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Nicholas D. Mazarakis - One of the best experts on this subject based on the ideXlab platform.
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Venezuelan equine encephalitis virus glycoprotein Pseudotyping confers neurotropism to lentiviral vectors
Gene Therapy, 2013Co-Authors: Antonio Trabalza, James N. Hislop, Ch. Georgiadis, Stuart M. Ellison, Ioanna Eleftheriadou, Maria Elizabeth Karavassilis, Nicholas D. MazarakisAbstract:We have produced high-titre HIV-1 green fluorescent protein-expressing lentiviral (LV) vectors pseudotyped with strain 3908 Venezuelan equine encephalitis virus glycoprotein (VEEV-G) and used them to study transduction of: (1) rat embryonic motor neuron (MN) and striatal neuron primary cultures, (2) differentiated MN cell line NSC-34 and (3) adult rat striatum. In primary neuronal cultures, transduction with VEEV-G-pseudotyped LV was more efficient and more neuronal than with vesicular stomatitis virus glycoprotein (VSV-G)-pseudotyped LV. In NSC-34 cells clear retrograde transport of VEEV-G vector particles was observed. In the striatum at the injection site, transduction with the VEEV-G vectors driven by cytomegalovirus or phosphoglycerate kinase promoters exhibited a distinct neuronal tropism with no microglial and only a minor astroglial component, superior to that obtained with VSV-G-pseudotyped LV, irrespective of the promoter used. Neuronal transduction efficiency increased over time. Distal to the injection site transduction of mitral cells in the olfactory bulb, thalamic neurons and dopaminergic neurons in the substantia nigra pars compacta was detected. This, together with observations of retrograde axonal trafficking in vitro indicates that these vectors also possess low level of retrograde neuronal transduction capability in vivo . In this study, we demonstrate both strong neurotropism as well as sustainability of expression and minimal host immune response in vivo , making the VEEV-G-pseudotyped LV vectors potentially useful for gene therapy of neurodegenerative diseases.
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Venezuelan equine encephalitis virus glycoprotein Pseudotyping confers neurotropism to lentiviral vectors
Gene Therapy, 2012Co-Authors: Antonio Trabalza, James N. Hislop, Ch. Georgiadis, Stuart M. Ellison, Ioanna Eleftheriadou, Maria Elizabeth Karavassilis, Nicholas D. MazarakisAbstract:Venezuelan equine encephalitis virus glycoprotein Pseudotyping confers neurotropism to lentiviral vectors
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Transduction Patterns of Pseudotyped Lentiviral Vectors in the Nervous System
Molecular therapy : the journal of the American Society of Gene Therapy, 2004Co-Authors: Liang-fong Wong, Susan M Kingsman, Kyriacos A Mitrophanous, Mimoun Azzouz, Lucy E. Walmsley, Zoe Askham, Fraser J. Wilkes, Nicholas D. MazarakisAbstract:We have developed a non-primate-based lentiviral vector based on the equine infectious anemia virus (EIAV) for efficient gene transfer to the central and peripheral nervous systems. Previously we have demonstrated that Pseudotyping lentiviral vectors with the rabies virus glycoprotein confers retrograde axonal transport to these vectors. In the present study we have successfully produced high-titer EIAV vectors pseudotyped with envelope glycoproteins from Rhabdovirus vesicular stomatitis virus (VSV) serotypes (Indiana and Chandipura strains); rabies virus [various Evelyn–Rokitnicki–Abelseth ERA strains and challenge virus standard (CVS)]; Lyssavirus Mokola virus, a rabies-related virus; and Arenavirus lymphocytic choriomeningitis virus (LCMV). These vectors were delivered to the striatum or spinal cord of adult rats or muscle of neonatal mice by direct injection. We report that the lentiviral vectors pseudotyped with envelopes from the VSV Indiana strain, wild-type ERA, and CVS strains resulted in strong transduction in the striatum, while Mokola- and LCMV-pseudotyped vectors exhibited moderate and weak transduction, respectively. Furthermore ERA- and CVS-pseudotyped lentiviral vectors demonstrated retrograde transport and expression in distal neurons after injection in brain, spinal cord, and muscle. The differences in transduction efficiencies and retrograde transport conferred by these envelope glycoproteins present novel opportunities in designing therapeutic strategies for different neurological diseases.
Antonio Trabalza - One of the best experts on this subject based on the ideXlab platform.
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Venezuelan equine encephalitis virus glycoprotein Pseudotyping confers neurotropism to lentiviral vectors
Gene Therapy, 2013Co-Authors: Antonio Trabalza, James N. Hislop, Ch. Georgiadis, Stuart M. Ellison, Ioanna Eleftheriadou, Maria Elizabeth Karavassilis, Nicholas D. MazarakisAbstract:We have produced high-titre HIV-1 green fluorescent protein-expressing lentiviral (LV) vectors pseudotyped with strain 3908 Venezuelan equine encephalitis virus glycoprotein (VEEV-G) and used them to study transduction of: (1) rat embryonic motor neuron (MN) and striatal neuron primary cultures, (2) differentiated MN cell line NSC-34 and (3) adult rat striatum. In primary neuronal cultures, transduction with VEEV-G-pseudotyped LV was more efficient and more neuronal than with vesicular stomatitis virus glycoprotein (VSV-G)-pseudotyped LV. In NSC-34 cells clear retrograde transport of VEEV-G vector particles was observed. In the striatum at the injection site, transduction with the VEEV-G vectors driven by cytomegalovirus or phosphoglycerate kinase promoters exhibited a distinct neuronal tropism with no microglial and only a minor astroglial component, superior to that obtained with VSV-G-pseudotyped LV, irrespective of the promoter used. Neuronal transduction efficiency increased over time. Distal to the injection site transduction of mitral cells in the olfactory bulb, thalamic neurons and dopaminergic neurons in the substantia nigra pars compacta was detected. This, together with observations of retrograde axonal trafficking in vitro indicates that these vectors also possess low level of retrograde neuronal transduction capability in vivo . In this study, we demonstrate both strong neurotropism as well as sustainability of expression and minimal host immune response in vivo , making the VEEV-G-pseudotyped LV vectors potentially useful for gene therapy of neurodegenerative diseases.
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Venezuelan equine encephalitis virus glycoprotein Pseudotyping confers neurotropism to lentiviral vectors
Gene Therapy, 2012Co-Authors: Antonio Trabalza, James N. Hislop, Ch. Georgiadis, Stuart M. Ellison, Ioanna Eleftheriadou, Maria Elizabeth Karavassilis, Nicholas D. MazarakisAbstract:Venezuelan equine encephalitis virus glycoprotein Pseudotyping confers neurotropism to lentiviral vectors
Ramesh Akkina - One of the best experts on this subject based on the ideXlab platform.
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Generation of High-Titer Pseudotyped Lentiviral Vectors.
Methods in molecular biology (Clifton N.J.), 2019Co-Authors: Ramesh AkkinaAbstract:Lentiviral vectors (LVs) are widely used in gene transfer protocols due to many advantages that include stable gene expression, higher transgene payloads, and, importantly, the ability to pseudotype the vectors with a diverse number of heterologous viral envelopes with broad or restricted cell tropism depending on the need. The Pseudotyping process also allows for incorporation of specific antibodies/ligands to engineer LVs. These features greatly facilitate customization of lentiviral vectors for cell/tissue specific gene delivery. The VSV-G protein containing envelope remains the most widely used among the viral glycoproteins used for LV Pseudotyping due to its versatile host range and stability. However, many other viral envelopes are being identified for special applications of LVs. Here we describe the methodology to generate pseudotyped LVs using a four-plasmid transient transfection system focusing on aspects to generate high-titer vector stocks.
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Pseudotyping of lentiviral vector with novel vesiculovirus envelope glycoproteins derived from chandipura and piry viruses
Virology, 2016Co-Authors: Dipu Mohan Kumar, Chelsea Sax, Clayton Schuler, Ramesh AkkinaAbstract:While the envelope glycoprotein of vesicular stomatitis virus (VSV-G) is widely used for Pseudotyping of lentiviral vectors, sub-optimal gene transfer into certain cell types and its sensitivity to inactivation by human complement hinders its broader applications. To find alternative candidates, here we evaluated two serologically distinct novel viral envelopes derived from Chandipura (CNV-G) and Piry (PRV-G) vesiculoviruses. Both permitted generation of high titer psuedotyped lentiviral vectors with a capacity for high efficiency gene transfer into various cell types from different species. In human lymphoid and hematopoietic stem cells, their transduction efficiency was significantly lower than that of VSV-G. However, both novel envelopes were found to be more resistant to inactivation by human serum complement compared to VSV-G. Thus CNV-G and PRV-G envelopes can be harnessed for multiple uses in the future based on the cell type that needs to be gene transduced and possibly for in vivo gene transfer.
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731 Pseudotyping of lentiviral vectors with new envelope g proteins from vesiculoviruses vsv new jersey chandipura and piry
Molecular Therapy, 2004Co-Authors: Akhil C Banerjea, Ramesh AkkinaAbstract:Retroviral and lentiviral vectors are highly efficient in gene transfer. Their tropism, stability and ability to be concentrated by ultracentrifugation were greatly increased by Pseudotyping with the vesicular stomatitis virus G (VSV-G) protein. This allowed expanded use of these vectors for many purposes. Although most current gene therapy studies focused on ex vivo gene transductions with these vectors, the next desirable step is to be able to inject them directly for systemic delivery. While vectors pseudotyped with VSV-G from Indiana serotype yielded high levels of gene transfer, successive rounds of their in vivo administration will have immunological limitations because this protein is known to elicit strong neutralizing antibodies. Hence, for the purpose of repeated administrations, additional non-crossreactive envelope proteins that have comparable value in Pseudotyping retro- and lentiviral vectors are needed. Furthermore, as VSV-G is highly fusogenic, there is cytotoxicity associated with its use via concentrated vector stocks. Accordingly, a less cytotoxic envelope protein for Pseudotyping is also highly desirable. Taking these criteria into consideration, we tested envelope proteins from three different vesiculoviruses, namely VSV-New Jersey, Chandipura and Piry. A third generation lentiviral vector encoding GFP reporter gene was pseudotyped with each of these envelope G proteins in a transient transfection packaging system. The respective pseudotyped lentiviral vectors were used to transduce different mammalian cells including BHK-21, HeLa, Cos, Prostate cancer cell lines, as well as primary human CD34+ hematopoietic cells and macrophages derived in vitro. The transfection efficiencies were compared to the vector pseudotyped with VSV-G Indiana. Our results showed that the three new G proteins used were effective in Pseudotyping the lentiviral vectors as assayed by GFP marker expression from transduced cells. Differential transduction levels were found between different cell types indicating variation in cell tropism between different envelopes. These studies established that envelope proteins from related vesiculoviruses are useful for Pseudotyping lentiviral vectors and provide alternative non-crossreactive envelope proteins for in vivo administration of gene transfer vectors.
James N. Hislop - One of the best experts on this subject based on the ideXlab platform.
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Venezuelan equine encephalitis virus glycoprotein Pseudotyping confers neurotropism to lentiviral vectors
Gene Therapy, 2013Co-Authors: Antonio Trabalza, James N. Hislop, Ch. Georgiadis, Stuart M. Ellison, Ioanna Eleftheriadou, Maria Elizabeth Karavassilis, Nicholas D. MazarakisAbstract:We have produced high-titre HIV-1 green fluorescent protein-expressing lentiviral (LV) vectors pseudotyped with strain 3908 Venezuelan equine encephalitis virus glycoprotein (VEEV-G) and used them to study transduction of: (1) rat embryonic motor neuron (MN) and striatal neuron primary cultures, (2) differentiated MN cell line NSC-34 and (3) adult rat striatum. In primary neuronal cultures, transduction with VEEV-G-pseudotyped LV was more efficient and more neuronal than with vesicular stomatitis virus glycoprotein (VSV-G)-pseudotyped LV. In NSC-34 cells clear retrograde transport of VEEV-G vector particles was observed. In the striatum at the injection site, transduction with the VEEV-G vectors driven by cytomegalovirus or phosphoglycerate kinase promoters exhibited a distinct neuronal tropism with no microglial and only a minor astroglial component, superior to that obtained with VSV-G-pseudotyped LV, irrespective of the promoter used. Neuronal transduction efficiency increased over time. Distal to the injection site transduction of mitral cells in the olfactory bulb, thalamic neurons and dopaminergic neurons in the substantia nigra pars compacta was detected. This, together with observations of retrograde axonal trafficking in vitro indicates that these vectors also possess low level of retrograde neuronal transduction capability in vivo . In this study, we demonstrate both strong neurotropism as well as sustainability of expression and minimal host immune response in vivo , making the VEEV-G-pseudotyped LV vectors potentially useful for gene therapy of neurodegenerative diseases.
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Venezuelan equine encephalitis virus glycoprotein Pseudotyping confers neurotropism to lentiviral vectors
Gene Therapy, 2012Co-Authors: Antonio Trabalza, James N. Hislop, Ch. Georgiadis, Stuart M. Ellison, Ioanna Eleftheriadou, Maria Elizabeth Karavassilis, Nicholas D. MazarakisAbstract:Venezuelan equine encephalitis virus glycoprotein Pseudotyping confers neurotropism to lentiviral vectors
Ioanna Eleftheriadou - One of the best experts on this subject based on the ideXlab platform.
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Venezuelan equine encephalitis virus glycoprotein Pseudotyping confers neurotropism to lentiviral vectors
Gene Therapy, 2013Co-Authors: Antonio Trabalza, James N. Hislop, Ch. Georgiadis, Stuart M. Ellison, Ioanna Eleftheriadou, Maria Elizabeth Karavassilis, Nicholas D. MazarakisAbstract:We have produced high-titre HIV-1 green fluorescent protein-expressing lentiviral (LV) vectors pseudotyped with strain 3908 Venezuelan equine encephalitis virus glycoprotein (VEEV-G) and used them to study transduction of: (1) rat embryonic motor neuron (MN) and striatal neuron primary cultures, (2) differentiated MN cell line NSC-34 and (3) adult rat striatum. In primary neuronal cultures, transduction with VEEV-G-pseudotyped LV was more efficient and more neuronal than with vesicular stomatitis virus glycoprotein (VSV-G)-pseudotyped LV. In NSC-34 cells clear retrograde transport of VEEV-G vector particles was observed. In the striatum at the injection site, transduction with the VEEV-G vectors driven by cytomegalovirus or phosphoglycerate kinase promoters exhibited a distinct neuronal tropism with no microglial and only a minor astroglial component, superior to that obtained with VSV-G-pseudotyped LV, irrespective of the promoter used. Neuronal transduction efficiency increased over time. Distal to the injection site transduction of mitral cells in the olfactory bulb, thalamic neurons and dopaminergic neurons in the substantia nigra pars compacta was detected. This, together with observations of retrograde axonal trafficking in vitro indicates that these vectors also possess low level of retrograde neuronal transduction capability in vivo . In this study, we demonstrate both strong neurotropism as well as sustainability of expression and minimal host immune response in vivo , making the VEEV-G-pseudotyped LV vectors potentially useful for gene therapy of neurodegenerative diseases.
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Venezuelan equine encephalitis virus glycoprotein Pseudotyping confers neurotropism to lentiviral vectors
Gene Therapy, 2012Co-Authors: Antonio Trabalza, James N. Hislop, Ch. Georgiadis, Stuart M. Ellison, Ioanna Eleftheriadou, Maria Elizabeth Karavassilis, Nicholas D. MazarakisAbstract:Venezuelan equine encephalitis virus glycoprotein Pseudotyping confers neurotropism to lentiviral vectors
