Engineered Exosomes with a Photoinducible Protein Delivery System Enable CRISPR-Cas–Based Epigenome Editing in Alzheimer’s Disease

Neurology Biochemistry Bioengineering Life Sciences Immunology Medicine Cell Biology
engineered exosomes photoinducible protein Alzheimer’s disease CRISPR-Cas epigenome editing delivery system memory improvement

CRISPR-Cas-Based Epigenome Editing in Alzheimer’s Disease: Engineered Exosomes with a Photoinducible Protein Delivery System

Background

Alzheimer’s Disease (AD) is a common neurodegenerative disorder characterized by progressive decline in cognitive function and memory, with currently limited treatment options. Genome editing technologies, such as CRISPR-Cas9, and epigenome editing techniques have shown great potential in treating various diseases. However, efficiently delivering these functional proteins into cells has been a significant challenge. Exosomes, naturally secreted nano-sized vesicles by cells, can serve as carriers for delivering therapeutic biomolecules, with advantages such as stability, high biocompatibility, and low immunogenicity.

Research Source

This article was published in Science Translational Medicine on August 7, 2024, titled “Engineered Exosomes with a Photoinducible Protein Delivery System Enable CRISPR-Cas–Based Epigenome Editing in Alzheimer’s Disease”. The research was conducted by Jihoon Han, Jae Hoon Sul, Jeongmi Lee, and others, affiliated with Sungkyunkwan University and several other research institutions.

Research Process

Engineering of Exosomes and Photoinducible Protein Release System

  1. Design of Engineered Exosomes:

    • The research team designed a system called mMaple3-mediated protein loading and release (MAPLEX). This system loads cargo proteins into exosomes by fusing the cargo protein with a photocleavable protein (mMaple3) and an exosome membrane marker (such as CD9).
    • mMaple3 can be cleaved under 405nm blue light, releasing the cargo protein.

  2. In Vitro Experiments:

    • In vitro, researchers first delivered transcription factors Octamer-binding transcription factor 4 (Oct4) and Sex determining region Y-box 2 (Sox2) to fibroblasts to observe their transcriptional regulation function.
    • Fluorescence imaging and immunoblotting results showed that the system could efficiently release proteins from exosomes, successfully enter recipient cells, and function in the nucleus.

  3. In Vivo Experiments in Mice:

    • In vivo, researchers delivered Cre recombinase to Cre reporter gene mice to verify its gene recombination function. Through intravenous injection and intranasal administration, they confirmed that the system could efficiently deliver Cre protein for gene recombination.

  4. Epigenome Editing:

    • Finally, the research team conducted experiments in Alzheimer’s disease mouse models (5xFAD and 3xTg-AD), embedding single guide RNA (sgRNA) targeting the β-site amyloid precursor protein cleaving enzyme 1 (BACE1) promoter into deactivated Cas9 (dCas9) nucleoprotein complexes, achieving epigenome editing through intranasal administration.
    • Results showed significantly increased DNA methylation levels at specific genomic sites, reduced BACE1 expression, improved cognitive memory deficits in mice, and reduced Aβ pathological features.

Research Results

  1. Targeted Protein Delivery:

    • The research team achieved precise protein release through blue light induction, with mMaple3 being cleaved under light exposure, releasing the cargo protein from exosomes.
    • Fluorescence imaging proved that proteins were successfully delivered to the cell nucleus and exerted their regulatory functions.

  2. Gene Recombination:

    • Cre recombinase was successfully delivered to the liver of Cre reporter gene mice, with fluorescence imaging showing tdTomato gene expression, proving successful gene recombination.
    • Intranasal administration also successfully delivered Cre to specific brain regions, indicating that the MAPLEX system can effectively cross the blood-brain barrier.

  3. Epigenome Editing:

    • The MAPLEX system successfully delivered dCas9-D3A complexes with sgRNA, increasing methylation levels of the BACE1 promoter.
    • Both intra-group and inter-group experiments showed reduced BACE1 expression, decreased Aβ pathological features in Alzheimer’s disease mouse models, and improved cognitive function.

Significance and Value of the Research

This study demonstrates for the first time the successful case of achieving epigenome editing in Alzheimer’s disease models using engineered exosomes and a photoinducible protein release system. This not only provides new ideas for the treatment of Alzheimer’s disease but also showcases the potential of the MAPLEX system in treating other diseases. The system enables efficient and precise protein delivery, which has important application value for future gene editing and epigenome editing.

Research Highlights

  1. Innovative Protein Delivery System:
    • Utilizing mMaple3 combined with photoinducible protein release solves the loading and release challenges in traditional protein delivery methods.
  2. Multifunctional Applications:
    • The system can not only deliver transcription factors but also achieve gene recombination and epigenome editing, providing possibilities for the treatment of various diseases in the future.
  3. Crossing the Blood-Brain Barrier:
    • Intranasal administration successfully delivered proteins to the brain, providing a new method for treating neurological diseases.

This paper demonstrates the broad application potential and important clinical translation prospects through an innovative photoinducible protein delivery system, achieving epigenome editing in Alzheimer’s disease mouse models.