Preclinical efficacy and safety of adeno-associated virus 5 alpha-galactosidase: A gene therapy for Fabry disease.

We developed a novel adeno-associated virus 5 gene therapy (AAV5-GLA) expressing human alpha-galactosidase A (GLA) under the control of a novel, small and strong, liver-restricted promoter. We assessed the preclinical potential of AAV5-GLA for treating Fabry disease, an X-linked hereditary metabolic disorder resulting from mutations in the gene encoding GLA that lead to accumulation of the substrates globotriaosylceramide and globotriaosylsphingosine, causing heart, kidney, and central nervous system dysfunction. Effects of intravenously administered AAV5-GLA were evaluated in (1) GLA-knockout mice aged 7-8 weeks (early in disease) and 20 weeks (nociception phenotype manifestation) and (2) cynomolgus macaques during an 8-week period.

Erratum: Proof-of-concept study for liver-directed miQURE technology in a dyslipidemic mouse model.

[This corrects the article DOI: 10.1016/j.omtn.

Proof-of-concept study for liver-directed miQURE technology in a dyslipidemic mouse model.

A gene-silencing platform (miQURE) has been developed and successfully used to deliver therapeutic microRNA (miRNA) to the brain, reducing levels of neurodegenerative disease-causing proteins/RNAs via RNA interference and improving the disease phenotype in animal models. This study evaluates the use of miQURE technology to deliver therapeutic miRNA for liver-specific indications. Angiopoietin-like 3 () was selected as the target mRNA because it is produced in the liver and because loss-of-function mutations and/or pharmacological inhibition of ANGPTL3 protein lowers lipid levels and reduces cardiovascular risk.

PhP.B Enhanced Adeno-Associated Virus Mediated-Expression Following Systemic Delivery or Direct Brain Administration.

Of the adeno-associated viruses (AAVs), AAV9 is known for its capability to cross the blood-brain barrier (BBB) and can, therefore, be used as a noninvasive method to target the central nervous system. Furthermore, the addition of the peptide PhP.B to AAV9 increases its transduction across the BBB by 40-fold.

Transduction patterns in the CNS following various routes of AAV-5-mediated gene delivery.

Various administration routes of adeno-associated virus (AAV)-based gene therapy have been examined to target the central nervous system to answer the question what the most optimal delivery route is for treatment of the brain with certain indications. In this study, we evaluated AAV5 vector system for its capability to target the central nervous system via intrastriatal, intrathalamic or intracerebroventricular delivery routes in rats. AAV5 is an ideal candidate for gene therapy because of its relatively low level of existing neutralizing antibodies compared to other serotypes, and its broad tissue and cell tropism.

In-Depth Characterization of a Mifepristone-Regulated Expression System for AAV5-Mediated Gene Therapy in the Liver.

Gene therapy is being developed for the treatment of inherited diseases, whereby a therapeutic gene is continuously expressed in patients after delivery via viral vectors such as adeno-associated virus (AAV). Depending on the transgene, there could be a limited therapeutic window, and regulating timing and levels of transgene expression is advantageous. To control transgene transcription, the regulatory system GeneSwitch (GS) was evaluated in detail both and .

Targeting RNA-Mediated Toxicity in C9orf72 ALS and/or FTD by RNAi-Based Gene Therapy.

A hexanucleotide GGGGCC expansion in intron 1 of chromosome 9 open reading frame 72 (C9orf72) gene is the most frequent cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). The corresponding repeat-containing sense and antisense transcripts cause a gain of toxicity through the accumulation of RNA foci in the nucleus and deposition of dipeptide-repeat (DPR) proteins in the cytoplasm of the affected cells. We have previously reported on the potential of engineered artificial anti-C9orf72-targeting miRNAs (miC) targeting C9orf72 to reduce the gain of toxicity caused by the repeat-containing transcripts.

Artificial MicroRNAs Targeting C9orf72 Can Reduce Accumulation of Intra-nuclear Transcripts in ALS and FTD Patients.

The most common pathogenic mutation in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) is an intronic GGGGCC (GC) repeat in the chromosome 9 open reading frame 72 (C9orf72) gene. Cellular toxicity due to RNA foci and dipeptide repeat (DPR) proteins produced by the sense and antisense repeat-containing transcripts is thought to underlie the pathogenesis of both diseases. RNA sequencing (RNA-seq) data of C9orf72-ALS patients and controls were analyzed to better understand the sequence conservation of C9orf72 in patients.

Therapeutic efficacy of regulable GDNF expression for Huntington's and Parkinson's disease by a high-induction, background-free "GeneSwitch" vector.

Gene therapy is currently an irreversible approach, without possibilities to fine-tune or halt the expression of a therapeutic gene product. Especially when expressing neurotrophic factors to treat neurodegenerative disorders, options to regulate transgene expression levels might be beneficial. We thus developed an advanced single-genome inducible AAV vector for expression of GDNF, under control of the approved small molecule drug mifepristone.

Viral genotype correlates with distinct liver gene transcription signatures in chronic hepatitis C virus infection.

Background: Chronic hepatitis C virus (HCV) infection of the liver with either genotype 1 or genotype 3 gives rise to distinct pathologies, and the two viral genotypes respond differently to antiviral therapy.

Methods: To understand these clinical differences, we compared gene transcription profiles in liver biopsies from patients infected with either gt1 or gt3, and uninfected controls.

Results: Gt1-infected biopsies displayed elevated levels of transcripts regulated by type I and type III interferons (IFN), including genes that predict response to IFN-α therapy.

Modulation of triglyceride and cholesterol ester synthesis impairs assembly of infectious hepatitis C virus.

In hepatitis C virus infection, replication of the viral genome and virion assembly are linked to cellular metabolic processes. In particular, lipid droplets, which store principally triacylglycerides (TAGs) and cholesterol esters (CEs), have been implicated in production of infectious virus. Here, we examine the effect on productive infection of triacsin C and YIC-C8-434, which inhibit synthesis of TAGs and CEs by targeting long-chain acyl-CoA synthetase and acyl-CoA:cholesterol acyltransferase, respectively.

The human collagen beta(1-O)galactosyltransferase, GLT25D1, is a soluble endoplasmic reticulum localized protein.

Background: Glycosyl transferases transfer glycosyl groups onto their substrate. Localization partially defines their function. Glycosyl transferase 25 domain 1 (GLT25D1) was recently shown to have galactosyltransferase activity towards collagens and another well known substrate, mannose binding lectin (MBL).

Characterization of hepatitis C virus NS3 modifications in the context of replication.

Post-translational modifications (PTMs) of viral proteins regulate various stages of infection. With only 10 proteins, hepatitis C virus (HCV) can orchestrate its complete viral life cycle. HCV non-structural protein 3 (NS3) has many functions.

Hepatitis C virus NS4B carboxy terminal domain is a membrane binding domain.

Background: Hepatitis C virus (HCV) induces membrane rearrangements during replication. All HCV proteins are associated to membranes, pointing out the importance of membranes for HCV. Non structural protein 4B (NS4B) has been reported to induce cellular membrane alterations like the membranous web.

Repair and polyadenylation of a naturally occurring hepatitis C virus 3' nontranslated region-shorter variant in selectable replicon cell lines.

The 3' nontranslated region (NTR) of the hepatitis C virus (HCV) genome is highly conserved and contains specific cis-acting RNA motifs that are essential in directing the viral replication machinery to initiate at the correct 3' end of the viral genome. Since the ends of viral genomes may be damaged by cellular RNases, preventing the initiation of viral RNA replication, stable RNA hairpin structures in the 3' NTR may also be essential in host defense against exoribonucleases. During 3'-terminal sequence analysis of serum samples of a patient with chronic hepatitis related to an HCV1b infection, a number of clones were obtained that were several nucleotides shorter at the extreme 3' end of the genome.

Exosomes contain ubiquitinated proteins.

Multivesicular bodies (MVB) are endosomal compartments that contain multiple vesicles, which derive from a delimiting membrane by inward budding. Incorporation of membrane proteins into the luminal vesicles requires, at least for some model proteins, monoubiquitination of their cytoplasmic domain. The ubiquitin tags are recognized by a sorting machinery, of which some components are also monoubiquitinated.