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<p>NK cells are innate lymphoid cells that can acquire a memory-like phenotype in vitro when stimulated with IL-12, IL-15, and IL-18. These cytokine-induced memory-like (CIML) NK cells exhibit prolonged lifespan and increased cytotoxicity, making them ideal for immunotherapy. This study characterizes two CIML NK cell subsets based on CD16 expression. NK cells were isolated from the peripheral blood of healthy donors and stimulated overnight to induce a memory-like phenotype. After seven days, we analyzed the phenotype and degranulation potential of CD16−/CD56 + and CD16+/CD56 + cells. The subsets were purified by fluorescence-activated cell sorting (FACS) and examined using high-throughput multiplexed quantitative proteomics. CD16 − cells showed higher levels of activating receptors, increased Granulysin expression, and lower inhibitory receptor expression compared to CD16 + cells. Functionally, CD16 − cells exhibited greater degranulation capacity, as determined by CD107a/b expression, when co-incubated with K562 and melanoma cells. Proteomic profiling identified 35 differentially expressed proteins out of 4,750, with 22 downregulated and 13 upregulated in the CD16 − subset. Key proteins included Granzyme family proteins, NCAM1, CALM1, CD247, and Fc receptors. This study provides a detailed characterization of CIML NK cells based on CD16 expression. Our findings highlight the molecular and functional diversity of CIML NK cells and may guide improved cancer immunotherapy strategies.</p>

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Phenotypic, proteomic, and functional analyses of cytokine-induced memory-like NK cells show two distinct subsets based on CD16 expression

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年10月23日

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  • Sofía Carreira-Santos,
  • Marina González-Sánchez,
  • Nelson López-Sejas,
  • Fakhri Hassouneh,
  • Lauro González-Fernández,
  • Inmaculada Jorge,
  • Esther Durán,
  • Alejandra Pera,
  • Jesús Vázquez,
  • Rafael Solana,
  • Raquel Tarazona,
  • Javier G. Casado

摘要

NK cells are innate lymphoid cells that can acquire a memory-like phenotype in vitro when stimulated with IL-12, IL-15, and IL-18. These cytokine-induced memory-like (CIML) NK cells exhibit prolonged lifespan and increased cytotoxicity, making them ideal for immunotherapy. This study characterizes two CIML NK cell subsets based on CD16 expression. NK cells were isolated from the peripheral blood of healthy donors and stimulated overnight to induce a memory-like phenotype. After seven days, we analyzed the phenotype and degranulation potential of CD16−/CD56 + and CD16+/CD56 + cells. The subsets were purified by fluorescence-activated cell sorting (FACS) and examined using high-throughput multiplexed quantitative proteomics. CD16 − cells showed higher levels of activating receptors, increased Granulysin expression, and lower inhibitory receptor expression compared to CD16 + cells. Functionally, CD16 − cells exhibited greater degranulation capacity, as determined by CD107a/b expression, when co-incubated with K562 and melanoma cells. Proteomic profiling identified 35 differentially expressed proteins out of 4,750, with 22 downregulated and 13 upregulated in the CD16 − subset. Key proteins included Granzyme family proteins, NCAM1, CALM1, CD247, and Fc receptors. This study provides a detailed characterization of CIML NK cells based on CD16 expression. Our findings highlight the molecular and functional diversity of CIML NK cells and may guide improved cancer immunotherapy strategies.