Abstract:
:The production of high-titer recombinant adeno-associated virus (rAAV) vector is essential for treatment of genetic diseases affecting the retina and choroid, where anatomical constraints may limit injectable volumes. Problematically, cytotoxicity arising from overexpression of the transgene during vector production frequently leads to a reduction in vector yield. Herein, we evaluate the use of microRNA (miRNA)-mediated silencing to limit overexpression of cytotoxic transgenes during packaging as a method of increasing vector yield. We examined if post-transcriptional regulation of transgenes during packaging via miRNA technology would lead to increased rAAV yields. Our results demonstrate that silencing of cytotoxic transgenes during production resulted in up to a 22-fold increase in vector yield. The inclusion of organ-specific miRNA sequences improved biosafety by limiting off-target expression following systemic rAAV administration. The small size (22-23 bp) of the target site allows for the inclusion of multiple copies into the vector with minimal impact on coding capacity. Taken together, our results suggest that inclusion of miRNA target sites into the 3'-untranslated region of the AAV cassette allow for silencing of cytotoxic transgenes during vector production leading to improved vector yield, in addition to increasing targeting specificity without reliance on cell-specific promoters.
journal_name
Gene Therjournal_title
Gene therapyauthors
Reid CA,Boye SL,Hauswirth WW,Lipinski DMdoi
10.1038/gt.2017.50subject
Has Abstractpub_date
2017-08-01 00:00:00pages
462-469issue
8eissn
0969-7128issn
1476-5462pii
gt201750journal_volume
24pub_type
杂志文章相关文献
GENE THERAPY文献大全abstract::Sindbis virus expression has been used for in vitro investigations of antigen processing, presentation and epitope mapping. The recent development of a replication-deficient recombinant Sindbis virus expression vector has made in vivo expression possible with minimal pathogenic risk. Advantages of Sindbis virus over o...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3300458
更新日期:1997-08-01 00:00:00
abstract::During the past decade, both in vitro and in vivo studies have provided new insights into the cellular and molecular mechanisms that govern angiogenesis and arteriogenesis. However, therapeutic angiogenesis clinical trials using recombinant protein or gene therapy formulations of single angiogenic growth factors have ...
journal_title:Gene therapy
pub_type: 杂志文章,评审
doi:10.1038/sj.gt.3302953
更新日期:2007-05-01 00:00:00
abstract::Bone marrow presents an attractive option for the treatment of articular cartilage defects as it is readily accessible, it contains mesenchymal progenitor cells that can undergo chondrogenic differentiation and, once coagulated, it provides a natural scaffold that contains the cells within the defect. This study was p...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/gt.2010.16
更新日期:2010-06-01 00:00:00
abstract::Short hairpin RNA (shRNA)-mediated gene regulation is a commonly used technique for gene manipulation. An efficient and safe delivery system is indispensable when shRNA is delivered into living organisms for gene therapy. Previous studies have proved that DNA and protein can be delivered into dendritic cells (DCs) by ...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/gt.2014.50
更新日期:2014-07-01 00:00:00
abstract::A retroviral vector constructed from the murine leukemia virus (MLV) can only express transgenes in cells undergoing mitosis, indicating its suitability as a delivery vehicle for cancer gene therapy. However, the transduction efficiency (TE) of retroviruses embedding endogenous envelope proteins in human cancer cells ...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3301390
更新日期:2001-02-01 00:00:00
abstract::Dendritic cell (DC)-based vaccine approaches are being actively evaluated for developing immunotherapeutic agents against cancers. In this study, we investigated the use of engineered DCs expressing transgenic tumor-associated antigen hgp100 and the regulatory cytokine interleukin-21, namely DC-hgp100/mIL-21, as a the...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/gt.2014.12
更新日期:2014-05-01 00:00:00
abstract::Silencing of gene expression by small interfering RNAs (siRNAs) is rapidly becoming a powerful tool for genetic analysis of mammalian cells. The use of DNA-based plasmid vectors to achieve transient and stable expression of siRNA has been developed to avoid the problems of double-stranded oligonucleotides transfection...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3302878
更新日期:2007-03-01 00:00:00
abstract::Interferon-alpha (IFN-alpha) or CD80 transduction of tumor cells individually reduces tumorigenicity and enhances antitumor responses. Here, we report that the combination of IFN-alpha and CD80 cancer gene therapy in poorly immunogenic murine tumor models, the colorectal adenocarcinoma cell line MC38, and the methylch...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3301034
更新日期:1999-12-01 00:00:00
abstract::Therapeutic RNA interference (RNAi) has emerged as a promising approach for the treatment of many incurable diseases, including cancer, infectious disease or neurodegenerative disorders. Demonstration of efficacy and safety in animal models is necessary before planning human application. Our group and others have prev...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/gt.2011.10
更新日期:2011-07-01 00:00:00
abstract::Late pregnant rats exhibit endogenous opioid restraint of oxytocin cells since i.v. naloxone (NLX opioid antagonist) increases oxytocin (OXT) secretion but OXT nerve terminals become desensitised to opioids. We have studied central opioid inhibition of OXT neurones in late pregnancy by measuring SON OXT neurones firin...
journal_title:Gene therapy
pub_type: 杂志文章
doi:
更新日期:1994-01-01 00:00:00
abstract::Gene transfer complexes containing poly-L-lysine (poly-K) and DNA with ligands directed at the serpin enzyme complex receptor (sec-R) deliver reporter genes to receptor-bearing cells in vivo. Expression lasts for about 30 days, when complexes containing long-chain poly-K are used. Extending the duration of expression ...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3302299
更新日期:2004-09-01 00:00:00
abstract::To facilitate the understanding of the complex process of target gene expression and its control, we report a modified inducible system for activation or repression of target gene expression in response to an exogenously administered compound. The main component of this inducible system is a chimeric transcriptional a...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3300402
更新日期:1997-05-01 00:00:00
abstract:UNLABELLED:The aim of this study was to determine the effect of RNA interference inhibition of mineralocorticoid receptor (MR) on cold-induced hypertension (CIH) and renal damage. Recombinant adeno-associated virus (AAV) carrying short hairpin small interference (si)RNA for MR (AAV.MR-shRNA) was constructed and tested ...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3302768
更新日期:2006-07-01 00:00:00
abstract::Nonhealing bone defects are difficult to treat. As the bone morphogenic protein and transforming growth factor beta pathways have been implicated in bone healing, we hypothesized that percutaneous Smad7 silencing would enhance signaling through both pathways and improve bone formation. Critical sized parietal trephine...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/gt.2013.15
更新日期:2013-09-01 00:00:00
abstract::Complexes of DNA and cationic lipid offer potential advantages for gene transfer to airway epithelia. However, we found that application of DNA-lipid (DMRIE-DOPE) complexes to primary cultures of human ciliated airway epithelia or explants of rabbit trachea generated only low levels of gene transfer. In contrast, when...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3300524
更新日期:1997-11-01 00:00:00
abstract::Advances in genetic analysis and a greater understanding of human immunodeficiency virus (HIV) molecular pathogenesis have identified critical viral targets for gene interference strategies. RNase P molecules have been proposed as a novel approach for gene targeting based upon their potent catalytic activity, as well ...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3301606
更新日期:2001-12-01 00:00:00
abstract::Alphavirus vectors based on Sindbis virus and Semliki Forest virus (SFV) were characterized as potential gene transfer vectors. Initial studies were performed using vectors engineered to transfer either lacZ or green fluorescent protein (GFP). High levels of gene transfer were achieved in human primary fibroblasts, BH...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3301122
更新日期:2000-03-01 00:00:00
abstract::Gene targeting by single-stranded oligodeoxyribonucleotides (ssODNs) is a promising technique for introducing site-specific sequence alterations without affecting the genomic organization of the target locus. Here, we discuss the significant progress that has been made over the last 5 years in unraveling the mechanism...
journal_title:Gene therapy
pub_type: 杂志文章,评审
doi:10.1038/gt.2010.161
更新日期:2011-03-01 00:00:00
abstract::We have comparatively evaluated the efficiency of a series of retroviral vectors transducing the gp91-phox gene, whose defects are responsible for impaired production of superoxide anion (O2-) by phagocytic cells and lead to the X-linked form of chronic granulomatous disease (X-CGD). These vectors included four constr...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3301057
更新日期:2000-01-01 00:00:00
abstract::VP22, a structural protein from herpes simplex virus type I, exhibits the unique property of intercellular trafficking. This protein is exported from primary expressing cells and subsequently imported into neighbouring cells. This property is conserved when VP22 is genetically fused to a protein, making it a promising...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3301904
更新日期:2003-02-01 00:00:00
abstract::Defensins are small cationic proteins that harbor broad-spectrum microbicidal activity against bacteria, fungi and viruses. This study examines the effects on pathogens of the epidermis engineered to express human beta-defensin 3 (HBD3) to combat bacterial infections. First, we examined the localization of HBD3 in the...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3302472
更新日期:2005-05-01 00:00:00
abstract::Retroviral vectors are effective shuttle systems by introducing therapeutically relevant genes stably into the genome of proliferating cells. The majority of vectors applied for research or clinical applications use neomycin for cell selection and identification. To circumvent the time consuming and potentially toxic ...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3300384
更新日期:1997-04-01 00:00:00
abstract::In 2012, we reported that 5 out of 11 subjects in a clinical trial (NCT00372320) administering AdhAQP1 to radiation-damaged parotid glands showed increased saliva flow rates and decreased symptoms over the initial 42 days. AdhAQP1 is a first-generation, E1-deleted, replication-defective, serotype 5 adenoviral vector e...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/gt.2015.55
更新日期:2015-09-01 00:00:00
abstract::Hyperammonemia is less severe in arginase 1 deficiency compared with other urea cycle defects. Affected patients manifest hyperargininemia and infrequent episodes of hyperammonemia. Patients typically suffer from neurological impairment with cortical and pyramidal tract deterioration, spasticity, loss of ambulation, s...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/gt.2014.106
更新日期:2015-02-01 00:00:00
abstract::Hydrodynamic gene delivery is an attractive option for non-viral liver gene therapy, but requires evaluation of efficacy, safety and clinically applicable techniques in large animal models. We have evaluated retrograde delivery of DNA to the whole liver via the isolated segment of inferior vena cava (IVC) draining the...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3303079
更新日期:2008-03-01 00:00:00
abstract::For many gene therapy applications the effective titre of retroviral vectors is a limiting factor both in vitro and in vivo. Purification and concentration of retrovirus from packaging cell supernatant can overcome this problem. To this end we have investigated a novel procedure which involves complexing retrovirus to...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3301201
更新日期:2000-06-01 00:00:00
abstract::The potential for gene therapy to be an effective treatment for cystic fibrosis has been hampered by the limited gene transfer efficiency of current vectors. We have shown that recombinant Sendai virus (SeV) is highly efficient in mediating gene transfer to differentiated airway epithelial cells, because of its capaci...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3302991
更新日期:2007-10-01 00:00:00
abstract::Adenovirus-mediated gene transfer of interferon gamma (AdIFN) elicits rejection of intracerebral Lewis lung carcinoma. In this system, gene transfer into brain parenchymal cells is both necessary and sufficient to generate the antitumor response. Despite persistent parenchymal inflammation and demyelination, wild-type...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3301346
更新日期:2000-12-01 00:00:00
abstract::The possibility of achieving multiple systemic expression of human interferon-beta in mice upon repeated intravenous administration of cationic liposome-DNA complex (lipoplex) was investigated. Lipoplexes containing the pentammonio lipid pcTG90 were first optimized by selecting the most efficient ratio of pcTG90 to ph...
journal_title:Gene therapy
pub_type: 杂志文章
doi:10.1038/sj.gt.3301289
更新日期:2000-09-01 00:00:00
abstract::The E1 deleted adenoviral vectors are efficient at gene transfer to cells in culture or in animals. However, their use is limited because of an immune-mediated loss of transduced cells. This immune response is believed to result from low-level production of viral antigens from these vectors after gene transfer. The ea...
journal_title:Gene therapy
pub_type: 杂志文章
doi:
更新日期:1995-06-01 00:00:00