Flow cytometry in acute leukemia diagnostics. Guidelines of Russian-Belarusian multicenter group for pediatric leukemia studies

Popov, Alexander; Verzhbitskaya, Tatiana; Movchan, Liudmila; Demina, Irina; Mikhailova, Ekaterina; Semchenkova, Alexandra; Permikin, Zhan; Шман, Т. В.; Karachunskiy, Alexander; Novichkova, Galina

doi:10.24287/1726-1708-2023-22-1-165-177

Cited by 10 publications

(7 citation statements)

References 49 publications

(51 reference statements)

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“…Three of ten patients were initially diagnosed with TI-ALL, and seven had TII-ALL. According to the current diagnostic standards [ 13 ], TII-ALL was diagnosed based on the presence of surface CD2 and/or CD5 (CD5 expression level < 75%). Thus, the TII-ALL group of ETP-ALL can be divided into two subgroups based on the absence/presence of CD5.…”

Section: Resultsmentioning

confidence: 99%

“…This is because early T-cell precursors are essentially immature thymocytes that retain stem cell-like features and can differentiate into both T lymphoid and myeloid cells [ 19 , 20 , 21 ]. Their assignment to ETP-ALL by immunophenotyping is based on the detection of intracellular CD3 expression and the presence of surface CD7 [ 10 , 13 ]. According to the classification, 10% expression of intracellular CD3 is sufficient to assign the whole population to the T-cell lineage of differentiation [ 10 ].…”

Section: Discussionmentioning

confidence: 99%

“…Leukemic cells were characterized at initial diagnosis and relapse using a range of techniques, including immunophenotyping, fluorescent in situ hybridization (FISH), conventional karyotyping, next-generation sequencing (NGS) of the clonal T-cell receptor (TCR) repertoire, reverse transcription PCR (RT−PCR), and RNA sequencing (RNA-seq). Immunophenotyping was performed according to established guidelines [ 13 ]. ALL subtypes were defined per EGIL [ 31 ] with later modifications [ 10 , 13 ].…”

Section: Methodsmentioning

confidence: 99%

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Immunophenotypic but Not Genetic Changes Reclassify the Majority of Relapsed/Refractory Pediatric Cases of Early T-Cell Precursor Acute Lymphoblastic Leukemia

Demina,

Dagestani,

Borkovskaia

et al. 2024

IJMS

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Early T-cell precursor acute lymphoblastic leukemia (ETP-ALL) develops from very early cells with the potential for both T-cell and myeloid differentiation. The ambiguous nature of leukemic blasts in ETP-ALL may lead to immunophenotypic alterations at relapse. Here, we address immunophenotypic alterations and related classification issues, as well as genetic features of relapsed pediatric ETP-ALL. Between 2017 and 2022, 7518 patients were diagnosed with acute leukemia (AL). In addition to conventional immunophenotyping, karyotyping, and FISH studies, we performed next-generation sequencing of the T-cell receptor clonal repertoire and reverse transcription PCR and RNA sequencing for patients with ETP-ALL at both initial diagnosis and relapse. Among a total of 534 patients diagnosed with T-cell ALL (7.1%), 60 had ETP-ALL (11.2%). Ten patients with ETP-ALL experienced relapse or progression on therapy (16.7%), with a median time to event of 5 months (ranging from two weeks to 5 years). Most relapses were classified as AL of ambiguous lineage (n = 5) and acute myeloid leukemia (AML) (n = 4). Major genetic markers of leukemic cells remained unchanged at relapse. Of the patients with relapse, four had polyclonal leukemic populations and a relapse with AML or bilineal mixed-phenotype AL (MPAL). Three patients had clonal TRD rearrangements and relapse with AML, undifferentiated AL, or retention of the ETP-ALL phenotype. ETP-ALL relapse requires careful clinical and laboratory diagnosis. Treatment decisions should rely mainly on initial examination data, taking into account both immunophenotypic and molecular/genetic characteristics.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Immunophenotypic but Not Genetic Changes Reclassify the Majority of Relapsed/Refractory Pediatric Cases of Early T-Cell Precursor Acute Lymphoblastic Leukemia

Demina,

Dagestani,

Borkovskaia

et al. 2024

IJMS

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“…As already described, the patients were divided into standard risk (SR) and intermediate risk (ImR) ( online supplemental figure S1 ) 9 10 and received the normal risk-adapted induction therapy in accordance with the ALL-MB protocol 2015 ( figure 1 ). 11 Before treatment, a centralized and standardized diagnosis including cytomorphology, immunophenotyping 12 and cyto- and molecular genetics 13 were routinely carried out in all patients. Using multicolor flow cytometry (MFC), MRD was measured at EOI, immediately after the treatment with blinatumomab and then four times at 3-month intervals during maintenance therapy ( figure 1 ).…”

Section: Methodsmentioning

confidence: 99%

Blinatumomab as postremission therapy replaces consolidation and substantial parts of maintenance chemotherapy and results in stable MRD negativity in children with newly diagnosed B-lineage ALL

Mikhailova,

Popov,

Roumiantseva

et al. 2024

J Immunother Cancer

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The bispecific T cell-binding antibody blinatumomab (CD19/CD3) is widely and successfully used for the treatment of children with relapsed or refractory B-cell precursor acute lymphoblastic leukemia (BCP-ALL). Here, we report the efficacy of a single course of blinatumomab instead of consolidation chemotherapy to eliminate minimal residual disease (MRD) and maintain stable MRD-negativity in children with primary BCP-ALL.Between February 2020 and November 2022, 177 children with non-high-risk BCP-ALL were enrolled in the ALL-MB 2019 pilot study (NCT04723342). Patients received the usual risk-adapted induction therapy according to the ALL-MB 2015 protocol. Those who achieved a complete remission at the end of induction (EOI) received treatment with blinatumomab immediately after induction at a dose of 5 μg/m2/day for 7 days and 21 days at a dose of 15 μg/m2/day, followed by 12 months of maintenance therapy. MRD was measured using multicolor flow cytometry (MFC) at the EOI, then immediately after blinatumomab treatment, and then four times during maintenance therapy at 3-month intervals.All 177 patients successfully completed induction therapy and achieved a complete hematological remission. In 174 of these, MFC-MRD was measured at the EOI. 143 patients (82.2%) were MFC-MRD negative and the remaining 31 patients had varying degrees of MFC-MRD positivity.MFC-MRD was assessed in all 176 patients who completed the blinatumomab course. With one exception, all patients achieved MFC-MRD negativity after blinatumomab, regardless of the MFC-MRD score at EOI. One adolescent girl with high MFC-MRD positivity at EOI remained MFC-MRD positive. Of 175 patients who had completed 6 months of maintenance therapy, MFC-MRD data were available for 156 children. Of these, 155 (99.4%) were MFC-MRD negative. Only one boy with t(12;21) (p13;q22)/ETV6::RUNX1became MFC-MRD positive again. The remaining 174 children had completed the entire therapy. MFC-MRD was examined in 154 of them, and 153 were MFC-MRD negative. A girl with hypodiploid BCP-ALL showed a reappearance of MFC-MRD with subsequent relapse.In summary, a single 28-day course of blinatumomab immediately after induction, followed by 12 months of maintenance therapy, is highly effective in achieving MRD-negativity in children with newly diagnosed non-high risk BCP-ALL and maintaining MRD-negative remission at least during the treatment period.

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“…The antigen expression profile of tumor blasts was investigated by 6–10-color MFC using three-laser flow cytometers: FACS Canto II (Becton Dickinson, BD, San Jose, CA, USA) or Navios (Beckman Coulter, BC, Indianapolis, IN, USA). EuroFlow guidelines for machine performance monitoring were used [ 18 ]. Cytometer Setup and Tracking Beads (BD) and Flow-Check Pro Fluorospheres (BC) were used for daily cytometer optimization.…”

Section: Methodsmentioning

confidence: 99%

Molecular Heterogeneity of Pediatric AML with Atypical Promyelocytes Accumulation in Children—A Single Center Experience

Borkovskaia

Bogacheva

Konyukhova

et al. 2023

Genes

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Acute promyelocytic leukemia (APL) pathogenesis is based on RARA gene translocations, which are of high importance in the diagnosis of and proper therapy selection for APL. However, in some cases acute myeloid leukemia (AML) demonstrates APL-like morphological features such as atypical promyelocytes accumulation. This type of AML is characterized by the involvement of other RAR family members or completely different genes. In the present study, we used conventional karyotyping, FISH and high-throughput sequencing in a group of 271 de novo AML with atypical promyelocytes accumulation. Of those, 255 cases were shown to carry a typical chromosomal translocation t(15;17)(q24;q21) with PML::RARA chimeric gene formation (94.1%). Other RARA-positive cases exhibited cryptic PML::RARA fusion without t(15;17)(q24;q21) (1.8%, n = 5) and variant t(5;17)(q35;q21) translocation with NPM1::RARA chimeric gene formation (1.5%, n = 4). However, 7 RARA-negative AMLs with atypical promyelocytes accumulation were also discovered. These cases exhibited TBL1XR1::RARB and KMT2A::SEPT6 fusions as well as mutations, e.g., NPM1 insertion and non-recurrent chromosomal aberrations. Our findings demonstrate the genetic diversity of AML with APL-like morphological features, which is of high importance for successful therapy implementation.

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Flow cytometry in acute leukemia diagnostics. Guidelines of Russian-Belarusian multicenter group for pediatric leukemia studies

Cited by 10 publications

References 49 publications

Immunophenotypic but Not Genetic Changes Reclassify the Majority of Relapsed/Refractory Pediatric Cases of Early T-Cell Precursor Acute Lymphoblastic Leukemia

Immunophenotypic but Not Genetic Changes Reclassify the Majority of Relapsed/Refractory Pediatric Cases of Early T-Cell Precursor Acute Lymphoblastic Leukemia

Blinatumomab as postremission therapy replaces consolidation and substantial parts of maintenance chemotherapy and results in stable MRD negativity in children with newly diagnosed B-lineage ALL

Molecular Heterogeneity of Pediatric AML with Atypical Promyelocytes Accumulation in Children—A Single Center Experience

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