Immunostimulatory cytokines and immune checkpoint inhibitors hold promise as cancer therapeutics; however, their use is often limited by reduced efficacy and significant toxicity. In this study, we developed small-format immunocytokines (ICKs) based on interleukin-12 (IL-12) and blocking nanobodies (Nbs) targeting mouse and human programmed cell death 1 (PD-1) and programmed cell death ligand 1 (PD-L1). Both PD-1- and PD-L1-targeted ICKs demonstrated similar in vitro performance, significantly increasing IL-12 tethering to immune cells and enhancing T cell cytotoxic activity compared with IL-12 alone. The antitumor efficacy of ICKs was evaluated by intratumoral delivery using self-amplifying RNA-based vectors or as recombinant proteins in mice. Despite effective PD-L1-mediated tumor anchoring and promising in vitro results, IL-12 antitumor activity was significantly enhanced only when specific targeting to intratumoral T cells was achieved via anti-PD-1 Nb. This effect was also observed when the PD-1 specific ICK was delivered by electroporation of a DNA/RNA layered vector. Our findings suggest that targeting the appropriate type of cell within the tumor microenvironment could outperform tumor-anchoring strategies in the context of IL-12 therapy. Human versions of these ICKs were also developed, which showed to be active in human immune cells, opening an opportunity for clinical translation.

中文翻译：

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用小尺寸免疫细胞因子靶向 PD-1+ T 细胞可增强 IL-12 抗肿瘤活性


免疫刺激细胞因子和免疫检查点抑制剂有望成为癌症治疗药物;然而，它们的使用通常受到疗效降低和显著毒性的限制。在这项研究中，我们开发了基于白细胞介素 12 （IL-12） 和靶向小鼠和人类程序性细胞死亡 1 （PD-1） 和程序性细胞死亡配体 1 （PD-L1） 的阻断纳米抗体 （Nb） 的小尺寸免疫细胞因子 （INK）。PD-1 和 PD-L1 靶向 ICK 均表现出相似 的体外性能，与单独使用 IL-12 相比，显著增加 IL-12 与免疫细胞的结合并增强 T 细胞的细胞毒活性。通过使用基于自扩增 RNA 的载体或作为小鼠重组蛋白的瘤内递送来评估 ICKs 的抗肿瘤疗效。尽管 PD-L1 介导的肿瘤锚定有效且体外结果有希望 ，但只有当通过抗 PD-1 Nb 实现对瘤内 T 细胞的特异性靶向时，IL-12 抗肿瘤活性才会显着增强。当通过 DNA/RNA 分层载体的电穿孔递送 PD-1 特异性 ICK 时，也观察到了这种效果。我们的研究结果表明，在 IL-12 治疗的背景下，靶向肿瘤微环境中适当类型的细胞可能优于肿瘤锚定策略。还开发了这些 ICK 的人类版本，这些 ICK 在人类免疫细胞中具有活性，为临床转化提供了机会。