A synthetic BBB-permeable tripeptide GCF confers neuroprotection by increasing glycine in the ischemic brain

Juan Chen; Yang Zhuang; Ya Zhang; Huabao Liao; Rui Liu; Jing Cheng; Zhifeng Zhang; Jiangdong Sun; Jingchen Gao; Xiyuran Wang; Shujun Chen; Liang Zhang; Fengyuan Che; Qi Wan

doi:10.3389/fphar.2022.950376

A synthetic BBB-permeable tripeptide GCF confers neuroprotection by increasing glycine in the ischemic brain

Front Pharmacol. 2022 Aug 15:13:950376. doi: 10.3389/fphar.2022.950376. eCollection 2022.

Authors

Juan Chen^{1

2}, Yang Zhuang², Ya Zhang², Huabao Liao², Rui Liu², Jing Cheng², Zhifeng Zhang², Jiangdong Sun³, Jingchen Gao³, Xiyuran Wang³, Shujun Chen³, Liang Zhang⁴, Fengyuan Che⁵, Qi Wan^{3

6}

Affiliations

¹ Department of Neurology, The Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
² Department of Physiology, School of Medicine, Wuhan University, Wuhan, China.
³ Department of Pathophysiology, School of Basic Medicine, Institute of Neuroregeneration and Neurorehabilitation, Qingdao University, Qingdao, China.
⁴ Krembil Research Institute, University Health Network, University of Toronto, Toronto, ON, Canada.
⁵ Central Laboratory, Department of Neurology, Linyi People's Hospital, Qingdao University, Linyi, China.
⁶ Qingdao Gui-Hong Intelligent Medical Technology Co., Ltd., Qingdao, China.

Abstract

Background: We and others have previously demonstrated that glycine is neuroprotective in cerebral ischemia-reperfusion injury. But glycine has low permeability to the blood-brain barrier (BBB). To deliver glycine into the ischemic brain to confer neuroprotection, we designed a novel glycine-containing and BBB-permeable tripeptide, the H-glycine-cysteine-phenylalanine-OH (GCF). Methods: For the synthesis of GCF, phenylalanine was included to increase the BBB permeability of the tripeptide. Cysteine was conjugated with glycine to enable the release of glycine from GCF. With the use of immunofluorescence labeling and HPLC assays, we measured the distribution and level of GCF. We used TTC labeling, LDH release, and MTT assays to evaluate the neuroprotective effect of GCF. Results: Following intravenous injection in a rat model of cerebral ischemia-reperfusion injury, GCF was intensively distributed in the ischemic neurons. Intravenous injection of GCF, but not the non-cleavable acetyl-GCF, resulted in the elevation of glycine in the ischemic brain. GCF but not acetyl-GC conferred neuroprotection in ischemic stroke animals. Conclusion: GCF protects against cerebral ischemia-reperfusion injury in the rat. In contrast to peptide drugs that exert therapeutic effect by interfering with signaling interaction, GCF acts as a BBB shuttle and prodrug to deliver glycine to confer neuroprotection, representing a novel therapeutic strategy for acute ischemic stroke.

Keywords: BBB; glycine; ischemic stroke; neuroprotection; tripeptide.