Viral-based animal models in polyglutamine disorders.

Polyglutamine disorders are a complex group of incurable neurodegenerative disorders caused by an abnormal expansion in the trinucleotide cytosine-adenine-guanine tract of the affected gene. To better understand these disorders, our dependence on animal models persists, primarily relying on transgenic models. In an effort to complement and deepen our knowledge, researchers have also developed animal models of polyglutamine disorders employing viral vectors. Viral vectors have been extensively used to deliver genes to the brain, not only for therapeutic purposes but also for the development of animal models, given their remarkable flexibility. In a time- and cost-effective manner, it is possible to use different transgenes, at varying doses, in diverse targeted tissues, at different ages, and in different species, to recreate polyglutamine pathology. This paper aims to showcase the utility of viral vectors in disease modelling, share essential considerations for developing animal models with viral vectors, and provide a comprehensive review of existing viral-based animal models for polyglutamine disorders.

Blood and cerebellar abundance of ATXN3 splice variants in spinocerebellar ataxia type 3/Machado-Joseph disease.

Spinocerebellar ataxia type 3 (SCA3)/Machado-Joseph disease (MJD) is a heritable proteinopathy disorder, whose causative gene, ATXN3, undergoes alternative splicing. Ataxin-3 protein isoforms differ in their toxicity, suggesting that certain ATXN3 splice variants may be crucial in driving the selective toxicity in SCA3. Using RNA-seq datasets we identified and determined the abundance of annotated ATXN3 transcripts in blood (n = 60) and cerebellum (n = 12) of SCA3 subjects and controls. The reference transcript (ATXN3-251), translating into an ataxin-3 isoform harbouring three ubiquitin-interacting motifs (UIMs), showed the highest abundance in blood, while the most abundant transcript in the cerebellum (ATXN3-208) was of unclear function. Noteworthy, two of the four transcripts that encode full-length ataxin-3 isoforms but differ in the C-terminus were strongly related with tissue expression specificity: ATXN3-251 (3UIM) was expressed in blood 50-fold more than in the cerebellum, whereas ATXN3-214 (2UIM) was expressed in the cerebellum 20-fold more than in the blood. These findings shed light on ATXN3 alternative splicing, aiding in the comprehension of SCA3 pathogenesis and providing guidance in the design of future ATXN3 mRNA-lowering therapies.

Modeling Lysosomal Storage Disorders in an Innovative Way: Establishment and Characterization of Stem Cell Lines from Human Exfoliated Deciduous Teeth of Mucopolysaccharidosis Type II Patients.

Among the many lysosomal storage disorders (LSDs) that would benefit from the establishment of novel cell models, either patient-derived or genetically engineered, is mucopolysaccharidosis type II (MPS II). Here, we present our results on the establishment and characterization of two MPS II patient-derived stem cell line(s) from deciduous baby teeth. To the best of our knowledge, this is the first time a stem cell population has been isolated from LSD patient samples obtained from the dental pulp. Taking into account our results on the molecular and biochemical characterization of those cells and the fact that they exhibit visible and measurable disease phenotypes, we consider these cells may qualify as a valuable disease model, which may be useful for both pathophysiological assessments and in vitro screenings. Ultimately, we believe that patient-derived dental pulp stem cells (DPSCs), particularly those isolated from human exfoliated deciduous teeth (SHEDs), may represent a feasible alternative to induced pluripotent stem cells (iPSCs) in many labs with standard cell culture conditions and limited (human and economic) resources.

Molecular therapy for polyQ disorders: from bench to clinical trials.

Polyglutamine (polyQ) disorders are monogenic neurodegenerative disorders. Currently, no therapies are available for this complex group of disorders. Here, we aim to provide an overview of recent promising preclinical studies and the ongoing clinical trials focusing on molecular therapies for polyQ disorders.

Exploring the potential of cell-derived vesicles for transient delivery of gene editing payloads.

Gene editing technologies have the potential to correct genetic disorders by modifying, inserting, or deleting specific DNA sequences or genes, paving the way for a new class of genetic therapies. While gene editing tools continue to be improved to increase their precision and efficiency, the limited efficacy of in vivo delivery remains a major hurdle for clinical use. An ideal delivery vehicle should be able to target a sufficient number of diseased cells in a transient time window to maximize on-target editing and mitigate off-target events and immunogenicity. Here, we review major advances in novel delivery platforms based on cell-derived vesicles - extracellular vesicles and virus-like particles - for transient delivery of gene editing payloads. We discuss major findings regarding packaging, in vivo biodistribution, therapeutic efficacy, and safety concerns of cell-derived vesicles delivery of gene editing cargos and their potential for clinical translation.

RNA-based liposomes for oral cancer: From biophysical characterization to biological evaluation.

Oral cancer incidence and mortality are increasing over time. The most common therapies for oral cancers are surgery and radiotherapy, either used alone or combined, and immunotherapy can be also an option. Although there are several therapeutic options, none of them are completely effective, and in addition, there are numerous associated side effects. To overcome these limitations, researchers have been trying to reduce these drawbacks by using drug delivery systems that carry drugs for specific delivery to cancer cells. For that purpose, RNA-coated liposomes to selectively deliver the ligands C8 (acridine orange derivative) and dexamethasone to oral cancer cells were produced, characterized, and biologically evaluated. Firstly, the RNA structure and binding interaction with ligands (C8 and dexamethasone) were evaluated by circular dichroism (CD), thermal difference spectroscopy (TDS), nuclear magnetic resonance (NMR) and fluorescence titrations. The biophysical assays evidenced the formation of an RNA hairpin and duplex structure. Moreover, steady-state and time-resolved fluorescence intensity and anisotropy experiments show that C8 forms a complex with RNA and adopts an open conformation upon RNA binding. Then, RNA-coated liposomes were characterized by dynamic light scattering, and diameters near 160 nm were observed. Time-resolved anisotropy measurements of C8 loaded in RNA-functionalized liposomes indicate the co-existence of free C8 in solution (inside the liposome) and C8 bound to RNA at the external liposome surface. The RNA-functionalized liposomes loaded with C8 or dexamethasone mediated a significant reduction in the cell viability of malignant UPCI-SCC-154 cells while maintaining viable non-malignant NHDF cells. Additionally, the liposomes were able to internalize the cells, with higher uptake by the malignant cell line. Overall, the results obtained in this work can contribute to the development of new drug delivery systems based on RNA-coated liposomes.