Flow cytometric immunophenotypic alterations of persistent clonal haematopoiesis in remission bone marrows of patients with NPM1‐mutated acute myeloid leukaemia

Abstract

Summary Clonal haematopoiesis (CH) in patients with acute myeloid leukaemia (AML) may persist beyond attaining complete remission. From a consecutive cohort of 67 patients with nucleophosmin 1‐mutated (NPM1mut) AML, we identified 50 who achieved NPM1mut clearance and had parallel multicolour flow cytometry (MFC) and next generation sequencing (NGS). In total, 13 (26%) cleared all mutations, 37 (74%) had persistent CH frequently involving DNA methyltransferase 3α (DNMT3A,70%), tet methylcytosine dioxygenase 2 (TET2, 27%), isocitrate dehydrogenase 2 (IDH2, 19%) and IDH1 (11%). A small number (<1%) of aberrant CD34+ myeloblasts, but immunophenotypically different from original AML blasts [herein referred to as a pre‐leukaemic (PL) phenotype], was detected in 17 (49%) patients with CH, but not in any patients with complete clearance of all mutations (P = 0·0037). A PL phenotype was associated with higher mutation burden (P = 0·005). Persistent IDH2 and serine and arginine‐rich splicing factor 2 (SRSF2) mutations were exclusively observed in PL+ CH+ cases (P = 0·016). Persistent dysplasia was seen exclusively in cases with a PL+ phenotype (29% vs. none; P = 0·04). The PL+ phenotype did not correlate with age, intensity of induction therapy or relapse‐free survival. Post‐remission CH in the setting of NPM1mut clearance is common and may result in immunophenotypic changes in myeloid progenitors. It is important to not misinterpret these cells as AML measurable residual disease (MRD).

2021 Update on MRD in acute myeloid leukemia: a consensus document from the European LeukemiaNet MRD Working Party

“…9 The panel advises the combined LAIP/DfN approach, but notes that some abnormal immunophenotypes may appear and/or disappear during monitoring, potentially due to transient expression on regenerating non-leukemic progenitors. [10][11][12] This phenomenon may affect the respective specificities of both LAIP and DfN MRD detection, in particular when the percentages of LAIPs at lower thresholds (e.g. <0.1%) are investigated.…”

Technical Aspects of Flow Cytometry-based Measurable Residual Disease Quantification in Acute Myeloid Leukemia: Experience of the European LeukemiaNet MRD Working Party

“… 39 , 40 However, there is some evidence that preleukemic clonal hematopoiesis may persist in AML patients who are in complete remission without any genetic evidence of MRD, and that the aberrant blast phenotypes may represent preleukemic clonal hematopoiesis leading to their misidentification as MRD. 41 , 42 Age-matched controls are therefore preferred to avoid possible age-related differences in marker expression to be identified as LAIPs. Furthermore, it is essential to know how the surface marker expression may be modified in the recovery phase after chemotherapy or transplantation.…”

Comparison of Multiple Clinical Testing Modalities for Assessment of NPM1-Mutant AML

“…FC has the potential advantages of quantitation and cell-sorting, but its main disadvantage is the challenge of phenotypic variability in AML due to disease heterogeneity ( 21 ), therapy-induced changes in protein expression, and phenotypic aberrations associated with background clonal hematopoiesis, which can confound the identification of residual disease, as we, and others ( 35 ) have observed. Thus, in the setting of post-therapy follow up for NPM1-mutated disease, priority can be given to RT-PCR, NGS and IHC testing.…”