Minimal Residual Disease Status In Patients Research Articles

Myeloid Derived Suppressor Cells and Their Potential Impact on Minimal Residual Disease Status and Disease Response in a Cohort of Patients with Plasma Cell Dyscrasias

Myeloid Derived Suppressor Cells (MDSCs) are known to promote immune-suppressive and tumorigenic effects in plasma cell dyscrasias (PCDs), including multiple myeloma, and may play a role in treatment response and minimal residual disease MRD status in patients undergoing cancer-directed therapies. MDSCs are classified by their granulocyte-like effects (g-MDSCs) or monocyte-like effects (m-MDSCs). To our knowledge, there are no published studies that correlate minimal residual disease (MRD) with the presence of MDSCs and immune effector-cell populations in the blood and bone marrow of patients with PCDs. In 14 patients with PCDs, we defined immune-cell populations in the blood and marrow and correlated them with MRD status, treatment regimens, disease status (stable vs. progressive) and autologous stem cell transplantation status. In the entire cohort, there were a higher proportion of m-MDSCs in blood (blood 5.56%, aspirate 2.47%). There were a higher proportion of g-MDSCs in aspirate (blood 0.04%, aspirate 0.16%). Interestingly, in patients who remained MRD positive (3/13), the proportions were reversed, and we identified more m-MDSC in aspirate than blood, as well as a low proportion of g-MDSCs in both the aspirate and bone marrow. In patients achieving undetectable MRD (10/13) there were higher proportions of CD8 and CD4 T-cells, with no differences in MDSC populations. Not surprisingly, in patients treated with the anti-CD38 antibody daratumumab (5/14), fewer m-MDSCs were isolated in the blood and aspirate, relative to daratumumab-untreated patients (9/14).Daratumumab treatment decreased activated CD4 and CD8 T cells (HLA-DR+/CD38-, HLA-DR-/CD38+ or HLA-DR+/CD38+ CD4 and CD8 T cells) and increased the proportion of inactive HLA-DR-/CD38- CD4 and CD8 T-cells. In patients with progressive disease (3/14), there was a paucity of all immune effector-cells, including MDSCs, T-cells and monocytes with the exception of neutrophils. MDSC populations were no different in pre-transplant patients (3/14) compared to post-transplant (11/14). In this preliminary study, we have enumerated the presence of MDSCs and their potential impact in patients with PCDs undergoing cancer-directed therapy.

Identifying Primary Refractory Disease and Early Relapses in Peripheral T-Cell Lymphoma Using Baseline Characteristics, Minimal Residual Disease, and Mutational Profiling

INTRODUCTION: Despite approvals in the relapsed/refractory setting for T-cell lymphomas, upfront advances have been more difficult to realize. A critical component in improving treatment paradigms is a greater ability to identify patients at diagnosis with high-risk for chemorefractory disease who may benefit from alternative induction programs and distinguish them from patients who fare well with standard approaches. In order to characterize such cases, we analyzed patients treated at Memorial Sloan Kettering (MSK) with curative intent to identify features associated with primary refractoriness or early relapse (<1 year). METHODS: We identified patients ≥ 18 years of age with PTCL-NOS, AITL, or ALCL who either were consecutively treated at our center at the time of diagnosis or were seen during initial therapy from 1/1/2001-10/1/2021. All patients had histologically-confirmed disease and were treated with curative intent. Patients were grouped into two sub-cohorts: (1) primary refractory disease/early relapse (< 1 year), or (2) relapse-free at 1 year from initiation of first-line therapy. Patients without relapse but < 1 year of followup were excluded. A univariate logistic regression was performed to compare baseline features between the sub-cohorts using the Wilcoxon rank sum test and Pearson's Chi-squared test. A multivariate model was then constructed using histology and baseline IPI apriori and any factor significant in univariate analyses to produce odds ratios (OR) for primary refractory disease/early relapse. In a separate analysis of patients who underwent autologous transplant (autoHCT) in 1st remission, the impact of bone marrow (BM) minimal residual disease (MRD) on PFS was evaluated. Finally, in a subset of patients who underwent targeted mutational profiling at diagnosis (Ptashkin et al., Cancer Res 2019), genomic results were analyzed. Genes were characterized into oncogenic pathways (Sanchez-Vega et al., Cell 2018) and included in analyses if mutations were present in ≥ 5% of patients. A Cox regression model was performed for altered genes/pathways to obtain hazard ratios (HR) for progression/death, with significance testing using the false discovery rate correction for multiple testing (q-value). RESULTS: A total of 349 patients were identified (Table 1). With a median followup time of 60 months (m) among survivors (IQR: 23-95), median PFS and OS of the entire cohort was 19 (95% CI 13-28) and 73 m (60-128), respectively. By histology, 2-year PFS for PTCL-NOS, AITL, ALK- ALCL, and ALK+ ALCL was 33%, 43%, 66%, and 79%, respectively (p<0.001); 5-year OS was 41%, 54%, 75%, and 92%, respectively (p<0.001, Figure 1). In comparing those with primary refractory disease/early relapse versus those relapse-free at 1-year, patients with PTCL-NOS, stage III/IV disease, >1 extranodal site, BM involvement, and cutaneous involvement were significantly more likely to have primary refractory disease/early relapse. On multivariable analysis, a diagnosis of PTCL-NOS (OR for AITL: 0.42, 95% CI 0.22-0.81), advanced IPI (OR for IPI 4-5: 4.88, 95% CI 1.56-16.8), BM involvement (OR: 2.28, 95% CI 1.14-4.68), and cutaneous involvement (OR: 2.47, 95% CI 1.04-6.21) all conferred increased risk of primary refractory disease/early relapse. In the subgroup analysis of BM MRD status in patients who underwent autoHCT, the presence of MRD did not significantly impact PFS (5-year PFS 45% MRD+ vs. 45% MRD, p=0.55). Finally, in evaluation of mutational profiles (n=86), mutations in TET2 (59%), RHOA (30%) and DNMT3A (23%) were the most commonly altered genes. Although rare (n=6), alterations in DNA damage response pathways significantly increased risk of progression (HR 5.08, 95% CI 2.05-12.6, q-value=0.007) and death (HR 9.9, 95% CI 3.22-30.8, q-value=0.001). CONCLUSIONS: This is one of the largest cohorts of nodal PTCL in the 21st century. Despite prolonged survival in some patients, primary refractory disease/early relapses remain common. On multivariate analyses, a histological diagnosis of PTCL-NOS, advanced IPI, BM involvement, and cutaneous involvement predicted primary refractoriness/early relapse. Efforts to improve frontline strategies in PTCL should focus on identifying and exploiting biological vulnerabilities in such patients, and trials of intensified regimens/alternative platforms should prioritize evaluation of these patients. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Abstract 2793: Minimal residual disease (MRD) as a predictive biomarker/endpoint for novel drug development

Abstract MRD is defined as the continual presence of cancer cells in patients post therapy that is below the threshold of detection by conventional morphologic or radiologic assessments. Monitoring patient status clinically as MRD negative or MRD positive has become the basis for predicting outcomes, remission or reoccurrence, as well as determining therapeutic options. The three primary methods to assess MRD status in patients include Flow Cytometry (FC), Polymerase Chain Reaction (PCR) and Next-Generation Sequencing (NGS). All three are highly quantitative measurements with a high degree of sensitivity to detect “new” circulating tumor cells or ctDNA that can potentially indicate disease recurrence before it happens. The choice of MRD methodology usually depends on the tumor type, liquid or solid. Flow Cytometry has been widely used to monitor MRD status in patients with hematologic malignancies. Specifically designed MRD panels have been used clinically in CLL, MM, CML and AML patients. In addition to its prognostic/diagnostic relevance, recent guidance by the FDA is now driving the use of MRD testing as a predictive biomarker/endpoint for novel cancer drug development. NeoGenomics Pharma Services has provided flow cytometry MRD expertise to multiple sponsors conducting clinical trials to evaluate the effectiveness of novel cancer therapy. In a CLL study, NeoGenomics employed the ERIC MRD assessment to monitor patient status during therapeutic intervention at various points throughout treatment. Representative patient data show progression from full, active disease prior to therapy to non-detectable disease levels below 0.01%MRD. These examples show patient progression through multiple cycles of treatment to full remission. Clearly, this is an example of how MRD can serve as the primary “biomarker” to evaluate a patient’s response to novel drug therapy in clinical trials. This presentation will further discuss the data and the relevance of MRD testing to support new drug development. Citation Format: Nicholas Jones, Brian Ngo, Floyd Davis, Benjamin Fancke. Minimal residual disease (MRD) as a predictive biomarker/endpoint for novel drug development [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2793.

Prognostic value of early bone marrow MRD status in CAR-T therapy for myeloma.

8022 Background: Bone marrow (BM) assessment of minimal residual disease (MRD) is being considered as a surrogate endpoint in clinical trials and is prognostic for survival in multiple myeloma (MM). Timing of BM assessment is variable across Chimeric Antigen Receptor T cell (CART) therapy trials and differs from standard of care practice. BM myeloma cell clearance can be detected by month 1 post CART, even before serum immunofixation becomes negative. BM is still hypocellular at month 1, thus prognostic value of MRD negative (MRDneg) at this timepoint is unclear. We examined impact of Day 30 MRD status in patients (pts) who received CART at Mayo Clinic. Methods: Medical records were reviewed retrospectively for MM pts who received CART between 8/2016 and 6/2021. PFS and OS were plotted by Kaplan-Meier method. Results: Sixty MM pts received CART and had BM biopsy at month 1. Median age was 62 yrs, 53% were male, and 78% were BM MRDneg by flow cytometry. Baseline demographics were similar between MRDneg and MRD+ (Table). Overall, 85% (40/47) who were month 1 BM MRDneg had i/u FLC&lt;normal. Patients who achieved CR/sCR had higher rates of BM MRDneg (100% vs 61%, p&lt;0.001) and i/u FLC&lt; normal (89% vs 58%, p&lt;0.001). At month 1, 24/60 (40%) pts had hypocellular BM. Serial BM samples at month 3 (n=35), 6 (n=28) and 12 (n=23) showed MRDneg rate of 93% (25/27), 56% (9/16) 58% (7/12), respectively.. Rate of hypocellularity was 54% (19/35), 32% (9/28) and 30% (7/23), respectively. Among the MRDneg/hypocellular pts at month 1, hypocellular BM was seen in 8/11 (73%) pts at month 3 and 2/4 (50%) pts at month 6 and 12. Compared to MRD+, pts who had BM MRDneg at months1 had longer PFS (Table). PFS was not statistically significantly different between pts who had BM MRDneg and were either hypocellular or not. MRDneg pts with i/u FLC&lt;normal at months1 had better median PFS compared to those who did not. (MRD+:2.9 months (1.2-NR). MRDneg/FLC&gt;normal: 4.9 months (2.3-NR). vs MRDneg/FLC&lt;normal:17.9 months (11.8-NR), p&lt;0.0001). Conclusions: Hypocellular BM is common in the 3 months post CART. Regardless of BM cellularity, BM MRDneg at month 1 correlates with deep response and prolonged PFS. Majority of BM MRDneg pts at month 1 also had FLC&lt;normal. BM MRDneg status and FLC normalization were associated with longer survival. Our data support the continued evaluation of BM early post CART infusion as a prognostic tool. [Table: see text]

Challenging the Prognostic Value of MRD: Sequential Tumor-Ablative Conditioning of FA5-Bucy for Ph-Positive Acute Lymphoblastic Leukemia

Background:Minimal residual disease (MRD) quantification prior to allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a powerful predictor for post-transplant outcome. Thus, new strategies with modified transplantation procedures are needed to reverse the outcomes of that remain MRD-positive despite multiple induction attempts.OBJECTIVE:To determine the prognostic relevance of MRD status in patients with acute lymphoblastic leukemia (ALL), especially those with Philadelphia chromosome-positive ALL (Ph+ ALL), receiving the conditioning regimen of FA5-Bucy, registered on http:// ClinicalTrials.gov (NCT02328950).METHODS:We retrospectively reviewed our HSCT experience using FA5-Bucy, i.e., 5-day salvage chemotherapy (Fludarabine/Ara-C) and conditioning (Busulfan/ Cyclophosphamide) for patients with ALL. A total of 48 consecutive patients from 2013 to 2016 are analyzed. 11 out of 48 (7 males and 4 females) were diagnosed with Ph+ ALL with the induction treatments including a tyrosine kinase inhibitor (TKI). Median age was 30 years (range, 8-52 years). Among whom four (36.4%) achieved complete remission(CR)at transplantation, Six (54.5%) with detectable residual P190 BCR-ABL positive leukaemic cells, while 1 (9.1%) not in the status of remission with 39.5% blasts in the bone marrow (BM). The other 37 cases were Ph-negative (Ph-) ALL, with the median age of 21 years (range, 2-61 years). Among whom 25 (67.6%) patients were in CR, 6 (16.2%) MRD-positive and 6 (16.2%) with 23.5%-98% BM blasts. The MRD status was monitored by four-color FCM in Ph- group, and by RT-PCR to detect the chimeric bcr-abl mRNA transcript in Ph+ group. Patients were transplanted either from an HLA-identical sibling or haplo-identical related donors. The GVHD prophylaxis consisted of ATG, CsA, MMF and short-term MTX. None of these patients had severe infection or organ failure before HSCT.RESULTS:All patients achieved successful engraftment and full donor chimerism. Patients with Ph+ ALL seems have a better outcome than that with Ph- ALL (3-year OS of 54.5% vs 37.8%, P=0.245). With a median follow-up of 666 (83-1276) days, the 1y-OS and 1y-DFS were 72.7%, 3y-OS and 3y-DFS were 54.5% in Ph+ group, respectively, and non-relapse mortality and relapse incidence were 18.2% and 10%. MRD monitoring is an important tool for relapse prediction in Ph- ALL patients (3-year OS and DFS of MRD-positive vs MRD-negative =16.7% vs 56%, P=0.021).However, the profile of OS and DFS was similar in Ph+ patients with bcr-abl positive to that with bcr-abl negative (3-year OS and DFS of bcr-abl positive vs bcr-abl negative= 57.1% vs 50%, P=0.744). Severe acute-GvHD (aGVHD) occurred in 3/11 patients. Few patients experienced mild-to-moderate toxicity, and main causes of death were aGVHD.CONCLUSION:We concluded that allo-HSCT with the sequential tumor-ablative conditioning of FA5-Bucy offers great benefits for the patients with Ph+ ALL, with the potentially capability of reducing or overcoming the negative impacts of MRD. Thus creates exciting transplant opportunities for patients not in remission to be cured. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Minimal Residual Disease (MRD) Detection By Flow Cytometry during Aplasia and Post-Induction Is an Independent Prognostic Parameter for Relapse-Free Survival in AML

The identification of patients at high risk of relapse is crucial for treatment stratification in acute myeloid leukemia (AML). Persistence and reoccurrence of AML cells with aberrant phenotypes can be assessed by flow cytometry at high sensitivity in more than 95% of patients. Therefore, assessment by flow cytometry enables determination of MRD status in patients without suitable molecular markers (e.g. NPM1). Previously, we demonstrated that early detection of MRD during aplasia is associated with shorter relapse-free survival (RFS) after intensive induction chemotherapy (Köhnke T et al., Leukemia 2015). Here, we aimed to validate these findings in an independent patient cohort. In addition, we assessed the prognostic value of a combined MRD determination during aplasia and post induction for improved prediction of relapse-free survival (RFS).Flow cytometry MRD was analyzed in patients with AML (excluding APL) diagnosed between 2012 and 2016 receiving intensive induction chemotherapy (sHAM or 7+3). MRD flow analysis was performed using a NAVIOS flow cytometer. Presence ≥0.1% Leukemia-associated immunophenotype (LAIP)-positive cells (identified by comparison with bone marrow from healthy donors) during aplasia or post-induction defined MRD positivity. Kaplan-Meier estimators and log-rank tests were used to retrospectively analyze survival data. Cox's proportional hazards regression model was used to determine the influence of individual factors in multivariate analyses. Pearson's chi square test was used to compare categorical variables between MRD groups.A total of 105 patients were included. In 3 cases (2.9% of all cases), no LAIP could be identified, and these patients were excluded from further analyses. MRD assessment during aplasia was available in 87 cases. Regarding flow MRD assessment post induction, patients refractory to induction therapy were excluded. Therefore, 72 patients had flow MRD assessments available post induction therapy. 59 patients achieving CR or CRi after induction therapy had flow MRD assessments available at both time points.MRD positivity during aplasia was associated with refractory disease (36.2% vs. 10.0% for MRD pos. vs. MRD neg., p=0.017) and shorter median event-free survival (EFS, 5.8 vs. 15.7 months, p=0.011). Importantly, early blast clearance by morphology had no prognostic value if flow MRD assessment was available (HR for EFS 2.0, p=0.03 for flow MRD vs. HR 1.6 and p=0.17 for Blast clearance by morphology). However, differences in median RFS for MRDpos vs. MRDneg patients (10.3 vs. 20.5 months) for assessment during aplasia have not reached significance yet (p=0.135). MRDpos patients post induction had significantly shorter median EFS (10.8 vs. 15.7 months, p=0.032) and RFS (7.7 months vs. not reached, p=0.007) than MRDneg patients. Using a combination strategy for aplasia and post induction assessments, we grouped patients with MRD positivity at both timepoints (confirmed MRD positivity) vs. all others (MRDneg or MRDuncertain). Confirmed MRD positivity was associated with significantly shorter median EFS (6.7 vs. 15.7 months, p=0.003) as well as RFS (4.4 months vs. not reached, p=0.002). Confirmed MRD positivity was an independent predictor of EFS and RFS (HR 2.7 and 2.8, p=0.017 and p=0.018, respectively).Taken together, patients with confirmed two-time flow MRD positivity showed lower EFS and RFS compared to MRDneg or MRDuncertain patients, respectively. Identification of patients with poor treatment response was improved in comparison with cytomorphologic assessment or independent flow MRD assessments at both timepoints alone. Using this approach, early identification of patients with adverse prognosis is possible in >95% of AML patients. MRD flow will be prospectively evaluated as a therapy stratification tool in patients with AML receiving intensive induction chemotherapy. DisclosuresNo relevant conflicts of interest to declare.

Intestinal Microbiota Composition Is Associated with Minimal Residual Disease Negativity in Patients with Multiple Myeloma

Background:Multiple myeloma (MM) patients who achieve minimal residual disease (MRD) negative status after upfront treatment have prolonged progression-free and overall survival compared with those who remain MRD(+) (Landgren CO, Devlin SM et al. Bone Marrow Transplant. 2016;51(12):1565-8). Commensal intestinal microbial composition has been associated with treatment outcomes in cancer patients. We sought to evaluate whether the composition of the intestinal microbiota is associated with MRD status in patients with MM.Methods:Stool samples were collected prospectively from 34 patients after completion of upfront therapy for MM at the time of MRD testing. MRD was assessed with next-generation flow cytometry of bone marrow aspirates (sensitivity 10-5). Microbial analysis was performed via sequencing of 16S rRNA V4-V5 regions using the Illumina MiSeq platform and sequence data was analyzed using UPARSE (Edgar RC, Nature Methods 2013;10(10):996-8). The linear discriminant effect size method (LEfSe) (Segata N et al. Genome Biol. 2011;12(6):R60.) was used to compare detected clades among all groups and evaluate for associations with outcomes, using MRD as class and autologous stem cell transplant (ASCT) as subclass. Alpha diversity was calculated by the Inverse Simpson index and differential relative abundance were calculated using the phyloseq package and compared using the Wilcoxon rank sum test on the R statistical computing platform.Results:Among 34 patients evaluable for microbiota composition and MRD status, the median age was 62.5 years and 16 (47.1%) were MRD(-) at time of stool collection. 24 (70.6%) were treated with carfilzomib, lenalidomide, and dexamethasone as induction therapy (MRD(-): 14 (87.5%), MRD(+):10 (55.5%). 4 (28.5%) MRD(-) patients had autologous stem cell transplant(ASCT), compared with 10 (55.5%) who were MRD(+). In the cohort's samples, we observed 19 phyla, 315 genera, 654 species, and 1549 operational taxonomic units (OTUs). There was no significant difference in alpha diversity between MRD(-) (median 12.24, IQR = 8.76-13.98) and MRD(+) patients (median 12.44, IQR = 8.36 -16.23), p=0.6 by Wilcoxon rank sum test. A positive association with MRD negativity was noted with two butyrate-producing organisms, Eubacterium hallii (p=0.001) and Faecalibacterium prausnitzii (p= 0.006). To further evaluate these relationships, we performed a differential abundance analysis of these selected taxa in MRD(+) and MRD(-) patients at the genus and species level. The relative abundance of the genera Eubacterium and Faecalibacterium were higher in fecal samples from MRD(-) patients than MRD(+) patients (Eubacterium MRD(-): median 4.51% (IQR = 2.83 - 7.32%) vs. MRD(+): median 3.07% (IQR = 1.35 - 3.87%), p=0.0326; Faecalibacterium MRD(-): median 1.68% (IQR = 0.69 - 7.48%) vs. MRD(+): median 0.003% (IQR = 0 - 3.19%), p=0.022. The relative abundance of both species of interest were higher in MRD(-) patients than in MRD(+) patients: E. hallii MRD(-): median 2.67% (IQR = 2.11 - 3.98%) vs. MRD(+): median 1.01% (IQR = 0 - 2.16%), p=0.001; F. prausnitzii MRD(-): median 1.43% (IQR = 0.53 - 7.28%) vs. MRD(+): median 0.3%, (IQR = 0 - 2.54%), p=0.022. Other species of Eubacterium and Faecalibacterium were not significantly differentially abundant between the two groups.Conclusions:Intestinal microbiota containing several butyrate-producing anaerobes appear to be associated with MRD-negativity in patients with myeloma, with higher relative abundance of Eubacterium hallii and Faecalibacterium prausnitzii in MRD(-) patients compared with MRD(+) patients. Butyrate and other short-chain fatty acids are biologically active metabolites formed during microbial fermentation of dietary or host-derived carbohydrates, which supply the host with energy and also modulate immunity, including exerting anti-inflammatory functions. Microbes of the genus Eubacterium have been associated with reduced risk of relapse in several hematologic cancers after allogeneic hematopoietic cell transplantation, including MM (Peled JU, Devlin SM et al. J Clin Oncol 2017;35(15):1650-9). This is first study to our knowledge to suggest an association between gut microbiota and MRD status in patients with myeloma and supports further investigation of a potential role for intestinal microbiota in the natural history and treatment of myeloma. DisclosuresPeled:Seres Therapeutics: Research Funding. Landgren:Karyopharm: Consultancy; Merck: Membership on an entity's Board of Directors or advisory committees; Pfizer: Consultancy; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Consultancy, Research Funding; Amgen: Consultancy, Research Funding; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. Lesokhin:Squibb: Consultancy, Honoraria; Genentech: Research Funding; Bristol-Myers Squibb: Consultancy, Honoraria, Research Funding; Serametrix, inc.: Patents & Royalties: Royalties; Takeda: Consultancy, Honoraria; Janssen: Research Funding.

Minimal Residual Disease (MRD) Detection By Flow Cytometry Complements Molecular MRD Assessment in AML

The prognosis of patients with AML is determined by a multitude of recurrent genetic alterations, and treatment algorithms heavily rely on risk stratification by genetic characterization of the disease at the time of diagnosis. However, this a priori risk stratification does not integrate information on treatment susceptibility of the individual patient. Assessment of Minimal Residual Disease (MRD) aims to implement this information in the patient-specific treatment management. AML cells with aberrant phenotypes can be detected at sensitivities below 1:104 by flow cytometry in the majority of patients. Therefore, flow cytometry MRD assessment (flow MRD) enables determination of MRD status in patients without suitable molecular markers (e.g. NPM1, CBFß-MYH11, and RUNX1-RUNX1T1). Here, we validate the role of flow MRD in AML patients receiving intensive chemotherapy with and without available molecular markers.Flow MRD was analyzed in patients with AML (excluding APL) diagnosed between 2012 and 2017 receiving intensive induction chemotherapy (sHAM or 7+3). Flow MRD analysis was performed during aplasia (on day 16 after treatment initiation) as well as post induction. Presence of ≥0.1% Leukemia-associated immunophenotype (LAIP)-positive cells was defined as flow MRD positivity. Molecular MRD was analyzed post induction for NPM1 and CBF, and post consolidation for RUNX1-RUNX1T1. Kaplan-Meier estimators and log-rank tests were used to analyze survival data. Cox's proportional hazards regression model was used to determine the influence of individual factors in multivariate analyses.A total of 161 patients were included. In 5 cases (3.1% of all cases), no LAIP could be identified, and these patients were excluded from further analyses. Flow MRD assessment during aplasia was available in 145 cases. 122 patients had flow MRD assessments available post induction. 114 patients achieving CR or CRi after induction therapy had flow MRD assessments available at both time points. Flow MRD positivity during aplasia was associated with shorter event free survival (EFS, 6.1 months vs. 19.1 months, p<0.001). Similarly, flow MRD positivity post induction was associated with shorter EFS (11.9 months vs. median not reached, p=0.007). For both timepoints, flow MRD was an independent risk factor in multivariate analysis compared to known risk factors such as age, genetic/molecular risk profile as determined by the ELN2017 risk categories as well as early blast clearance by morphology. Persistent flow MRD positivity at both timepoints (combined flow MRD) identified patients with particularly short EFS (8.2 months), whereas patients with flow MRD negativity at both time points had the best outcome in our cohort (median not reached, p=0.002). Combined flow MRD status was an independent predictor of EFS and RFS (HR 1.9 and 1.8, p=0.001 and p=0.007, respectively), whereas blast clearance by morphology had no significant prognostic impact (p>0.05 for all endpoints).64/161 patients (39%) had molecular MRD assessment available for analysis. In these patients, molecular MRD positivity predicted a significantly shorter EFS (9.3 months vs. median not reached, p=0.01). Indeed, molecular MRD positivity was an independent risk factor for adverse EFS and RFS (HR 1.7 and 1.6, p=0.008 and p=0.018, respectively). In this subgroup, flow MRD was not an independent prognostic factor. However, for patients without available molecular MRD marker (97/161 patients), flow MRD positivity at aplasia (p=0.004), post induction (p=0.015) or as combined status (p=0.004) was associated with a significantly shorter EFS and remained an independent risk factor in multivariate analysis (HR 2.5 and 2.6, p=0.016 and p=0.012 for EFS and RFS, respectively).Taken together, we demonstrate that both flow MRD as well as molecular MRD strongly correlate with survival. While molecular MRD assessment was only available in 39% of patients, MRD assessment by flow cytometry was feasible in >95% of AML patients. Flow MRD positivity both during aplasia and post induction was an independent risk factor, confirming the superiority of flow MRD compared to early morphologic response assessment. In conclusion, molecular and flow cytometric MRD assessment are complementing methods for the estimation of treatment response, and will be integrated in clinical trials to validate their significance for patient-specific treatment management. DisclosuresMetzeler:Celgene: Consultancy, Research Funding; Novartis: Consultancy. Hiddemann:F. Hoffman-La Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bayer: Consultancy, Research Funding; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Subklewe:Gilead Sciences: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Membership on an entity's Board of Directors or advisory committees; Roche AG: Research Funding; AMGEN: Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees.

Association of Minimal Residual Disease With Clinical Outcome in Pediatric and Adult Acute Lymphoblastic Leukemia

Minimal residual disease (MRD) refers to the presence of disease in cases deemed to be in complete remission by conventional pathologic analysis. Assessing the association of MRD status following induction therapy in patients with acute lymphoblastic leukemia (ALL) with relapse and mortality may improve the efficiency of clinical trials and accelerate drug development. To quantify the relationships between event-free survival (EFS) and overall survival (OS) with MRD status in pediatric and adult ALL using publications of clinical trials and other databases. Clinical studies in ALL identified via searches of PubMed, MEDLINE, and clinicaltrials.gov. Our search and study screening process adhered to the PRISMA Guidelines. Studies that addressed EFS or OS by MRD status in patients with ALL were included; reviews, abstracts, and studies with fewer than 30 patients or insufficient MRD description were excluded. Study sample size, patient age, follow-up time, timing of MRD assessment (postinduction or consolidation), MRD detection method, phenotype/genotype (B cell, T cell, Philadelphia chromosome), and EFS and OS. Searches of PubMed and MEDLINE identified 566 articles. A parallel search on clinicaltrials.gov found 67 closed trials and 62 open trials as of 2014. Merging results of 2 independent searches and applying exclusions gave 39 publications in 3 arms of patient populations (adult, pediatric, and mixed). We performed separate meta-analyses for each of these 3 subpopulations. The 39 publications comprised 13 637 patients: 16 adult studies (2076 patients), 20 pediatric (11 249 patients), and 3 mixed (312 patients). The EFS hazard ratio (HR) for achieving MRD negativity is 0.23 (95% Bayesian credible interval [BCI] 0.18-0.28) for pediatric patients and 0.28 (95% BCI, 0.24-0.33) for adults. The respective HRs in OS are 0.28 (95% BCI, 0.19-0.41) and 0.28 (95% BCI, 0.20-0.39). The effect was similar across all subgroups and covariates. The value of having achieved MRD negativity is substantial in both pediatric and adult patients with ALL. These results are consistent across therapies, methods of and times of MRD assessment, cutoff levels, and disease subtypes. Minimal residual disease status warrants consideration as an early measure of disease response for evaluating new therapies, improving the efficiency of clinical trials, accelerating drug development, and for regulatory approval. A caveat is that an accelerated approval of a particular new drug using an intermediate end point, such as MRD, would require confirmation using traditional efficacy end points.

The Relationship of Acute Gvhd and Pre- and Post-Transplant Flow-MRD to the Incidence and Timing of Relapse in Children Undergoing Allogeneic Transplantation for High Risk ALL: Defining a Target Population and Window for Immunological Intervention to Prevent Relapse

AbstractAbstract ▪470▪This icon denotes a clinically relevant abstractWe previously reported lower rates of grade II-IV aGVHD, but no improvement in survival with the addition of sirolimus to tacrolimus/methotrexate GVHD prophylaxis in a randomized phase III Children’s Oncology Group/Pediatric Blood and Marrow Transplant Consortium trial, ASCT 0431. We performed further analysis of this cohort to test whether the presence or absence of aGVHD combined with detection of pre- and/or post-HCT minimal residual disease (MRD) would allow definition of high-risk populations amenable to early intervention to prevent relapse. MRD was measured on patient bone marrow samples with multi-channel flow cytometry at a single reference laboratory (detection threshold 0.01%). The trial included children ages 1–21yrs with high-risk CR1 and CR2 ALL receiving related and unrelated donor TBI-based allogeneic HCT. The analysis included 144 eligible patients with an estimated median follow up of 757d (interquartile range 558–1022d).Both the absence of grade II-IV aGVHD at day 100 and positive MRD pre-and/or post-HCT were correlated with relapse. To illustrate the effect of aGVHD on relapse, Table 1 shows the cumulative incidence of TRM, relapse, and EFS at 100 days, 1 yr, and 2 yrs after HCT in patients with or without aGVHD.Table 1Effect of grade II-IV aGVHD by day 100 on TRM, Relapse, and EFSTime post-HCT# of Pts at RiskTRMRelapseEFSyes aGVHDno aGVHDyes aGVHDno aGVHDyes aGVHDno aGVHDyes aGVHDno aGVHDDay 10069443.0%6.6%0.8%3.7%35%51%1 year32554.6%7.4%5.0%24%31%28%2 years17197.0%7.4%6.4%30%27%22%Thirty eight percent of patients developed grade II-IV aGVHD (most (89%) by day +50). Analysis of patients who did or did not experience aGVHD by day 100 showed that patients who developed aGVHD rarely experienced TRM or relapse from that time forward (7.0 and 6.4% with TRM and relapse at 2 yrs post-HCT, respectively). Conversely, patients who did not develop aGVHD had a small risk of early relapse (3.7% at d100) but a steadily increasing rate of relapse starting day 200 post-HCT. Patients who did not develop grade II-IV aGVHD relapsed nearly 5 times more than those with aGVHD (30% vs. 6.4 at 2 yrs), with the result that EFS at 2 years was 19% (17/89) in those with no aGVHD vs. 35% (19/55) in those with aGVHD (p=0.001). It is important to note that the occurrence of aGVHD did not impact the rate of TRM.To assess the effect of the MRD status on relapse, we measured MRD by flow cytometry of BM pre-HCT, within 2 wks of engraftment, and at 3, 6, 9, or 12m after HCT and correlated the presence of MRD in these samples with relapse rates. More than 400 samples were analyzed. For patients positive for pre-HCT MRD, the rate ratio (RR) for relapse was 3.7 (1.6–8.2) with an estimated two-year relapse risk of 27.4% (17.4–43.1) and 70.8% (50.4–99.4) in pre-HCT MRD negative and positive patients, respectively. For patients positive for post-HCT MRD, we constructed a Cox regression model to assess the effect of positive post-HCT MRD in the context of other transplant-related variables. In the post-HCT period, an MRD positive result was associated with a 14-fold increase in relapse rate over an MRD negative result (RR=14.3, 95% CI:6.1–33.6). These findings did not change after controlling for risk group, immunophenotype, donor type and pre-HCT MRD status.Finally, we did an analysis of the combined effect of aGVHD and MRD status. Patients who developed grade II-IV aGVHD had low rates of post-HCT MRD and relapse, and a combined predictive effect of MRD status in patients with aGVHD could not be assessed. However, among patients who did not develop aGVHD, pre- and post-HCT MRD statuses were found to be independent risk factors for relapse (Table 2).Table 2Effect of Pre- and Post-HCT MRD on Relapse risk of Patients without aGHVDPre-HCT MRDPost HCT MRDRR for Relaspe (CI)--1 (reference)+-2.5 (1.0-5.8)-+12.7 (4.2-38.2)++31.2 (7.5-129.3)In summary, measurement of flow MRD on BM samples pre- and early post-HCT is feasible and positive results along with absence of grade II-IV aGVHD is highly predictive of relapse. There is a brief “window of opportunity” after day 50 and before day 200 post-HCT when most patients who will get aGVHD have experienced it, and the large majority of high-risk patients identified by MRD and aGHVD status have yet to relapse. It is imperative that novel post-HCT interventions be developed and given during this window in order to decrease relapse in the very high-risk patients we have defined. Disclosures:No relevant conflicts of interest to declare.