Nanoparticle-mediated delivery of non-viral gene editing technology to the brain

Abstract

Neurological disorders pose a significant burden on individuals and society, affecting millions worldwide. These disorders, including but not limited to Alzheimer's disease, Parkinson's disease, and Huntington's disease, often have limited treatment options and can lead to progressive degeneration and disability. Gene editing technologies, including Zinc Finger Nucleases (ZFN), Transcription Activator-Like Effector Nucleases (TALEN), and Clustered Regularly Interspaced Short Palindromic Repeats-associated Protein 9 (CRISPR-Cas9), offer a promising avenue for potential cures by targeting and correcting the underlying genetic mutations responsible for neurologic disorders. However, efficient delivery methods are crucial for the successful application of gene editing technologies in the context of neurological disorders. The central nervous system presents unique challenges to treatment development due to the blood-brain barrier, which restricts the entry of large molecules. While viral vectors are traditionally used for gene delivery, nonviral delivery methods, such as nanoparticle-mediated delivery, offer safer alternatives that can efficiently transport gene editing components. Herein we aim to introduce the three main gene editing nucleases as nonviral treatments for neurologic disorders, the delivery barriers associated with brain targeting, and the current nonviral techniques used for brain-specific delivery. We highlight the challenges and opportunities for future research in this exciting and growing field that could lead to blood-brain barrier bypassing therapeutic gene editing.

Keywords: Blood-brain barrier; Gene editing technologies; Genome editing; Non-viral vectors.

Figure 1.

Types of Non-Viral Gene Editing Nucleases. A.) Zinc Finger Nucleases (ZFNs) use two FokI nucleases that must dimerize at the target sequence to induce a DSB. Each zinc finger is uniquely colored and is represented by binding to three DNA base pairs. B.) Transcription Activator-Like Effector Nucleases (TALENs) also use two FokI nucleases that must dimerize at the target sequences to induce a DSB. Each TALE is uniquely colored, and they are shown binding to individual nucleotides. C.) Cas9 protein in its ribonucleoprotein form is presented. The figure displays the Cas9 protein, single-guide RNA (sgRNA), and the PAM (Protospacer Adjacent Motif) sequence, highlighting the components involved in the CRISPR-Cas9 gene-editing system to induce a DSB. This figure is reproduced from source 10, which is published in Acta Pharmacoligica Sinica is licensed under CC BY-NC 4.0

Figure 2.

Payload options for Cas9 as a non-viral gene editing tool include dual delivery of Cas9 protein and sgRNA, Cas9 in a ribonucleoprotein (RNP) form, or Cas9 encoding plasmids. All payloads can be delivered independently to induce NHEJ, while the addition of donor DNA leads to HDR. Similar repair behavior will occur when delivering either TALENs or ZFNs with or without donor DNA. This figure is reproduced from source 17, which is reprinted with permission from ACS Macro Lett. 2021, 10, 12, 1576–1589. Copyright 2021 American Chemical Society.

Figure 3

Schematic of the blood-brain barrier showing the main cell types involved in the complexity. Figure made using BioRender.

Figure 4.

Representation of Primary Brain Regions Affected by Various Diseases. This figure highlights the importance of understanding brain heterogeneity when focusing on therapeutic development. Figure made in Biorender.

Figure 5.

Nanoparticle delivery of Cas9 involves many decisions, with a focus on the large parameter space available associated with materials selection. This figure is reproduced from source 17, which is reprinted with permission from ACS Macro Lett. 2021, 10, 12, 1576–1589. Copyright 2021 American Chemical Society.

Figure 6:

Clinical Trial Count for Gene Editing Technologies in Brain and Brain-Related Disorders—Zinc Finger Nucleases (ZFN), Transcription Activator-Like Effector Nucleases (TALEN), and Clustered Regularly Interspaced Short Palindromic Repeats-associated Protein 9 (Cas9). Data obtained on clinicaltrials.gov