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JournalISSN: 1043-0342

Human Gene Therapy 

Mary Ann Liebert, Inc.
About: Human Gene Therapy is an academic journal published by Mary Ann Liebert, Inc.. The journal publishes majorly in the area(s): Genetic enhancement & Viral vector. It has an ISSN identifier of 1043-0342. Over the lifetime, 4519 publications have been published receiving 232892 citations.


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Journal ArticleDOI
TL;DR: A recombinant adeno-associated virus serotype 2 vector, altered to carry the human RPE65 gene (rAAV2-CBSB-hRPE65) restored vision in animal models with R PE65 deficiency, and Comparisons are drawn between the present work and two other studies of ocular gene therapy for RPE 65-LCA that were carried out contemporaneously and reported.
Abstract: Leber congenital amaurosis (LCA) is a group of autosomal recessive blinding retinal diseases that are incurable. One molecular form is caused by mutations in the RPE65 (retinal pigment epithelium-specific 65-kDa) gene. A recombinant adeno-associated virus serotype 2 (rAAV2) vector, altered to carry the human RPE65 gene (rAAV2-CBSB-hRPE65), restored vision in animal models with RPE65 deficiency. A clinical trial was designed to assess the safety of rAAV2-CBSB-hRPE65 in subjects with RPE65-LCA. Three young adults (ages 21–24 years) with RPE65-LCA received a uniocular subretinal injection of 5.96 × 1010 vector genomes in 150 μl and were studied with follow-up examinations for 90 days. Ocular safety, the primary outcome, was assessed by clinical eye examination. Visual function was measured by visual acuity and dark-adapted full-field sensitivity testing (FST); central retinal structure was monitored by optical coherence tomography (OCT). Neither vector-related serious adverse events nor systemic tox...

971 citations

Journal ArticleDOI
TL;DR: It is demonstrated that high levels of plasmid DNA expression in hepatocytes can be easily obtained by tail vein injections and has great potential for a wide variety of laboratory studies.
Abstract: We have previously shown that the intramuscular injection of naked plasmid DNA enables foreign gene expression in muscle. Further studies showed that the intravascular delivery of naked plasmid DNA enables high levels of expression not only in muscle but also in hepatocytes. For the liver, this technique required injection directly into the liver vessels (portal vein, hepatic vein, or bile duct) and occlusion of outflow. The present study now demonstrates that high levels of plasmid DNA expression in hepatocytes can be easily obtained by tail vein injections. The highest levels of expression are achieved by rapidly injecting the plasmid DNA in large volumes, ~2.5 ml. This technique has great potential for a wide variety of laboratory studies.

959 citations

Journal ArticleDOI
TL;DR: The nuclear replication and retention functions of the Epstein-Barr virus have been utilized here to maintain Retroviral constructs episomally within human cell-based retroviral packaging lines, affording reproducibly rapid, large-scale, stable, and high-titer retrovirus production.
Abstract: The nuclear replication and retention functions of the Epstein-Barr virus (EBV) have been utilized here to maintain retroviral constructs episomally within human cell-based retroviral packaging lines. These hybrid EBV/retroviral constructs are capable of producing helper-free recombinant retrovirus as soon as 48 hr and for at least 30 days after transfection into 293T-based ecotropic and/or amphotropic retroviral packaging cells. Viral titers greater than 10(7) TU/ml were obtained after puromycin selection of transfected retroviral packaging cells. This episomal approach to retroviral production circumvents some limitations inherent in transient and chromosomally stable retroviral producer systems, affording reproducibly rapid, large-scale, stable, and high-titer retrovirus production.

815 citations

Journal ArticleDOI
TL;DR: Characterization of the preexisting humoral responses to the AAV capsid and cross-reactivity will allow development of new strategies to circumvent AAV acquired immune responses, and vectors based on AAV5, AAV8, and AAV9 may have an advantage for gene therapy in humans.
Abstract: Adeno-associated viruses (AAVs) are small, nonenveloped single-stranded DNA viruses that require helper viruses to facilitate efficient replication. Despite the presence of humoral responses to the wild-type AAV in humans, AAV remains one of the most promising candidates for therapeutic gene transfer to treat many genetic and acquired diseases. Characterization of the IgG subclass responses to AAV and study of the prevalence of both IgG and neutralizing factors to AAV types 1, 2, 5, 6, 8, and 9 in the human population are of importance for the development of new strategies to overcome these immune responses. Natural exposure to AAV types 1, 2, 5, 6, 8, and 9 can result in the production of antibodies from all four IgG subclasses, with a predominant IgG1 response and low IgG2, IgG3, and IgG4 responses. Prevalences of anti-AAV1 and -AAV2 total IgG determined by enzyme-linked immunosorbent assay were higher (67 and 72%) than those of anti-AAV5 (40%), anti-AAV6 (46%), anti-AAV8 (38%), and anti-AAV9 (47%). Furthermore, data showed that cross-reactions are important. The two highest neutralizing factor seroprevalences were observed for AAV2 (59%) and AAV1 (50.5%) and the lowest were observed for AAV8 (19%) and AAV5 (3.2%). Vectors based on AAV5, AAV8, and AAV9 may have an advantage for gene therapy in humans. Furthermore, among individuals seropositive for AAV5, AAV8, and AAV9, about 70-100% present low titers. Better characterization of the preexisting humoral responses to the AAV capsid and cross-reactivity will allow development of new strategies to circumvent AAV acquired immune responses.

755 citations

Journal ArticleDOI
TL;DR: Using a monoclonal antibody highly specific for AAV-2 capsids (A20), an rAAV affinity purification procedure protocol was established and led to a significant improvement in recombinant AAV vector production and purification.
Abstract: Standard protocols for the generation of adenoassociated virus type 2 (AAV-2)-based vectors for human gene therapy applications require cotransfection of cells with a recombinant AAV (rAAV) vector plasmid and a packaging plasmid that provides the AAV rep and cap genes. The transfected cells must also be overinfected with a helper virus, e.g., adenovirus (Ad), which delivers multiple helper functions necessary for rAAV production. Therefore, rAAV stocks produced using these protocols are contaminated with helper adenovirus. The generation of a novel packaging/helper plasmid, pDG, containing all AAV and Ad functions required for amplification and packaging of AAV vector plasmids, is described here. Cotransfection of cells with pDG and an AAV vector plasmid was sufficient for production of infectious rAAV, resulting in helper virus-free rAAV stocks. The rAAV titers obtained using pDG as packaging plasmid were up to 10-fold higher than those achieved using conventional protocols for rAAV production. Replacement of the AAV-2 p5 promoter by an MMTV-LTR promoter in pDG led to reduced expression of Rep78/68; however, expression of the VP proteins was significantly increased compared with VP levels from standard packaging plasmids. Immunofluorescence analyses showed that the strong accumulation of VP proteins in pDG-transfected cells resulted in enhanced AAV capsid assembly, which is limiting for efficient rAAV production. Furthermore, using a monoclonal antibody highly specific for AAV-2 capsids (A20), an rAAV affinity purification procedure protocol was established. The application of the tools described here led to a significant improvement in recombinant AAV vector production and purification.

730 citations

Performance
Metrics
No. of papers from the Journal in previous years
YearPapers
202371
2022152
2021143
2020126
2019130
2018103