Presentation of an endogenous bioactive ligand in its native form is a key factor in controlling and determining its bioactivity, stability, and therapeutic efficacy. In this study, we developed a novel strategy for presenting trimeric ligands on nanocages by designing, optimizing and testing based on the rational design, high-resolution structural analysis and agonistic activity assays in vitro and in vivo. We successfully designed a nanocage that presents the TNF superfamily member, TRAIL (TNF-related apoptosis-inducing ligand) in its native-like trimeric structure. The native structure of TRAIL complexes was mimicked on the resulting TRAIL-presenting nanocages (TTPNs) by inserting sufficient spacing, determined from three-dimensional structural models, to provide optimal access to the corresponding receptors. The efficacy of TTPNs as an anti-tumor agent was confirmed in preclinical studies, which revealed up to 330-fold increased affinity, 62.5-fold enhanced apoptotic activity, and improved pharmacokinetic characteristics and stability compared with the monomeric form of TRAIL (mTRAIL). In this latter context, TTPNs exhibited greater than 90% stability over 1 mo, whereas ∼50% of mTRAIL aggregated within 2 d. Consistent with their enhanced stability and ultra-high affinity for the TRAIL receptor, TTPNs effectively induced apoptosis of tumor cells in vivo, leading to effective inhibition of tumor growth. Although TRAIL was used here as a proof-of-concept, all members of the TNF superfamily share the TNF homology domain (THD) and have similar distances between ecto-domain C-termini. Thus, other TNF superfamily ligands could be genetically substituted for the TRAIL ligand on the surface of this biomimetic delivery platform.

呈天然形式的内源性生物活性配体的呈递是控制和确定其生物活性，稳定性和治疗功效的关键因素。在这项研究中，我们开发了一种新颖的策略，可通过在合理设计，高分辨率结构分析和体内和体外激动活性分析的基础上进行设计，优化和测试，在纳米笼上展示三聚体配体。我们成功设计了一个纳米笼，该笼以其天然的三聚体结构展示了TNF超家族成员TRAIL（TNF相关的凋亡诱导配体）。通过插入足够的间距（由三维结构模型确定），可以在生成的TRAIL提呈纳米笼（TTPN）上模拟TRAIL配合物的天然结构，从而提供对相应受体的最佳通道。临床前研究证实了TTPNs作为抗肿瘤药的功效，与TRAIL的单体形式（mTRAIL）相比，它揭示了高达330倍的亲和力提高，62.5倍的凋亡活性提高，药代动力学特性和稳定性的提高。在后一种情况下，TTPN在1个月内表现出大于90％的稳定性，而约50％的mTRAIL在2天内聚集。在体内，导致有效抑制肿瘤生长。尽管在这里使用TRAIL作为概念验证，但TNF超家族的所有成员都共享TNF同源域（THD），并且在外部域C末端之间的距离相似。因此，其他TNF超家族配体可以被该仿生递送平台表面上的TRAIL配体遗传取代。