Human-derived NLS enhance the gene transfer efficiency of chitosan

Bitoque, Diogo B.; Morais, Joana; Oliveira, Ana V.; Sequeira, Raquel L.; Fortunato, Tiago Moderno; Simão, Sónia; Costa, Ana M. Rosa da; Silva, Gabriela A.

doi:10.1042/bsr20201026

Cited by 9 publications

(4 citation statements)

References 54 publications

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“…There have been very mixed results when NLS have been incorporated into polyplexes or plasmids. In many cases, NLS have very little or no positive effect on transfection that cannot be attributed to confounding factors (such as the nominal NLS stabilizing the nanoparticle compared to the unligated particle). − When NLS is conjugated to the DNA directly, there is sometimes an increase in transfection, and sometimes not . The principle investigator of the former report, J-P Behr, noted that the NLS only enhanced transfection with fast growing cell lines …”

Section: Nuclear Import and Expressionmentioning

confidence: 99%

Polycation-Mediated Transfection: Mechanisms of Internalization and Intracellular Trafficking

Monnery

2021

Biomacromolecules

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Polyplex-mediated gene transfection is now in its' fourth decade of serious research, but the promise of polyplex-mediated gene therapy has yet to fully materialize. Only approximately one in a million applied plasmids actually expresses. A large part of this is due to an incomplete understanding of the mechanism of polyplex transfection. There is an assumption that internalization must follow a canonical mechanism of receptor mediated endocytosis. Herein, we present arguments that untargeted (and most targeted) polyplexes do not utilize these routes. By incorporating knowledge of syndecan-polyplex interactions, we can show that syndecans are the "target" for polyplexes. Further, it is known that free polycations (which disrupt cell-membranes by acid-catalyzed hydrolysis of phospholipid esters) are necessary for (untargeted) endocytosis. This can be incorporated into the model to produce a novel mechanism of endocytosis, which fits the observed phenomenology. After membrane translocation, polyplex containing vesicles reach the endosome after diffusing through the actin mesh below the cell membrane. From there, they are acidified and trafficked toward the lysosome. Some polyplexes are capable of escaping the endosome and unpacking, while others are not. Herein, it is argued that for some polycations, as acidification proceeds the polyplexes excluding free polycations, which disrupt the endosomal membrane by acid-catalyzed hydrolysis, allowing the polyplex to escape. The polyplex's internal charge ratio is now insufficient for stability and it releases plasmids which diffuse to the nucleus. A small proportion of these plasmids diffuse through the nuclear pore complex (NPC), with aggregation being the major cause of loss. Those plasmids that have diffused through the NPC will also aggregate, and this appears to be the reason such a small proportion of nuclear plasmids express mRNA. Thus, the structural features which promote unpacking in the endosome and allow for endosomal escape can be determined, and better polycations can be designed.

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Section: Nuclear Import and Expressionmentioning

confidence: 99%

Polycation-Mediated Transfection: Mechanisms of Internalization and Intracellular Trafficking

Monnery

2021

Biomacromolecules

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“…117 It is now well established that incorporating NLS into non-viral vectors significantly increases gene transfer efficiency. [118][119][120][121] Another approach to facilitate nuclear entry is attachment of dexamethasone to non-viral vehicles or directly to pDNA. Dexamethasone interacts with a glucocorticoid receptor that changes the receptor's conformation and exposes its NLS, with subsequent nuclear localization of the vector.…”

Section: Nuclear Entry and Transgene Expressionmentioning

confidence: 99%

The menace of severe adverse events and deaths associated with viral gene therapy and its potential solution

Kachanov,

Kostyusheva,

Brezgin

et al. 2024

Medicinal Research Reviews

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Over the past decade, in vivo gene replacement therapy has significantly advanced, resulting in market approval of numerous therapeutics predominantly relying on adeno‐associated viral vectors (AAV). While viral vectors have undeniably addressed several critical healthcare challenges, their clinical application has unveiled a range of limitations and safety concerns. This review highlights the emerging challenges in the field of gene therapy. At first, we discuss both the role of biological barriers in viral gene therapy with a focus on AAVs, and review current landscape of in vivo human gene therapy. We delineate advantages and disadvantages of AAVs as gene delivery vehicles, mostly from the safety perspective (hepatotoxicity, cardiotoxicity, neurotoxicity, inflammatory responses etc.), and outline the mechanisms of adverse events in response to AAV. Contribution of every aspect of AAV vectors (genomic structure, capsid proteins) and host responses to injected AAV is considered and substantiated by basic, translational and clinical studies. The updated evaluation of recent AAV clinical trials and current medical experience clearly shows the risks of AAVs that sometimes overshadow the hopes for curing a hereditary disease. At last, a set of established and new molecular and nanotechnology tools and approaches are provided as potential solutions for mitigating or eliminating side effects. The increasing number of severe adverse reactions and, sadly deaths, demands decisive actions to resolve the issue of immune responses and extremely high doses of viral vectors used for gene therapy. In response to these challenges, various strategies are under development, including approaches aimed at augmenting characteristics of viral vectors and others focused on creating secure and efficacious non‐viral vectors. This comprehensive review offers an overarching perspective on the present state of gene therapy utilizing both viral and non‐viral vectors.

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“…In the case that the genetic cargo of a nanoparticle is DNA, the delivery voyage is not over in the cytosol, and the nucleic acid must next enter the nucleus (Shahryari et al, 2021). One commonly explored approach to facilitate this import utilizes nuclear localization sequences (NLS) to direct a plasmid to the nucleus (Bitoque et al, 2021; Maggio et al, 2020). NLSs are recognized by importins, which shuttle the NLS and associated cargo through the nuclear pore.…”

Section: Mechanism Of Nanoparticle‐mediated Intracellular Deliverymentioning

confidence: 99%

Nonviral nanoparticle gene delivery into the CNS for neurological disorders and brain cancer applications

Yang

Luly

Green

2022

WIREs Nanomed Nanobiotechnol

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Nonviral nanoparticles have emerged as an attractive alternative to viral vectors for gene therapy applications, utilizing a range of lipid‐based, polymeric, and inorganic materials. These materials can either encapsulate or be functionalized to bind nucleic acids and protect them from degradation. To effectively elicit changes to gene expression, the nanoparticle carrier needs to undergo a series of steps intracellularly, from interacting with the cellular membrane to facilitate cellular uptake to endosomal escape and nucleic acid release. Adjusting physiochemical properties of the nanoparticles, such as size, charge, and targeting ligands, can improve cellular uptake and ultimately gene delivery. Applications in the central nervous system (CNS; i.e., neurological diseases, brain cancers) face further extracellular barriers for a gene‐carrying nanoparticle to surpass, with the most significant being the blood–brain barrier (BBB). Approaches to overcome these extracellular challenges to deliver nanoparticles into the CNS include systemic, intracerebroventricular, intrathecal, and intranasal administration. This review describes and compares different biomaterials for nonviral nanoparticle‐mediated gene therapy to the CNS and explores challenges and recent preclinical and clinical developments in overcoming barriers to nanoparticle‐mediated delivery to the brain. This article is categorized under: Therapeutic Approaches and Drug Discovery > Nanomedicine for Neurological Disease Therapeutic Approaches and Drug Discovery > Emerging Technologies Nanotechnology Approaches to Biology > Nanoscale Systems in Biology

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Human-derived NLS enhance the gene transfer efficiency of chitosan

Cited by 9 publications

References 54 publications

Polycation-Mediated Transfection: Mechanisms of Internalization and Intracellular Trafficking

Polycation-Mediated Transfection: Mechanisms of Internalization and Intracellular Trafficking

The menace of severe adverse events and deaths associated with viral gene therapy and its potential solution

Nonviral nanoparticle gene delivery into the CNS for neurological disorders and brain cancer applications

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