An international research team headed by scientists at New York University (NYU) Abu Dhabi has developed small proteins called cell-penetrating peptides (CPPs), which can prevent formation of the amyloid-β (Aβ) protein aggregates that are characteristic of Alzheimer’s disease (AD), and so inhibit Aβ-induced neurotoxicity.
The CPPs effectively target Aβ both outside and inside neurons, protecting them against the damage caused by Aβ aggregation. “The designed CPPs represent a novel potential treatment strategy for Alzheimer’s disease,” said NYU Abu Dhabi assistant professor of biology, Mazin Magzoub, PhD. “These findings also reveal a general underlying principle for inhibition of pathogenic protein aggregation that will facilitate the design of even more potent CPP-based therapeutics for various neurodegenerative diseases.”
The investigators reported on their developments and results in Cell Reports Physical Science, in a paper titled, “Designed Cell-Penetrating Peptide Inhibitors of Amyloid-beta Aggregation and Cytotoxicity.”
Alzheimer’s disease, Huntington’s disease, Parkinson’s disease, and prion diseases are among the broad range of incurable degenerative disorders that are associated with the misfolding of proteins or peptides into aggregate known as amyloids, the authors explained. Of these, Alzheimer’s disease is an irreversible and progressive brain disorder, and the most common cause of dementia. AD is characterized by the formation of neurofibrillary tangles, and aggregates, or plaques that contain misfolded Aβ peptides derived from the amyloid-β precursor protein (AβPP), in a process that is toxic to the brain’s neurons. The disorder causes progressive death of neurons, which destroys memory and cognitive function.
Proteins and peptides represent promising classes of therapeutics for many types of diseases because they are biocompatible, biodegradable, and can selectively bind to specific targets, which reduces the potential for toxicity, the researchers noted. Proteins also offer greater chemical diversity than other biological molecule classes, and they can be produced relatively easily and at relatively low cost. “Consequently, there has been a concerted effort to develop peptide-based amyloid inhibitors, which fall into two broad classes; rationally designed peptides, which include sequences derived from the target amyloid protein, and randomly generated peptides, which are often identified from library screens,” the investigators noted.
There are still some hurdles to be overcome, however. One significant obstacle to the successful application of most proteins as therapeutics is their poor delivery to target organs and cells. “This has necessitated the use of drug-delivery systems, such as CPPs, in order to overcome major physiological obstacles, e.g., the blood-brain and blood-cerebrospinal fluid barriers, and facilitate efficient delivery of the amyloid inhibitor peptides.”
The Magzoub lab researchers, together with collaborators in the lab of NYU president Andrew Hamilton, PhD, at NYU New York, and a team at the lab of Astrid Gräslund, PhD, at Stockholm University, reported on their development of CPPs that can be readily delivered to the brain, and which effectively prevent the aggregation of Aβ. CPPs are short peptides, typically 5–40 residues, the authors explained. “CPPs alone, or coupled to cargoes many times their own molecular mass, enter cells with high efficiency and low toxicity in vitro and in vivo and target specific intracellular organelles. Notably, CPPs also readily cross the blood-brain barrier.” The constructs effectively combine beneficial properties of proteins with potent therapeutic effects and highly efficient delivery to target cells.
The team had previously demonstrated the ability of a prion protein (PrP)-targeting CPP to protect against abnormal and harmful forms of the protein associated with prion diseases, a class of neurodegenerative disorders that includes mad cow disease in cattle and Creutzfeldt-Jakob disease (CJD) in humans.
The second inhibitor model comprised the NCAM1 peptide conjugated to a sequence derived from Aβ (Aβ16-20) that is analogous to the PrP sequence.
Using a range of techniques, they then extensively characterized the interactions of the designed CPPs with Aβ. Specifically, they used established aggregation and cell viability assays to determine the effects of the designed CPPs on the aggregation and associated neurotoxicity of Aβ. Simultaneously, the scientists used confocal fluorescence microscopy to probe the cellular uptake and intracellular distribution of Aβ in the presence of the CPPs. Combining experimental techniques with computer simulations shed further light on the mechanism of binding of the CPPs to Aβ. The results showed that the designed CPPs targeted both extracellular and intracellular Aβ protein, stabilizing it in a non-aggregated, non-abnormal state, and inhibited Aβ induced neurotoxicity.
The authors reported that the CPPs effectively inhibited Aβ oligomerization, fiber formation, and the associated neurotoxicity. “Based on these studies on two diverse amyloid systems, and given that the polycationic sequences target highly conserved molecular features of amyloids we propose that these constructs are general amyloid inhibitors, with potential applications in many amyloid-related diseases,” they concluded. “The CPP property of the constructs ensures their efficient delivery to all target tissues including the brain, and all target cells and subcellular organelles (e.g., mitochondria). At these locations, the CPPs will interact with the target amyloid protein/peptide and effectively inhibit its self-assembly and downstream toxic effects.”
