Determination Of Minimal Residual Disease Research Articles

Molecular pathology for cholangiocarcinoma: a review of actionable genetic targets and their relevance to adjuvant & neoadjuvant therapy, staging, follow-up, and determination of minimal residual disease.

Cholangiocarcinoma (CCA) represents a group of epithelial cell tumors classified based on their anatomic location along the biliary tree. This rare malignancy is often diagnosed at an advanced stage and deemed unresectable. Even for those patients who are surgical candidates, recurrence rates are high and survival rates low. The mainstay of therapy for advanced CCA remains cisplatin plus gemcitabine, with a median overall survival (mOS) under 12 months, although the TOPAZ-1 trial showed a survival benefit with the addition of programmed cell death ligand 1 (PD-L1) blockade. In recent years, molecular profiling has revealed a wealth of potentially targetable genetic alterations, including fibroblast growth factor receptor (FGFR) fusions, isocitrate dehydrogenase 1 (IDH1) mutations, human epidermal growth factor receptor 2 (HER2) amplification and overexpression, and microsatellite instability (MSI). These discoveries have prompted numerous clinical trials employing drugs against these specific genetic changes. The foundation laid by early clinical studies and the landscape of ongoing trials are both summarized here. While the role of adjuvant therapy has yet to be defined in this disease, we emphasize the importance of employing targeted therapies in trials in the adjuvant and neoadjuvant spaces and discuss ways to overcome challenges due to low incidence of targetable mutations. Personalized medicine for this disease promises significant clinical benefit to patients, but further investigation is needed.

Molecular pathology and protein markers for pancreatic cancer: relevance in staging, in adjuvant therapy, in determination of minimal residual disease, and follow-up.

The diagnosis and monitoring of disease through the detection of circulating protein biomarkers is a growing field in the practice of oncology. The search for more effective protein biomarkers to aid in the diagnosis and treatment of patients with pancreatic ductal adenocarcinoma (PDAC) remains a valuable area of study, given the aggressive and often occult nature of this malignancy. Liquid biopsies are attractive, as they offer a minimally invasive and cost-effective approach when compared to traditional biopsy methods and imaging modalities used for diagnosis and surveillance. Carbohydrate antigen (CA) 19-9 is currently the most commonly used serum protein biomarker for the diagnosis and monitoring of patients with PDAC, but due to its sensitivity and specificity, its utility remains limited. In this review, we examine how circulating protein biomarkers are used in the diagnosis, prognostication, and surveillance of PDAC. We also highlight protein biomarkers that are currently under investigation that have the potential to enhance our ability to detect early-stage malignancies, predict response to therapy, and monitor for recurrence, but these markers require larger prospective validation studies before they can be widely implemented. Continued efforts to identify and validate novel biomarkers will be crucial for improving the management and outcomes of patients with this challenging disease.

Existing and emerging biomarkers in hepatocellular carcinoma: relevance in staging, determination of minimal residual disease, and monitoring treatment response: a narrative review.

With the development of novel active systemic therapies, the landscape of hepatocellular carcinoma (HCC) management is rapidly changing. However, HCC lacks sensitive and specific biomarkers to predict prognosis, monitor for minimal residual disease after locoregional therapy, and predict treatment response. In this review, we aim to summarize the best supporting evidence for refining existing, and development of novel biomarkers for staging, prognosis, determination of minimal residual disease and monitoring treatment response in HCC, focusing on those with evidence in clinical trials. PubMed and Embase databases were searched using the keywords; hepatocellular carcinoma, biomarker, minimal residual disease, surveillance, prognosis, staging, alpha-fetoprotein (AFP), liquid biopsy, treatment response, adjuvant, immunotherapy. Relevant clinical studies were included. AFP remains the major workhorse as the most widely used biomarker in HCC, however, its lack of wide applicability due to the high proportion of patients with HCC who are AFP negative, limits its value throughout all stages of HCC management. Significant work has been done to combine AFP with other clinical and serologic factors to increase its accuracy and utility as a biomarkers. However, it is likely that other more novel biomarkers such as those obtained through liquid biopsy will provide the prognostic power necessary for applications such as detecting recurrence and predicting treatment response. Liquid biopsy provides not only a wealth of potential biomarkers including circulating tumor cells and cell-free RNA/DNA, but also the ability to examine the mutational characteristics of the tumor with next generation sequencing. While early evidence supports the potential impact of many new biomarkers, validation in large clinical trials is lacking. This review highlights the paucity of sensitive and specific, widely applicable biomarkers, throughout all phases of management of HCC and summarizes evidence on biomarkers currently in use, as well as those in development and validation. Inclusion of biomarker analysis through clinical trials in HCC is critical to development of optimal therapeutic regimens, and improve patient outcomes.

Circulating Tumor DNA Analysis Associates with Progression-Free Survival (PFS) with Odronextamab Monotherapy in Relapsed/Refractory (R/R) Follicular Lymphoma (FL) and Diffuse Large B-Cell Lymphoma (DLBCL): Identification of Minimal Residual Disease Status and High-Risk Subgroups from the Phase 2 ELM-2 Study

Background Molecular characterization of B-cell non-Hodgkin lymphoma (B-NHL), through circulating tumor DNA (ctDNA) assessment of minimal residual disease (MRD) has been proposed as a tool to predict clinical outcome. Odronextamab, a CD20×CD3 bispecific antibody, demonstrated deep and durable responses and a generally manageable safety profile in patients (pts) with R/R FL or DLBCL in the Phase 2 ELM-2 study (NCT03888105; Kim TM et al. and Kim WS et al. ASH. 2022). In the overall populations, 12-month PFS rates were 64% and 29%, respectively. Using ctDNA from ELM-2, we show that these assessments associate with clinical outcomes following odronextamab treatment. Methods In ELM-2, pts received IV odronextamab in 21-day cycles, with step-up dosing in Cycle (C) 1, 80 mg (FL)/160 mg (DLBCL) QW in C2-4, then 160 mg (FL)/320 mg (DLBCL) Q2W until disease progression or unacceptable toxicity. Baseline (BL) ctDNA and tumor biopsies were used for molecular profiling. BL and on-treatment ctDNA were used for MRD determination in the biomarker population (BP; pts required ≥1 available plasma biomarker sample to be included in the BP). The first post-BL ctDNA sample was collected on C5 Day (D) 1, at the time of positron emission tomography-computed tomography (PET-CT). A modified AVENIO ctDNA analysis workflow (Roche; research only) was used for next-generation sequencing, based on the cancer personalized profiling by deep sequencing technique (Kurtz et al. J Clin Oncol. 2018). Whole blood cell pellets were used to filter out germline allele variants. MRD negativity was reported when the P value for variant allele frequency was >0.005. Results The BP comprised 53 FL pts and 63 DLBCL pts; at BL, all FL pts and all but one DLBCL pt were MRD(+). Pts remaining on study until C5D1 had similar PFS regardless of whether they were in the BP or overall population (FL, n=111; DLBCL, n=83). Pts who were MRD(-) at C5D1 had significantly longer PFS vs. those who remained MRD(+) (FL: HR 0.27 [95% CI 0.09-0.84], P=0.024 [Fig. 1a]; DLBCL: HR 0.29 [95% CI 0.12-0.71], P=0.007). In pts with FL achieving complete response (CR) by PET-CT at C5D1, a trend for prolonged PFS was observed in those who were MRD(-) (n=26) at this timepoint vs. those who were MRD(+) (n=17; HR 0.29 [95% CI 0.07-1.1], P=0.072). Pts with DLBCL who were MRD(-) at C5D1 (n=20) had similar PFS benefit regardless of PET-CT CR status (HR 0.56 [95% CI 0.10-3.11], P=0.511). Four DLBCL pts who were MRD(-) and did not achieve CR by PET-CT at C5D1 (partial response, n=3; stable disease, n=1) went on to achieve PET-CT CR at a later timepoint. Mutational analyses of BL ctDNA identified TP53 as the most frequent mutation (FL, n=34/53 [64%] mutation-evaluable pts; DLBCL, n=44/58 [76%]). In FL pts, TP53 mutations were associated with MRD(+) status at C5D1 (Fisher's exact test, P=0.002) and predicted significantly shorter PFS (HR 3.57 [95% CI 1.01-12.69]; P=0.049); this association was not observed in DLBCL. LymphGen classification (Wright et al. Cancer Cell. 2020) of BL ctDNA in pts with DLBCL identified mostly MCD and EZB subtypes (MCD, n=15; EZB, n=14; ST2, n=1; BN2, n=1; other, n=26). EZB-subtype pts treated with odronextamab had significantly longer PFS compared with MCD-subtype pts (HR 0.23 [95% CI 0.07-0.83]; P=0.025 [Fig. 1b]). Using BL ctDNA, cell of origin was determined in 43/58 pts with DLBCL; odronextamab treatment led to similar PFS in higher-risk non-germinal-center B-cell-like (non-GCB, n=18 [42%]) pts vs. GCB pts (n=25 [58%]; HR 1.62 [95% CI 0.72-3.62]; P=0.244). Further molecular assessment focused on gene fusions in DLBCL pts in the BP (39 with local laboratory gene fusion and ctDNA analyses, 24 with only ctDNA analyses). Odronextamab led to similar PFS in pts with double-hit (n=15) or triple-hit (n=5) fusions compared to those without (n=43; P=0.343 and P=0.140, respectively). Conclusions This study is among the first to analyze ctDNA in pts with R/R FL and DLBCL in a pivotal trial. This non-invasive method allows molecular characterization of pts with no available tissue, enabling identification of high-risk subgroups. CtDNA MRD status at C5D1 of odronextamab treatment was highly associated with PFS in pts with FL and DLBCL and in the future could form the basis of response-directed treatment paradigms. Molecular characterization in tumor biopsies, including CD20, will be presented. Ongoing monitoring of ctDNA may serve as an important early progression marker in R/R FL and DLBCL.

The role of minimal residual disease in therapy of pediatric acute lymphoblastic leukemia: a prospective cohort study

Background. During last 50 years there was a significant progress in understanding the nature of pediatric acute lymphoblastic leukemia (ALL). There were developed effective chemoradiotherapy regimens, new methods of diagnosis and emerged evaluation of treatment results. Determination of minimal residual disease (MRD) has become the most important factor in the patients stratification for risk-adapted treatment in the ALL IC-BFM 2009.
 Aim. To evaluate the survival rates of children with ALL according to the ALL IC-BFM 2009 protocol.
 Materials and methods. There were 136 people in the study of evaluating the effectiveness of therapy according to the ALL IC-BFM 2009 protocol: 69 boys and 67 girls (ratio 1.03:1). The median age is 4 years and 10 months. The observation period is from 26.01.2010 to 06.11.2022.
 Results. High survival rates of children with newly diagnosed ALL are achieved: overall survival (OS) is 91.2%, event-free (EFS) 82.4%, relapse-free (RFS) 88.6%. The best results are among patients of the standard risk group: OS, EFS and RFS reach 96.6%. In the intermediate group OS 96.2%, EFS 84.8% and RFS 88.3%. Worse results of survival are in the high risk group: OS 76.9%, EFS 65.4% and RFS 80.7%. Outcome analysis depending on the linear reveals a statistically insignificant difference in survival rates (for B-ALL OS 92.4%, EFS 83.1% and RFS 89.5%, for T-ALL OS 83.3%, EFS 77.8% and RFS 83.3%). It determines the tendency of improving the prognosis of pediatric T-ALL by optimizing the stratification of patients based on the indicators of MRD and the best direction of protocol.
 Conclusion. Results of survival rates of patients with ALL confirm high effectiveness of treatment according to the ALL IC-BFM 2009 protocol. MRD level on day 15 makes it possible to stratify patients and choose the optimal risk-adapted therapy.

Efficacy of the combination of venetoclax and hypomethylating agents in the treatment of patients with primary, relapsed and/or refractory acute myeloid leukemia

Relevance and goals. Treatment of patients with acute myeloid leukemia has traditionally been highly intensive and includes induction therapy using cytarabine and anathracyclines. In addition to new opportunities in the treatment of acute myeloid leukemia, the problem of toxicity of high-intensity therapy in elderly and young somatically burdened patients is quite acute. International clinical trials of phases I-III demonstrated a combination of high efficacy and acceptable hematological toxicity of combinations of hypomethylating agents and venetoclax in the first line of therapy, as well as in the treatment of resistant forms and relapses of acute myeloid leukemia in the older age group, which contributed to the study of the effectiveness of combinations of hypomethylating agents and venetoclax in the treatment of similar groups of young comorbid patients. In this work, we evaluated the efficiency of a combination of hypomethylating agents and venetoclax and overall and disease-free survival in patients with acute myeloid leukemia in routine practice.Methods. In the period from October 2017 to December 2021 on the basis of the Department of Hematology No. 11 and the Department of Bone Marrow and Hematopoietic Stem Cell Transplantation No. 56 of the Botkin Hospital (Moscow, Russia) 33 patients with acute myeloid leukemia received venetoclax therapy in combination with decitabine or azacitidine: 14 (42%) men and 19 (58%) women, median age was 60 years (23–83 years). In 42% (14 of 33) of cases, the regimen was prescribed for resistant course or relapse of acute myeloid leukemia and in 61% (20 of 33) as induction therapy. Three patients (15%) out of 19 from the group of newly diagnosed acute myeloid leukemia received this treatment regimen in the first line, taking into account the ECOG status 3-4. By August 2022, 13 (39%) patients are alive, 20 (61%) people have died. Overall survival, the rate of complete remission and complete remission with incomplete recovery, the rate of achieving negativity of minimal residual disease, the frequency of hematological toxicity and infectious complications were assessed. Statistical data processing used: frequency analysis using contingency tables (Fisher's exact test), survival analysis using the Kaplan-Meier method.Results. Complete remission and complete remission with incomplete recovery were achieved in 72.72% (24 of 33) of patients. In the group of primary acute myeloid leukemia, remissions were observed in 80% (16 out of 20) of cases, in the group with resistant course or recurrence of acute myeloid leukemia in 67% (8 out of 12) (p = 0.3). Determination of minimal residual disease by flow cytometry after the 1st course was performed for 54.54% (18 of 33) patients, while negativity was stated in 84.2% (14 of 18 patients) cases. In both groups, the incidence of hematological toxicity and infectious complications are comparable to those according to the literature data. The median follow-up was 9.5 months (1–47). Median overall survival was 39 months, 2-year overall survival was 63%, and overall 4-year survival was 39%. The disease-free survival rate was 33%.Conclusion. The combination of hypomethylating agents and venetoclax showed good efficacy and fairly high overall survival in patients of all age groups, both for primary acute myeloid leukemia and for relapses and resistant forms, regardless of previous therapy. Given the moderate hematological toxicity, as well as the relatively low rates of infectious complications during therapy, including the rather low mortality rates in case of COVID-19 infection in comparison with those on the background of high-intensity courses of therapy for acute myeloid leukemia, this scheme can be widely used not only in patients of the older age group, but also in young comorbid patients.

Flow Cytometric Analysis of Bone Marrow Particle Cells for Measuring Minimal Residual Disease in Multiple Myeloma.

Simple SummaryThe hemodilution of bone marrow aspirates is the main factor affecting the reliability of the determination of the minimal residual disease (MRD) resulting in multiple myeloma (MM). A bone marrow particle cell (BMPL) enrichment assay may be applied to overcome the problems caused by the hemodilution of bone marrow aspirates and to improve the rate of myeloma cell detection. Assessment of BMPL samples could decrease the false-negative rate of the MRD assessed with routine bone marrow aspirates. This study validated that the BMPL was superior to the routine bone marrow aspirate samples in the MM and MRD analysis.Minimal residual disease (MRD) is one of the most relevant prognostic factors in patients with multiple myeloma (MM). However, the hemodilution of bone marrow (BM) aspirates, the most common preanalytical problem, is known to affect MRD detection. In the present study, we analyzed a preanalytical method for routine BM aspirates and a bone marrow particle cell (BMPL) enrichment assay and validated it as a reliable preanalytical method for flow cytometric MRD determination. A total of 120 BM samples were taken from 103 MM patients consecutively recruited; 77 BM samples had BMPL enrichment analysis and 99 BM samples were routinely analyzed. Then, the two different samples from patients with MM were sent for MRD detection using an eight-color flow cytometry. Our data showed that assessment of the BMPL enrichment samples attenuated the overestimation of MRD-negative assessed in the routine BM samples, which was mainly caused by hemodilution. In conclusion, the BMPL enrichment assay is a functional and practical preanalytical method for flow cytometric MRD analysis.

Genomic Alterations in Acute Myeloid Leukemia in Routine Clinical Practice

Acute Myeloid Leukemia (AML), the most common acute leukemia in adults, is a genetically heterogeneous disease. Genomic alterations condition the pathophysiology of AML. Nowadays, these changes are considered to be important biomarkers for risk stratification, treatment decisions (including new targeted drugs) and Minimal Residual Disease (MRD) monitoring during followup of AML. Positive MRD in AML patients is associated with a higher risk of relapse and shorter overall survival compared to MRD negative individuals. Nevertheless, MRD-targets, diagnostic techniques, time points of MRD determination, and analyzed material are not standardized. Thus, integration of MRD testing in individual AML patients in routine practice is still a workin- progress. This review article comprehensively focuses on key molecular biomarkers in AML and their utilization in clinical practice, especially regarding risk assessment and MRD testing. Moreover, new AML molecular-targeted therapies are briefly summarized here.

Minimal residual disease detection by next-generation sequencing in multiple myeloma: Promise and challenges for response-adapted therapy.

Assessment of minimal residual disease (MRD) is becoming a standard diagnostic tool for curable hematological malignancies such as chronic and acute myeloid leukemia. Multiple myeloma (MM) remains an incurable disease, as a major portion of patients even in complete response eventually relapse, suggesting that residual disease remains. Over the past decade, the treatment landscape of MM has radically changed with the introduction of new effective drugs and the availability of immunotherapy, including targeted antibodies and adoptive cell therapy. Therefore, conventional serological and morphological techniques have become suboptimal for the evaluation of depth of response. Recently, the International Myeloma Working Group (IMWG) introduced the definition of MRD negativity as the absence of clonal Plasma cells (PC) with a minimum sensitivity of <10−5 either by next-generation sequencing (NGS) using the LymphoSIGHT platform (Sequenta/Adaptative) or by next-generation flow cytometry (NGF) using EuroFlow approaches as the reference methods. While the definition of the LymphoSIGHT platform (Sequenta/Adaptive) as the standard method derives from its large use and validation in clinical studies on the prognostic value of NGS-based MRD, other commercially available options exist. Recently, the LymphoTrack assay has been evaluated in MM, demonstrating a sensitivity level of 10−5, hence qualifying as an alternative effective tool for MRD monitoring in MM. Here, we will review state-of-the-art methods for MRD assessment by NGS. We will summarize how MRD testing supports clinical trials as a useful tool in dynamic risk-adapted therapy. Finally, we will also discuss future promise and challenges of NGS-based MRD determination for clinical decision-making. In addition, we will present our real-life single-center experience with the commercially available NGS strategy LymphoTrack-MiSeq. Even with the limitation of a limited number of patients, our results confirm the LymphoTrack-MiSeq platform as a cost-effective, readily available, and standardized workflow with a sensitivity of 10−5. Our real-life data also confirm that achieving MRD negativity is an important prognostic factor in MM.

Igh-V(D)J NGS-MRD Measurement in Pediatric B-Acute Lymphoblastic Leukemia

IntroductionB-acute lymphoblastic leukemia (B-ALL) is the most common cancer of childhood. Early response to induction chemotherapy is one of the important prognostic factors in B-ALL. However, the analytic sensitivity for flow cytometry (FC) is only 10 -4. The feasibility of using next-generation sequencing (NGS) of immunoglobulin for the determination of minimal residual disease (MRD) in B-ALL has been demonstrated. This study aimed to investigate the performance of NGS techniques measuring immunoglobulin heavy chain (IgH)-variable, diversity, and joining (V[D]J) clonal rearrangements compared with FC in detecting MRD for children with B-ALL and to predict the clinical outcome of B-ALL patients.MethodsNewly diagnosed younger than 18 years old B-ALL patients who received the treatment strategy of South China children's leukemia Group (SCCLG)-ALL 2016 were recruited. DNA extracted from bone marrow cells at all available time points for each patient was submitted to Simcere diagnostics for sequencing using Illumina NovaSeq platform. We performed IgH V(D)J NGS and FCM on the bone marrow serially obtained at diagnosis (D0), 15 days at induction therapy (D15), 33 days at induction therapy (D33) and then at the end of induction therapy (EOI). We defined MRD positive (MRD +) by IgH V(D)J NGS and FCM as more than 1 blast cell among 10 4 and 10 6 bone marrow cells, respectively. The sensitivity of MRD detection by IgH V(D)J NGS and FCM, and the association of MRD status with clinicopathological characteristics were investigated. Statistical analysis was performed through SPSS Statistics 22. Enumeration data and correlation between MRD data and clinicopathological characteristics were compared by Chi-square test or Fisher's exact test. This trial was registered at www.clinicaltrials.gov as # NCT04977895.ResultsAs of July 27, 2021, 22 patients (median age, 4.5 years; range, 3.0-7.3) were enrolled in the study. Three patients (13.6%) had a t (9;22) translocation consistent with Philadelphia chromosome positive disease. According to risk stratifications, 8 (36.4%), 8 (36.4%), and 2 (9.1%) patients were classified as low risk (LR), intermediate risk (IR), and high risk (HR) groups, respectively. The remaining 4 patients are still under treatment and have not been classified. We identified leukemic IgH clones in 100% of the diagnostic samples and 68.2% (15/22) of the patients were polyclonal. In 11 patients whose samples of all the four timepoints (D0, D15, D33, EOI) have been tested in parallel by FCM and IgH V(D)J NGS, the frequencies of patients with MRD + were 30.4% vs. 90.9% at D15 (P＜0.05) by FCM and IgH V(D)J NGS. IgH V(D)J NGS MRD monitoring could identify MRD + patients with frequency of 45.5% and 18.2% among patients achieved MRD negativity by FCM at D33 (P＜0.05) and EOI (P = 0.46). With an MRD detection limit of 10 -6, 90.9% (10/11), 36.4% (4/11) and 18.2% (2/11) patients were MRD negative by FCM but positive by the NGS test at D15, D33 and EOI, respectively. This suggested that the sensitivity of IgH V(D)J NGS was significantly higher than that of FCM. Correlation of the measured MRD between the two methods in the entire cohort (r = 0.7934, P < 0.0001) as well as in the concordant cases (r = 0.5558, P = 0.0032) was very high. There was a high discordant rate with NGS identifying more patients MRD + at this threshold. Furthermore, NGS MRD was positive but the FCM MRD was negative in 13 samples (P < 0.0001). In addition, positive MRD status of D33 by NGS was significantly associated with the age of B-ALL patients, patients under 6 years more frequently harbored detectable MRD compared with those ≥ 6 years old (87.5% vs. 11.1%, P < 0.01). There was no patient relapsed after a medium follow-up of 10.5 months.ConclusionsWe demonstrated the higher sensitivity of IgH-V(D)J NGS in MRD detection of B-ALL, which implies that NGS MRD monitoring could be helpful for more accurate risk stratifications and more precise treatment according to risk stratifications. Further study with a larger sample size and a longer follow-up period is need. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Performance of Next Generation Sequencing for Minimal Residual Disease Detection for Pediatric Patients with Acute Lymphoblastic Leukemia: Results from the Prospective Clinical Trial DFCI 16-001

Performance of Next Generation Sequencing for Minimal Residual Disease Detection for Pediatric Patients with Acute Lymphoblastic Leukemia: Results from the Prospective Clinical Trial DFCI 16-001

Monitoring Leukemic Mutations Using Ultrasensitive IBSAFE Digital Droplet Analysis Predicts Relapse of Acute Myeloid Leukemia

IntroductionOptimal management of patients with acute myeloid leukemia (AML) depends on accurate monitoring of minimal residual disease (MRD) after treatment. The presence of MRD predicts outcome, with implications for risk stratification and treatment decisions. Therefore, improvements in MRD determination are desirable. Current MRD methods include multicolor flow cytometry (MFC), real-time quantitative polymerase chain reaction (QPCR), and more recently next generation sequencing (NGS) and droplet digital PCR (ddPCR). MFC can be applied on most patients, but suffers from limited sensitivity (rarely below 0.1% leukemic cells). QPCR is more sensitive but can only be applied on leukemias carrying the targeted mutation. NGS can track several mutations simultaneously but with limited sensitivity. We recently developed the method IBSAFE, which utilizes compartmentalized ddPCR micelles with a limit of detection of 0.001% mutant allele frequency (MAF). Given that even patients with negative MFC-MRD results (<0.1%) often have poor prognosis, we set out to analyse relapsing AML patients using IBSAFE-ddPCR to investigate if this method would be helpful for predicting the relapses.Materials and MethodsIn this pilot study, 10 relapsing AML patients were retrospectively tested for MRD with IBSAFE, a patent-pending method, in bone marrow aspirates taken at 3 to 13 (average 8) follow-up time points between 145 and 2607 days after diagnosis. First, the mutational profile of each leukemia was determined at diagnosis and relapse by whole-exome sequencing and targeted AML gene panel sequencing. Skin fibroblasts were used as germline controls.Based on the identified mutations, candidates for MRD monitoring were selected with priority towards recurrent mutations and mutations present at both diagnosis and relapse. IBSAFE assays were developed for selected mutations and used to analyse the diagnostic and follow-up samples.ResultsNGS of AML samples revealed between 10 and 36 somatic mutations at diagnosis (average 18) in each patient of which 0 to 7 (average 4) were known recurrent AML mutations including NPM1 , DMNT3A and TET2 . For a total of 66 selected mutations, IBSAFE assays were designed and validated, representing between 5 to 9 mutations (average 7) for each patient. The concordance between MAF by IBSAFE and NGS was excellent, corroborating the precision of our IBSAFE method. For MRD monitoring, molecular evidence of MRD/relapse was apparent prior to clinical relapse in all 10 patients, and three different patterns of emerging and disappearing mutations could be discerned. In some patients (pattern one; n=5), it was possible to monitor two or more separate clones, all of which reappeared at relapse. In these, some mutations were undetectable at certain time points, whereas others (e.g. TET2 ) were present at all time points at low levels, possibly representing preleukemic clones. In other patients (pattern two; n=4), most monitored mutations were undetectable (<0.001% MAF) at certain time points. Interestingly, in this second group, the emerging relapsing leukemia carried only some of the mutations monitored, demonstrating clonal heterogeneity with selection of one relapsing clone, which often showed additional new mutations. In one patient (pattern three; n=1) there was no distinct decrease before stem cell transplantation for several of the mutations (including DNMT3A and TET2 ) despite morphological remission and negative MFC-MRD, demonstrating the preleukemic nature of the regenerating hematopoiesis. For 21 samples (affecting 9 out of 10 patients), mutations were detected at MAF between 0.1% and 0.001%.ConclusionsWe demonstrate that our IBSAFE method can track early recurrence of leukemic clones. The advantage as compared to other MRD methods is the high sensitivity down to 0.001% MAF and simplified workflow to follow several mutations and track different emerging clones. One drawback is that our method does not allow tracking of acquired new mutations after treatment. However, developed IBSAFE assays can quickly be applied on follow-up samples and easily utilized for other patients carrying the same mutation. Of course, it may still be necessary to design some new assays for each patient, given the mutational heterogeneity of AML. In conclusion, these pilot results demonstrate the power of the IBSAFE method with possible applications in clinical routine AML-MRD analyses. DisclosuresChen:SAGA Diagnostics AB: Employment. George:SAGA Diagnostics AB: Employment, Equity Ownership, Patents & Royalties. Rigo:SAGA Diagnostics AB: Employment. Saal:SAGA Diagnostics AB: Employment, Equity Ownership, Patents & Royalties.

Presence of Minimal Residual Disease at End of Induction in Acute Myeloid Leukemia Is Predictive of Poor Outcome

Introduction: One of the mainstays of chemotherapy in acute myeloid leukemia (AML), other than acute promyelocytic leukemia, is induction with a goal to achieve morphological complete remission (CR) as evident by less than 5% blasts. Post remission strategies then focus on either consolidation chemotherapy or allogeneic bone marrow transplantation (aBMT). However, not all patients by this remission criterion achieve long term remission and a subset of patients relapse. This relapse originates from further expansion chemoresistant clones. Detection of minimal residual disease (MRD) following chemotherapy, early in the course of treatment, is highly predictive of outcome and offers a window of opportunity to intensify treatment and prevent relapse. Here, we describe assessment of immunophenotypic MRD using a 10-colour two tube assay and a “difference from normal” approach.Methods: We accrued 280 consecutive patients of adult (>18 years) AML, other than with t(15;17), over a 30 month period after obtaining consent. All patients received “3+7” induction therapy with daunorubicin and cytarabine. If patients were in morphological CR at end of induction, they either received 3 courses of 12-18 gm/m2 high dose cytarabine (HiDAC) or aBMT if feasible.MRD testing was done at end of induction and if feasible, post 1st Cycle HiDac using a two tube 10 colour assay. (CD15, CD13, CD19, CD34, CD56, CD7, CD45, CD11b, HLA-DR, CD117, CD14, CD123, CD64, CD33, CD36 & CD38). A minimum of 1.6 million events were acquired per tube on a Navios flow cytometer. Identical panel was used at MRD time points as well as on the diagnostic sample. Analysis of MRD was done using Kaluza 1.3 by a difference from normal approach that focused on the development of progenitors to monocytes. Conventional karyotyping and FISH was done as per standard recommendations and patients were classified into favorable, intermediate and poor cytogenetic risk. The presence of FLT3 -ITD, NPM1 and CEBPA mutations was detected by a fragment length analysis based assay. Overall survival (OS) was calculated from date of diagnosis to time of last follow up or death. Relapse free survival (RFS) was calculated after achieving of 1st remission (CR) till relapse or death or last follow up if in CR. Results of the MRD assays, cytogenetic and molecular risk groups were analyzed for their impact on OS and DFS using Log Rank test of Kaplan-Meier survival analysis.Results: A total of 280 AML patients were treated and followed up till maximum of 30 months period. Based on cytogenetics, 28.6% were classified as favorable risk whereas 61.4% and 10% were intermediate and poor risk respectively. FLT3-ITD, NPM1 and CEBPA mutations were harbored by 20.7%, 26.4% and 10.3% of patients respectively. The median follow up was of 9 months (Ranging from 17 days to 30 months). Of these, 35 had induction death and 59 had refractory disease. Median OS was 50.3% and RFS was 50.9%. Post induction MRD was assessed in 235 patients of which 108 (45.9%) had detectable residual disease (range 0.01-66%, median:1.38%). Post consolidation MRD was assessed in 98 patients of which 22 (22.4%) were MRD positive (range 0.002-7.7%, median: 0.05%). Favorable risk cytogenetics in cases excluding induction deaths was predictive of better OS than intermediate (p= 0.031) and poor risk (p=0.003) cytogenetic profiles but not RFS. Patients harboring MRD at the end of induction were associated with worse OS (p=0.04) but not RFS, whereas post consolidation positive MRD status did not show significant change in OS and RFS.Conclusion: Our data is in agreement with other studies that determination of immunophenotypic MRD is extremely important in predicting outcome. AML MRD is a very useful guide for guiding post remission strategies in AML and should be incorporated into routine treatment algorithms. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Utility of Leukemic Stem Cells Markers in Determining Minimal Residual Disease in Patients with AML

Introduction: Minimal residual disease (MRD) assessments are being increasingly used to define prognosis in patients with acute myeloid leukemia (AML). However, data suggests that a proportion of patients found to be MRD negative experience leukemia relapse. These relapses are thought to be related to the persistence of leukemia stem cells (LSC) that survive chemotherapy. Recent studies have identified a number of LSC antigens that may be useful to follow LSC populations including CLL1, CD123 and CD47. We hypothesized that the use of a routine antibody panel in conjunction with CD123 and CLL1 specific antibodies will allow for more accurate determination of MRD using multiparametric flow cytometry (MPFC) and better identification of patients destined to relapse.Methods: In this single center prospective study we evaluated the diagnostic bone marrow samples of all newly diagnosed patients with AML (excluding APL) for leukemia associated immunophenotypes (LAIP's) using MPFC and a panel of antibodies targeted towards myeloid, lymphoid and LSC markers. LAIP's were scored as a percentage of the bulk blast population which was defined using plots of Side Scatter (SCC) vs. CD45. LSC compartment was defined as CD34+CD38-CD123+ or CD34+CD38-CLL1+. Bone marrow samples from non-leukemic patients were used to identify the background level of CD123 and CLL1 positivity. Samples were considered MRD positive if having >0.1% of a specific LAIP as a function of the bulk blast population.Results: Over the study duration, 88 patients with newly diagnosed AML received intensive chemotherapy (Table1). Complete MRD data (routine and LSC markers) was available for 58 patients had at the end of induction and 31 patients at the end consolidation (EOC). For the 58 patients with end-of-induction (EOI) MRD data the diagnostic sample revealed CD123+ LSC and CLL1+ LSC at a frequency of >0.1% in 30 (51.7%) and 28 (48.3%) patients respectively and at a frequency of <0.1% in 24 (25.9%) and 26 (44.8%) patients respectively. Data was not available for 4 patients.Following a single induction chemotherapy cycle 52/58 (89.7%) patients were in complete remission. At the EOI, 26/58 (44.8%) patients were found to be MRD negative using the standard antibody panel. Only 3/30 patients (10%) who had CD123+ LSC and 2/28 patients who had CLL1+ LSC at diagnosis were found to be negative at the EOI. However, 21/24 (87.5%) patients that were negative for CD123+ LSC and 22/24 (85%) patients that were negative for CLL1+ LSC at diagnosis were found to be positive for CD123+ LSC and CLL1+ LSC at the EOI, respectively. At the EOC, 31 patients were assessed for routine and LSC based MRD. Amongst these patients 16 (51.6%) were MRD negative using routine markers, 13 (41.9%) were negative CD123 and only 6 (19.4%) were negative for CLL1.There was no relationship between relapse rates and the MRD status at the EOI or EOC using either standard MRD markers, CD123 or CLL1. Although not statistically significant we observed that only 1 of 6 (16.7%) patients that was negative for CLL1+ LSC at the end of consolidation relapsed in contrast to 11 of 25 (44%) patients that were positive for CLL1+ LSC (p=0.4).After a median follow-up of 11.3 months, the median relapse free survival for patients that were positive for CLL1+ LSC at the EOC was 13.8 months and was not achieved for patients negative for CLL1+ LSC (p=0.4). Similarly, there was no significant difference in the RFS for patients that were MRD positive or negative for routine MRD markers (16.8 vs. 13.8 months, p=0.94) or for CD123 (13.8 vs. 16.8 months, p=0.73). The median overall survival (OS) for the entire cohort was not achieved. There was no significant difference in the OS for patients based upon MRD status at the EOC when using standard MRD markers, CLL1 (Figure 1) or CD123 (Figure 2).Conclusions: This proof-of-concept study suggests that the CLL1+ and CD123+ LSC can be identified at diagnosis at very low levels relative to the bulk blast population. Surprisingly, patients who were negative for CD123+ and CLL1+ LSC at diagnosis were often found to be positive at EOI. This may be related to a reduction in the overall blast population due to chemotherapy, but persistence of the original LSC population. Larger studies are necessary to explore the impact of persistent LSC markers on clinical outcomes. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

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.

Prognostic Implications of Minimal Residual Disease Detection in Acute Myeloid Leukemia (AML) with Cytarabine Based Therapy of Differing Dose Intensities

Prognostic Implications of Minimal Residual Disease Detection in Acute Myeloid Leukemia (AML) with Cytarabine Based Therapy of Differing Dose Intensities

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.

Prognostic Value of Minimal Residual Disease in Children with Acute Lymphoblastic Leukemia.

Purpose To assess the prognostic value of determination of minimal residual disease in the bone marrow after multiagent remission induction therapy in children with acute lymphoblastic leukemia. Methods From September 2001 to December 2004, 102 patients with newly diagnosed B-cell precursor acute lymphoblastic leukemia were enrolled on protocol ALL-XH-99. Minimal residual disease in the bone marrow at the end of remission induction therapy, before high-dose methotrexate, early intensification as well as 1 year, 2 year of maintains therapy was detected by multi-parameter flow cytometry with various combinations of monoclonal antibodies for leukemia-associated immunophenotype. The probability of event-free survival was estimated by Kaplan-Meier analysis and the distributions of probability of event-free survival (pEFS) were compared using the log-rank test. Chi-square analysis or Fisher exact test were used to compare differences in the distribution of biologic presenting features. A Cox proportional hazards model was used to identify independent prognostic factors. Results [circ1] The probability of 39-month event-free survival (EFS) was significantly worse for patients who achieved a minimal residual level ≥ 0.01% in the bone marrow on the remission date as compared to that of other patients with a minimal residual level < 0.01% (0.00% vs 83.00±9.90%, P=0.0000). [circ2]Univariate analysis indicated that there were no relationship between level of minimal residual disease at the date of complete remission and biologic presenting features (gender, age, white blood cells), but did Philadelphia chromosome, ALL-XH-99 risk groups as well as lymphoblasts in the bone marrow on day 19 (P<0.05). [circ3] Multivariate analysis suggests patients with high level minimal residual disease after the first induction course was an independent prognostic factor (hazard ratio, 5.381; 95% confidence interval 0.004 to 0.624; P=0.020). Conclusions Early treatment response as assessed by minimal residual leukemia determination by flow cytometry has important prognostic significance and can be performed in a resource-poor patient population. Patients with level of minimal residual disease ≥ 0.01% at end of remission induction ought to be used an novel and more intentive chemotherapy.

Relationship of genetic deviations with laboratory indicators of peripheral blood and bone marrow in children with acute lymphoblastic leukemia

The aim of this study was to evaluate the prognostic value of the relationship between genetic abnormalities and clinical and laboratory parameters of peripheral blood and bone marrow in children with acute lymphoblastic leukemia (ALL). Material and methods. 105 children diagnosed with ALL were examined (average age 6 years). To detect chromosomal translocations AF4/MLL t(4; 11) (q23; p23), BCR/ABL t (9; 22) (q34; q11), E2A/PBX1 t (1; 19) (q23; p13) and TEL/AML t(12; 21) (q13; q22) the method of polymerase chain reaction with reverse transcription (RT-PCR) was applicated. PCR was performed with specific primers for the appropriate chromosomal aberrations. Detection of PCR products was performed by electrophoresis in 2% agarose gel. Determination of minimal residual disease (MRD) was performed by multiparameter flow cytofluorimetry using monoclonal antibodies. Results. Among patients, the incidence of ALL is most pronounced in children aged 3 to 6 years - 37 people (35.2%) and aged 6 to 9 years - 26 people (24.8%). The highest accidence was found among patients with chromosomal translocation TEL/AML - 22 (21%) of patients with a median age of 5 years. In the second place, the frequency of mutations is the translocation of E2A / PBX1. BCR / ABL translocation was less common - 1.9% of patients, but the expression of this gene indicates a bad course of the disease, as patients after cytostatic therapy under the ALLIC BFM 2009 program had a recurrence. Recurrence has also been observed in patients with TEL/AML chromosomal translocation. Determination of MRD showed its increased level in patients with chromosomal aberrations BCR / ABL and TEL/AML throughout the treatment phase. In addition, patients in these groups were diagnosed with initial leukocytosis followed by leukopenia after a course of chemotherapy. Patients of all groups showed a decrease in hemoglobin. Conclusion. The most significant changes in clinical and laboratory parameters were found among patients with chromosomal translocations BCR/ABL and TEL/AML, as evidenced by the development of relapses in patients of these groups. The low level of association between karyotype disorders, with the formation of AF4/MLL and E2A/PBX1, and clinical and laboratory parameters in patients with GLL may indicate that the isolated clonal disorders are independent prognostic factors for the course of the disease.

MOLECULAR AND GENETIC MECHANISMS OF DEVELOPMENT OF CHILDHOOD ACUTE LYMPHOBLASTIC LEUKEMIA AND THEIR CORRECTION ALLIC BFM 2009 BY CYTOSTATIC THERAPY

The aim of this study was to evaluate the prognostic value of the relationship between genetic abnormalities and clinical and laboratory parameters of peripheral blood and bone marrow in children with acute lymphoblastic leukemia (ALL).Material and methods. 105 children diagnosed with ALL were examined (average age 6 years). To detect chromosomal translocations AF4/MLL t(4; 11) (q23; p23), BCR/ABL t (9; 22) (q34; q11), E2A/PBX1 t (1; 19) (q23; p13) and TEL/AML t(12; 21) (q13; q22) the method of polymerase chain reaction with reverse transcription (RT-PCR) was applicated. PCR was performed with specific primers for the appropriate chromosomal aberrations. Detection of PCR products was performed by electrophoresis in 2% agarose gel. Determination of minimal residual disease (MRD) was performed by multiparameter flow cytofluorimetry using monoclonal antibodies.Results. Among patients, the incidence of ALL is most pronounced in children aged 3 to 6 years - 37 people (35.2%) and aged 6 to 9 years - 26 people (24.8%). The highest accidence was found among patients with chromosomal translocation TEL / AML - 22 (21%) of patients with a median age 5 years. In second place, the frequency of mutations is the translocation of E2A / PBX1. BCR / ABL translocation was less common - 1.9% of patients, but the expression of this gene indicates a bad course of the disease, as patients after cytostatic therapy under the ALLIC BFM 2009 program had a recurrence. Recurrence has also been observed in patients with TEL/AML chromosomal translocation.Determination of MRD showed its increased level in patients with chromosomal aberrations BCR / ABL and TEL/AML throughout the treatment phase. In addition, patients in these groups were diagnosed with initial leukocytosis followed by leukopenia after a course of chemotherapy. Patients of all groups showed a decrease in hemoglobin.Conclusion. The biggest changes in clinical and laboratory parameters were found between patients with chromosomal translocations BCR/ABL and TEL/AML, as evidenced by the development of relapses in patients of these groups. The low level of association between karyotype disorders, with the formation of AF4/MLL and E2A/PBX1, and clinical and laboratory parameters in patients with GLL may indicate that the isolated clonal disorders are independent prognostic factors for the course of the disease.