Characterization of aberrant phenotypes in acute myeloblastic leukemia

Abstract

The existence of leukemic-associated phenotypes has been suggested to be a valuable tool for the detection of minimal residual disease (MRD) in AML patients, as they would allow to distinguish leukemic blast cells from normal hematopoietic progenitors. The present study was designed to analyze in which proportion of AML patients the immunological detection of MRD is feasible, based on the presence of aberrant phenotypes that allow the distinction of leukemic from normal cells. For this purpose we have prospectively investigated the blast cells from 40 AML patients at diagnosis with a large panel of MoAb in double and triple staining combinations analyzed at flow cytometry, in order to detect aberrant phenotypes on blast cells (lineage infidelity, antigenic overexpression, and asynchronous antigenic expression, as well as aberrant light-scatter pattern). In the analysis of the 40 AML cases more than one blast cell subset, distinguished by its different antigenic expression, was detected in 85% of the patients: five different phenotypic blast cell subsets were observed in six cases, four in 13 patients, three subsets in three cases, and two in 12 patients; only six cases showed a homogeneous phenotypical blast cell population. Twenty-nine of the 40 AML cases analyzed (73%) showed the existence of at least one aberrant phenotype: in 15 cases the myeloid blast cells co-expressed lymphoid-associated antigens (CD2, CD5, CD7, and/or CD19)--lineage infidelity--; asynchronous antigen expression was detected in 25 patients (CD34+CD56+, CD34+CD11b+, CD34+CD14+, CD117+CD15+, CD33-CD13+, CD13-CD15+, HLADR + CD15 , HLADR-CD14+CD11b+ CD4+); seven cases displayed antigen overexpression (CD13, CD33, CD15, or CD14); and in 13 patients leukemic cells had an abnormal FSC/SSC distribution according to their phenotype. These results suggest that immunological methods for the detection of MRD based on the existence of aberrant phenotypes could be used in the majority of AML patients.

Universal monitoring of minimal residual disease in acute myeloid leukemia

“…Those overexpressed genes included WT1 (in 84.7% of cases) (34,35), CD56 (46.5%) (36), CD7 (38.2%) (37,38), CD33 (36.9%) (39), CD4 (36.3%) (40), CD14 (30.6%) (39), and CD19 (28.0%) (40), while CD34 was underexpressed (36.3%) (41). Interestingly, genes previously reported to be leukemia stem cell specific had also emerged in our screening.…”

EuroFlow antibody panels for standardized n-dimensional flow cytometric immunophenotyping of normal, reactive and malignant leukocytes

“…Immunophenotypic similarities between the suspected cells and their potential normal counterparts allow the assignment of such cells to a given hematopoietic cell lineage and maturational stage, as well as the identification of aberrant phenotypes, such as leukemia-associated immunophenotypes. 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 Such immunophenotyping requires careful selection of unique combinations of individual markers based on their degree of specificity for the identification of a given cell lineage, maturation stage and aberrant phenotype, as well as the selection of appropriate antibody clones and fluorochrome conjugates to be used in multicolor combinations; the performance of these marker combinations is even more relevant than that of the individual markers. Consequently, such careful selection of reagents is essential for the design of standardized multicolor antibody combinations that provide unique staining patterns for each normal or aberrant cell population in a given sample.…”

“…Frequently occurring cross-lineage markers are CD19 in AML with t(8;21), CD7, and CD56. 4, 210, 211 Because cross-lineage expression can be weak, CD7–APC and CD56–PE were selected. NuTdT is expressed in many AML, but has limited added value for AML diagnosis.…”

The presence of leukaemia‐associated phenotypes is an independent predictor of induction failure in acute myeloid leukaemia

“…Immunophenotyping using multiparameter flow cytometry (MFC) campana & Coustan- Smith, 1999;San Miguel et al, 1999) enables the frequency and distribution of aberrant antigen expression on AML cells to be identified (Campana & Pui, 1995;San Miguel et al, 1986). Comparison with normal bone marrow cells has revealed five patterns of aberrant expressions: (i) asynchronous antigen expression (simultaneous expression of early and late markers in one cell such as the co-expression of CD34 and CD15 antigens), (ii) lineage infidelity (expression of the lymphoid-associated markers such as CD2, CD3, CD5, CD7, CD10 and CD19 on myeloid blast cells), (iii) antigen overexpression (abnormally increased expression of a certain antigen per cell), (iv) aberrant light-scatter properties (the expression of the lymphoid-associated antigens in blast cells displaying a relatively high forward scatter (FSC) and side scatter (SSC), corresponding to normal myeloid cells) and (v) absence of expected lineage specific antigens: absence of antigen expression such as CD13 and CD33 on myeloid blasts (Campana & Pui, 1995;Macedo et al, 1995b;San Miguel et al, 1997;Voskova et al, 2003). Early reports showed no correlation between leukaemic and normal cells of early myelopoiesis, and identified the presence of antigens not associated with normal myeloid differentiation (Andrews, Torok-Storb & Bernstein, 1983;Griffin et al, 1983).…”