Novel Cdk5i Peptide Shows Promise Against Alzheimer’s Neurodegeneration

A promising peptide capable of reversing Alzheimer’s Disease (AD) symptoms and neurodegeneration has been developed recently.^[2] Cdk5i peptide is a 12-amino acids chain derived from a protein called cyclin-dependent kinase 5 (Cdk5) (see fig. 1.B).^[2] The way Cdk5 is triggered is through its activators p35 and p39, forming two types of complexes (Cdk5/p35 and Cdk5/p39).^[2,3,8] Cdk5 speeds up chemical reactions in the brain (enzymatic activity).^[2] Also, it plays a crucial role in some of the most critical processes, like axonal direction, synaptic plasticity, memory, learning, synaptic growth, neuronal maturation, vesicular transportation, neuronal differentiation, and cell migration, among others.^[2] Cdk5 has a direct relationship with AD pathology because amyloid beta peptides induce the activity of this kinase.^[1-3] Several studies show that an imbalance of Cdk5 activity leads to neurodegenerative disorders like Alzheimer’s (AD), Frontotemporal dementia, and Parkinson’s Disease.^[2] Neurodegenerative stimuli cause drastic changes in calcium levels inside the cells, activating a protease (calpain) that cuts a regulatory fragment in p35, converting p35 into a smaller activator called p25.^[2] In addition, calpain cuts a piece of the Cdk5/p35 complex that binds it with the cell membrane, liberating the complex to the cytosol and other nuclear compartments.^[2] Once the conversion is done, the molecule p25 activates Cdk5 forming the Cdk5/p25 complex (see fig.1.A).^[2] The Cdk5/p25 complex has a longer half-life than the others, thus accumulating Cdk5 over time, affecting the well-functioning of brain cells.^[2,3]

(A) Cdk5/p25 complex protein structure with the T-loop region (in red). (B) The comparison of the T-loop sequences in Cdk5 of humans with the other three organisms. (C) Human Cdks (Cdk5, Cdk1, Cdk2, Cdk3) sequences.^[2]

Figure 1.

The abnormal aggregation of the Tau protein is well known to be one of the hallmarks of AD pathology. ^[1,3] In recent studies, researchers manipulated the p35 genetically, creating the mutation called Δp35 that prevents p35 from being converted into p25 to evaluate the effect of decreasing the formation of p25 (Cdk5/p25 complex) in neuronal cells.^[2] This study demonstrated reduced AD pathology progression and a decline in neurodegeneration in vivo mice models.^[2] Avoiding the p25 formation also suggests a defense against the Tau protein overexpression in the mice and humans used in this study.^[2] These findings open a new door for developing therapies for AD and other neurodegenerative diseases.^[2,4,5] Two Cdk5 inhibitors were created in 2002, peptides P5 and CIP, P5 being the most effective with 24 amino acids and a TAT modification, a cell-penetrating sequence commonly used to deliver small molecules inside the brain and cell membranes.^[2,4,5] P5 proves to decrease the activity of Cdk5 in brain cells and decrease neurodegeneration.^[2,4]

Nevertheless, the size of the peptide is crucial according to research in terms of transportation and specificity. ^[2,6] This characteristic inspires the creation of a novel short peptide called Cdk5i (12-amino acids), capable of inhibiting the interaction of Cdk5 with p25 and decreasing the enzymatic activity of the Cdk5/p25 complex significantly.^[2] To test the effectiveness of Cdk5i in mice, researchers administered Cdk5i via intraperitoneal injection (body cavity).^[2] Cdk5i were also tested in mammalian cell cultures, where they were exhibited to reverse the neurodegeneration, decreasing the accumulation of Cdk5 by blocking the Cdk5/p25 interaction.^[2] The sequence used for creating the Cdk5i peptide was determined after the analysis of the T-loop zone, where Cdk5 and p25 interact for binding (see fig. 1.A).^[2] Cdk5i exhibited more affinity to bind cdk5/p25 complexes than cdk5 alone.^[2] Therefore, this peptide is particular in targeting the Cdk5/p25 complex principally, which is one of the leading causes of the advance of AD.^[1,3]

The TAT sequence delivers small molecules inside the brain, while the FITC is for attaching fluorescent labels to proteins (see Appendix fig. 2).^2,6,7] Several studies demonstrated promising results in decreasing the Cdk5/p25 complex interaction by 22%.^[2] In fact, by modifying the Cdk5i peptide with TAT and a FITC having; as a result, Cdk5i-TF, investigators found a special delivery of the peptide inside the brain, and no effect on neuron viability was recorded.^[2] Different experiments testing the Cdk5i peptide with these new modifications (Cdk5i-TF) show a significant improvement by better decreasing the enzymatic activity of Cdk5 and reducing induced mitochondrial dysfunction.^[2] Cdk5i-TF also decreased the upregulation of Cdk5 by 35% (see Appendix fig.3).^[2] Dropping the upregulation of Cdk5 is very significant since neurofibrillary tangles form because of an excessive upregulation of Cdk5 in neuron cells.^[1] Cdk5i-TF is a great candidate for AD therapy, decreasing one of the principles responsible for the progression of the disease, which is Cdk5.^[2] This peptide improves cognition, increases cell survival, promotes neuroprotection, reduces neurodegeneration, and decreases neuroinflammation and other symptoms of AD and neurodegenerative disorders.^[2] In addition, adding the TAT modification to the original peptide (Cdk5i) improves the peptide capacity to cross the BBB and cell membranes.^[2,6] Cdk5i combines the best characteristics needed for an AD treatment because of its short chain, high specificity, and affinity to the Cdk5/p25 complex, decreasing neurodegeneration and reversing the progression of the disease.^[2,5]

Benefits of using Cdk5i peptide (↑increase, ↓decrease)

• ↑ Neuroprotection
• ↓ Cdk5/p25 complex interaction
• ↓ Tau aggregation (neurofibrillary tangles)
• ↓ Neurodegeneration
• ↓ AD progression
• ↓ Mitochondrial dysfunction
• ↓ Cdk5 abnormal upregulation
• ↑ Cognition
• ↑ Cell survival
• ↑ Learning and memory

References

1. Lee, M. S., & Tsai, L. H. (2003). Cdk5: one of the links between senile plaques and neurofibrillary tangles?.Journal of Alzheimer’s Disease, 5(2), 127-137.
2. Pao, P. C., Seo, J., Lee, A., Kritskiy, O., Patnaik, D., Penney, J., … & Tsai, L. H. (2023). A Cdk5-derived peptide inhibits Cdk5/p25 activity and improves neurodegenerative phenotypes. Proceedings of the National Academy of Sciences, 120(16), e2217864120.
3. Lu, T. T., Wan, C., Yang, W., & Cai, Z. (2019). Role of Cdk5 in amyloid-beta pathology of Alzheimer’s disease. Current Alzheimer Research, 16(13), 1206-1215.
4. Zhang, J., Zhang, Y., Xu, M., Miao, Z., & Tian, Y. (2021). Inhibition of the CDK5/caspase-3 pathway by p5-TAT protects hippocampal neurogenesis and alleviates radiation-induced cognitive dysfunction. Neuroscience, 463, 204-215.
5. Huang, Y., Huang, W., Huang, Y., Song, P., Zhang, M., Zhang, H. T., … & Hu, Y. (2020). Cdk5 inhibitory peptide prevents loss of neurons and alleviates behavioral changes in p25 transgenic mice. Journal of Alzheimer’s Disease, 74(4), 1231-1242.
6. Frain, K. M., Robinson, C., & van Dijl, J. M. (2019). Transport of folded proteins by the Tat system. The Protein Journal, 38, 377-388.
7. Gonzalez-Carter, D., Liu, X., Tockary, T. A., Dirisala, A., Toh, K., Anraku, Y., & Kataoka, K. (2020). Targeting nanoparticles to the brain by exploiting the blood–brain barrier impermeability to selectively label the brain endothelium. Proceedings of the National Academy of Sciences, 117(32), 19141-19150.
8. Zheng, C., & Tang, Y. D. (2022). The emerging roles of the CDK/cyclin complexes in antiviral innate immunity. Journal of Medical Virology, 94(6), 2384-2387.

Appendix

Figure 2. FITC signals (fluorescent marker) in nuclear compartments and the cytosolic, indicating a successful peptide delivery in the brain.

Figure 3. Cdk5i-FT peptide reduced Cdk5 upregulation by 35%