Bi- und tri-functional IL-15 based cytokine fusion proteins for targeted cancer immunotherapy

调节免疫应答的细胞因子/配体-受体相互作用的抗肿瘤潜力已经成为癌症治疗策略的焦点。在这里，我们提出了一种多功能抗体融合蛋白的方法，用于靶向递送不同的细胞因子和配体到肿瘤部位，以支持有效的抗肿瘤免疫应答。靶向应降低全身毒性和自身免疫效应的风险。此外，不同免疫调节分子的组合使用应允许克服肿瘤逃避机制。在前面的项目中，我们已经成功地产生了双功能抗体融合蛋白，其中细胞因子IL-15连接到IL-15 R α链的一部分，在体外和体内获得了改善的活性。与共刺激配体4-1BBL的额外组合成三功能融合蛋白，在体外以靶向形式呈现增强的免疫刺激能力。鉴于这些令人鼓舞的结果，我们的目标是开发第二代三功能抗体融合蛋白，满足简单构型、最小尺寸和可溶性和靶向形式的所有化合物的良好可及性的标准。因此，我们将关注IL-15与4-螺旋束细胞因子家族（IL-7、IL-21）、TNF家族的共刺激配体（OX 40 L、GITRL、LIGHT）或趋化因子CCL家族（CCL 16、CCL 21）中的不同配体的组合。我们将分析这些分子的构型对以可溶性和靶向形式显示的活性的影响，以及它们对体外T细胞和NK细胞的免疫应答的影响。随后，将在肺转移肿瘤小鼠模型中体内评估最有前途的三功能融合蛋白的抗肿瘤潜力。因此，深入了解这种创新方法的实施和前景。

The antitumor potential of cytokine/ligand-receptor interactions that modulate an immune response has moved into the spotlight of cancer treatment strategies. Here, we propose an approach with multifunctional antibody-fusion proteins for target-directed delivery of different cytokines and ligands to the tumor site in order to support an efficient antitumor immune response. Targeting should reduce the risk of systemic toxicity and autoimmune effects. Furthermore, the combinatorial use of different immune-modulatory molecules should allow to overcome tumor evasion mechanisms. In the preceding project we have already successfully generated bi-functional antibody-fusion proteins with the cytokine IL-15 linked to part of the IL-15Ralpha chain, obtaining improved activity in vitro and in vivo. Additional combination with the costimulatory ligand 4-1BBL into a tri-functional fusion protein rendered enhanced immune-stimulatory capacity in targeted form in vitro. In light of these encouraging results, we aim to develop second generation tri-functional antibody-fusion proteins, meeting criteria of simple configuration, minimal size and good accessibility of all compounds in soluble and targeted form. Therefore we will focus on the combination of IL-15 with different ligands either of the 4-helix-bundle cytokine family (IL-7, IL-21), costimulatory ligands of the TNF-family (OX40L, GITRL, LIGHT) or the chemokine CCL-family (CCL16, CCL21). We will analyze the influence of the configuration of these molecules on the activity displayed in soluble and targeted form and their impact on the immune response of T cells and NK cells in vitro. Subsequently the antitumor potential of the most promising tri-functional fusion proteins will be evaluated in a lung metastasis tumor mouse model in vivo. Thus, insight into the implementation and prospect of this innovative approach is expected to be gained.