Sezary syndrome (SS) is a rare leukemic form of cutaneous T-cell lymphoma. Diagnosis mainly depends on flow cytometry, but results are not specific enough to be unequivocal. The difficulty in defining a single marker that could characterize Sezary cells may be the consequence of different pathological subtypes. In this study, we used multivariate flow cytometry analyses. We chose to investigate the expression of classical CD3, CD4, CD7, and CD26 and the 2 new association of 2 markers CD158k and PD-1. We performed lymphocyte computational phenotypic analyses during diagnosis and follow-up of SS patients to define new SS classes and improve the sensitivity of the diagnosis and the follow-up flow cytometry method. Three classes of SS, defined by different immunophenotypic profiles, CD158k-positive SS, CD158k-negative PD-1-positive SS, CD158k and PD-1 double-negative SS, showed different CD8+ and B-cells environments. Such a study could help to diagnose and define biological markers of susceptibility/resistance to treatment including immunotherapy. -

Purpose: Acute myeloid leukemias (AML) are clonal diseases that develop from leukemic stem cells (LSCs) that carry an independent prognostic impact on the initial response to induction chemotherapy, demonstrating the clinical relevance of LSC abundance in AML. In 2018, the European LeukemiaNet has published recommendations for the detection of measurable residual disease (Bulk MRD) and suggested the exploration of LSC MRD and the use of multiparametric displays. Experimental design: We evaluated the performance of unsupervised clustering for the post-induction assessment of bulk and LSC MRD in 155 AML patients who received intensive conventional chemotherapy treatment. Results: The median overall survival (OS) for Bulk+ MRD patients was 16.7 months and was not reached for negative patients (Hazard Ratio, HR:3.82, p<0.0001). The median OS of LSC+ MRD patients was 25.0 months and not reached for negative patients (HR:2.84, p=0.001). Interestingly, one-year (y) and 3-y OS were 60% and 39% in Bulk+, 91% and 52% in Bulk-LSC+ and 92% and 88% in Bulk-LSC-. Conclusion: In this study, we confirm the prognostic impact of post-induction MFC Bulk MRD in AML patients. Focusing on LSCs, we identified a group of patients with negative Bulk MRD but positive LSC MRD (25.8% of our cohort) with an intermediate prognosis, demonstrating the interest of MRD analysis focusing on leukemic chemoresistant subpopulations.

Classifications of acute myeloid leukemia (AML) patients rely on morphologic, cytogenetic, and molecular features. Here we have established a novel flow cytometry-based immunophenotypic stratification showing that AML blasts are blocked at specific stages of differentiation where features of normal myelopoiesis are preserved. Six stages of leukemia differentiation-arrest categories based on CD34, CD117, CD13, CD33, MPO, and HLA-DR expression were identified in two independent cohorts of 2087 and 1209 AML patients. Hematopoietic stem cell/multipotent progenitor-like AMLs display low proliferation rate, inv(3) or RUNX1 mutations, and high leukemic stem cell frequency as well as poor outcome, whereas granulocyte–monocyte progenitor-like AMLs have CEBPA mutations, RUNX1-RUNX1T1 or CBFB-MYH11 translocations, lower leukemic stem cell frequency, higher chemosensitivity, and better outcome. NPM1 mutations correlate with most mature stages of leukemia arrest together with TET2 or IDH mutations in granulocyte progenitors-like AML or with DNMT3A mutations in monocyte progenitors-like AML. Overall, we demonstrate that AML is arrested at specific stages of myeloid differentiation (SLA classification) that significantly correlate with AML genetic lesions, clinical presentation, stem cell properties, chemosensitivity, response to therapy, and outcome.

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