将特定细胞因子有效荷载输送到肿瘤环境中，利用能够选择性聚集于肿瘤部位的抗体手段，是一种激发对癌症免疫反应的有效策略。尽管基于单一有效荷载的传统抗体-细胞因子融合蛋白已经显示出强大的抗癌活性，但同时输送两种细胞因子荷载可能会进一步提高治疗效果，因为免疫系统通常采用多个信号来强化抗肿瘤策略。我们在这里描述了一种功效匹配的双细胞因子抗体融合蛋白，其中包含靶向于人类成纤维母细胞激活蛋白（FAP）特异性的肿瘤靶向抗体片段，同时与白细胞介素-2（IL2）和肿瘤坏死因子（TNF）突变体连接。得到的融合蛋白被称为IL2-7NP2-TNFmut，由肿瘤坏死因子次单元的相互作用驱动形成稳定的非共价三聚体。如体外细胞实验所示，两种细胞因子荷载在融合蛋白中保留了其生物活性。在移植人类FAP基因的SKRC52细胞的免疫缺陷小鼠中，研究了IL2-7NP2-TNFmut的肿瘤靶向特性和抗癌活性。融合蛋白优先定位于癌症部位，并诱导了部分肿瘤缓解。

The delivery of specific cytokine payloads to a neoplastic environment employing antibodies able to selectively accumulate at the tumor site represents an attractive strategy to stimulate an immune response to cancer. Whilst conventional antibody-cytokine fusions based on a single payload have shown potent anticancer activity, the concomitant delivery of two cytokine payloads may further improve the therapeutic outcome as the immune system typically adopts multiple signals to reinforce an antitumor strategy. We here describe a potency-matched dual-cytokine antibody fusion protein containing a tumor-targeting antibody fragment specific to human fibroblast activation protein (FAP), simultaneously linked to both interleukin-2 (IL2) and a tumor necrosis factor (TNF) mutant. The resulting fusion protein, termed IL2-7NP2-TNFmut, formed stable non-covalent trimers driven by the interaction of the tumor necrosis factor subunits. Both cytokine payloads retained their biological activity within the fusion protein, as shown by in vitro cellular assays. The tumor-targeting properties and the anticancer activity of IL2-7NP2-TNFmut were investigated in vivo in immunocompromised mice bearing SKRC52 cells transduced with human FAP. The fusion protein preferentially localized to the cancer site and induced partial tumor retardation.