Background: The significance of very early chimerism assessment before day + 28, which is considered the moment of engraftment, is still unclear. In this retrospective study, we evaluated the clinical impact of very early chimerism on the clinical outcome after allogeneic haematopoietic stem cell transplantation (allo-HSCT) in children with acute lymphoblastic leukaemia (ALL). Methods: The study group included 38 boys and 18 girls. Very early chimerism was evaluated on days + 7, + 14, + 21 and + 28 after the transplant. Short tandem repeat polymerase chain reaction (STR PCR) was used to analyse chimerism. Results: Overall survival (OS) and event-free survival (EFS) were 84 and 80%, respectively. The OS in the group of 24 patients with complete donor chimerism on day + 14 was 83%, and it did not differ statistically compared to the 32 patients with mixed chimerism on day + 14 (OS was 84%). In our cohort of patients, the matched unrelated donor, male gender of donor, number of transplanted cells above 4.47 × 10⁶ kg and no serotherapy with anti-thymocyte globulin (ATG) were statistically related to a higher level of donor chimerism. The immunophenotypes of disease, age of patient at time HSCT, recipient sex, stem cell source (peripheral blood/bone marrow) and conditioning regimen had no impact on early chimerism. Acute graft versus host disease grades II-IV was diagnosed in 23 patients who presented with donor chimerism levels above 60% on day 7. Conclusions: The data presented in this study provide valuable insight into the analysis of very early chimerism in children with ALL treated with HSCT.

The classical BCR::ABL1-negative myeloproliferative neoplasms such as polycythemia vera (PV), essential thrombocythemia (ET), and myelofibrosis (MF) are clonal diseases with the presence of characteristic “driver mutations” in one of the genes: JAK2, CALR, or MPL. The search for mutations in these three genes is required for the diagnosis of MPNs. Nevertheless, the progress that has been made in the field of molecular genetics has opened a new era in medicine. The search for additional mutations in MPNs is helpful in assessing the risk stratification, disease progression, transformation to acute myeloid leukemia (AML), or choosing the right treatment. In some cases, advanced technologies are needed to find a clonal marker of the disease and establish a diagnosis. This review focuses on how the use of new technologies like next-generation sequencing (NGS) helps in the diagnosis of BCR::ABL1-negative myeloproliferative neoplasms.

Background: Acute lymphoblastic leukemia is the most common type of cancer in children. Most often it affects the age group between 2 and 5 years of age. Studies have shown an improvement in general survivability, more than 90% 5-year overall survival (OS). Current treatment protocols for acute lymphoblastic leukemia require verification of the presence of favorable and unfavorable genetic abnormalities, which help qualify patients to the appropriate risk group and select a more suitable treatment. The presence of the BCR/ABL1 fusion gene stratifies the patient into a high-risk group and requires special treatment with tyrosine kinase inhibitors (TKI). The three dominant mRNA transcripts are e1a2, e13a2, and e14a2. Nevertheless, cases of atypical BCR/ABL1 transcripts have also been reported. Case presentation: This paper presents the case of a pediatric patient with Ph + B-cell precursor acute lymphoblastic leukemia with rare atypical e8a2 BCR/ABL1 fusion transcript. Our patient achieved complete remission after 33 days of treatment. Molecular and cytogenetic studies in TP1 did not reveal the presence of the BCR/ABL1 transcript. The PCR-MRD test in TP1b was negative, the patient did not require hematopoietic stem cell transplantation. Conclusion: Genetic evaluation of the bone marrow sample is crucial in the initial stage of the diagnosis. Fluorescent in situ hybridization and reverse transcriptase polymerase chain reaction with Sanger sequencing are the appropriate methods used in the detection of rare variants of BCR/ABL1 transcripts.

Essential thrombocythemia and primary myelofibrosis belong to the group of BCR::ABL1 -negative myeloproliferative neoplasms. The presence of mutations in the JAK2 , CALR , and MPL genes is essential for the diagnosis of myeloproliferative neoplasms. These mutations are called “driver” mutations. However, not only leading mutations have been identified in patients with MPN, but also more than half of individuals with essential thrombocythemia and more than 80% of patients with myelofibrosis have additional mutations. One technique that makes it possible to find prognostic, predictive, and diagnostic indicators is next-generation sequencing. Coexisting mutations are associated with reduced response to therapy, shortened overall survival, and a higher risk of transformation to acute myeloid leukemia or myelofibrosis. The study group consisted of 42 patients with the diagnosis of BCR::ABL1 -negative MPN and the presence of a mutation in the CALR gene. The research material was archival, and DNA was obtained from patients’ peripheral blood. Forty genes (17 genes, 23 hotspots) were sequenced using the commercial kit AmpliSeq for Illumina Myeloid Panel applying the targeted next-generation sequencing approach. For the study, the Illumina MiniSeq platform was used. The analysis of the obtained genetic results was carried out using bioinformatics tools and genetic databases. We studied 42 CALR -positive ET ( n = 28) and MF ( n = 14) patients with NGS panel testing. The median age at diagnosis of the entire patient series was 58 years. Additional mutations were detected in 48% of patients in the whole cohort. The most frequently mutated genes in the study population were ASXL1 , TET2 , and DNMT3A , which are largely associated with epigenetic regulatory mechanisms. NGS panel studies represent a breakthrough in the diagnostic and prognostic evaluation of MPNs with CALR mutations. The ability to perform such a comprehensive study provides valuable information on the biology of the disease and the selection of the appropriate treatment regimen. The use of new technologies shows that not only driver mutations have clinical significance for the patient. NGS has the potential to increase the precision and effectiveness of diagnosis and prognosis.

Https://doi.org/10.18632/oncotarget.28686 Aleksandra Mroczkowska-Bękarciak, Tomasz Wróbel

Acute lymphoblastic leukemia (ALL) is the most common malignancy of childhood. The presence or absence of a characteristic genetic abnormality usually observed in childhood ALL plays a very important role in determining the prognosis and stratification for treatment. Intrachromosomal amplification of chromosome 21 (iAMP21) is an uncommon high-risk chromosomal abnormality than can occur only in 2% of childhood B-cell precursor lymphoblastic leukemia. Molecular genetic analysis and the fluorescence in situ hybridization (FISH) technique are the basic methods used to detect the presence of the most common genetic abnormalities, the presence or absence of which has an impact on the patient’s classification into the appropriate risk group. This work presents 3 BCP-ALL iAMP21-positive patients who were detected during routine genetic diagnostics using the FISH method and microarray test. iAMP21 is associated with a poor prognosis and high risk for relapse. Children with B-cell precursor lymphoblastic leukemia with this genetic entity are associated with a delayed treatment response. The FISH method and single-nucleotide polymorphism array provides a useful method to detect characteristic genetic changes.