Looking into subgroups, MRD-positive patients among the ELN2017 adverse-category (P < 0.0001), myeloablative and reduced-intensity conditioning regimens (P < 0.0001, P = 0.005), < 60-year patients (P < 0.001) and AML patients (P < 0.001) were associated with lower DFS. This difference was not found in ≥ 60-year patients (P = 0.27) and MDS patients (P = 0.70). MRD-positive patients within the favorable/intermediate ELN2017 category trended toward lower DFS (P = 0.05). We confirmed that MRD status prior to alloHSCT is a strong prognostic factor for OS, DFS and CIR. Combining MFC and molecular-PCR techniques to assess MRD seems primordial as inter-method discordance can be consequential.

Long-Term Survival and Immune Reconstitution of Donor-Derived Chimeric Antigen Receptor T-Cell Therapy for Childhood Molecular Relapse of B-Cell Acute Lymphoblastic Leukemia After Allogeneic Hematopoietic Stem Cell Transplantation.

Measurable residual disease (MRD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT) is an independent risk factor for relapse in patients with acute lymphoblastic leukemia (ALL). This study aimed to assess the efficacy, safety, and immune reconstitution of chimeric antigen receptor T-cell (CAR-T) therapy in patients with molecular relapse after allo-HSCT. Eleven patients with molecular relapse of B-cell-ALL who underwent CAR-T therapy after allo-HSCT were enrolled. The rate of MRD negativity after a month of CAR-T infusion was 81.8%. Patients who bridged to second-HSCT after CAR-T therapy (n = 3) showed a trend of higher 3-year leukemia-free survival and 3-year overall survival than those who did not (n = 8; 100% vs. 75.0%; 95% CI, 45.0-104.9%; p = 0.370). No treatment-related mortalities were observed. Among patients who did not bridge to second-HSCT and remained in complete remission until the last follow-up (n = 6), five of them had not recovered normal immunoglobulin concentrations with a median follow-up of 43 months. CAR-T therapy may be a safe and effective treatment strategy to improve survival after allo-HSCT; however, the problem of prolonged hypogammaglobulinemia in patients who do not bridge to second-HSCT is worth noting.

Detection methods and prognosis implications of measurable residual disease in acute myeloid leukemia.

Measurable residual disease (MRD) in acute myeloid leukemia (AML) refers to the quantity of residual leukemic cells in a patient after treatment.According to the latest agreements, MRD in AML offering essential prognostic insights. However, there is ongoing debate regarding MRD-based monitoring and treatment strategies. There are multiple platforms for detecting MRD, each varying in sensitivity and suitability for different patients. MRD not only predicts treatment outcomes but also serves as an indicator of treatment effectiveness and a prognostic biomarker. In AML, most retrospective studies indicate that patients who are MRD-positive or show increasing MRD levels at specific time points during remission have significantly higher risks of relapse and mortality compared to MRD-negative patients. Although achieving MRD-negative status can improve patient prognosis, the possibility of relapse remains. Despite the correlation between MRD and clinical outcomes, MRD assessment methods are not yet standardized, leading to discrepancies in results across different techniques. To provide reliable MRD results, it is essential to optimize and standardize MRD detection methods. Methods for assessing MRD include multiparameter flow cytometry (MFC) and molecular assays, chosen based on disease characteristics. This review focuses on currently available MRD detection methods and discusses how the prognostic value of MRD test results informs personalized treatment strategies for AML patients.

Only FLT3-ITD co-mutation did not have a deleterious effect on acute myeloid leukemia patients with NPM1 mutation, but concomitant with DNMT3A co-mutation or a < 3log reduction of MRD2 predicted poor survival.

Co-occurring mutations are frequently observed in acute myeloid leukemia (AML) with NPM1 mutation, and NPM1 measurable residual disease (MRD) is an effective prognostic biomarker. This retrospective study investigated the impact of gene co-mutations and NPM1 MRD on outcomes in these patients. Among 234 patients, 11.5% carried the rare type NPM1 mutation (NPM1RT). The median age was 49 years (IQR 36-58), with a median follow-up of 30.4 months (IQR 12.1-55.7). Nine genes were mutated in > 10%, with DNMT3A (53.8%) and FLT3-ITD (44.4%) being most prevalent. Univariable analysis in 137 patients showed FLT3-ITD, DNMT3A co-mutations, and MRD2 < 3 log reduction predicted poorer survival. FLT3-ITD and DNMT3A co-mutations correlated with the lowest event-free (EFS) and overall survival (OS) (3-year EFS 30.0%; 3-year OS 34.4%; both p < 0.001). FLT3-ITD alone did not worsen survival compared to patients without FLT3-ITD. Multivariable analysis identified DNMT3A co-mutation [EFS, HR = 1.9, p = 0.021; OS, HR = 2.2, p = 0.023] and MRD2 ≥ 3 log reduction (EFS, HR = 0.2; OS, HR = 0.1, both p < 0.001) as independent survival predictors. Patients with FLT3-ITD and DNMT3A co-mutations or a MRD2 < 3 log reduction were identified as high risk, but allogeneic hematopoietic stem cell transplantation (allo-HSCT) improved survival significantly compared to chemotherapy only (3-year EFS, 57.9% vs. 30.0%, p = 0.012; 3-year OS, 72.9% vs. 34.4%, p = 0.001). In AML patients with NPM1 mutation, the detrimental impact of FLT3-ITD mutation was exacerbated by DNMT3A co-mutation. Poor-risk younger patients identified by FLT3-ITD and DNMT3A co-mutations or MRD2 < 3 log reduction benefit from allo-HSCT.

The significance of CD49f expression in pediatric B-cell acute lymphoblastic leukemia.

OBJECTIVES: CD49f is an adhesion molecule present on malignant lymphoblasts in B-cell acute lymphoblastic leukemia; it is associated with a poor prognosis. CD49f expression has been proposed as a marker for measurable residual disease (MRD) marker, but this marker has yet to be implemented in clinical practice. METHODS: In this study, we used flow cytometry to detect CD49f expression by leukemic blasts in paired bone marrow and cerebrospinal fluid samples at diagnosis and bone marrow at day 15 of treatment. RESULTS: At diagnosis, 93% of bone marrow and 100% of cerebrospinal fluid lymphoblasts expressed CD49f. The intensity of CD49f expression statistically significantly increased during treatment (P < .001). In MRD-negative end-of-treatment samples, only a small population of hematogones expressed CD49f. Interestingly, the intensity of CD49f expression varied among the different groups of recurrent genetic abnormalities. The ETV6::RUNX1 fusion and ETV6::RUNX1 combined with the high hyperdiploid group were associated with increased expression, whereas the Philadelphia-like group showed low CD49f expression. The lower CD49f expression at diagnosis predicted a lower MRD rate at day 15 of treatment. CONCLUSIONS: We concluded that CD49f can be used as an MRD marker and possible prognostic factor in B-cell acute lymphoblastic leukemia.

Factors impeding organ recovery despite a deep haematological response in patients with systemic AL amyloidosis.

Patients with AL amyloidosis can have persistent organ dysfunction despite achieving a haematological complete response (hemCR). We aimed to identify factors for organ non-response among 143 patients who achieved hemCR for ≥6 months. Kidney, heart and liver non-response were observed in 40/117 (34%), 19/68 (28%) and 3/17 (18%) patients respectively. Predisposing factors varied by organ system. Kidney non-responders had more advanced organ dysfunction at diagnosis, whereas heart non-responders had disproportionately more lambda-typic amyloidogenic light chains. Most patients without an apparent reason for organ non-response had detectable residual clonal disease. The interplay of factors impeding organ recovery in AL amyloidosis is complex.

Cerebrospinal fluid liquid biopsy by low-pass whole genome sequencing for clinical disease monitoring in pediatric embryonal tumors.

Background: Liquid biopsy assays that detect cell-free DNA (cfDNA) in cerebrospinal fluid (CSF) are a promising tool for disease monitoring in pediatric patients with primary central nervous system (CNS) tumors. As a compliment to tissue-derived molecular analyses, CSF liquid biopsy has the potential to transform risk stratification, prognostication, and precision medicine approaches. Methods: In this pilot study, we evaluated a clinical pipeline to determine feasibility and sensitivity of low-pass whole genome sequencing (LP-WGS) of CSF-derived cfDNA from patients with CNS embryonal tumors. Thirty-two longitudinal CSF samples collected from 17 patients with molecularly characterized medulloblastoma (12), embryonal tumor with multilayered rosettes (2), CNS embryonal tumor, not elsewhere classified (NEC) (2), and atypical teratoid/rhabdoid tumor (1) were analyzed. Results: Adequate CSF-derived cfDNA for LP-WGS analysis was obtained in 94% of samples (30/32). Copy number variants compatible with neoplasia were detected in 90% (27/30) and included key alterations, such as isodicentric ch17, monosomy 6, and MYCN amplification, among others. Compared to tissue specimens, LP-WGS detected additional aberrations in CSF not previously identified in corresponding primary tumor specimens, suggesting a more comprehensive profile of tumor heterogeneity or evolution of cfDNA profiles over time. Among the 12 CSF samples obtained at initial staging, only 2 (17%) were cytologically positive, compared to 11 (92%) that were copy number positive by LP-WGS. Conclusions: LP-WGS of CSF-derived cfDNA is feasible using a clinical platform, with greater sensitivity for tumor detection compared to conventional CSF cytologic analysis at initial staging. Large prospective studies are needed to further evaluate LP-WGS as a predictive biomarker.

Measurable residual disease (MRD)-testing in haematological cancers: A giant leap forward or sideways?

Measurable residual disease (MRD)-testing is used in many haematological cancers to estimate relapse risk and to direct therapy. Sometimes MRD-test results are used for regulatory approval. However, some people including regulators wrongfully believe results of MRD-testing are highly accurate and of proven efficacy in directing therapy. We review MRD-testing technologies and evaluate the accuracy of MRD-testing for predicting relapse and the strength of evidence supporting efficacy of MRD-guided therapy. We show that at the individual level MRD-test results are often an inaccurate relapse predictor. Also, no convincing data indicate that increasing therapy-intensity based on a positive MRD-test reduces relapse risk or improves survival. We caution against adjusting therapy-intensity based solely on results of MRD-testing.

The role of the primitive marker CD133 in CD34-negative acute myeloid leukemia for the detection of leukemia stem cells.

The most important reason for dismal outcomes in acute myeloid leukemia (AML) is the development of relapse. Leukemia stem cells (LSCs) are hypothesized to initiate relapse, and high CD34+CD38- LSC load is associated with poor prognosis. In 10% of AML patients, CD34 is not or is low expressed on the leukemic cells (<1%), and CD34+CD38- LSCs are absent. These patients are classified as CD34-negative. We aimed to determine whether the primitive marker CD133 can detect LSCs in CD34-negative AML. We retrospectively quantified 148 CD34-negative patients for proportions of CD34-CD133+ and CD133+CD38- cell fractions in the diagnostic samples of CD34-negative patients in the HOVON102 and HOVON132 trials. No prognostic difference was found between patients with high or low proportions of CD34-CD133+, which is found to be aberrantly expressed in AML. A high level of CD133+CD38- cells was not associated with poor overall survival, and expression in AML was similar to normal bone marrow. To conclude, CD133 is useful as an additional primitive marker for the detection of leukemic blast cells in CD34-negative AML. However, CD133+CD38 alone is not suitable for the detection of LSCs at diagnosis.

Laboratory validation and clinical utility of next-generation sequencing-based IGH/TCR clonality testing for the monitoring of measurable residual disease in acute lymphoblastic leukaemia: real-world experience at Austin Pathology.

Measurable residual disease (MRD) testing is an essential aspect of disease prognostication in acute lymphoblastic leukaemia (ALL) and informs clinical decisions. The depth of MRD clearance is highly relevant and requires assays with sufficient sensitivity. Austin Pathology is one of the few laboratories in Australia currently utilising a fully validated and National Association of Testing Authorities (NATA)-accredited ultrasensitive next-generation sequencing (NGS) platform for MRD monitoring in ALL. This technology is based on the detection of clonal rearrangement of immunoglobulin and T cell receptor genes in leukaemic cells, and is capable of achieving a limit of detection at least one to two logs below that of multiparametric flow cytometry (MFC). In this retrospective analysis, we report a clonotype detection rate of up to 85.7% at diagnosis, and a concordance rate of 78.7% in MRD results between NGS and MFC. Of the discordant samples, nearly all were NGS+/MFC-, highlighting the superior sensitivity of NGS. The enhanced sensitivity is clinically relevant, as discordant MRD results often heralded fulminant relapse, and therefore offer clinicians additional lead time and a window of opportunity to initiate pre-emptive therapy. Notwithstanding a small and heterogeneous cohort, our real-world survival data indicate an intermediate relapse risk for NGS+/MFC- patients. In light of recent approval of Medicare rebatable ALL MRD testing, we discuss how NGS can complement other techniques such as MFC in personalising management strategies. We recommend routine clonality testing by NGS at diagnosis and use a multi-modality approach for subsequent MRD monitoring.

Single-cell immune landscape of measurable residual disease in acute myeloid leukemia.

Measurable residual disease (MRD) is a powerful prognostic factor of relapse in acute myeloid leukemia (AML). We applied the single-cell RNA sequencing to bone marrow (BM) samples from patients with (n=20) and without (n=12) MRD after allogeneic hematopoietic stem cell transplantation. A comprehensive immune landscape with 184,231 cells was created. Compared with CD8+ T cells enriched in the MRD-negative group (MRD-_CD8), those enriched in the MRD-positive group (MRD+_CD8) showed lower expression levels of cytotoxicity-related genes. Three monocyte clusters (i.e., MRD+_M) and three B-cell clusters (i.e., MRD+_B) were enriched in the MRD-positive group. Conversion from an MRD-positive state to an MRD-negative state was accompanied by an increase in MRD-_CD8 clusters and vice versa. MRD-enriched cell clusters employed the macrophage migration inhibitory factor pathway to regulate MRD-_CD8 clusters. These findings revealed the characteristics of the immune cell landscape in MRD positivity, which will allow for a better understanding of the immune mechanisms for MRD conversion.

Immunophenotypic Profile and Measurable Residual Disease Monitoring in Multiple Myeloma: A Prospective Study From a Tertiary Care Centre in Southern India.

BACKGROUND: Multiple myeloma (MM) immunophenotyping (IPT) and measurable residual disease (MRD) monitoring by flow cytometry is a surrogate for progression-free survival and overall survival in clinical trials. However, plasma cell enumeration is challenging owing to morphological discrepancies and plasma cell (PC) loss during the sample processing. METHODS: In (n=87) newly diagnosed MM patients, we evaluated the immunophenotype of PCs at baseline, and for a subset of 35 patients MRD at post-induction was quantified and analyzed for association with outcomes and survival. The software Statistical Package for Social Sciences (SPSS), version 16.0 (SPSS Inc., Chicago, IL, USA) was used for all the statistical analysis. RESULTS: Immunophenotyping showed strong positive expression of CD56 (83%), CD200 (94%), CD38 (92%), and CD117 (91%) and negative/weak expression of CD19 (83%), CD45 (89%), CD27 (74%), and CD81 (90%) respectively. Negative/weak expression of CD19 was significantly associated with age ≥56 years (p<0.048), with lower albumin (<3.4g/dL, p<0.001). Strong positive CD56 expression was significantly associated with the presence of M-protein (p<0.03). Strong positive CD117 expression was significantly associated with lower albumin (p<0.02). Strong positive CD200 expression was significantly associated with a good response (p<0.02). The median (IQR) value of bone marrow (BM)-MRD% was 0.005 (0.002-0.034). We found that there was no significant difference in the correlation, association, and survival outcomes with MRD%. CONCLUSION: This study sheds light on the utility of IPT as an invaluable diagnostic tool in disease management. The findings of this study could be important when it comes to modifying the criteria for high-risk diseases and implementing a risk-adapted first therapy in clinical practice.

Indeterminate measurable residual disease by multiparameter flow cytometry is associated with an intermediate risk of clinical relapse in adult patients with acute leukaemia.

Measurable residual disease (MRD) is useful for prognostication and for monitoring response to treatment in patients with acute leukaemia. MRD by multiparametric flow cytometry (MFC-MRD) utilises the leukaemia-associated immunophenotype (LAIP) and difference from normal (DfN) strategies to identify the leukaemic clone. Difficulties arise when the LAIP overlaps with normal regeneration, there is clonal evolution, or when the abnormal clone population is exceptionally small e.g., <0.01% of CD45+ cells. Such cases are reported as 'indeterminate'; however, there is little international consensus on this reporting. The relationship between clinical outcomes and indeterminate MFC-MRD is unknown. Here we determine the rate of indeterminate MFC-MRD reporting, its relationship to concurrent molecular MRD results when available, and to clinical outcomes to 12 months. We performed an internal audit of all adult testing for MFC-MRD between January and December 2021. A total of 153 consecutive patients with a diagnosis of acute leukaemia were included. Successive MFC-MRD results and clinical outcomes were recorded over a 12-month period from time of inclusion into the study. In total, 460 MFC-MRD tests from 153 patients were reviewed and 73 (16%) MFC-MRD tests from 54 (35%) patients were reported as indeterminate. The majority (70%) were at low levels between 0.01-0.1% of CD45+ cells. Compared to patients with a negative result, acute myeloid leukaemia (AML) was more frequent in patients who had an indeterminate MFC-MRD (70% vs 36%), and B-cell acute lymphoblastic leukaemia was less common (20% vs 55%). In patients with indeterminate MFC-MRD results, one-third had received either chemotherapy or allogeneic haemopoietic stem cell transplant (aHSCT) within the preceding 3 months. Agreement between MFC and molecular MRD testing was low. Patients with indeterminate MFC-MRD had leukaemia relapse rates below patients with a positive MFC-MRD, but greater than those with negative MFC-MRD (positive 33% vs indeterminate 21% vs negative 8%, p = 0.038). Overall, these findings indicate that indeterminate MFC-MRD results are more common in adults with AML and also in those who have received chemotherapy or aHSCT within the previous 3 months. We report for the first time that indeterminate MFC-MRD is a finding of potential clinical significance, which associates with a numerically higher median relapse rate within 12 months when compared to a negative MFC-MRD result.

Optimizing maintenance therapy in acute myeloid leukemia: where do we stand in the year 2024?

INTRODUCTION: Despite the prognosis of patients affected by acute myeloid leukemia (AML) improved in the last decade, most patients relapse. Maintenance therapy after a chemotherapy approach with or without allogeneic stem cell transplantation could be a way to control the undetectable residual burden of leukemic cells. Several studies are being carried out as maintenance therapy in AML. Some critical points need to be defined, how the physician can choose among the various drugs available. AREAS COVERED: This review discusses the advances and controversies surrounding maintenance therapy for AML patients. EXPERT OPINION: Patients withFLT3-positive AML should receive midostaurin or quizartinib in the first-linesetting. For a patient initially receiving midostaurin, consider switching to sorafenib in the post-transplant setting. Because of the improved safety profile and potency, many experts will lean toward using a second-generation FLT3 inhibitor such as quizartinib or gilteritinib. Finally, no data indicate whether maintenance therapy should be prolonged until progression or for a defined period.

Measurable residual disease testing by next generation sequencing is more accurate compared with multiparameter flow cytometry in adults with B-cell acute lymphoblastic leukemia.

Results of measurable residual disease (MRD)-testing by next-generation sequencing (NGS) correlate with relapse risk in adults with B-cell acute lymphoblastic leukemia (ALL) receiving chemotherapy or an allotransplant from a human leukocyte antigen (HLA)-identical relative or HLA-matched unrelated donor. We studied cumulative incidence of relapse (CIR) and survival prediction accuracy using a NGS-based MRD-assay targeting immunoglobulin genes after 2 courses of consolidation chemotherapy cycles in 93 adults with B-cell ALL most receiving HLA-haplotype-matched related transplants. Prediction accuracy was compared with MRD-testing using multi-parameter flow cytometry (MPFC). NGS-based MRD-testing detected residual leukemia in 28 of 65 subjects with a negative MPFC-based MRD-test. In Cox regression multi-variable analyses subjects with a positive NGS-based MRD-test had a higher 3-year CIR (Hazard Ratio [HR] = 3.37; 95 % Confidence Interval [CI], 1.34-8.5; P = 0.01) and worse survival (HR = 4.87 [1.53-15.53]; P = 0.007). Some data suggest a lower CIR and better survival in NGS-MRD-test-positive transplant recipients but allocation to transplant was not random. Our data indicate MRD-testing by NGS is more accurate compared with testing by MPFC in adults with B-cell ALL in predicting CIR and survival. (Registered in the Beijing Municipal Health Bureau Registration N 2007-1007 and in the Chinese Clinical Trial Registry [ChiCTR-OCH-10000940 and ChiCTROPC-14005546]).

Flow Cytometry-Based Detection of Minimal/Measurable Residual Disease Predicts Survival Outcomes in Pediatrics, Adolescents, and Young Adults With T-acute Lymphoblastic Leukemia.

BACKGROUND: Measurable/minimal residual disease (MRD) is considered the single most powerful high-risk factor in acute leukemia, including T-cell acute lymphoblastic leukemia (T-ALL). In this study, we evaluated the impact of flow cytometry (FC)-based detection of MRD on survival outcomes in pediatrics, adolescents, and young adults (AYA) with T-ALL. METHODS: We included 139 patients, 88 pediatric patients between the ages of one and 14 years, and 51 AYA patients between 15 and 39 years of age, over a period of three years and were treated with the Indian Collaborative Childhood Leukemia Group (ICiCLe) protocol. MRD assessment was performed on post-induction (PI) bone marrow aspirate samples using a 10-color 11-antibody MRD panel on a Gallios instrument (Beckman Coulter, Miami, FL, USA). MRD value > 0.01% was considered positive. PI-MRD status was available in 131 patients. RESULTS: The five-year event-free survival (5-year EFS) in PI-MRD positive patients was inferior to those of negative patients (13.56% vs 79.06%), which was statistically significant (P < 0.001). However, the five-year overall survival (5-year OS) did not show any statistically significant difference between PI-MRD positive and negative T-ALL patients (92.93% vs 94.28%). The hazard ratio (HR) for 5-year EFS and MRD positivity was 8.03 (p-value < 0.0001). HR for 5-year EFS and early T-cell precursor ALL (ETP-ALL) was 2.63 (p = -0.02). CONCLUSIONS: PI-MRD detected using FC is a strong predictive factor of inferior survival outcomes in pediatrics, AYA patients with T-ALL. PI-MRD positivity can be used to modify the treatment of T-ALL patients, especially in resource-constrained developing countries where molecular tests are not widely available.

Monitoring measurable residual disease in paediatric acute lymphoblastic leukaemia using immunoglobulin gene clonality based on next-generation sequencing.

BACKGROUND: Assessment of measurable residual disease (MRD) is an essential prognostic tool for B-lymphoblastic leukaemia (B-ALL). In this study, we evaluated the utility of next-generation sequencing (NGS)-based MRD assessment in real-world clinical practice. METHOD: The study included 93 paediatric patients with B-ALL treated at our institution between January 2017 and June 2022. Clonality for IGH or IGK rearrangements was identified in most bone marrow samples (91/93, 97.8%) obtained at diagnosis. RESULTS: In 421 monitoring samples, concordance was 74.8% between NGS and multiparameter flow cytometry and 70.7% between NGS and reverse transcription-PCR. Elevated quantities of clones of IGH alone (P < 0.001; hazard ratio [HR], 22.2; 95% confidence interval [CI], 7.1-69.1), IGK alone (P = 0.011; HR, 5.8; 95% CI, 1.5-22.5), and IGH or IGK (P < 0.001; HR, 7.2; 95% CI, 2.6-20.0) were associated with an increased risk of relapse. Detection of new clone(s) in NGS was also associated with inferior relapse-free survival (P < 0.001; HR, 18.1; 95% CI, 3.0-108.6). Multivariable analysis confirmed age at diagnosis, BCR::ABL1-like mutation, TCF3::PBX1 mutation, and increased quantity of IGH or IGK clones during monitoring as unfavourable factors. CONCLUSION: In conclusion, this study highlights the usefulness of NGS-based MRD as a routine assessment tool for prognostication of paediatric patients with B-ALL.

Blinatumomab in Children with MRD-Positive B-Cell Precursor Acute Lymphoblastic Leukemia: A Report of 11 Cases.

Background/Objectives: Relapsed B-cell acute lymphoblastic leukemia (B-ALL) remains an unresolved matter of concern regarding adverse outcomes. This case study aimed to evaluate the effectiveness of blinatumomab, with or without door lymphocyte infusion (DLI), in treating measurable residual disease (MRD)-positive B-ALL. Methods: All patients who received blinatumomab salvage therapy were included in this study. Eleven patients were included in the study. All patients were evaluated for MRD-negativity. Results: Before starting blinatumomab therapy, seven patients tested positive for MRD, three tested negative, and one had refractory disease. Hematopoietic cell transplantation (HCT) was reserved for five patients with persistent MRD. Six patients became MRD-negative and subsequent HCT was not performed. Only two patients relapsed; one patient died of relapse, and the other one received carfilzomib-based therapy and was MRD-negative thereafter. Nine patients were MRD-negative at a median follow-up of 28 months (15-52 months). Two of three MRD-positive post-transplant patients remained in complete molecular remission after preemptive DLI at the last follow-up date. In the first salvage, blinatumomab may achieve complete remission and bridging to HCT in pediatric patients with end-of-induction MRD-positive B-cell precursor ALL. Conclusions: The decision on how to treat post-transplant relapse continues to affect survival outcomes. Blinatumomab combined with DLI may extend the armamentarium of release options for high-risk pediatric patients. This approach is encouraging for high-risk ALL patients who are MRD-positive post-transplantation.

Monitoring molecular changes in the management of myelodysplastic syndromes.

The ongoing or anticipated therapeutic advances as well as previous experience in other malignancies, including acute myeloid leukaemia, have made molecular monitoring a potential interesting tool for predicting outcomes and demonstrating treatment efficacy in patients with myelodysplastic syndromes (MDS). The important genetic heterogeneity in MDS has made challenging the establishment of recommendations. In this context, high-throughput/next-generation sequencing (NGS) has emerged as an attractive tool, especially in patients with high-risk diseases. However, its implementation in clinical practice still suffers from a lack of standardization in terms of sensitivity, bioinformatics and result interpretation. Data from literature, mostly gleaned from retrospective cohorts, show NGS monitoring when used appropriately could help clinicians to guide therapy, detect early relapse and predict disease evolution. Translating these observations into personalized patient management requires a prospective evaluation in clinical research and remains a major challenge for the next years.

FLT3-TKD Measurable Residual Disease Detection Using Droplet Digital PCR and Clinical Applications in Acute Myeloid Leukemia.

The tyrosine kinase domain of the FMS-Like tyrosine kinase 3 (FLT3-TKD) is recurrently mutated in acute myeloid leukemia (AML). Common molecular techniques used in its detection include PCR and capillary electrophoresis, Sanger sequencing and next-generation sequencing with recognized sensitivity limitations. This study aims to validate the use of droplet digital PCR (ddPCR) in the detection of measurable residual disease (MRD) involving the common FLT3-TKD mutations (D835Y, D835H, D835V, D835E). Twenty-two diagnostic samples, six donor controls, and a commercial D835Y positive control were tested using a commercial Bio-rad® ddPCR assay. All known variants were identified, and no false positives were detected in the wild-type control (100% specificity and sensitivity). The assays achieved a limit of detection suitable for MRD testing at 0.01% variant allelic fraction. Serial samples from seven intensively-treated patients with FLT3-TKD variants at diagnosis were tested. Five patients demonstrated clearance of FLT3-TKD clones, but two patients had FLT3-TKD persistence in the context of primary refractory disease. In conclusion, ddPCR is suitable for the detection and quantification of FLT3-TKD mutations in the MRD setting; however, the clinical significance and optimal management of MRD positivity require further exploration.