Subset‐specific and tissue‐specific expression of NK ‐cell markers and killer immunoglobulin‐like receptors: Implications for NK ‐cell clonality assessment

Abstract Accurate immunophenotypic evaluation of natural killer (NK)‐cell neoplasms is challenging because NK‐cells lack somatically rearranged antigen receptors, are comprised of phenotypically distinct subsets, and exhibit tissue‐specific subset distribution. We developed a flow cytometry (FC) panel for NK‐cell immunophenotypic analysis and clonality assessment and applied it to 120 specimens, including normal peripheral blood (PB), bone marrow (BM), lymph node (LN), and body fluids, as well as a series of NK‐cell neoplasms and NK‐cell clones of uncertain significance (NK‐CUS). The expression ranges for NK‐cell receptors (NKR) including killer immunoglobulin‐like receptors (KIR, CD158) in the PB were consistent with those reported previously by other groups. The distribution of NK‐cell subsets differed by the specimen type. While PB and BM NK‐cells showed predominance of the CD56 dim subset, LN and fluid samples were enriched for the CD56 bright subset, resulting in higher CD94/NKG2A and lower KIR expression compared to PB. Epstein–Barr‐virus (EBV)‐positive NK/T‐cell neoplasms exhibited a relatively consistent CD56 bright CD16 − CD94/NKG2A + CD57 − KIR − immunophenotype that resembled normal CD56 bright NK‐cells, emphasizing the importance of immunophenotypic aberrancies and increased scatter properties rather than restricted KIR expression in diagnosing such neoplasms. NK‐large granular lymphocytic leukemia (NK‐LGLL) demonstrated a mature CD56 dim phenotype with restricted KIR expression. Twelve cases fulfilled the proposed criteria for NK‐CUS, comprising both putative cytomegalovirus (CMV)‐related NK‐cell clones and phenotypically distinct (likely) CMV‐unrelated clones with more pronounced immunophenotypic abnormalities. Our findings suggest that interpretation of NK‐cell clonality should take into account NK‐cell subset composition and tissue‐specific receptor expression. These results provide practical considerations for NK‐cell immunophenotypic analysis and contribute to a better understanding of the role of FC in diagnosing such neoplasms.