A novel flow cytometric method for enhancing acute promyelocytic leukemia screening by multidimensional dot-plots

Abstract

Acute promyelocytic leukemia (APL) is generally characterized by t(15;17)(q24;q21). In some cases, the classic translocation cannot be identified by conventional methods, since the PML-RARA fusion protein results from complex, variant, or cryptic translocation. The diagnostic algorithm of APL starts with screening methods, such as flow cytometry (FC), followed by fluorescence in situ hybridization or polymerase chain reaction to confirm the diagnosis. Our aim was to develop a novel protocol for analyzing APL samples based on multidimensional dot-plots that can provide comprehensive information about several markers at the same time. The protocol included four optimized multidimensional dot-plots, which were tested by retrospective reanalysis of FC results in APL ( n = 8) and non-APL ( n = 12) acute myeloid leukemia (AML) cases. After predicting the potential position of hypergranular- and microgranular-type aberrant promyelocytes, the percentages of blast populations were examined within the gates in all AML cases. The percentage of blasts in each predefined gate was well above the cut-off value (95%) in APL cases in all tubes. In non-APL AML cases, the percentage of blasts in the same gates never reached the cut-off value in all investigated tubes, and even when it did in a single tube, the pattern was markedly different from that observed in APL cases. In conclusion, multidimensional dot-plots can be used for screening APL even in cryptic APL cases, although reproducibility across several laboratories would require standardization of antibodies and fluorochromes. This easy-to-use and quick method can support the diagnosis of APL and the prompt initiation of the appropriate treatment. Electronic supplementary material The online version of this article (10.1007/s00277-019-03642-w) contains supplementary material, which is available to authorized users.

Recent Advances in the Management of Pediatric Acute Myeloid Leukemia—Report of the Hungarian Pediatric Oncology-Hematology Group

“…However, results of the multidimensional flow cytometric analysis suggested unequivocally that both patients had APL. More than 95% of the pathological cells were detected in the APL gate pre-defined in our previous study in all multidimensional dot-plots in both cases (Figure 4) [17]. This means that the positions of pathological cells were typical for APL.…”

“…One dot-plot was developed for each of the four tubes. During the analysis, the position of pathological cells was compared to the expected position of pathological promyelocytes in hypergranular and hypogranular APL, as defined in the previous study [17].…”

Radar plots facilitate differential diagnosis of acute promyelocytic leukemia and NPM1+ acute myeloid leukemia by flow cytometry

“…One previous study described a unique localization of APL on RPs applying a four‐tube 8‐color FCM panel including cytoplasmic myeloperoxidase. A limited number of APL ( n = 8) and NPM1+ AML samples were compared ( n = 12) (Karai et al, 2019 ). The authors defined a characteristic location of APL on created RPs using merged files for hypergranular ( n = 6) and hypogranular ( n = 2) APLs.…”

CD markers polymorphisms as prognostic biomarkers in hematological malignancies

“…The phenotype of the involved cells, often characterized by the expression of surface CD markers, is another feature playing an essential role in the diagnosis and prognosis of hematological malignancies and solid tumors. 3 , 71 , 72 Although flow cytometry is a practical technique to analyze the expression levels of CD markers, genetic alterations (including SNPs) affecting expression patterns of CD markers may cause significant limitations in the classification and precise prognosis of hematological malignancies. 73-75 As an example, CD38 rs6449182 is a known polymorphism in CLL and it has been shown that patients carrying the GG genotype of this polymorphism have more aggressive disease.…”

“…Therefore, the evaluation of CD markers’ expressions is used as the first diagnostic strategy that is followed by an appropriate approach to monitoring the clinical course of leukemia. 3 Despite the fact that specific immunophenotypic patterns have been defined for the diagnosis of leukemia and lymphoma, results of studies in recent years indicate the heterogeneous expressions of CD markers in these malignancies. Numerous environmental and genetic factors like single nucleotide polymorphisms (SNPs) can alter the expression patterns of these surface markers.…”