APP C-Terminal CRISPR Truncation to Suppress Beta-Amyloid Production
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Solution Overview
Problem
Current gene editing technologies, such as CRISPR/Cas9, have limited efficacy and scope in treating neurodegenerative diseases like Alzheimer's disease, particularly in cases not inherited, and have not effectively targeted the amyloidogenic pathway to reduce amyloid plaque formation.
Innovation Solution
A method using CRISPR/Cas9 constructs to truncate the amyloid precursor protein (APP) at its C-terminus, specifically at residues 659, 670, or 676, delivered via adeno-associated viral vectors, to decrease the production of amyloid beta peptides, thereby reducing plaque formation.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If CRISPR/Cas9 gene editing is used to disrupt disease-causing genes, then the expression of disease-causing genes can be disrupted, but the editing efficiency is low and in vivo validation is limited
Solution Approach 1:
The patent segments the Cas9 nuclease into two separate polypeptide chains (N-terminal and C-terminal fragments) that can be delivered independently. This segmentation allows each fragment to be packaged in separate AAV vectors, overcoming the limitation of single-vector delivery and improving editing efficiency while reducing delivery complexity
Solution Approach 2:
The patent uses adeno-associated viral (AAV) vectors as intermediaries to deliver the Cas9 fragments and guide RNA to target cells. The AAV vectors serve as safe and effective delivery vehicles that can cross the blood-brain barrier, thereby improving in vivo delivery efficiency and reducing technical caveats associated with direct gene editing approaches
2Adaptability or versatility
If canonical gene editing approaches are used, then triplet-repeat expansions in Huntington's and Fragile X syndrome can be edited, but these approaches are only applicable to the small fraction of inherited cases
Solution Approach 1:
The patent develops a universal gene editing platform using segmented Cas9 delivered via AAV vectors that can target multiple disease mechanisms beyond inherited triplet-repeat expansions. The system can be applied to sporadic Alzheimer's disease by targeting APP C-terminal truncation, demonstrating versatility across different disease etiologies and significantly expanding treatment coverage from only inherited cases to include sporadic neurodegenerative diseases
3Reliability
If the amyloid precursor protein (APP) is truncated at the C-terminus using gene editing, then production of Aβ peptides is decreased, but delivery to the brain requires crossing the blood-brain barrier
Solution Approach 1:
The patent employs adeno-associated viral (AAV) vectors as intermediaries to deliver the segmented Cas9 nuclease and guide RNA across the blood-brain barrier to target cells in the brain. The AAV vectors are specifically engineered to cross this biological barrier, thereby simplifying the delivery process while maintaining reliable Aβ production reduction through APP C-terminal truncation
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
Robust APP editing is achieved in cell lines, human stem cells, and mouse brains, effectively reducing amyloid plaque formation without affecting physiological parameters, and selectively modulating APP/BACE-1 interaction to inhibit Aβ production.
Implementation Method 1
the Cas9 nuclease—guided by a short guide RNA (sgRNA)—generates double-stranded breaks (DSB) at targeted genomic loci
Implementation Method 2
the construct and the nuclease are delivered in a composition comprising an adeno-associated viral vector
Data Source
AI summary
Described herein are CRISPR/Cas9 constructs designed for the C-terminal truncation of human amyloid precursor protein (APP) as well as methods of making and using such a construct.