(M1230-06-36) A New Neurolysin Activator Ameliorates Brain Injury in Ischemic Stroke Mice

Purpose: Peptidase neurolysin (Nln) has been identified as a potential therapeutic target for stroke, a leading cause of death and disability globally. Previous studies have shown that increased expression of Nln reduces brain injury in ischemic stroke animals, while inhibition of Nln worsens the post-stroke outcomes. These findings spark interest in exploring the pharmacological activation of Nln in the central neural system (CNS) for post-stroke treatment. This study seeks new Nln activators with optimal brain disposition and therapeutic potential.

Methods: Compound KS73, a synthesized non-peptidomimetic activator of Nln, was evaluated for its activity, BBB permeability, pharmacokinetics, and therapeutic potential against ischemic stroke in a mouse transient middle cerebral artery occlusion (tMCAO) model.

Results: Compound KS73 showed approximately 100-fold higher proteinase activity and 9-fold greater BBB penetration than the endogenous Nln activator His-Tyr, indicated by the decreased minimal concentration required for enhancing 50% peptidase activity (A₅₀, 0.74 µM vs. 85.5 µM) and increased BBB permeability coefficient (Pe, 0.6 x10^-3 cm²/min vs. 0.07 x10^-3 cm²/min), respectively. After a single administration of 30 mg/kg via intraperitoneal (i.p.) injection, the plasma level of KS73 was 14787.67 ± 3839.65 to 26405.83 ± 793.63 ng/ml, and the brain level was 23042.83 ± 6315.47 to 42803.83 ± 1554.92 ng/ml within 1 hour, with peak levels at 30 minutes. At each time point, the KS73 level in the brain was higher than in the plasma (p < 0.05). The peak concentrations of brain and plasma were equivalent to 125.38 ± 4.55 µM and 77.35 ± 2.32 µM, respectively, significantly higher than its sub-molar A₅₀ (p < 0.0001). Furthermore, administration of KS73 (30 mg/kg, i.p. two doses) reduced brain infarction size and brain water content in the treatment group compared to the control group (p < 0.05), suggesting it may ameliorate brain injury after stroke onset.

Conclusion: This study presents a new Nln activator with CNS drug-like pharmacokinetic properties and therapeutic potential, supporting it as a promising lead for developing therapeutical agents for post-stroke treatment.

References: 1. Rashid M, Wangler NJ, Yang L, Shah K, Arumugam TV, Abbruscato TJ, Karamyan VT. Functional up-regulation of endopeptidase neurolysin during post-acute and early recovery phases of experimental stroke in mouse brain. J Neurochem. 2014;129(1):179-89.
2. Jayaraman S, Al Shoyaib A, Kocot J, Villalba H, Alamri FF, Rashid M, Wangler NJ, Chowdhury EA, German N, Arumugam TV, Abbruscato TJ, Karamyan VT. Peptidase neurolysin functions to preserve the brain after ischemic stroke in male mice. J Neurochem. 2020;153(1):120-137.
3. Rahman MS, Kumari S, Esfahani SH, Nozohouri S, Jayaraman S, Kinarivala N, Kocot J, Baez A, Farris D, Abbruscato TJ, Karamyan VT, Trippier PC. Discovery of First-in-Class Peptidomimetic Neurolysin Activators Possessing Enhanced Brain Penetration and Stability. J Med Chem. 2021;64(17):12705-12722.
4. Zhang Y, Sharma S, Jonnalagadda S, Kumari S, Queen A, Esfahani SH, Archie SR, Nozohouri S, Patel D, Trippier PC, Karamyan VT, Abbruscato TJ. Discovery of the Next Generation of Non-peptidomimetic Neurolysin Activators with High Blood-Brain Barrier Permeability: a Pharmacokinetics Study in Healthy and Stroke Animals. Pharm Res. 2023;40(11):2747-2758.

Acknowledgements: This work is supported by NIH grant R01NS106879.