Technical Considerations for the Generation of Novel Pseudotyped Viruses

King, Barnabas; Temperton, Nigel; Grehan, Keith; Scott, Simon D.; Wright, Edward; Tarr, Alexander W.; Daly, Janet M.

doi:10.2217/fvl.15.106

Cited by 12 publications

(15 citation statements)

References 127 publications

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“…This could lead to bias in the phenotype of HCV strains that are selected for analysis of entry inhibitors and therefore provide an unrepresentative view of the properties of glycoproteins representing circulating HCV strains. While the methods for producing HCVpp are generally standardized ( Bartosch et al , 2003a ; Hsu et al , 2003 ; Tarr et al , 2007 ), successful generation of infectious pseudotypes is dependent on many factors [reviewed in King et al . (2016) ].…”

Section: Introductionmentioning

confidence: 99%

Novel functional hepatitis C virus glycoprotein isolates identified using an optimized viral pseudotype entry assay

Urbanowicz

McClure

King

et al. 2016

Journal of General Virology

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Retrovirus pseudotypes are a highly tractable model used to study the entry pathways of enveloped viruses. This model has been extensively applied to the study of the hepatitis C virus (HCV) entry pathway, preclinical screening of antiviral antibodies and for assessing the phenotype of patient-derived viruses using HCV pseudoparticles (HCVpp) possessing the HCV E1 and E2 glycoproteins. However, not all patient-isolated clones produce particles that are infectious in this model. This study investigated factors that might limit phenotyping of patient-isolated HCV glycoproteins. Genetically related HCV glycoproteins from quasispecies in individual patients were discovered to behave very differently in this entry model. Empirical optimization of the ratio of packaging construct and glycoprotein-encoding plasmid was required for successful HCVpp genesis for different clones. The selection of retroviral packaging construct also influenced the function of HCV pseudoparticles. Some glycoprotein constructs tolerated a wide range of assay parameters, while others were much more sensitive to alterations. Furthermore, glycoproteins previously characterized as unable to mediate entry were found to be functional. These findings were validated using chimeric cell-cultured HCV bearing these glycoproteins. Using the same empirical approach we demonstrated that generation of infectious ebolavirus pseudoviruses (EBOVpv) was also sensitive to the amount and ratio of plasmids used, and that protocols for optimal production of these pseudoviruses are dependent on the exact virus glycoprotein construct. These findings demonstrate that it is crucial for studies utilizing pseudoviruses to conduct empirical optimization of pseudotype production for each specific glycoprotein sequence to achieve optimal titres and facilitate accurate phenotyping.

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Section: Introductionmentioning

confidence: 99%

Novel functional hepatitis C virus glycoprotein isolates identified using an optimized viral pseudotype entry assay

Urbanowicz

McClure

King

et al. 2016

Journal of General Virology

Self Cite

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“…A major application of rVSV-ΔG pseudotypes has been enabling wide accessibility to study properties of high containment level 3 or 4 viruses at a low containment level [101][102][103]. It is particularly valuable in the study of novel emerging viruses, where a pseudotype can be rapidly produced once the sequence of the envelope glycoprotein is known.…”

Section: Vsv-based Vectorsmentioning

confidence: 99%

A tool with many applications: vesicular stomatitis virus in research and medicine

Munis

Bentley

Takeuchi

2020

Expert Opinion on Biological Therapy

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Introduction: Vesicular stomatitis virus (VSV) has long been a useful research tool in virology and recently become an essential part of medicinal products. Vesiculovirus research is growing quickly following its adaptation to clinical gene and cell therapy and oncolytic virotherapy.Areas covered: This article reviews the versatility of VSV as a research tool and biological reagent, its use as a viral and vaccine vector delivering therapeutic and immunogenic transgenes and an oncolytic virus aiding cancer treatment. Challenges such as the immune response against such advanced therapeutic medicinal products and manufacturing constraints are also discussed. Expert opinion: The field of in vivo gene and cell therapy is advancing rapidly with VSV used in many ways. Comparison of VSV's use as a versatile therapeutic reagent unveils further prospects and problems for each application. Overcoming immunological challenges to aid repeated administration of viral vectors and minimizing harmful host-vector interactions remains one of the major challenges. In the future, exploitation of reverse genetic tools may assist the creation of recombinant viral variants that have improved onco-selectivity and more efficient vaccine vector activity. This will add to the preferential features of VSV as an excellent advanced therapy medicinal product (ATMP) platform.

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“…Efforts at altering the host range of retroviral vectors―also known as pseudotyping, which results in the incorporation, on viral vectors, of glycoproteins from other enveloped viruses―have been also extensively studied [ 138 ]. Common methodologies for viral pseudotyping have been described at length [ 139 ], alongside the pseudotyping of emerging viruses [ 140 ]. Most commonly, pseudotyping has been carried out with vesicular stomatitis virus GP (VSV-G).…”

Section: Strategies For the Genetic Modification Of Nk Cellsmentioning

confidence: 99%

Viral and Nonviral Engineering of Natural Killer Cells as Emerging Adoptive Cancer Immunotherapies

Matosevic

2018

Journal of Immunology Research

100

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Natural killer (NK) cells are powerful immune effectors whose antitumor activity is regulated through a sophisticated network of activating and inhibitory receptors. As effectors of cancer immunotherapy, NK cells are attractive as they do not attack healthy self-tissues nor do they induce T cell-driven inflammatory cytokine storm, enabling their use as allogeneic adoptive cellular therapies. Clinical responses to adoptive NK-based immunotherapy have been thwarted, however, by the profound immunosuppression induced by the tumor microenvironment, particularly severe in the context of solid tumors. In addition, the short postinfusion persistence of NK cells in vivo has limited their clinical efficacy. Enhancing the antitumor immunity of NK cells through genetic engineering has been fueled by the promise that impaired cytotoxic functionality can be restored or augmented with the use of synthetic genetic approaches. Alongside expressing chimeric antigen receptors to overcome immune escape by cancer cells, enhance their recognition, and mediate their killing, NK cells have been genetically modified to enhance their persistence in vivo by the expression of cytokines such as IL-15, avoid functional and metabolic tumor microenvironment suppression, or improve their homing ability, enabling enhanced targeting of solid tumors. However, NK cells are notoriously adverse to endogenous gene uptake, resulting in low gene uptake and transgene expression with many vector systems. Though viral vectors have achieved the highest gene transfer efficiencies with NK cells, nonviral vectors and gene transfer approaches—electroporation, lipofection, nanoparticles, and trogocytosis—are emerging. And while the use of NK cell lines has achieved improved gene transfer efficiencies particularly with viral vectors, challenges with primary NK cells remain. Here, we discuss the genetic engineering of NK cells as they relate to NK immunobiology within the context of cancer immunotherapy, highlighting the most recent breakthroughs in viral vectors and nonviral approaches aimed at genetic reprogramming of NK cells for improved adoptive immunotherapy of cancer, and, finally, address their clinical status.

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Technical Considerations for the Generation of Novel Pseudotyped Viruses

Cited by 12 publications

References 127 publications

Novel functional hepatitis C virus glycoprotein isolates identified using an optimized viral pseudotype entry assay

Novel functional hepatitis C virus glycoprotein isolates identified using an optimized viral pseudotype entry assay

A tool with many applications: vesicular stomatitis virus in research and medicine

Viral and Nonviral Engineering of Natural Killer Cells as Emerging Adoptive Cancer Immunotherapies

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