Abstract
Background:Tumor cell genome aberrations are often detected by various methods. Even while monitoring chimerism after bone marrow transplantation being done for patients with hematological malignancies, one could came across the loss of heterozygosity (LOH) in the loci of short tandem repeats (STR).Aims:To evaluate the frequency of LOH in relapsed and non‐relapsed patients with AML and ALL and possible association of LOH in STR loci with other genomic alterations in malignant cells. terozygosity (LOH) in the loci of short tandem repeats (STR).Methods:This study represents the analysis of STR profiles of 46 patients (27 AML, 17 ALL (4B‐ALL, 8 T‐ALL, 5 Ph + ALL), 2 mixed phenotype acute leukemia) who received allogeneic BMT between 2012 and 2018 in the National Research Center for Hematology, Moscow. The patients were divided in two cohorts: 26 patients without relapses (18AML, 4 Ph + ALL, 3 B‐ALL, 2 T‐ALL, 1 mixed phenotype acute leukemia) and 20 patients having bone marrow or central nervous system relapses (9 AML, 10 ALL, 1 mixed phenotype acute leukemia). DNA samples were isolated from bone marrow in onset of the disease (except six relapsed patients, for whom “debut” samples were not available), in relapse and from cerebrospinal fluid during CNS relapse. The presence of blast cells in bone marrow samples and cerebrospinal fluid was confirmed morphologically and cytogenetically. The control DNA samples were isolated from bone marrow of the same patients in remission. STR‐profiles were assessed by PCR with COrDIS Plus multiplex kit for amplification of 19 polymorphic STR‐markers and amelogenin loci (Gordiz Ltd, Russia). The frequencies of heterozygosity for STR loci D3S1358, TH01, D12S391, D1S1656, D10S1248, D22S1045, D2S441, D7S820, D13S317, FGA, TPOX, D18S51, D16S539, D8S1179, CSF1PO, D5S818, VWA, D21S11, SE33 was evaluated on extended patients and donors groups, 120 persons each. The fragment analysis was performed on ABI3130 Genetic Analyzer. The data processing was accomplished using GeneMapper v.4‐0 software. Whole‐genome chromosomal microarray (CMA) was performed on Affimetrix CytoscanTM HD array platform. Data processing was carried out using Chromosome Analysis Suite v.4.0 software, genome assembly version hg19, GRCh37 Genome Reference Consortium Human Reference 37, https://genome.ucsc.edu/.Results:In preliminary experiments on extended cohorts the frequency of heterozygosity for each of STR loci was established between 70% to 95%. Three patients (11.5%) were identified with a loss of heterozygosity at the STR loci in the “no relapses” cohort, 2 from 18 AML (11%) and one T‐ALL patient. For “patients in relapse” LOH by STR analysis was detected in 10 cases from 20, and additionally three patients were identified as LOH‐positive by CMA (65%): 5 from 9 AML patients (55.5%) and 7 from10 ALL patients (70%) and one mixed phenotype acute leukemia patient (Table1). One B‐ALL patient with four CNS relapses and three bone marrow relapses had LOH in 10 from 15 heterozygous loci (Table1, patient#3). The CMA showed that LOH of STR loci often is a result of uniparental disomy (UPD). For two AML relapsed patients (Table1, patients#2,#5) UPD of 13 chromosome resulted into FLT3/ITD mutation (13q12.11) homozygocity. Therefore these patients shoud be considered candidates for target therapy against FLT3.Summary/Conclusion:LOH frequency in STR loci is significantly higher for relapsed patients versus those relapse‐free. CMA confirmes and expands STR analysis data,evaluates genes crucial for disease progression in certain patient and may possibly help to select effective target therapy.image
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