Novel protein variants and compositions for treating neurodegenerative diseases using the same

Protein variants that slow amyloid-beta fibrillation by substituting residues 19 and 32 with polar amino acids

This technology involves protein variants in which the 19th and 32nd amino acids of the amyloid-beta 42 sequence are replaced with the same type of polar uncharged amino acids—asparagine, glutamine, serine, or threonine—as well as the polynucleotides and vectors that encode them. Treating cells with these variants can control amyloid-beta fibrillation.

Amyloid-beta self-assembles into long, beta-sheet-structured fibrils, acting as a key pathology in Alzheimer's disease. While antibody therapies to remove fibrillar amyloid-beta have been introduced, they have not met expectations because monomers, intermediates, and fibrils coexist in vivo, and assembling molecules promote each other's fibrillation. Attempts to correct the Swedish mutation using CRISPR have also had limited application, as the disease can still occur with normal levels of amyloid-beta.

This technology uses point mutations designed based on protein structure and dynamics analysis to slow the rate of fibril formation. Depending on the need, residues 34 and 42 can also be substituted with the same amino acids to form variants of SEQ ID NOs 2–8. When mixed with wild-type amyloid-beta, these variants interact to delay fibrillation and reduce the cytotoxicity caused by fibrils in neuroblastoma cells. Applicable to protein-based Alzheimer's therapeutics, gene therapy vectors for inhibiting amyloid aggregation, and standard proteins for studying aggregation mechanisms, this approach intervenes in the fibril growth process itself to slow the aggregation of wild-type proteins, rather than simply chasing and removing existing fibrils.

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Key Features:

• A protein variant in which the 19th and 32nd amino acids from the N-terminus of SEQ ID NO: 1 are substituted with the same asparagine, glutamine, serine, or threonine
• A protein variant in which, in addition to the 19th and 32nd, one or more amino acids at positions 34 and 42 are also substituted with the same polar uncharged amino acids
• A polynucleotide encoding the protein variant and a vector containing it for introduction into cells
• A step of controlling amyloid-beta fibrillation by treating isolated neuroblastoma cells with the protein variant or vector

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This invention was developed with support from the Ministry of Science and ICT for research on kinetic control based on quantitative analysis of amyloid protein aggregation in complex systems.

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