Institute of Neuroimmunology, Slovak Academy of Sciences, Dubravska cesta 9, 845 10 Bratislava, Slovakia.
Department of Microbiology and Immunology, The University of Veterinary Medicine and Pharmacy, Komenskeho 73, 04181 Kosice, Slovakia.
Molecules. 2020 Feb 17;25(4):874. doi: 10.3390/molecules25040874.
Delivery of therapeutic agents into the brain is a major challenge in central nervous system drug development. The blood-brain barrier (BBB) prevents access of biotherapeutics to their targets in the central nervous system and, therefore, prohibits the effective treatment of many neurological disorders. To find blood-brain barrier shuttle peptides that could target therapeutics to the brain, we applied a phage display technology on a primary endothelial rat cellular model. Two identified peptides from a 12 mer phage library, GLHTSATNLYLH and VAARTGEIYVPW, were selected and their permeability was validated using the in vitro BBB model. The permeability of peptides through the BBB was measured by ultra-performance liquid chromatography-tandem mass spectrometry coupled to a triple-quadrupole mass spectrometer (UHPLC-MS/MS). We showed higher permeability for both peptides compared to N-C reversed-sequence peptides through in vitro BBB: for peptide GLHTSATNLYLH 3.3 × 10 cm/s and for peptide VAARTGEIYVPW 1.5 × 10 cm/s. The results indicate that the peptides identified by the in vitro phage display technology could serve as transporters for the administration of biopharmaceuticals into the brain. Our results also demonstrated the importance of proper BBB model for the discovery of shuttle peptides through phage display libraries.
将治疗剂递送到大脑是中枢神经系统药物开发中的一个主要挑战。血脑屏障 (BBB) 阻止生物治疗剂进入中枢神经系统的靶标，因此，禁止了许多神经疾病的有效治疗。为了寻找能够将治疗剂靶向大脑的血脑屏障穿梭肽，我们在原代内皮大鼠细胞模型上应用噬菌体展示技术。从 12 肽噬菌体文库中鉴定出两种肽，GLHTSATNLYLH 和 VAARTGEIYVPW，并使用体外 BBB 模型验证其通透性。通过与三重四极杆质谱联用的超高效液相色谱-串联质谱 (UHPLC-MS/MS) 测量肽通过 BBB 的通透性。与 N-C 反向序列肽相比，我们显示这两种肽通过体外 BBB 的通透性更高：肽 GLHTSATNLYLH 的通透性为 3.3 × 10 cm/s，肽 VAARTGEIYVPW 的通透性为 1.5 × 10 cm/s。结果表明，通过体外噬菌体展示技术鉴定的肽可作为将生物制药递送到大脑的载体。我们的结果还表明，对于通过噬菌体展示文库发现穿梭肽，适当的 BBB 模型非常重要。
