Key Laboratory of Drug Targeting and Drug Delivery System, Ministry of Education, West China School of Pharmacy , Sichuan University , No. 17, Block 3, Southern Renmin Road , Chengdu 610041 , P.R. China.
ACS Appl Mater Interfaces. 2018 Oct 10;10(40):34039-34049. doi: 10.1021/acsami.8b13707. Epub 2018 Sep 26.
In oral delivery of biomacromolecules, ligand-modified nanoparticles (NPs) have emerged as a promising tool to improve the epithelial uptake of the loaded protein/peptide. Unfortunately, the stability and the transport mechanisms of the biotherapeutics during the intracellular transportation still remained unclear, leading to the poor transepithelial efficiency. Additionally, developing novel approaches to simultaneously monitor the payload bioactivity during the transport processes is veritably benefit for keeping their bioactivity. In the present study, EGP peptide (KRKKKGKGLGKKRDPCLRKYK), a ligand with high affinity to heparan sulfate proteoglycans (HSPGs), was found remarkably increasing the cellular uptake (4.5-fold) and also surprisingly achieving high transcytosis efficiency (4.2-fold) of poly(lactide- co-glycolide) NPs on Caco-2 cell monolayer. Compared with unmodified NPs (C NPs), EGP modified NPs (EGP NPs) exhibited more desirable colloidal stability within epithelia. In the subsequent study, the bioactivity of encapsulated insulin during the cellular transportation was innovatively monitored by a glucose consumption assay. Inspiringly, EGP NPs could mostly retain the bioactivity of loaded insulin whereas insulin from INS-C NPs was significantly degraded. Then the detailed mechanism study revealed that the binding of EGP to HSPGs played a vital role on NP transportation. Unlike C NPs being delivered in the endo/lysosomal pathway, EGP NPs were involved in caveolae-mediated transport, which contributes to the efficient avoidance of the lysosomal entrapment and sequentially facilitates the direct apical-to-basolateral transcytosis. The enhanced absorption of EGP NPs was confirmed in in situ intestinal loop models. Most importantly, oral administrated INS-EGP NPs generated a strong hypoglycemic response on diabetic rats with 10.2-fold and 2.6-fold increase in bioavailability compared with free insulin and INS-C NPs, respectively. The work provided an innovative strategy to monitor the payload bioactivity during the transport processes and proposed a novel aspect to increase oral bioavailability of biomacromolecules via improving payload stability and regulating the transcytosis pathway of nanocarriers.
在生物大分子的口服递送中，配体修饰的纳米颗粒（NPs）已成为提高负载蛋白/肽上皮摄取的有前途的工具。不幸的是，生物治疗剂在细胞内运输过程中的稳定性和转运机制仍不清楚，导致跨上皮效率低下。此外，开发同时监测转运过程中有效载荷生物活性的新方法对于保持其生物活性确实是有益的。在本研究中，发现具有高亲和力肝素硫酸蛋白多糖（HSPGs）的 EGP 肽（KRKKKGKGLGKKRDPCLRKYK）可显著增加细胞摄取（4.5 倍），并出人意料地实现了聚（乳酸-共- 乙交酯）NPs 在 Caco-2 细胞单层上的高转胞作用效率（4.2 倍）。与未修饰的 NPs（C NPs）相比，EGP 修饰的 NPs（EGP NPs）在细胞内具有更理想的胶体稳定性。在随后的研究中，通过葡萄糖消耗测定法创新地监测了包封胰岛素在细胞运输过程中的生物活性。令人鼓舞的是，EGP NPs 可以保留负载胰岛素的大部分生物活性，而 INS-C NPs 中的胰岛素则明显降解。然后，详细的机制研究表明，EGP 与 HSPGs 的结合对 NP 转运起着至关重要的作用。与 C NPs 被递送至内体/溶酶体途径不同，EGP NPs 参与了穴样内陷介导的运输，这有助于有效地避免溶酶体捕获，并随后促进直接顶侧至基底外侧的转胞作用。在原位肠袢模型中证实了 EGP NPs 的吸收增强。最重要的是，口服给予 INS-EGP NPs 可使糖尿病大鼠产生强烈的降血糖反应，与游离胰岛素和 INS-C NPs 相比，生物利用度分别提高了 10.2 倍和 2.6 倍。该工作提供了一种在转运过程中监测有效载荷生物活性的创新策略，并提出了一种通过提高有效载荷稳定性和调节纳米载体转胞作用途径来提高生物大分子口服生物利用度的新方法。
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