Podcasts about aav2

From the European Society for Clinical Virology 2022 Conference in Manchester UK, Vincent speaks with Emma Thomson about the recent outbreak of pediatric hepatitis of unknown etiology and the finding that it is linked to infection by adenovirus-associated virus 2. Hosts: Vincent Racaniello Guest: Emma Thomson Subscribe (free): Apple Podcasts, Google Podcasts, RSS, email Become a patron of TWiV! Links for this episode Thomson laboratory Emma on TWiV 188 and TWiV 341 AAV2-associated pediatric hepatitis (bioRxiv) Timestamps by Jolene. Thanks! Intro music is by Ronald Jenkees Send your virology questions and comments to twiv@microbe.tv

Powered by synthetic biology, Pierce Ogden makes ALL possible mutations to an adeno-associated virus (AAV) outer shell and rapidly screens them to dissect their attributes. Pierce discusses the technological advances that make this breakthrough screen possible and the novel properties that were discovered. AAVs are rapidly becoming the prefered way to perform gene therapy, correcting cells that carry disease-causing mutations through genetic modification. This technology forms the basis for company Dyno Therapeutics.About the AuthorPierce performed this work as a postdoc at Harvard University in the lab of Professor George Church. Professor Church is one of the founding fathers of synthetic biology and the lab is renowned for developing high throughput methods to design, build, and test bioengineered parts.In his role as Co-Founder & CSO at Manifold Bio, Pierce utilizes his multiplexing expertise to uncover the design principles of protein therapeutics and make new drugs faster than ever before.Key TakeawaysGene therapy uses genetic information as a drug, correcting cells that carry disease-causing mutations.The inability to deliver these genes to the correct cells limits the widespread adoption of gene therapy.Adeno-associated viruses (AAVs) are an extremely promising way to deliver DNA to human cells. Their outer shell, or capsid, can be engineered for increased safety, specificity, and shelf-life.Using advances in DNA synthesis technology, all possible single mutations to the AAV capsid are generated.With a DNA barcode read through next generation sequencing, this AAV library was simultaneously tested cheaply and quickly to find mutations with improved properties.Increased thermal stability, evasion of immune responses, and specificity toward the brain were all found. TranslationPierce demonstrates that smart usage of our synthetic biology toolbox can allow millions of protein variants to be tested simultaneously, in direct opposition to the “tested in parallel” model that has dominated high-throughput biology.Manifold Bio takes this idea of DNA barcodes coupled with simultaneous screening and points it toward the field of protein therapeutics.First Author: Pierce OgdenPaper: Comprehensive AAV capsid fitness landscape reveals a viral gene and enables machine-guided design. Science, 2020.Follow Fifty Years on Twitter!If you’re an author of an upcoming paper in bio or know of any interesting papers dropping soon and want to hear from the authors, drop us an email at translation [AT] fifty [DOT] vc.

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.09.11.293894v1?rss=1 Authors: Zhai, J., Kim, H., Han, S. B., Manire, M., Yoo, R., Pang, S., Smith, G., Son, Y.-J. Abstract: A major barrier to intraspinal regeneration after dorsal root (DR) injury is the DR entry zone (DREZ), the CNS/PNS interface. DR axons stop regenerating at the DREZ, even if regenerative capacity is increased by a conditioning lesion. This potent blockade has long been attributed to myelin-associated inhibitors and CSPGs, but incomplete lesions and conflicting reports have hampered conclusive agreement. Here we evaluated DR regeneration in adult mice, using novel strategies to facilitate complete lesions and comprehensive analyses, selective tracing of proprio-/mechanoreceptive axons with AAV2, and genetic or viral targeting of Nogo, MAG, OMgp, CSPGs and GDNF. Simultaneously eliminating Nogo/MAG/OMgp elicited little intraspinal penetration of DR axons, even with additional removal of CSPGs and a conditioning lesion. Their absence, however, synergistically enhanced GDNF-elicited intraspinal regeneration. We conclude that myelin inhibitors and CSPGs constrain intraspinal regrowth of DR axons, but that they are not the primary mechanism(s) stopping axons at the DREZ. Copy rights belong to original authors. Visit the link for more info

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.05.17.100230v1?rss=1 Authors: Gauvain, G., Akolkar, H., Chaffiol, A., Arcizet, F., Khoei, M., Desrosiers, M., Jaillard, C., Caplette, R., Marre, O., Bertin, S., Fovet, C.-M., Demilly, J., Forster, V., Brazhnikova, E., Hantraye, P., Pouget, P., Douar, A., Pruneau, D., Chavas, J., Sahel, J.-A., Dalkara, D., Duebel, J., Benosman, R., Picaud, S. Abstract: Restoring vision using optogenetics is an ideal medical application because the eye offers a direct window to access and stimulate the pathological area: the retina. Optogenetic therapy could be applied to diseases with photoreceptor degeneration such as retinitis pigmentosa. Here, we select the specific optogenetic construct that is now used in the clinical trial and assess the opsin functional efficacy on non-human primates retinal ganglion cells (RGCs). We chose the microbial opsin ChrimsonR and showed that the vector AAV2.7m8 produced greater transfection in RGCs compared to AAV2, and that ChrimsonR attached to tdTomato (ChR-tdT) is more efficiently expressed than ChrimsonR. The 600 nm light activates the RGCs transfected with the vector AAV2.7m8-ChR-tdT from an irradiance of 1015 photons.cm-2.s-1. Vector doses of 5.1010 and 5.1011 vg/eye transfect up to 7000 RGCs/mm2 in the perifovea, with no significant immune reaction. Furthermore, using a multielectrode array we recorded RGCs responses starting from 1ms stimulus duration. Using the recorded activity we were able to decode stimulus information and estimate a theoretical visual acuity of 20/249, above legal blindness. Altogether, our results pave the way for the ongoing clinical trial with the AAV2.7m8-ChrimsonR-tdT vector for vision restoration in patients affected by retinitis pigmentosa. Copy rights belong to original authors. Visit the link for more info

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.05.11.089078v1?rss=1 Authors: De Miranda, B. R., Rocha, E. M., Castro, S., Greenamyre, J. T. Abstract: Dopaminergic neurons of the substantia nigra are selectively vulnerable to mitochondrial dysfunction, which is hypothesized to be an early and fundamental pathogenic mechanism in Parkinson's disease (PD). Mitochondrial function depends on the successful import of nuclear-encoded proteins, many of which are transported through the TOM20-TOM22 outer mitochondrial membrane import receptor machinery. Recent data suggests that post-translational modifications of -synuclein promote its interaction with TOM20 at the outer mitochondrial membrane and thereby inhibit normal protein import, which leads to dysfunction and death of dopaminergic neurons. As such, preservation of mitochondrial import in the face of -synuclein accumulation might be a strategy to prevent dopaminergic neurodegeneration, however, this is difficult to assess using current in vivo models of PD. To this end, we established an exogenous co-expression system, utilizing AAV2 vectors to overexpress human -synuclein and TOM20, individually or together, in the adult Lewis rat substantia nigra in order to assess whether TOM20 overexpression attenuates -synuclein-induced dopaminergic neurodegeneration. Twelve weeks after viral injection, we observed that AAV2-TOM20 expression was sufficient to prevent loss of nigral dopaminergic neurons caused by AAV2-Syn overexpression. The observed TOM20-mediated dopaminergic neuron preservation appeared to be due, in part, to the rescued import of nuclear-encoded mitochondrial electron transport chain proteins that were inhibited by -synuclein overexpression. In addition, TOM20 overexpression rescued the import of the chaperone protein GRP75/mtHSP70/mortalin, a stress-response protein involved in -synuclein-induced injury. Collectively, these data indicate that TOM20 expression prevents -synuclein-induced mitochondrial dysfunction, which is sufficient to rescue dopaminergic neurons in the adult rat brain. Copy rights belong to original authors. Visit the link for more info

[Frame - 00:55] Liz Parrish – Introduction[Frame - 06:10] Gene therapy & deliver mechanisms - AAV2 For Part 1 go here --> http://bit.ly/2tR9vdQ For Part 3 go here --> http://bit.ly/2SCETX1 This communication is available for information purposes only and does not constitute an offer or sale or any form of general solicitation or general advertising of interests in any fund or investment vehicle.  Any such offer will only be made in compliance with applicable state and federal securities laws pursuant to an offering memorandum and related offering documents which will be provided to qualified prospective investors upon request.  Prospective investors should review a Fund’s offering memorandum carefully, which includes important disclosures and risk factors associated with an investment in a Fund.   The views and strategies described may not be suitable for all investors. They also do not include all fees or expenses that may be incurred by investing in specific products. Past performance is no guarantee of future results. Investments will fluctuate and when redeemed may be worth more or less than when originally invested. You cannot invest directly in an index. The opinions expressed are subject to change as subsequent conditions vary. Reliance upon information in this material is at the sole discretion of the reader. Advisory services offered through ACG Wealth Inc.  ACG Wealth Inc. is an affiliate of ACG Investment

At the University of Zürich, Vincent speaks with virologists Cornel Fraefel, Urs Greber, and Silke Stertz about their careers and their work on AAV2, adenovirus entry, and influenza virus. Hosts: Vincent Racaniello Guests: Cornel Fraefel, Urs Greber, and Silke Stertz Subscribe (free): iTunes, Google Podcasts, RSS, email Become a patron of TWiV! Links for this episode AAV2 and HSV1 co-infections (J Virol) Movies of HeLa Fucci cells (J Virol) Cathepsin W and influenza virus entry (mBio) Plaque 2.0 (PLoS One) Video of this episode (YouTube) Timestamps by Jolene. Thanks! Intro music is by Ronald Jenkees. Send your virology questions and comments to twiv@microbe.tv

Eduard Ayuso, INSERM 1089, University of Nantes, Nantes, France speaks on "Unraveling mechanisms and biology of recombinant AAV vectors produced in insect cells". Recombinant adeno-associated vectors (rAAV) are viral vectors of choice for gene therapy of many inherited diseases. Medicinal products based on rAAV are predominantly manufactured by transient transfection of mammalian cells or baculovirus expressing vectors (BEV) infection of insect cells, being the latter method more suitable for large-scale production. Although AAVs are mammalian viruses they can be assembled in insect cells, but the biology of the system has been poorly investigated. Here, we have studied the role of the assembly-activating protein (AAP) in insect cells and we found that this protein is expressed in a similar manner as in mammalian cells. By knocking down the AAP, it was confirmed that AAP is required for the assembly of AAV2 particles in insect cells. Next, we aimed to identify and characterize DNA species encapsidated in rAAV stocks produced in insect cells. To this end, we developed a single-strand virus sequencing protocol based on Illumina high-throughput sequencing technology (HTS). Preliminary data obtained from rAAV stocks puri ed by CsCl ultracentrifugation or immunoaffnity chromatography revealed that baculoviral and cellular DNA correspond to ≤1.5% and ≤0.02% of the total reads, respectively. Moreover, the sequencing coverage showed that the proximity to the ITRs increases progressively the probability for baculoviral DNA to be encapsidated. Nonetheless, these baculovirus-derived reads are found at a frequency of 2-3 logs lower than the rAAV genome reads. The development of accurate quality control methods is not only critical for fulfilling regulatory requirements, but will also provide novel insights into the biological mechanism of rAAV assembly in insect cells. This movie has been recorded by ICGEB Trieste.

Dr. Craig Meyers, professor of microbiology and immunology, Penn State College of Medicine, discusses his research using the virus AAV2 to kill breast cancer cells in the laboratory. Click on the Pod icon above or the direct download link below to hear the show. Right-click to save the file to your system.

Thu, 12 Nov 2009 12:00:00 +0100 https://edoc.ub.uni-muenchen.de/10823/ https://edoc.ub.uni-muenchen.de/10823/1/Schulz_Alexandra.pdf Schulz, Alexandra ddc:610, ddc:600, Medizinische F

Gentherapie ist ein vielversprechender neuer Ansatz für die Tumortherapie. Daher ist die Entwicklung eines effizienten Vektorsystems von großer Bedeutung. Bisher wurden unterschiedliche Serotypen des Adeno-assoziierten Virus (AAV) beschrieben, welche einen erheblichen Unterschied im Tropismus für bestimmte Gewebearten und Zelltypen aufweisen. Daraufhin haben wir systematisch den effizientesten Serotyp unter den AAV Serotypen 1 bis 5 bezüglich der Effektivität des Gentransfers in humanen Tumorzellen untereinander verglichen. Um alle Einflüsse außer denen, die auf das Kapsid zurückzuführen sind, auszuschließen, enthalten alle fünf Serotypen die gleiche Transgen-Kassette, einheitlich flankiert von der ITR (Invertierte terminale Wiederholung = inverted terminal repeat) des AAV-Serotyp 2. AAV2 war der effizienteste Serotyp in einer Reihe klinisch relevanter Tumorzelllinien mit einer Transduktionseffizienz von über 99,5±0,2 % bei nur 1000 genomische Partikeln / Zelle. Transduktionseffizienzen um die 70 % konnte durch den Serotyp 1 in Prostata- und Zervixkarzinom und durch den Serotyp 3 in Prostata-, Mamma-, Kolon- und Zervixkarzinom beim Einsatz von 1000 genomischen Partikeln pro Zelle erzielt werden. AAV4 und AAV5 haben durchgehend schlecht transduziert. Die höchste Transduktionseffizienz unter den humanen Tumorzelllinien betrug für AAV4, 40,6±3,1 % und 25,4±2,2 % für AAV5 beim Zervixkarzinom. Die jüngsten Fortschritte in der AAV-Vektor-Technologie weisen darauf hin, dass AAV- basierte Vektoren für die Tumorgentherapie eingesetzt werden können. Unsere Vergleichsanalysen zeigen, dass AAV2 zwar der am vielversprechenste Kandidat für eine solche Anwendung ist, aber AAV1 und AAV3 auch als gute Alternativen verwendet werden können. Dies ist besonders dann von Bedeutung, wenn eine Anwendung mit AAV2 durch neutralisierende Antikörper verhindert wird. Gao et al. konnten zeigen, dass diese Serotypen trotz des Vorhandensein neutralisierender Antikörper gegen AAV2 zum effizienten Gentransfer befähigt sind. Die Transduktionseffizienz von AAV4 und AAV5 ist generell zu niedrig, was eine effiziente Anwendung in der Tumorgentherapie derzeit unmöglich macht. Bei Untersuchungen einiger muriner Zelllinien stellten wir fest, dass sie sich am besten von AAV1 transduzieren lassen. Um nachzuweisen, ob AAV1 generell murine Zellen besser transduziert als AAV2, müssten weitere Untesuchungen durchgeführt werden. Dies ist für die zukünftige AAV-Forschung von Bedeutung, da eine Bestätigung unserer Beobachtung bedeuten würde, dass die Ergebnisse aus den zahlreichen in-vivo-Experimenten mit Mäusen nicht auf den Menschen übertragbar wären.

Background: The atheroprotective effects of systemic delivery of either apolipoprotein A-I (wtApoA-I) or the naturally occurring mutant ApoA-I Milano (ApoA-I(M)) have been established in animal and human trials, but direct comparison studies evaluating the phenotype of ApoA-I or ApoAI-Milano knock-in mice or bone marrow transplantated animals with selectively ApoA-I or ApoAI-Milano transduced macrophages give conflicting results regarding the superior performance of either one. We therefore sought to compare the two forms of apoA-I using liver-directed somatic gene transfer in hypercholesterinemic mice - a model which is most adequately mimicking the clinical setting. Methods and results: Vectors based on AAV serotype 8 (AAV2.8) encoding wtApoA-I, ApoA-I(M) or green fluorescent protein (GFP) as control were constructed. LDL receptor deficient mice were fed a Western Diet. After 8 weeks the AAV vectors were injected, and 6 weeks later atherosclerotic lesion size was determined by aortic en face analysis. Expression of wtApoA-I reduced progression of atherosclerosis by 32% compared with control (p = 0.02) and of ApoA-I(M) by 24% (p = 0.04). There was no significant difference between the two forms of ApoA-I in inhibiting atherosclerosis progression. Conclusion: Liver-directed AAV2.8-mediated gene transfer of wtApoA-I and ApoA-I(M) each significantly reduced atherosclerosis progression to a similar extent.