Flow cytometric detection of minimal residual disease (MRD) in children with B-cell precursor acute lymphoblastic leukaemia (BCP-ALL) requires immunophenotypic discrimination between residual leukaemic cells and B-cell precursors (BCPs) which regenerate during therapy intervals. In this study, EuroFlow-based 8-colour flow cytometry and innovative analysis tools were used to first characterize the immunophenotypic maturation of normal BCPs in bone marrow (BM) from healthy children, resulting in a continuous multiparametric pathway including transition stages. This pathway was subsequently used as a reference to characterize the immunophenotypic maturation of regenerating BCPs in BM from children treated for BCP-ALL. We identified pre-B-I cells that expressed low or dim CD34 levels, in contrast to the classical CD34high pre-B-I cell immunophenotype. These CD34-dim pre-B-I cells were relatively abundant in regenerating BM (11-85% within pre-B-I subset), while hardly present in healthy control BM (9-13% within pre-B-I subset; P = 0·0037). Furthermore, we showed that some of the BCP-ALL diagnosis immunophenotypes (23%) overlapped with CD34-dim pre-B-I cells. Our results indicate that newly identified CD34-dim pre-B-I cells can be mistaken for residual BCP-ALL cells, potentially resulting in false-positive MRD outcomes. Therefore, regenerating BM, in which CD34-dim pre-B-I cells are relatively abundant, should be used as reference frame in flow cytometric MRD measurements.

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Keywords Acute leukaemia, B cells, Flow cytometry, Minimal residual disease
Persistent URL dx.doi.org/10.1111/bjh.14682, hdl.handle.net/1765/99306
Journal British Journal of Haematology
Theunissen, P.M.J, Sedek, L, de Haas, V, Szczepanski, T, van der Sluijs, A.J, Mejstříková, E, … van der Velden, V.H.J. (2017). Detailed immunophenotyping of B-cell precursors in regenerating bone marrow of acute lymphoblastic leukaemia patients. British Journal of Haematology, 178(2), 257–266. doi:10.1111/bjh.14682