Selection of the proper target is crucial for clinically relevant monitoring of minimal residual disease (MRD) in patients with acute lymphoblastic leukemia using the quantitation of clonal-specific immunoreceptor (immunoglobulin/T cell receptor) gene rearrangements. Consequently, correct interpretation of the results of the entire analysis is of utmost importance. Here we present an overview of the quality control measures that need to be implemented into the process of marker identification, selection, and subsequent quantitation of the MRD level.

• Klíčová slova
• Acute lymphoblastic leukemia, Minimal residual disease, Next-generation sequencing, PCR, Quality control,
• MeSH
• akutní lymfatická leukemie * diagnóza   genetika   MeSH
• biologické markery MeSH
• imunoglobuliny genetika   MeSH
• lidé MeSH
• reziduální nádor diagnóza   genetika   MeSH
• řízení kvality MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• biologické markery MeSH
• imunoglobuliny MeSH

Minimal residual disease (MRD) monitoring via quantitative PCR (qPCR) detection of Ag receptor gene rearrangements has been the most sensitive method for predicting prognosis and making post-transplant treatment decisions for patients with ALL. Despite the broad clinical usefulness and standardization of this method, we and others have repeatedly reported the possibility of false-positive MRD results caused by massive B-lymphocyte regeneration after stem cell transplantation (SCT). Next-generation sequencing (NGS) enables precise and sensitive detection of multiple Ag receptor rearrangements, thus providing a more specific readout compared to qPCR. We investigated two cohorts of children with ALL who underwent SCT (30 patients and 228 samples). The first cohort consisted of 17 patients who remained in long-term CR after SCT despite having low MRD positivity (<0.01%) at least once during post-SCT monitoring using qPCR. Only one of 27 qPCR-positive samples was confirmed to be positive by NGS. Conversely, 10 of 15 samples with low qPCR-detected MRD positivity from 13 patients who subsequently relapsed were also confirmed to be positive by NGS (P=0.002). These data show that NGS has a better specificity in post-SCT ALL management and indicate that treatment interventions aimed at reverting impending relapse should not be based on qPCR only.

• MeSH
• akutní lymfatická leukemie * krev   diagnóza   genetika   terapie   MeSH
• dítě MeSH
• falešně pozitivní reakce MeSH
• lidé MeSH
• mladiství MeSH
• polymerázová řetězová reakce * MeSH
• předškolní dítě MeSH
• prognóza MeSH
• reziduální nádor MeSH
• transplantace hematopoetických kmenových buněk * MeSH
• vysoce účinné nukleotidové sekvenování * MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• klinické zkoušky MeSH
• multicentrická studie MeSH

The prognosis for children with high-risk relapsed acute lymphoblastic leukemia (ALL) is poor. Here, we assessed the prognostic importance of response during induction and consolidation treatment prior to hematopoietic stem cell transplantation (HSCT) aiming to evaluate the best time to assess minimal residual disease (MRD) for intervention strategies and in future trials in high-risk ALL relapse patients. Included patients (n=125) were treated uniformly according to the ALL-REZ BFM (Berlin-Frankfurt-Münster) 2002 relapse trial (median follow-up time=4.8 years). Patients with MRD ⩾10(-3) after induction treatment (76/119, 64%) or immediately preceding HSCT (19/71, 27%) had a significantly worse probability of disease-free survival 10 years after relapse treatment begin, with 26% (±6%) or 23% (±7%), respectively, compared with 58% (±8%) or 48% (±7%) for patients with MRD <10(-3). Conventional intensive consolidation treatment reduced MRD to <10(-3) before HSCT in 63% of patients, whereas MRD remained high or increased in the rest of this patient group. Our data support that MRD after induction treatment can be used to quantify the activity of different induction treatment strategies in phase II trials. MRD persistence at ⩾10(-3) before HSCT reflects a disease highly resistant to conventional intensive chemotherapy and requiring prospective controlled investigation of new treatment strategies and drugs.

• MeSH
• akutní lymfatická leukemie farmakoterapie   mortalita   patologie   MeSH
• dítě MeSH
• indukční chemoterapie MeSH
• lidé MeSH
• lokální recidiva nádoru farmakoterapie   mortalita   patologie   MeSH
• míra přežití MeSH
• mladiství MeSH
• monitorování fyziologických funkcí * MeSH
• následné studie MeSH
• prognóza MeSH
• prospektivní studie MeSH
• protokoly protinádorové kombinované chemoterapie terapeutické užití   MeSH
• reziduální nádor farmakoterapie   mortalita   patologie   MeSH
• rizikové faktory MeSH
• staging nádorů MeSH
• transplantace hematopoetických kmenových buněk MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• multicentrická studie MeSH
• práce podpořená grantem MeSH

BACKGROUND: Most minimal residual disease-directed treatment interventions in current treatment protocols for acute lymphoblastic leukemia are based on bone marrow testing, which is a consequence of previous studies showing the superiority of bone marrow over peripheral blood as an investigational material. Those studies typically did not explore the prognostic impact of peripheral blood involvement and lacked samples from very early time points of induction. DESIGN AND METHODS: In this study, we employed real-time quantitative polymerase chain reaction analysis to examine minimal residual disease in 398 pairs of blood and bone marrow follow-up samples taken from 95 children with B-cell precursor acute lymphoblastic leukemia treated with the ALL IC-BFM 2002 protocol. RESULTS: We confirmed the previously published poor correlation between minimal residual disease in blood and marrow at early treatment time points, with levels in bone marrow being higher than in blood in most samples (median 7.9-fold, range 0.04-8,293-fold). A greater involvement of peripheral blood at diagnosis was associated with a higher white blood cell count at diagnosis (P=0.003) and with enlargement of the spleen (P=0.0004) and liver (P=0.05). At day 15, a level of minimal residual disease in blood lower than 10(-4) was associated with an excellent 5-year relapse-free survival in 78 investigated patients (100% versus 69 ± 7%; P=0.0003). Subgroups defined by the level of minimal residual disease in blood at day 15 (high-risk: ≥ 10(-2), intermediate-risk: <10(-2) and ≥ 10(-4), standard-risk: <10(-4)) partially correlated with bone marrow-based stratification described previously, but the risk groups did not match completely. No other time point analyses were predictive of outcome in peripheral blood, except for a weak association at day 8. CONCLUSIONS: Minimal residual disease in peripheral blood at day 15 identified a large group of patients with an excellent prognosis and added prognostic information to the risk stratification based on minimal residual disease at day 33 and week 12.

• MeSH
• časové faktory MeSH
• dítě MeSH
• kojenec MeSH
• kostní dřeň metabolismus   patologie   MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• lidé MeSH
• mladiství MeSH
• pre-B-buněčná leukemie krev   diagnóza   farmakoterapie   patologie   MeSH
• předškolní dítě MeSH
• prognóza MeSH
• protokoly protinádorové kombinované chemoterapie aplikace a dávkování   MeSH
• reziduální nádor MeSH
• Check Tag
• dítě MeSH
• kojenec MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• Publikační typ
• časopisecké články MeSH
• klinické zkoušky MeSH
• multicentrická studie MeSH
• práce podpořená grantem MeSH

BACKGROUND: While there is enough convincing evidence in childhood acute lymphoblastic leukemia (ALL), the data on the pre-natal origin in childhood acute myeloid leukemia (AML) are less comprehensive. Our study aimed to screen Guthrie cards (neonatal blood spots) of non-infant childhood AML and ALL patients for the presence of their respective leukemic markers. METHODS: We analysed Guthrie cards of 12 ALL patients aged 2-6 years using immunoglobulin (Ig) and T-cell receptor (TCR) gene rearrangements (n = 15) and/or intronic breakpoints of TEL/AML1 fusion gene (n = 3). In AML patients (n = 13, age 1-14 years) PML/RARalpha (n = 4), CBFbeta/MYH11 (n = 3), AML1/ETO (n = 2), MLL/AF6 (n = 1), MLL/AF9 (n = 1) and MLL/AF10 (n = 1) fusion genes and/or internal tandem duplication of FLT3 gene (FLT3/ITD) (n = 2) were used as clonotypic markers. Assay sensitivity determined using serial dilutions of patient DNA into the DNA of a healthy donor allowed us to detect the pre-leukemic clone in Guthrie card providing 1-3 positive cells were present in the neonatal blood spot. RESULTS: In 3 patients with ALL (25%) we reproducibly detected their leukemic markers (Ig/TCR n = 2; TEL/AML1 n = 1) in the Guthrie card. We did not find patient-specific molecular markers in any patient with AML. CONCLUSION: In the largest cohort examined so far we used identical approach for the backtracking of non-infant childhood ALL and AML. Our data suggest that either the prenatal origin of AML is less frequent or the load of pre-leukemic cells is significantly lower at birth in AML compared to ALL cases.

• MeSH
• akutní lymfatická leukemie krev   embryologie   epidemiologie   genetika   MeSH
• buněčné klony chemie   MeSH
• buňky kostní dřeně chemie   MeSH
• dítě MeSH
• DNA nádorová krev   MeSH
• duplikace genu MeSH
• fetální krev chemie   MeSH
• fúzní onkogenní proteiny krev   genetika   MeSH
• genová přestavba B-lymfocytů * MeSH
• genová přestavba T-lymfocytů * MeSH
• kohortové studie MeSH
• kojenec MeSH
• lidé MeSH
• myeloidní leukemie krev   embryologie   epidemiologie   genetika   MeSH
• nádorové biomarkery krev   MeSH
• nádorové proteiny krev   genetika   MeSH
• novorozenec MeSH
• novorozenecký screening MeSH
• polymerázová řetězová reakce MeSH
• předškolní dítě MeSH
• protein PEBP2A2 krev   genetika   MeSH
• protein RUNX1T1 MeSH
• protoonkogenní protein MLL krev   genetika   MeSH
• tandemové repetitivní sekvence MeSH
• tyrosinkinasa 3 podobná fms krev   genetika   MeSH
• Check Tag
• dítě MeSH
• kojenec MeSH
• lidé MeSH
• mužské pohlaví MeSH
• novorozenec MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• AML1-ETO fusion protein, human MeSH Prohlížeč
• CBFbeta-MYH11 fusion protein MeSH Prohlížeč
• DNA nádorová MeSH
• FLT3 protein, human MeSH Prohlížeč
• fúzní onkogenní proteiny MeSH
• MLL-AF10 fusion protein, human MeSH Prohlížeč
• MLL-AF6 fusion protein, human MeSH Prohlížeč
• nádorové biomarkery MeSH
• nádorové proteiny MeSH
• promyelocytic leukemia-retinoic acid receptor alpha fusion oncoprotein MeSH Prohlížeč
• protein PEBP2A2 MeSH
• protein RUNX1T1 MeSH
• protoonkogenní protein MLL MeSH
• TEL-AML1 fusion protein MeSH Prohlížeč
• tyrosinkinasa 3 podobná fms MeSH