Study Of Minimal Residual Disease Research Articles

Minimal Residual Disease Status in Multiple Myeloma 1 Year After Autologous Hematopoietic Cell Transplantation and Lenalidomide Maintenance Are Associated With Long-Term Overall Survival.

Prognostic Immunophenotyping in Myeloma Response (PRIMeR) is an ancillary study of minimal residual disease (MRD) assessment for multiple myeloma by next-generation multiparameter flow cytometry (MFC). Patients were enrolled on a three-arm randomized control trial (Blood and Marrow Transplants Clinical Trials Network 0702 Stem Cell Transplant for Myeloma in Combination of Novel Agents [STaMINA]; ClinicalTrials.gov identifier: NCT01109004). Four hundred and thirty-five patients consented to the MRD panel, which included 10 monoclonal antibodies measured via six-color MFC. MRD was measured at baseline/preautologous hematopoietic cell transplant (BL/preAutoHCT), premaintenance (PM), and 1 year (Y1) after AutoHCT with a sensitivity of 10-5 to 10-6. The primary objective was to assess MRD-negative (MRDneg) at 1 year after AutoHCT and progression-free survival and overall survival (PFS/OS). Similar to the STaMINA results, at a median follow-up of 70 months, there was no significant difference in PFS/OS by treatment arm in the PRIMeR patients. MRDneg at all three time points was associated with significantly improved PFS, and MRDneg at Y1 had significantly longer OS. Multivariate analysis of PFS, adjusting for disease risk and treatment arm, demonstrated hazard ratios (HRs) in MRD-positive patients compared with MRDneg patients at BL, PM, and Y1 of 1.55 (P = .0074), 1.83 (P = .0007), and 3.61 (P < .0001), respectively. Corresponding HRs for OS were 1.19 (P = .48), 0.88 (P = .68), and 3.36 (P < .001). Patients with sustained MRDneg or who converted to MRDneg by Y1 had similar PFS/OS. To our knowledge, this first, prospective US cooperative group, multicenter study demonstrates that MRDneg at Y1 after AutoHCT with lenalidomide maintenance is prognostic for improved 6-year PFS and OS. Serial MRD measurements may direct trials to test how further therapy may improve long-term PFS and OS.

A Phase 2 Study of Minimal Residual Disease (MRD)-Adapted Front Line Venetoclax and Obinutuzumab in Fit Patients with Chronic Lymphocytic Leukemia (CLL): Effect of Obinutuzumab on Tumor Lysis Syndrome (TLS) Risk and Safety of Outpatient Venetoclax Dose Escalation

Introduction: Venetoclax (V) / Obinutuzumab (O) is an effective one-year fixed duration therapy for patients (pts) requiring first-line CLL treatment (tx). In CLL14, tumor lysis syndrome (TLS) risk assessment (including lymph node measurements by imaging and ALC) was required once prior to tx initiation, and O's impact on TLS risk prior to introduction of V was not a pre-planned study endpoint. Therefore, TLS prophylaxis (ppx) and tx setting for V dose escalation did not account for the possibility that O may decrease TLS risk prior to V escalation. Based on this approach, 22% of pts in the CLL14 study were high risk for TLS, thus requiring hospitalization for intensive laboratory monitoring during V dose escalation. (Fischer, NEJM 2019) We hypothesized that cycle (C) 1 of O would significantly decrease TLS risk prior to V escalation and that all pts, regardless of TLS risk category, could safely undergo a standard 5-week V dose escalation in the outpatient setting. Methods: We are conducting a Phase 2 study (NCT04447768) designed to examine the efficacy of a minimal residual disease (MRD)-adapted approach to V/O combination for fit pts in the front-line setting (study schema, Figure 1). A planned secondary study endpoint is to evaluate the effect of O monotherapy in C1 on TLS risk. Here we present interim results of TLS risk modification for the first 80 patients. Per protocol, TLS risk assessment including CT imaging and ALC measurement was performed at two defined timepoints: (1) prior to C1 day (D) 1 and (2) immediately prior to V initiation on C1D22. Key eligibility criteria: adult pts with CLL requiring first line tx per iwCLL guidelines, CIRS score ≤ 6, adequate heme function (Hgb ≥8 g/dL, ANC ≥1 x 10 9/L, Plt ≥30 x 10 9/L), creatinine clearance &amp;gt;50 mL/min. O was administered on C1 D1/2, 8 and 15. Per protocol, pts underwent V dose escalation in the outpatient setting, regardless of TLS risk. For pts with low and medium (med) tumor burden, TLS ppx was per FDA label. For pts who remained high TLS risk at the time of V initiation, TLS ppx throughout dose escalation included allopurinol to start at least 3 days prior to V initiation, sevelamer to start 12 hours prior to V initiation, and oral intake of ≥2 L water daily. On days of dose escalation, ppx additionally included kayexalate, rasburicase, and IV normal saline on D1 and D2 of each dose escalation. TLS labs were monitored according to FDA label guidance. Statistical analysis of TLS risk and adverse events was descriptive. Results: As of 5/2023, 80 pts have enrolled in this study. Seventy-eight pts completed the 5-week V ramp-up. Two pts were removed after receiving at least one dose of O but before repeat assessment of TLS risk and initiation of V (1 death unrelated to tx/disease, 1 patient with recurrent grade 3 (G3) O infusion-related reaction (IRR) leading to consent withdrawal). Median pt age was 57.5 years, 68% male, 95% white. Prior to tx initiation, TLS risk categories were low: 16%, med: 58%, and high: 26% (n=80). Change in TLS risk from baseline to C1D22 is detailed in Figure 2. Of the 20 pts who had sequential assessments with high baseline TLS risk, TLS risk after C1 O (C1D22) was low: 55%, med: 35%, high: 10%. For pts with med TLS risk at baseline with sequential assessments (n=45), TLS risk at C1D22 was low: 82%, med: 18%. One patient developed laboratory TLS on C1D3 following administration of O; no clinical TLS was observed. All pts accomplished V dose escalation in the outpatient setting with no observed laboratory or clinical TLS as defined by Howard Criteria. Grade 3 / 4 tx emergent AEs that occurred in &amp;gt;5% between C1D1 and C3D1 included neutropenia (35% G3, 11% G4), IRR (11% G3, 0% G4), and thrombocytopenia (9% G3, 3% G4). Conclusions: A single cycle of O effectively reduced TLS risk prior to introduction of V escalation with most pts achieving low or med TLS risk by C1D22. Notably, 90% of pts with high pre-tx TLS risk converted to low or med risk following C1 of O allowing for outpatient V escalation per FDA label. Further, we demonstrate that V dose escalation can be safely accomplished in the outpatient setting with close monitoring regardless of TLS risk. With this strategy, we observed no laboratory or clinical TLS during V dose escalation. Eliminating the need for hospitalization during V dose escalation may enhance patient and provider convenience and decrease the economic burden of this regimen. This study is ongoing; efficacy results of this MRD-guided approach to V/O are forthcoming.

POSTER: TCL-054 A Multi-Institutional Prospective Cohort Study of Minimal Residual Disease in Peripheral T-Cell Lymphoma: Impact of Autologous Stem Cell Transplant (ASCT)

POSTER: TCL-054 A Multi-Institutional Prospective Cohort Study of Minimal Residual Disease in Peripheral T-Cell Lymphoma: Impact of Autologous Stem Cell Transplant (ASCT)

TCL-054 A Multi-Institutional Prospective Cohort Study of Minimal Residual Disease in Peripheral T-Cell Lymphoma: Impact of Autologous Stem Cell Transplant (ASCT)

TCL-054 A Multi-Institutional Prospective Cohort Study of Minimal Residual Disease in Peripheral T-Cell Lymphoma: Impact of Autologous Stem Cell Transplant (ASCT)

A multi-institutional prospective cohort study of minimal residual disease in peripheral T-cell lymphoma: Impact of autologous stem cell transplant.

7563 Background: Roughly 80% of pts with peripheral T-cell lymphomas (PTCL) respond to CHOP-based therapy but 5-yr survival ranges between 20-40%. The high relapse rate after complete response (CR) suggests more sensitive response assessment, such as minimal residual disease (MRD) testing, could have prognostic or predictive value. Next generation sequencing (NGS) of the T-cell receptor (TCR) can detect a known TCR clonotype at 10-5. In a prospective study, we evaluated the utility of MRD testing via TCR NGS in PTCL (NCT03297697). Here we report results for pts pre- and post-autologous stem cell transplant (ASCT). Methods: This investigator-initiated, multi-institutional, prospective cohort study assessed MRD testing by TCR NGS in PTCL. Pts with previously untreated PTCL not otherwise specified (NOS), angioimmunoblastic T-cell lymphoma (AITL), anaplastic large cell lymphoma (ALCL), PTCL with T-follicular helper (TFH) phenotype, or monomorphic epitheliotropic intestinal T-cell lymphoma (MEITL) treated with anthracycline-based curative-intent therapy were eligible. Baseline TCR (TRG or TRB) clonotypes were established from diagnostic tissue or peripheral blood if tissue was not available. TCR clonality was determined positive via the LymphoTrack TRG/TRB Assays – MiSeq (Invivoscribe, San Diego, CA) when top % reads were ≥2.5% and ≥2x above background. Samples were obtained at diagnosis (pre-treatment), following induction therapy (EOT/pre-ASCT), and serially post-ASCT. Results: Of 43 enrolled pts, 19 underwent consolidative ASCT, of which 10 had post-ASCT samples available for MRD testing (Table). Median age at ASCT was 62 years (range: 38-77). Baseline TCR clonotypes were identified in 10/10 pts. At pre-ASCT evaluation, 7/10 pts had detectable TCR MRD (‘pos’) and 3/10 pts had TCR MRD below the level of detection (‘neg‘). Of the 3 MRD neg pts pre-ASCT, 2 had CR and 1 had PR by PET-CT. All 7 TCR MRD pos pts pre-ASCT had CR by PET-CT. Of 7 MRD pos pts pre-ASCT, 6 remained MRD pos and 1 became MRD neg post-ASCT. Of the 4 MRD neg pts post-ASCT, none have relapsed at 52, 31, 34, and 31 mo post-ASCT. Of 7 MRD pos pts pre-ASCT, 3 have relapsed following ASCT (at 3, 9, and 29 mo post-ASCT). Conclusions: In this cohort of patients with PTCL receiving consolidative ASCT in CR1, all 4 pts with negative TCR MRD post-ASCT remain in remission at median follow up of 32.5 mo post-ASCT. The negative predictive value of TCR NGS MRD post-ASCT should be further evaluated. Clinical trial information: NCT03297697 . [Table: see text]

Free from Maintenance Drug Therapy in Multiple Myeloma (The FREEDMM Trial): A Pilot Study of Minimal Residual Disease (MRD)-Driven Discontinuation of Maintenance

Background Induction therapy followed by autologous stem cell transplant (ASCT) and maintenance is the current standard of care for the treatment of newly diagnosed multiple myeloma (NDMM). However, indefinite maintenance therapy after ASCT carries both significant clinical and financial toxicities. While minimal residual disease (MRD) testing has become part of routine standard of care, its use in guiding treatment decisions, in particular cessation of therapy, has not been established in the maintenance setting. We hypothesize that MRD monitoring could be used to guide clinical decision making, justifying cessation of maintenance therapy in a sub-group of patients with sustained MRD-negative remission. Study Design and Methods This is a phase 2 single-arm adaptive design study (NCT 05192122) for decision-making regarding maintenance therapy (Figure 1). Patients are eligible if they have a diagnosis of active MM, have completed at least 2 years of maintenance therapy post-ASCT, and meet International Myeloma Working Group (IMWG) criteria for very good partial response (VGPR) or complete response (CR). MRD testing is performed twice, at least one year apart, on bone marrow aspirate using next-generation sequencing (NGS) and MRD-negative status is defined at a threshold of 10-6. Patients with negative PET scan in VGPR or CR and sustained (i.e., twice, 1 year apart) MRD negative status discontinue maintenance therapy. MRD-positive patients continue continue maintenance and are taken off study. In patients who are MRD-negative and discontinue maintenance therapy, MRD testing will be re-assessed yearly for 3 years. At each time of MRD testing, MALDI-TOF spectrometry and measurement of circulating multiple myeloma cells (CMMCs) will be done in parallel to compare against MRD results. Health-related quality of life (HRQoL) will be assessed throughout and results on maintenance compared with those after discontinuation using the EORTC QLQ-MY20 questionnaire. This study will enroll 50 patients to address the primary objective of assessment for sustained MRD-negative status at 12 months after stopping maintenance therapy. For all patients, a 5-mL peripheral blood and bone marrow sample is collected and stored at the time of each standard of care bone marrow biopsy and will be used to characterize the immune microenvironment by Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-seq) of paired peripheral blood and bone marrow in MRD-negative and MRD-positive patients as well as of patients progressing after stopping therapy. Preliminary Results Enrollment into FREEDMM trial began in December 2021, and to date 20 patients in CR have undergone a first bone marrow for MRD assessment, with results pending in 9 patients. The median age is 66.5 (range: 39-76) years, and the majority of patients are either Black (n=7; 35%) or Hispanic (n=7; 35%). One patient has high risk disease (17p deletion). The median number of years of maintenance therapy completed is 4.2 (range: 2.14-7.9) years. 17 (85%) patients had a negative MALDI-TOF test at the time of the first marrow. Seven (63.6%) of 11 patients were found to be MRD-negative. MALDI-TOF failed to accurately correlate with MRD results in 2 patients. 40 paired peripheral blood (n=20) and bone marrow aspirate (n=20) samples have been collected for CITE-seq analysis. Pilot CITE-seq experiments captured 6,700-11,000 cells with between 38,000-68,000 reads per cell on one MRD-negative and one MRD-positive bone marrow aspirate and paired peripheral blood sample. CITE-seq results for these 20 patients will be presented at the conference. Conclusion: This phase 2 trial applies a novel clinical trial design with an adaptive strategy to explore use of MRD testing as a guide to discontinuing maintenance therapy in myeloma and represents one of the first studies to explore in parallel various measures of disease monitoring (i.e., MRD, MALDI-TOF, CMMCs, CITE-seq). Ongoing results will be reported at the conference. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Digital Droplet PCR for Minimal Residual Disease Assessment in Philadelphia-Positive Acute Lymphoblastic Leukemia Using IG/TR Genes and BCR/ABL1 as Markers. Preliminary Results of a Comparative Analysis

Beldinanzi M, Della Starza I, contributed equally Background. The Philadelphia (Ph) chromosome is the most frequent genetic abnormality in adult acute lymphoblastic leukemia (ALL). BCR/ABL1 fusion transcript levels and clonal immunoglobulin/T-cell receptor (IG/TR) gene rearrangements are used as biomarkers for the study of minimal residual disease (MRD) in Ph+ ALL, with IG/TR used mainly in pediatric cohorts. There is evidence suggesting a poor concordance between these two molecular markers and there is a growing interest in evaluating MRD using a "double-hit strategy", especially in the adult setting where data are lacking. The real-time quantitative polymerase chain reaction (RQ-PCR) represents the current gold standard for MRD monitoring. It carries, however, some intrinsic limitations that digital droplet PCR (ddPCR), an innovative evolution of standard method, could overcome. Aim of this study. To carry out in adult Ph+ ALL patients a parallel MRD monitoring at early time points by both molecular targets - BCR/ABL1 gene fusion and clonal IG/TR gene rearrangements - by ddPCR. For BCR/ABL1 this was carried out by both RQ-PCR and ddPCR, in order to determine the concordance between the MRD values. Materials and Methods. The study comprised samples derived from 17 adults with newly diagnosed Ph+ ALL enrolled in the ongoing phase III GIMEMA LAL2820 clinical trial. Bone marrow (BM) samples were collected at the end of the induction phase (day +70) with ponatinib in the experimental arm or with a chemotherapy backbone plus imatinib in the control arm. At diagnosis, all patients underwent also a IG/TR gene rearrangements screening and MRD monitoring was performed by RQ-PCR and ddPCR for the quantification of BCR/ABL1 transcript levels and by ddPCR for IG/TR rearrangements, as described. MRD results were considered discordant in case of a difference greater than one log10 or in case of positivity/negativity. Results. The comparison between MRD values obtained in all samples by RQ-PCR and ddPCR using BCR/ABL1 mRNA level as biomarker showed an excellent correlation degree (R2=0.99). At variance, by IG/TR screening, it was possible to identify a clonal rearrangement only in 11/17 cases (65%), while 6/17 (35%) had no marker (NM) (3 p190, 3 p210; 2 IKZF1plus, 2 IKZF1 loss only and 2 IKZF1 WT). Moreover, of the 11 patients with a clonal rearrangement, in 3 (27%) (3 p190; 1 IKZF1plus, 2 IKZF1 WT) it was not possible to design a sensitive allele-specific oligonucleotide (ASO) for MRD monitoring. Overall, 8 samples were evaluated using both molecular markers by ddPCR, showing a poor correlation (R2=-0.21) and a low concordance rate. Indeed, 3/8 (37.5%) samples were concordant by both targets, either having highly positive (2/3) or negative (1/3) MRD values, while 5/8 (62.5%) were discordant; among the discordant cases, 1/5 was MRD-positive by both targets but outside the concordance range, while in 4/5 IG/TR MRD monitoring was negative, while BCR/ABL1 MRD monitoring was positive (range 10-3-10-1). Finally, in the 9 patients who failed IG/TR MRD monitoring, the BCR/ABL1 RQ-PCR analysis revealed a quantifiable MRD positivity in 6 cases (ranged 10-1-10-3), 2 proved "positive non-quantifiable" (PNQ) and 1 was MRD-negative (Figure 1). Conclusions. In adult Ph+ ALL, ddPCR with BCR/ABL1 as marker proved to be a sensitive and reliable tool for MRD monitoring at least comparable to RQ-PCR, showing a very high concordance rate. At variance, at least at early time points, the degree of concordance between BCR/ABL1 fusion transcript levels and IG/TR clonal rearrangements appeared suboptimal. Although IG/TR gene rearrangements are the most widely used biomarker for MRD monitoring in pediatric patients, the rate of failure in IG/TR marker identification in the adult Ph+ ALL setting is higher than in other subtypes. These findings suggest that the BCR/ABL1 fusion transcript remains the main sensitive target in adult Ph+ ALL and lead to hypothesize that in in this setting IG/TR clonal rearrangements represent a surrogate marker. Cohort expansion is warranted and clinical correlations are ongoing. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Methodological aspects of the detection of minimal residual disease in patients with acute leukemia

Introduction. The study of minimal residual disease (MRD) has become an integral part of various treatment protocols for acute leukemia. Methods of polymerase chain reaction (PCR) and multicolor fl ow cytometry (MFC) are most widely used to assess MRD.Aim — to characterize the main technologies for the detection of residual tumor cells in acute leukemiaMain findings. Various approaches for detecting MRD are described: PCR with patient-specifi c primers for rearranged genes of immunoglobulin and/or T-cell receptors, reverse transcription PCR for detecting chimeric transcripts and assessing the expression of overexpressed genes, as well as the basics of detecting MRD by MFC. Each of these approaches has its own advantages, disadvantages, and limitations of use.

Genome-Wide Association Study (GWAS) of High-Risk Minimal Residual Disease and Relapse in B-Cell Acute Lymphoblastic Leukemia on Trial AIEOP-BFM ALL 2000

▪A significant proportion of patients with pediatric acute lymphoblastic leukemia (ALL) still suffers from relapse and therapy-related toxicities. Thus, the focus of further improvement of therapy for them is not only cure, but minimizing both short-term and long-term therapy-associated toxicity. This can be achieved by better personalized adjustment of therapy to the risk of relapse. Continuing research has identified numerous features with prognostic potential, including clinical and genetic variables as well as early response to chemotherapy being associated with treatment outcome. While most research activities on genetic prognostic factors have focused on somatic features, relatively little is known on hereditary contributions to treatment response and outcome in pediatric ALL. Therefore, we conducted a genome-wide association study (GWAS) of high-risk minimal residual disease (MRD) and relapse in B cell precursor (BCP) ALL patients treated on trial AIEOP-BFM ALL 2000.On this trial, patients were stratified into three risk groups, mainly by MRD analyses. MRD standard-risk (MRD-SR) patients were MRD-negative on treatment days 33 and 78, MRD high-risk (MRD-HR) patients had residual disease of ≥5x10-4 on treatment day 78. All the remaining MRD results were considered MRD intermediate-risk (MRD-IR). Besides MRD, established high-risk parameters were also retained for stratification: prednisone poor-response, induction failure, positivity for t(4;11)/ MLL-AF4 . Treatment used standard drugs and, in some of the patients, cranial irradiation and/or hematopoietic stem cell transplantation. DNA isolated from remission samples was genotyped using Illumina Human1M-Duo BeadChips containing 1.048.711 single SNV markers. For GWAS analyses gPLINK v2.050, PLINK v.1.09 and RStatistics v3.3.1 were used. The analysis on MRD was conducted in 69 MRD-HR patients (cases) and in 1034 patients of the MRD-SR and MRD-IR groups (controls). The analysis on relapse employed 145 relapsed (cases) and 1007 non-relapsed patients (controls). After quality control assessment, the final analyses included 749.262 SNPs among 62 MRD-HR patients compared to 927 controls and 129 relapsed patients compared to 925 controls.In the analysis on MRD-HR five SNVs reached genome-wide significance at P ≤5×10−8. An intronic SNV (rs1570765) in STARD13 demonstrated the strongest association (P =1.73x10-10; odds ratio, OR=5.06). Two additional SNVs with P ≤5×10−8 in STARD13 were in linkage disequilibrium with rs1570765. The two remaining SNVs with P ≤5×10−8 were located in FEZ2 and NRIP1. Five additional SNVs demonstrated significant levels of ≤1×10−6 and were associatedwith the following genes: STARD13 , VIT and FICD . STARD13 is located on chromosome 13, belongs to the family of RhoGap proteins and is frequently lost in hepatocellular and breast carcinomas. Its RhoGAP domain activates members of the Ras superfamily which leads to a suppression of cell growth and differentiation (Lin et al. Oncogene 2010). In the GWAS approach on relapse three SNVs reached significant levels of ≤1×10−6 and were associatedwith the genes ASB3 (rs1549749, P =3.51x10-7; rs12465316, P =4.26x10-7) on chromosome 2 and NDN (rs9972595, P =1.04x10-6) on chromosome 15. Twenty additional SNVs demonstrated significance levels of ≤1×10−5. ASB3 codes for a member of the ankyrin repeat and SOCS box-containing (ASB) family of proteins containing ankyrin repeat sequence and a SOCS box domain. The latter serves to couple suppressor of cytokine signalling (SOCS) proteins and their binding partners with the elongin B and C complex, possibly targeting them for degradation. NDN , a p53 target gene, codes for Necdin which was previously shown to regulate the quiescence and response to genotoxic stress of hematopoietic stem/progenitor cells (Asai et al., Blood 2012). Further replication results, an analysis of additive effects and multivariate analyses will be presented.In conclusion, our results demonstrate that germline genetic variation is associated with high levels of MRD after induction/consolidation treatment for ALL and relapse on an AIEOP-BFM ALL protocol. In line with previous studies (Yang et al., JAMA 2009), our data suggest that genome-wide approaches may be useful to identify novel and clinically relevant SNVs for further refinement of risk-adapted treatment strategies in pediatric ALL. DisclosuresSchrappe:JAZZ Pharma: Consultancy, Research Funding; SigmaTau: Consultancy, Research Funding; Medac: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Baxalta: Consultancy, Research Funding.

Negative Minimal Residual Disease Associated with Extended Response to Moxetumomab Pasudotox in Patients with Relapsed/Refractory Hairy Cell Leukemia: Long-Term Follow-up of Bone Marrow Immunohistochemistry Analyses from a Phase 1 Study

The clinical significance of minimal residual disease (MRD) in patients with relapsed/refractory hairy cell leukemia (r/r HCL) who have achieved complete response (CR) has not been demonstrated. In other hematologic malignancies, MRD eradication may be associated with prolonged clinical response (Hourigan et al. Leukemia 2017;31:1482-90; Lahuerta et al. J Clin Oncol 2017;epub ahead of print). Moxetumomab pasudotox (formerly CAT-8015), a recombinant immunotoxin fusion protein that specifically binds to B lymphocyte-specific surface antigen CD22, has been shown by flow cytometry to eradicate MRD in a significant (P= 0.04) percentage of patients with r/r HCL (Kreitman et al. ASCO 2015). However, a detailed study of MRD by bone marrow immunohistochemistry (IHC) was not previously presented. We analyzed data from a phase 1 dose escalation study of moxetumomab pasudotox in r/r HCL patients to test the hypothesis that eradication of MRD in bone marrow, assessed by IHC, may be associated with prolonged remission.CAT-8015-1001 (NCT00586924) was a phase 1 study evaluating the safety and efficacy of moxetumomab pasudotox in patients with r/r HCL who previously had received at least 2 courses of treatment, at least 1 of which included a purine analog; initial results from the dose escalation phase in 28 patients have been reported (Kreitman et al. J Clin Oncol 2012;30:1822-8). Moxetumomab pasudotox was administered on days 1, 3, and 5 of each 28-day cycle until 2 cycles past complete remission (CR), disease progression, unacceptable toxicity, or neutralizing antibodies. In the dose escalation phase, 28 patients received moxetumomab pasudotox 5, 10, 20, 30, 40, or 50 µg/kg. An additional 21 patients were enrolled in the 50-µg/kg cohort.Bone marrow samples were obtained at baseline and at multiple time points during and after treatment. A central pathology laboratory performed hematoxylin and eosin and IHC staining for HCL/B-cell antigens CD20, CD79a, Annexin A1, DBA.44, and PAX-5. A blinded independent central pathologist reviewed (BICR) and assessed stained slides as IHC MRD+ or IHC MRD− for each time point.The best BICR IHC MRD response was compared with the best investigator-reported (IR) disease response. Duration of CR, duration of response (DOR), progression-free survival (PFS), and overall survival were evaluated for patients with a best response of BICR IHC MRD+ vs MRD−.Thirty-seven patients were evaluated for BICR IHC MRD; posttreatment bone marrow biopsies were not available for 11 patients, and biopsies from one patient were non-evaluable. The BICR IHC MRD status appears to be associated with the IR response; 17 of 18 BICR IHC MRD− patients had a best response of CR vs 9 of 19 MRD+ patients. Among patients found to be BICR IHC MRD− at 1 or more posttreatment evaluations, median duration of CR and median PFS were not reached after 84 months of follow-up vs 13.08 and 82.07 months, respectively, for patients who were consistently MRD+ at all time points. DOR was 82.7 vs 54.7 months for the BICR IHC MRD− and MRD+ groups, respectively.Cladribine monotherapy and cladribine followed by rituximab eliminated IHC MRD in some newly diagnosed HCL patients (Sigal et. al, Blood 2010;115:1893-6; Ravandi et. al Blood 2011;118:3818-23). Although vemurafenib demonstrated 96% to 100% overall response rates and 35% to 42% CR rates in patients with r/r HCL, all CR patients were IHC MRD+, and duration of MRD+ CRs was 19 months (Tiacci et al. N Engl J Med 2015;373:1733-47). Moxetumomab pasudotox is, to our knowledge, the only non-chemotherapy regimen shown to eradicate MRD in a meaningful percentage of patients with r/r HCL. Here, BICR findings confirmed elimination of IHC MRD in 49% of evaluable patients. MRD− patients experienced a statistically significantly longer duration of CR (P=0.0002) and PFS (P=0.0031) vs MRD+ patients.Based on the results of this retrospective analysis, together with observations made in the vemurafenib study, BICR IHC MRD− vs MRD+ status is associated with extended DOR and PFS in patients with r/r HCL treated with moxetumomab pasudotox. If this finding can be replicated in additional clinical studies, including the phase 3 pivotal study of moxetumomab pasudotox, IHC MRD determinations may be useful in evaluating the efficacy of therapies in the r/r HCL population. [Display omitted] DisclosuresKreitman:NIH: Patents & Royalties: Co-inventor on pending Moxetumomab Pasudotox patent owned by NIH. Robak:Akari Therapeutics Plc: Research Funding; Pharmacyclics LLC, an AbbVie Company: Research Funding; Janssen: Consultancy, Honoraria, Research Funding; AbbVie: Consultancy, Honoraria, Other: Travel, Accommodations, Expenses, Research Funding. Coutre:Pharmacyclics LLC, an AbbVie Company: Consultancy, Research Funding; Janssen: Consultancy; Novartis: Consultancy, Research Funding; AbbVie: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Gilead: Consultancy, Research Funding. Marshall:MedImmune: Employment. Yang:MedImmune: Employment, Equity Ownership, Other: Travel expenses. Pastan:NIH: Patents & Royalties: Pastan is an inventor on several patents on immunotoxins that have all been assigned to the NIH. Yao:MedImmune: Employment, Equity Ownership.

MRD-Based Therapeutic Decisions in Genetically Defined Subsets of Adolescents and Young Adult Philadelphia-Negative ALL.

Simple SummaryIn acute lymphoblastic leukemia (ALL), once a complete remission is achieved following induction chemotherapy, the study of submicroscopic minimal residual disease (MRD) represents a highly sensitive tool to assess the efficacy of early chemotherapy courses and predict outcome. Because of the significant therapeutic progress occurred in adolescent and young adult (AYA) ALL, the importance of MRD in this peculiar age setting has grown considerably, to refine individual prognostic scores within different genetic subsets and support specific risk and MRD-oriented programs. The evidence coming from the most recent MRD-based studies and the new therapeutic directions for AYA ALL are critically reviewed according to ALL subset and risk category.In many clinical studies published over the past 20 years, adolescents and young adults (AYA) with Philadelphia chromosome negative acute lymphoblastic leukemia (Ph− ALL) were considered as a rather homogeneous clinico-prognostic group of patients suitable to receive intensive pediatric-like regimens with an improved outcome compared with the use of traditional adult ALL protocols. The AYA group was defined in most studies by an age range of 18–40 years, with some exceptions (up to 45 years). The experience collected in pediatric ALL with the study of post-induction minimal residual disease (MRD) was rapidly duplicated in AYA ALL, making MRD a widely accepted key factor for risk stratification and risk-oriented therapy with or without allogeneic stem cell transplantation and experimental new drugs for patients with MRD detectable after highly intensive chemotherapy. This combined strategy has resulted in long-term survival rates of AYA patients of 60–80%. The present review examines the evidence for MRD-guided therapies in AYA’s Ph− ALL, provides a critical appraisal of current treatment pitfalls and illustrates the ways of achieving further therapeutic improvement according to the massive knowledge recently generated in the field of ALL biology and MRD/risk/subset-specific therapy

Modeling dynamic correlation in zero-inflated bivariate count data with applications to single-cell RNA sequencing data.

Interactions between biological molecules in a cell are tightly coordinated and often highly dynamic. As a result of these varying signaling activities, changes in gene coexpression patterns could often be observed. The advancements in next‐generation sequencing technologies bring new statistical challenges for studying these dynamic changes of gene coexpression. In recent years, methods have been developed to examine genomic information from individual cells. Single‐cell RNA sequencing (scRNA‐seq) data are count‐based, and often exhibit characteristics such as overdispersion and zero inflation. To explore the dynamic dependence structure in scRNA‐seq data and other zero‐inflated count data, new approaches are needed. In this paper, we consider overdispersion and zero inflation in count outcomes and propose a ZEro‐inflated negative binomial dynamic COrrelation model (ZENCO). The observed count data are modeled as a mixture of two components: success amplifications and dropout events in ZENCO. A latent variable is incorporated into ZENCO to model the covariate‐dependent correlation structure. We conduct simulation studies to evaluate the performance of our proposed method and to compare it with existing approaches. We also illustrate the implementation of our proposed approach using scRNA‐seq data from a study of minimal residual disease in melanoma.

Circulating tumour DNA from the cerebrospinal fluid allows the characterisation and monitoring of medulloblastoma

The molecular characterisation of medulloblastoma, the most common paediatric brain tumour, is crucial for the correct management and treatment of this heterogenous disease. However, insufficient tissue sample, the presence of tumour heterogeneity, or disseminated disease can challenge its diagnosis and monitoring. Here, we report that the cerebrospinal fluid (CSF) circulating tumour DNA (ctDNA) recapitulates the genomic alterations of the tumour and facilitates subgrouping and risk stratification, providing valuable information about diagnosis and prognosis. CSF ctDNA also characterises the intra-tumour genomic heterogeneity identifying small subclones. ctDNA is abundant in the CSF but barely present in plasma and longitudinal analysis of CSF ctDNA allows the study of minimal residual disease, genomic evolution and the characterisation of tumours at recurrence. Ultimately, CSF ctDNA analysis could facilitate the clinical management of medulloblastoma patients and help the design of tailored therapeutic strategies, increasing treatment efficacy while reducing excessive treatment to prevent long-term secondary effects.

Ficolled bone marrow is superior to bone marrow buffy coat for detection of minimal residual disease in multiple myeloma

ABSTRACTObjective: Buffy coat and ficoll of bone marrow (BM) are viable options for the study of minimal residual disease (MRD) in multiple myeloma (MM). As yet, there is no data about the superiority of either sample types. Herein, we aimed to address this issue.Methods: Forty pairs of ficolled BMs and BM buffy coats of 19 MM patients were studied for MRD by allele-specific oligonucleotide real-time quantitative PCR, with patient-specific primers/probes whenever appropriate.Results: There were 41 pairs of MRD data for comparison analysis due to one patient with biclonal disease. MRD levels in ficolls and buffy coats were highly concordant (rs = 0.936, P < 0.0001), with 31 (76%) and seven (17%) pairs being concomitantly MRD-positive or -negative. On the other hand, apart from the 16 pairs being both MRD-negative, or -positive but not quantifiable in ficolls and buffy coats, majority (n = 22, 88%) had higher MRD levels in ficolled BMs than BM buffy coats. Furthermore, in 17 pairs, in which MRD was quantifiable in both, MRD levels in ficolled BMs were 3.1 times those of BM buffy coats (median, 567/105 vs. 184/105, P = 0.001).Conclusion: Taken together, ficolled BM is more sensitive than BM buffy coat for MRD detection in MM, hence should be recommended.

Whole Exome Sequencing of Residual Disease in Multiple Myeloma: Searching for Novel Therapeutic Targets

Introduction:Multiple myeloma (MM) is a plasma cell dyscrasia causing damage of multiple organs with fatal consequences for patients. Despite the success of modern therapies eliminating a vast bulk of the aberrant cells, surviving residual clones eventually lead to the relapse of the disease. Accumulation of genomic alterations during the stage of minimal residual disease (MRD) likely contributes to a selective grow advantage and survival under the drug pressure. Identification of specific mutations in MM patients with MRD can provide unique opportunities to target the residual plasma cell clones. Here we present the first whole exome sequencing (WES) analysis of 22 MM samples of patients with MRD that identified 814 mutated genes with 4% of genes previously implicated in the pathogenesis of MM.Methods:Aberrant plasma cells (A-PCs) and peripheral blood (PB) were collected from patients after signing informed consent form. Presence of MRD was assessed with EuroFlow protocol and A-PCs were sorted out from bone marrow according to their pathological immunophenotype based on the expression of antigens CD38, CD45, CD19 and CD56. DNA from A-PCs was isolated and amplified by Repli-g Single cell kit (QIAGEN). Sequencing libraries were prepared using SureSelect Human All Exon V6 Kit (Agilent Technologies) and sequenced by Macrogen Inc. on Illumina HiSeq 4000 platform with average coverage 50x and 2x 100bp read length. Sequencing data were processed using the Bcbio framework following the standard workflow for tumor-matched-normal studies. Specifically, reads were mapped to the human reference genome GRCh37, successively marking duplicates using Picard. Germline mutations were identified using GATK HaplotypeCaller, whereas somatic mutations were identified using MuTect2 reporting as significant variants observed in at least 5 reads and minimum allele frequency of 10%. Variants in homopolymer regions longer than 5 nucleotides were filtered out. The final set of calls were further characterised by assessing their functional impact using snpEff and by annotating each variant using data from 1000 Genomes Phase 3, ExAC, and ClinVar. We then used OncodriveCLUST to identify putative oncogenic genes, and later compared these results with a literature curated list of MM driver genes (Weaver & Tariman, 2017).Results:Our dataset comprises 22 samples from 21 patients (one patient was sampled in two time points) with MM MRD, who received bortezomib-based regimen (age 41-71, average 59 years, 11/22 males, 10/22 females). 8 patients reached complete response, 9 patients had very good partial response and 4 patients had partial response. In our analysis, we detected 1,014 tumour somatic variants (8-287 per sample, median 36), most of them being missense mutations (676/1014), splice site mutations (145/1014) and frameshift insertions (134/1014). The variants affected a total of 814 genes, 97 genes were shared in at least two samples. The most frequently mutated genes were KIAA1211 (11/22), the immunoglobulin gene IGLV3-1 (8/22), apoptotic chromatin condensation inducer ACIN1 (7/22) and CCR4-associated factor 3 CNOT3 (7/22). We also identified 32 genes known to be mutated in MM in 64% of our samples (14/22). We found mutations shared by at least 2 samples in KRAS (4/22), DIS3 (3/22), TRAF3 (3/22), NRAS (2/22), ANK2 (2/22), BRAF (2/22) and RBM15 (2/22). Further analysis with OncodriveCLUST identified 18 putative oncogenic genes (FDR < 0.1), including KRAS, DIS3, ACIN1.Conclusion:We presented the first whole exome study of MM MRD, providing a characterisation of the mutations observed in A-PCs. We overcame problem with low amount of A-PCs in this disease stage by using whole genome amplification and a highly customised bioinformatic analysis pipeline. Our study suggests that A-PCs are characterised by new MM MRD specific set of mutated genes, along with the presence of mutations in well-known multiple myeloma cancer driver genes. This offers a great potential for design of novel precise treatments targeting MRD after standard MM therapies.Supported by Ministry of Health of the Czech Republic (17-30089A, CZ-DRO-FNOs/2016) and Ministry of Education of the Czech Republic (SGS18/PřF/2017-2018) DisclosuresKryukov:JSC BIOCAD: Employment. Maisnar:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees. Hajek:Celgene: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding; Bristol Myers Squibb: Consultancy, Honoraria; Amgen: Consultancy, Honoraria, Research Funding; Novartis: Research Funding.

Distinct Gene Expression Patterns of Minimal Residual Disease (MRD) Cells in High-Risk AML Patients Identified By RNA-Sequencing

INTRODUCTIONEvolving techniques have made possible the direct detection, physical isolation, and study of AML minimal residual disease (MRD) after treatment. This could allow for better identification of therapeutic vulnerabilities in AML. Prior studies have focused on cells that initiate leukemia in mouse models, known as leukemia-initiating cells (LIC), generally with a foundational CD34+CD38- immunophenotype. LIC are typically derived from diagnostic samples of untreated patients. Such stem-like cells do not necessarily represent the residual fraction of AML after treatment. Relapse may originate from non-LIC, and the presence of phenotypically and molecularly defined MRD is now firmly established as a critical prognostic factor for patients. High-risk AML is characterized by relapse, despite morphologic complete remission with initial therapy in most cases. RNA-sequencing was performed on pre- and post-treatment AML subpopulations, including MRD, from high-risk patients, to determine differences in gene expression.METHODSMatched primary AML samples were collected from marrow and peripheral blood of patients with high-risk AML (including patients with unfavorable karyotype and/or TP53 mutation) at diagnosis and after treatment. Mononuclear cells were flow-sorted for bulk (CD45dim) and LIC (Lin-CD34+CD38-CD123+) from diagnostic samples. Post-treatment samples were sorted for bulk mononuclear cells (MNC) and MRD, based on difference-from-normal/MRD immunophenotype specific for each patient as determined from established 20-marker clinical flow cytometry analysis. RNA was isolated using low-input methodology, and RNA-sequencing was performed using Illumina HiSeq 2000. Gene expression was assessed using GO-Elite, and differences between patients and subpopulations were assessed using rank product method.RESULTSGene expression in MRD was analyzed by RNA-sequencing in comparison to diagnostic samples in eight patients with high-risk AML. Four patients had unfavorable karyotype, including two with TP53 mutations. Patients had additional high-risk features, such as FLT3-ITD or RUNX1 mutations, or secondary/therapy-related AML. Treatment consisted of chemotherapy (6/8) or hypomethylating agents (2/8), with or without other targeted drugs. Residual leukemia was detected in post-treatment samples in all study patients. Significant differences in gene expression were detected between MRD and other sorted populations, including diagnostic bulk AML and LIC. Relevant MRD pathways included those with strong interactions with the microenvironment. Anti-apoptotic mechanisms, cytoskeletal, and cell adhesion related genes, WNT/beta-catenin signaling, and TGFbeta signaling ranked among the most relevant processes in AML MRD subpopulations (Figure 1A, GO-Elite interactome of highly expressed genes in AML MRD). To identify potentially critical and unique MRD-specific genes, rank product method was applied using 1) the most highly expressed genes in AML MRD, 2) the most differential expressed genes between MRD and bulk AML at diagnosis, and 3) the most differentially expressed genes between MRD and bulk MNC after treatment. Among the top 50 scoring genes using this approach (Figure 1B), 16 genes were among the top 5% of genes expressed in MRD among all patients and 20 genes have cell surface gene-products (shown in yellow). Several potential leukemia- and cancer-related genes of interest were identified (shown in bold).CONCLUSIONSKey differences exist between the gene expression profiles of post-treatment MRD from high-risk AML patients, in comparison to other populations and subpopulations of sorted cells before and after treatment. The highlighted differences suggest that MRD relies on specific intrinsic gene expression changes and microenvironmental interactions, and therefore may be targetable after elimination of bulk AML with initial therapy. Accessible surfacesome targets are among top hits. [Display omitted] DisclosuresKonopleva:cellectis: Research Funding; Immunogen: Research Funding; abbvie: Research Funding; Stemline Therapeutics: Research Funding. Andreeff:Astra Zeneca: Research Funding; Amgen: Consultancy, Research Funding; Jazz Pharma: Consultancy; Celgene: Consultancy; Reata: Equity Ownership; SentiBio: Equity Ownership; Aptose: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; United Therapeutics: Patents & Royalties: GD2 inhibition in breast cancer ; Eutropics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Oncolyze: Equity Ownership; Daiichi-Sankyo: Consultancy, Patents & Royalties: MDM2 inhibitor activity patent, Research Funding; Oncoceutics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees.

The role of automated cytometry in the new era of cancer immunotherapy.

The introduction in the clinical practice of several new approaches to cancer immunotherapy has greatly increased the interest in analytical methodologies that can define the immunological profile of patients in the clinical setting. This requires huge effort to obtain reliable monitoring tools that could be used to improve the patient's clinical outcome. The clinical applications of flow cytometry (FCM) in oncology started with the measurement of DNA content for the evaluation of both ploidy and cell cycle profile as potential prognostic parameters in the majority of human solid cancer types. The availability of monoclonal antibodies widely broadened the spectrum of clinical applications of this technique, which rapidly became a fundamental tool for the diagnosis and prognosis of malignant hematological diseases. Among the emerging clinical applications of FCM, the study of minimal residual disease in hematological malignancies, the quantification of blood dendritic cells in various types of tumors, the study of metastatic spread in solid tumors throughout both the analysis of circulating endothelial progenitor cells and the identification and characterization of circulating tumor cells, all appear very promising. More recently, an advanced single cell analysis technique has been developed that combines the precision of mass spectrometry with the unique advantages of FCM. This approach, termed mass cytometry, utilizes antibodies conjugated to heavy metal ions for the analysis of cellular proteins by a mass spectrometer. It provides measurement of over 100 simultaneous cellular parameters in a single sample and has evolved from a promising technology to a high recognized platform for multi-dimensional single-cell analysis. Should a careful standardization of the analytical procedures be reached, both FCM and mass cytometry could effectively become ideal tools for the optimization of new immunotherapeutic approaches in cancer patients.

Abstract 4878: Sensitive and specific DNA and RNA sequencing techniques for detecting minimal residual disease

Abstract The goal of this study was to assess the clinical applicability of molecular genetic techniques in the study of minimal residual disease (MRD) in pediatric acute myeloid leukemia (AML) via DNA and RNA-based methods. Currently, multi-parametric flow cytometry (MPFC) for surface immunophenotypes is the gold standard for MRD with a limit of detection between 0.001-0.002. However, approximately one-third of children with no detectable MRD by MPFC after first induction still relapse and suffer inferior outcomes. One potential reason is that a majority of refractory or recurrent AML clones will present with altered immunophenotypes compared to diagnostic specimens. In contrast, DNA mutations present at diagnosis will remain in refractory disease and therefore, a gene-based MRD platform surveying many genes could enable targeted, gene-specific therapy. Recently, the heterogeneous and unique genomic landscape of pediatric AML was characterized in the TARGET project and highlights the potential for leveraging companion molecular screens in the analysis of AML MRD. Contrary to adult AML, pediatric AML is not primarily characterized by single nucleotide variants (SNVs), but rather by complex structural variants (SV) that are difficult to detect using standard next generation sequencing techniques and analysis pipelines. The need to detect these complex SV and SNVs, all of which are at low allelic ratios (AR) at the remission state, demands assays that are capable of analyzing both DNA and RNA molecules at levels of detection comparable to MPFC. As proof of principal, we leveraged a unique capture technique (ArcherDx; CoreAML) to detect an important SV in pediatric AML, the internal tandem duplication in Fms related tyrosine Kinase (FLT3-ITD). Via a serial dilution of cells with known FLT3-ITD allelic ratio (MV4-11 cells), to determine a limit of detection, we were able to detect the mutation as low as 0.001. Next, we bench-marked the technique using diagnostic and relapse samples, and successfully detected FLT3-ITD at AR &amp;lt;0.01. To complement the DNA-based approach, we compared two error-corrected (EC) RNA assays (a Druley lab developed protocol and the ArcherDx; HemeV2 panel) to further assay SVs that cause novel gene fusions and aberrant exon usage, which are not detectable via DNA assays. Compared to existing cytogenetic and RNA-seq data, this new platform was capable of detecting known and novel cryptic translocations. Finally, we integrated the results from the FLT3-ITD assay and the RNA assay with our custom error-corrected sequencing data to create a novel bioinformatics workflow for assessing the biological implications of MRD clones detected. Collectively, our data support that EC sequencing, at both the DNA and RNA level, enable accurate detection of low allelic variants that could be used for improved clinical MRD diagnostics, prognostication and therapeutic selection. Citation Format: Erin L. Crowgey, Nitin Mahajan, Wing H. Wong, Edward A. Kolb, Todd Druley. Sensitive and specific DNA and RNA sequencing techniques for detecting minimal residual disease [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4878. doi:10.1158/1538-7445.AM2017-4878

Minimal Residual Disease Assessment and Risk-based Therapy in Acute Lymphoblastic Leukemia.

The study of minimal residual disease (MRD) in adult patients with acute lymphoblastic leukemia (ALL) allows a greater refinement of the individual risk classification and is the best support for risk-specific therapy with or without allogeneic hematopoietic cell transplantation (HCT). Using case-specific sensitive molecular probes or multiparametric flow cytometry on marrow samples obtained from the end of induction until midconsolidation, MRD assays can detect up to 1 leukemic cell of 10,000 total mononuclear cells (sensitivity, 0.01%; ie,≥104). This cutoff, presently bound to technical limitations and subject to improvement, reflects the individual chemosensitivity and is strongly correlated with treatment outcome. The chance for cure is approximately 70% in the MRD-negative subset but only 20% to 30% in MRD-positive patients, in any diagnostic and risk subset. As shown by prospective trials from Germany, Italy, Spain, and France-Switzerland-Belgium, approximately 50% to 70% of unselected adult patients with Philadelphia-negative ALL achieve and maintain an early MRD response, whereas the remainder do not, including a substantial proportion of clinically standard-risk patients, and require an HCT to avert at least partially the risk of relapse. Along with the diffusion of more effective "pediatric-inspired" chemotherapy programs, the MRD analysis is an integral part of a modern management strategy, guiding the decision process to transplant or not, in which case nonrelapse mortality using HCT in first remission-still 10% to 20%-is totally abolished. The use of new agents such as monoclonal antibodies, small inhibitors, and chimeric antigen receptor T cells is opening a new era of MRD-directed therapies, that will further increase survival rates.

Study of Minimal Residual Disease in Adults with B-Lineage Acute Lymphoblastic Leukemia by Flowcytometry

Background: After achieving morphological remission, existence of few number of leukemic cells in the patient’s blood represents the minimal residual disease (MRD) and its monitoring helps in evaluating early treatment response and future relapse. Patients and methods: Eighty seven newly diagnosed (B-ALL) cases were enrolled in the present study in the time period from October 2013 to October 2016. A panel of 4 monoclonal antibodies (CD10FITC, CD19PE, CD34PercP and CD45APC) were defined at diagnosis and after morphological remission for tracing of minimal residual disease (MRD). Results: Eighty seven newly diagnosed B-ALL cases were included in the present study of which 73 (84%) showed positive expression to CD45 in combination with (CD10, CD19 and CD34) at diagnosis, which allow us to use this combination for further assessment of MRD after morphological remission. In our study 65% of patients had negative MRD ( Conclusion: MRD detection by flow cytometry using the combination of CD45 with CD10, CD19 & CD34 is an easy and reliable method. Patients with positive MRD are at higher risk of relapse and have inferior overall survival rates compared to those with MRD-ve. Future studies focusing on treatment intensification for the group of patients with +ve MRD aiming to improve the treatment outcome are warranted.