细胞因子诱导的记忆杀伤细胞（CIML NK）已被发现具有强大的抗肿瘤反应，并可在白血病患者中诱导完全缓解。然而，移植NK细胞的低浸润率是发展实体瘤细胞免疫治疗的主要障碍。在我们的研究中，我们探索了利用穿透肿瘤的肽iRGD，将激活的CIML NK细胞深入肿瘤组织中的潜力。经过短暂的白细胞介素-12（IL-12）、IL-15和IL-18刺激后，用流式细胞仪评估了CIML NK细胞的表型和功能。荧光共聚焦显微镜揭示了iRGD修饰的CIML NK细胞在肿瘤球体中的穿透和杀伤能力。这些修饰后的CIML NK细胞的抗肿瘤功效在肝细胞癌移植小鼠模型中得到了测试。细胞因子处理可导致NK细胞的显著活化，这一点在CD25和CD137的上调中得到了证明。经过6天的休息期后，CIML NK细胞仍能在针对HepG2和SK-Hep-1肝细胞癌细胞系时表现出强烈的活化。此外，CIML NK细胞产生的细胞因子（γ干扰素和肿瘤坏死因子α）增加，并对肝细胞癌细胞系显示出增强的细胞毒性。iRGD修饰能使CIML NK细胞渗透到多细胞球体中，并因此对目标癌细胞产生细胞毒作用。此外，iRGD修饰的CIML NK细胞显著降低了肝细胞癌移植小鼠模型的肿瘤生长。我们的发现表明，CIML NK细胞在体外具有增强的强度和持久性，对肝细胞癌细胞系具有增强的抗性。此外，我们发现将iRGD加入CIML NK细胞可以促进其渗透和定向摧毁多细胞球体。此外，应用iRGD修饰的CIML NK细胞在体内对肝细胞癌表现出显著的抗肿瘤效果。 © 2023作者。

Cytokine-induced memory-like natural killer (CIML NK) cells have been found to possess potent antitumor responses and induce complete remissions in patients with leukemia. However, the poor infiltration of transferred NK cells is a major obstacle in developing adoptive cell immunotherapy for solid tumors. In our study, we explored the potential of using the tumor-penetrating peptide iRGD to deliver activated CIML NK cells deep into tumor tissues.After being briefly stimulated with interleukin-12 (IL-12), IL-15, and IL-18, CIML NK cells were assessed for their phenotype and function with flow cytometry. The penetrating and killing capability of iRGD-modified CIML NK cells in tumor spheroids was revealed by confocal microscopy. The anti-tumor efficacy of these modified CIML NK cells was tested in hepatocellular carcinoma (HCC) xenograft mouse models.Treating NK cells with cytokines led to a substantial activation, which was evidenced by the upregulation of CD25 and CD137. After a resting period of six days, CIML NK cells were still able to display strong activation when targeting HepG2 and SK-Hep-1 HCC cell lines. Additionally, CIML NK cells produced increased amounts of cytokines (interferon-gamma and tumor necrosis factor alpha) and exhibited heightened cytotoxicity towards HCC cell lines. The iRGD modification enabled CIML NK cells to infiltrate multicellular spheroids (MCSs) and, consequently, to induce cytotoxicity against the target cancer cells. Moreover, the CIML NK cells modified with iRGD significantly decreased tumor growth in a HCC xenograft mouse model.Our findings demonstrate that CIML NK cells possess augmented potency and durability against HCC cell lines in vitro. Additionally, we have seen that the incorporation of iRGD to CIML NK cells facilitates enhanced infiltration and targeted destruction of MCSs. Moreover, the application of iRGD-modified CIML NK cells reveal remarkable anti-tumor efficacy against HCC in vivo.© 2023. The Author(s).