Chromosomal rearrangements of the human MLL (mixed lineage leukemia) gene are associated with high-risk infant, pediatric, adult and therapy-induced acute leukemias. We used long-distance inverse-polymerase chain reaction to characterize the chromosomal rearrangement of individual acute leukemia patients. We present data of the molecular characterization of 1590 MLL-rearranged biopsy samples obtained from acute leukemia patients. The precise localization of genomic breakpoints within the MLL gene and the involved translocation partner genes (TPGs) were determined and novel TPGs identified. All patients were classified according to their gender (852 females and 745 males), age at diagnosis (558 infant, 416 pediatric and 616 adult leukemia patients) and other clinical criteria. Combined data of our study and recently published data revealed a total of 121 different MLL rearrangements, of which 79 TPGs are now characterized at the molecular level. However, only seven rearrangements seem to be predominantly associated with illegitimate recombinations of the MLL gene ( ∼ 90%): AFF1/AF4, MLLT3/AF9, MLLT1/ENL, MLLT10/AF10, ELL, partial tandem duplications (MLL PTDs) and MLLT4/AF6, respectively. The MLL breakpoint distributions for all clinical relevant subtypes (gender, disease type, age at diagnosis, reciprocal, complex and therapy-induced translocations) are presented. Finally, we present the extending network of reciprocal MLL fusions deriving from complex rearrangements.

Chromosomal rearrangements of the human MLL gene are associated with high-risk pediatric, adult and therapy-associated acute leukemias. These patients need to be identified, treated appropriately and minimal residual disease was monitored by quantitative PCR techniques. Genomic DNA was isolated from individual acute leukemia patients to identify and characterize chromosomal rearrangements involving the human MLL gene. A total of 760 MLL-rearranged biopsy samples obtained from 384 pediatric and 376 adult leukemia patients were characterized at the molecular level. The distribution of MLL breakpoints for clinical subtypes (acute lymphoblastic leukemia, acute myeloid leukemia, pediatric and adult) and fused translocation partner genes (TPGs) will be presented, including novel MLL fusion genes. Combined data of our study and recently published data revealed 104 different MLL rearrangements of which 64 TPGs are now characterized on the molecular level. Nine TPGs seem to be predominantly involved in genetic recombinations of MLL: AFF1/AF4, MLLT3/AF9, MLLT1/ENL, MLLT10/AF10, MLLT4/AF6, ELL, EPS15/AF1P, MLLT6/AF17 and SEPT6, respectively. Moreover, we describe for the first time the genetic network of reciprocal MLL gene fusions deriving from complex rearrangements.

We report incidence and deep molecular characteristics of lineage switch in 182 pediatric patients affected by B-cell precursor acute lymphoblastic leukemia (BCP-ALL), who were treated with blinatumomab. We documented six cases of lineage switch that occurred after or during blinatumomab exposure. Therefore, lineage conversion was found in 17.4% of all resistance cases (4/27) and 3.2% of relapses (2/63). Half of patients switched completely from BCP-ALL to CD19-negative acute myeloid leukemia, others retained CD19-positive B-blasts and acquired an additional CD19-negative blast population: myeloid or unclassifiable. Five patients had KMT2A gene rearrangements; one had TCF3::ZNF384 translocation. The presented cases showed consistency of gene rearrangements and fusion transcripts across initially diagnosed leukemia and lineage switch. In two of six patients, the clonal architecture assessed by IG/TR gene rearrangements was stable, while in others, loss of clones or gain of new clones was noted. KMT2A-r patients demonstrated very few additional mutations, while in the TCF3::ZNF384 case, lineage switch was accompanied by a large set of additional mutations. The immunophenotype of an existing leukemia sometimes changes via different mechanisms and with different additional molecular changes. Careful investigation of all BM compartments together with all molecular –minimal residual disease studies can lead to reliable identification of lineage switch.

Background. Flow cytometry is one of the key technologies for acute lymphoblastic leukemia (ALL) diagnostics. Nevertheless lack of technological standards hampers implementation of immunophenotyping data in treatment protocols.Objective: development of harmonized guidelines for flow cytometric diagnostics of childhood ALL. Materials and methods. Three reference laboratories of the immunophenotyping group “Moscow–Berlin” took part in the development and standardization of approaches to the cytometric determination of the minimal residual disease. The sequence of steps for staining with monoclonal antibodies, selection of reagents for immunophenotyping, flow cytometer adjustment algorithms were discussed.Results. We developed and implemented in multicenter setting the harmonized approach for immunophenotyping flow cytometric childhood ALL. Successful integration of this protocol in the multicenter group has shown good level of our approach reproducibility. Conclusion. These results will allow implementing diagnostic standards in stratification system of pediatric ALL treatment protocols of russianbelarusian multicenter group.

   Introduction. Currently, there is no unequivocal opinion on the optimal list of studies for the genetic diagnosis of oncohematological disorders in children and adults. These discrepancies are due to the limited technological capabilities of laboratories, the rapid development of science, and a significant expansion of the range of new molecular markers, that are attractive, but only for a limited group of patients. Moreover, in modern conditions of limited access to resources, it seems important to bring desires, interests and opportunities to a common denominator.   Aim — to develop unified approaches to the cytogenetic and molecular genetic diagnosis of oncohematological diseases in children and adults based on the consensus opinion of the panel of experts.   Main findings. The review proposes the arrangement of cytogenetic and molecular genetic diagnostic tests in oncohematological disorders in children and adults into 3 categories depending on the frequency of genetic aberrations, the study complexity and the prognostic impact. Based on this and taking into account the diagnosis and age of patients, the minimal and optimal lists of clinically significant parameters and research markers were identified. The basic preanalytical principles for conducting cytogenetic and molecular genetic studies in oncohematology are pointed out. A brief description of a conventional cytogenetic study and a polymerase chain reaction for the diagnosis of oncohematological diseases is given. The paper also focused on the need for reference diagnostics of cytogenetic and molecular genetic studies in oncohematology. The article is addressed to the specialists in the field of laboratory genetics, clinical laboratory diagnostics, but may also be of interest to hematologists, pediatric oncologists and doctors of related branches.