PhenoGraph and viSNE facilitate the identification of abnormal T-cell populations in routine clinical flow cytometric data

Abstract

Background: Flow cytometric identification of neoplastic T-cell populations is complicated by the wide range of phenotypic abnormalities in T-cell neoplasia, and the diverse repertoire of reactive T-cell phenotypes. We evaluated whether a recently described clustering algorithm, PhenoGraph, and dimensionality-reduction algorithm, viSNE, might facilitate the identification of abnormal T-cell populations in routine clinical flow cytometric data. Methods: We applied PhenoGraph and viSNE to peripheral blood mononuclear cells labeled with a single 8-color T/NK-cell antibody combination. Individual peripheral blood samples containing either a T-cell neoplasm or reactive lymphocytosis were analyzed together with a cohort of 10 normal samples, which established the location and identity of normal mononuclear-cell subsets in viSNE displays. Results: PhenoGraph-derived subpopulations from the normal samples formed regions of phenotypic similarity in the viSNE display describing normal mononuclear-cell subsets, which correlated with those obtained by manual gating (r 2 = 0.99, P < 0.0001). In 24 of 24 cases of T-cell neoplasia with an aberrant phenotype, compared with 4 of 17 cases of reactive lymphocytosis (P = 1.4 × 10 −7 , Fisher Exact test), PhenoGraph-derived subpopulations originating exclusively from the abnormal sample formed one or more distinct phenotypic regions in the viSNE display, which represented the neoplastic T cells, and reactive T-cell subpopulations not present in the normal cohort, respectively. The numbers of neoplastic T cells identified using PhenoGraph/viSNE correlated with those obtained by manual gating (r 2 = 0.99; P < 0.0001). Conclusions: PhenoGraph and viSNE may facilitate the identification of abnormal T-cell populations in routine clinical flow cytometric data. © 2017 Clinical Cytometry Society.