Clades of Adeno-Associated Viruses Are Widely Disseminated in Human Tissues

ABSTRACT The potential for using Adeno-associated virus (AAV) as a vector for human gene therapy has stimulated interest in the Dependovirus genus. Serologic data suggest that AAV infections are prevalent in humans, although analyses of viruses and viral sequences from clinical samples are extremely limited. Molecular techniques were used in this study to successfully detect endogenous AAV sequences in 18% of all human tissues screened, with the liver and bone marrow being the most predominant sites. Sequence characterization of rescued AAV DNAs indicated a diverse array of molecular forms which segregate into clades whose members share functional and serologic similarities. One of the most predominant human clades is a hybrid of two previously described AAV serotypes, while another clade was found in humans and several species of nonhuman primates, suggesting a cross-species transmission of this virus. These data provide important information regarding the biology of parvoviruses in humans and their use as gene therapy vectors.

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Quantitation of Trace Levels of DNA Released from Disrupted Adeno-Associated Virus Gene Therapy Vectors

Journal of Pharmaceutical Sciences ◽

10.1016/j.xphs.2021.06.010 ◽

2021 ◽

Author(s):

Jared S. Bee ◽

Kristin O'Berry ◽

Yu (Zoe) Zhang ◽

Megan Kuhn Phillippi ◽

Akanksha Kaushal ◽

...

Keyword(s):

Gene Therapy ◽

Adeno Associated Virus ◽

Virus Gene ◽

Gene Therapy Vectors

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Characterization of Ocular Surface Microbial Profiles Revealed Discrepancies between Conjunctival and Corneal Microbiota

Pathogens ◽

10.3390/pathogens10040405 ◽

2021 ◽

Vol 10 (4) ◽

pp. 405

Author(s):

Anna Matysiak ◽

Michal Kabza ◽

Justyna A. Karolak ◽

Marcelina M. Jaworska ◽

Malgorzata Rydzanicz ◽

...

Keyword(s):

Microbial Communities ◽

Ocular Surface ◽

Molecular Techniques ◽

Corneal Tissue ◽

Rna Seq ◽

Microbiome Composition ◽

Viral Sequences ◽

Pcr Techniques ◽

Unmapped Reads

The ocular microbiome composition has only been partially characterized. Here, we used RNA-sequencing (RNA-Seq) data to assess microbial diversity in human corneal tissue. Additionally, conjunctival swab samples were examined to characterize ocular surface microbiota. Short RNA-Seq reads, obtained from a previous transcriptome study of 50 corneal tissues, were mapped to the human reference genome GRCh38 to remove sequences of human origin. The unmapped reads were then used for taxonomic classification by comparing them with known bacterial, archaeal, and viral sequences from public databases. The components of microbial communities were identified and characterized using both conventional microbiology and polymerase chain reaction (PCR) techniques in 36 conjunctival swabs. The majority of ocular samples examined by conventional and molecular techniques showed very similar microbial taxonomic profiles, with most of the microorganisms being classified into Proteobacteria, Firmicutes, and Actinobacteria phyla. Only 50% of conjunctival samples exhibited bacterial growth. The PCR detection provided a broader overview of positive results for conjunctival materials. The RNA-Seq assessment revealed significant variability of the corneal microbial communities, including fastidious bacteria and viruses. The use of the combined techniques allowed for a comprehensive characterization of the eye microbiome’s elements, especially in aspects of microbiota diversity.

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In Vivo Potency Assay for Adeno-Associated Virus–Based Gene Therapy Vectors Using AAVrh.10 as an Example

Human Gene Therapy Methods ◽

10.1089/hgtb.2017.246 ◽

2018 ◽

Vol 29 (3) ◽

pp. 146-155 ◽

Cited By ~ 5

Author(s):

Bishnu P. De ◽

Alvin Chen ◽

Christiana O. Salami ◽

Benjamin Van de Graaf ◽

Jonathan B. Rosenberg ◽

...

Keyword(s):

Gene Therapy ◽

Potency Assay ◽

Adeno Associated Virus ◽

Gene Therapy Vectors

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Enhancement of Adeno-Associated Virus-Mediated Gene Therapy Using Hydroxychloroquine in Murine and Human Tissues

Molecular Therapy — Methods & Clinical Development ◽

10.1016/j.omtm.2019.05.012 ◽

2019 ◽

Vol 14 ◽

pp. 77-89 ◽

Cited By ~ 6

Author(s):

Laurel C. Chandler ◽

Alun R. Barnard ◽

Sarah L. Caddy ◽

Maria I. Patrício ◽

Michelle E. McClements ◽

...

Keyword(s):

Gene Therapy ◽

Human Tissues ◽

Adeno Associated Virus

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The packaging capacity of adeno-associated virus (AAV) and the potential forwild-type-plusAAV gene therapy vectors

FEBS Letters ◽

10.1016/s0014-5793(97)00311-6 ◽

1997 ◽

Vol 407 (1) ◽

pp. 78-84 ◽

Cited By ~ 57

Author(s):

Paul L. Hermonat ◽

J.Gerald Quirk ◽

Brian M. Bishop ◽

Li Han

Keyword(s):

Gene Therapy ◽

Adeno Associated Virus ◽

Gene Therapy Vectors ◽

Packaging Capacity

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Overcoming the Cystic Fibrosis Sputum Barrier to Leading Adeno-associated Virus Gene Therapy Vectors

Molecular Therapy ◽

10.1038/mt.2014.89 ◽

2014 ◽

Vol 22 (8) ◽

pp. 1484-1493 ◽

Cited By ~ 51

Author(s):

Benjamin S Schuster ◽

Anthony J Kim ◽

Joshua C Kays ◽

Mia M Kanzawa ◽

William B Guggino ◽

...

Keyword(s):

Cystic Fibrosis ◽

Gene Therapy ◽

Adeno Associated Virus ◽

Virus Gene ◽

Cystic Fibrosis Sputum ◽

Gene Therapy Vectors

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346. Characterization of Adeno-Associated Virus Sequences in Human Tissues

Molecular Therapy ◽

10.1016/j.ymthe.2004.06.299 ◽

2004 ◽

Vol 9 ◽

pp. S132

Keyword(s):

Human Tissues ◽

Adeno Associated Virus

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Scientific and Regulatory Policy Committee Points to Consider: Nonclinical Research and Development of In Vivo Gene Therapy Products, Emphasizing Adeno-Associated Virus Vectors

Toxicologic Pathology ◽

10.1177/01926233211041962 ◽

2021 ◽

pp. 019262332110419

Author(s):

Julie A. Hutt ◽

Basel T. Assaf ◽

Brad Bolon ◽

Joy Cavagnaro ◽

Elizabeth Galbreath ◽

...

Keyword(s):

Gene Therapy ◽

Research And Development ◽

Molecular Techniques ◽

Therapeutic Modality ◽

Adeno Associated Virus ◽

Genetic Medicine ◽

Case Basis ◽

Regulatory Landscape ◽

Support Research

Sequencing of the human genome and numerous advances in molecular techniques have launched the era of genetic medicine. Increasingly precise technologies for genetic modification, manufacturing, and administration of pharmaceutical-grade biologics have proved the viability of in vivo gene therapy (GTx) as a therapeutic modality as shown in several thousand clinical trials and recent approval of several GTx products for treating rare diseases and cancers. In recognition of the rapidly advancing knowledge in this field, the regulatory landscape has evolved considerably to maintain appropriate monitoring of safety concerns associated with this modality. Nonetheless, GTx safety assessment remains complex and is designed on a case-by-case basis that is determined by the disease indication and product attributes. This article describes our current understanding of fundamental biological principles and possible procedures (emphasizing those related to toxicology and toxicologic pathology) needed to support research and development of in vivo GTx products. This article is not intended to provide comprehensive guidance on all GTx modalities but instead provides an overview relevant to in vivo GTx generally by utilizing recombinant adeno-associated virus-based GTx—the most common in vivo GTx platform—to exemplify the main points to be considered in nonclinical research and development of GTx products.

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Nucleotide sequence-based typing of meningococci directly from clinical samples

Journal of Medical Microbiology ◽

10.1099/jmm.0.05078-0 ◽

2003 ◽

Vol 52 (6) ◽

pp. 505-508 ◽

Cited By ~ 15

Author(s):

Mathew A. Diggle ◽

Carolyn M. Bell ◽

Stuart C. Clarke

Keyword(s):

Molecular Techniques ◽

Clinical Samples ◽

Nucleotide Sequencing ◽

Sequencing Data ◽

Conjugate Vaccines ◽

Laboratory Confirmation ◽

Highly Sensitive ◽

Culture Isolate ◽

Micro Organisms

The unpredictable characteristics of meningococcal disease (MD) make outbreaks complicated to monitor and consequently lead to high levels of public anxiety. Traditional molecular techniques have been utilized in order to understand better the epidemiology of MD, but some have disadvantages such as being highly specialized and labour-intensive, with low reproducibility. Some of these problems have been overcome by using multilocus sequence typing (MLST). This technique exploits the unambiguous nature and electronic portability of nucleotide sequencing data for the characterization of micro-organisms. The need for enhanced surveillance of MD after the introduction of serogroup C conjugate vaccines means that it is important to gain typing information from the infecting organism in the absence of a culture isolate. Here, the application of MLST for the laboratory confirmation and characterization of Neisseria meningitidis directly from clinical samples is described. This involved using a newly designed set of primers that were complementary to nucleotide sequences external to the existing MLST primers already in use for culture-based MLST of meningococci. This combination has produced a highly sensitive procedure to allow the efficient genotypic characterization of meningococci directly from clinical samples.

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