Re-structuring lentiviral vectors to express genomic RNA via cap-dependent translation

Lentiviral (LV) vectors based on human immunodeficiency virus type I (HIV-1) package two copies of their single-stranded RNA into vector particles. Normally, this RNA genome is reverse transcribed into a double-stranded DNA provirus that integrates into the cell genome, providing permanent gene tran...

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Main Authors:	John R. Counsell, Guillaume De Brabandere, Rajvinder Karda, Marc Moore, Antonio Greco, Alysha Bray, Juan Antinao Diaz, Dany P. Perocheau, Ulrike Mock, Simon N. Waddington
Format:	Article
Language:	English
Published:	Elsevier 2021-03-01
Series:	Molecular Therapy: Methods & Clinical Development
Online Access:	http://www.sciencedirect.com/science/article/pii/S2329050120302539

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spelling	doaj-91dfe80765324ae6b172bc6c3bfd05e52021-03-13T04:23:42ZengElsevierMolecular Therapy: Methods & Clinical Development2329-05012021-03-0120357365Re-structuring lentiviral vectors to express genomic RNA via cap-dependent translationJohn R. Counsell0Guillaume De Brabandere1Rajvinder Karda2Marc Moore3Antonio Greco4Alysha Bray5Juan Antinao Diaz6Dany P. Perocheau7Ulrike Mock8Simon N. Waddington9Dubowitz Neuromuscular Centre, Molecular Neurosciences Section, Developmental Neurosciences Programme, UCL Great Ormond Street Institute of Child Health, 30 Guilford Street, London, UK; NIHR Great Ormond Street Hospital Biomedical Research Centre, 30 Guilford Street, London WC1N 1EH, UK; Corresponding author: John R. Counsell, Dubowitz Neuromuscular Centre, Molecular Neurosciences Section, Developmental Neurosciences Programme, UCL Great Ormond Street Institute of Child Health, 30 Guilford Street, London, UK.Dubowitz Neuromuscular Centre, Molecular Neurosciences Section, Developmental Neurosciences Programme, UCL Great Ormond Street Institute of Child Health, 30 Guilford Street, London, UK; NIHR Great Ormond Street Hospital Biomedical Research Centre, 30 Guilford Street, London WC1N 1EH, UKGene Transfer Technology Group, Institute for Women’s Health, University College London, 86-96 Chenies Mews, London, UKDubowitz Neuromuscular Centre, Molecular Neurosciences Section, Developmental Neurosciences Programme, UCL Great Ormond Street Institute of Child Health, 30 Guilford Street, London, UK; NIHR Great Ormond Street Hospital Biomedical Research Centre, 30 Guilford Street, London WC1N 1EH, UKDubowitz Neuromuscular Centre, Molecular Neurosciences Section, Developmental Neurosciences Programme, UCL Great Ormond Street Institute of Child Health, 30 Guilford Street, London, UK; NIHR Great Ormond Street Hospital Biomedical Research Centre, 30 Guilford Street, London WC1N 1EH, UKDubowitz Neuromuscular Centre, Molecular Neurosciences Section, Developmental Neurosciences Programme, UCL Great Ormond Street Institute of Child Health, 30 Guilford Street, London, UK; NIHR Great Ormond Street Hospital Biomedical Research Centre, 30 Guilford Street, London WC1N 1EH, UKGene Transfer Technology Group, Institute for Women’s Health, University College London, 86-96 Chenies Mews, London, UKGene Transfer Technology Group, Institute for Women’s Health, University College London, 86-96 Chenies Mews, London, UKNIHR Great Ormond Street Hospital Biomedical Research Centre, 30 Guilford Street, London WC1N 1EH, UKGene Transfer Technology Group, Institute for Women’s Health, University College London, 86-96 Chenies Mews, London, UK; MRC Antiviral Gene Therapy Research Unit, Faculty of Health Sciences, University of the Witswatersrand, Johannesburg, South AfricaLentiviral (LV) vectors based on human immunodeficiency virus type I (HIV-1) package two copies of their single-stranded RNA into vector particles. Normally, this RNA genome is reverse transcribed into a double-stranded DNA provirus that integrates into the cell genome, providing permanent gene transfer and long-term expression. Integration-deficient LV vectors have been developed to reduce the frequency of genomic integration and thereby limit their persistence in dividing cells. Here, we describe optimization of a reverse-transcriptase-deficient LV vector, which enables direct translation of LV RNA genomes upon cell entry, for transient expression of vector payloads as mRNA without a DNA intermediate. We have engineered a novel LV genome arrangement in which HIV-1 sequences are removed from the 5′ end, to enable ribosomal entry from the 5′ 7-methylguanylate cap for efficient translation of the vector payload. We have shown that this LV-mediated mRNA delivery platform provides transient transgene expression in vitro and in vivo. This has a potential application in gene and cell therapy scenarios requiring temporary payload expression in cells and tissues that can be targeted with pseudotyped LV vectors.http://www.sciencedirect.com/science/article/pii/S2329050120302539
collection	DOAJ
language	English
format	Article
sources	DOAJ
author	John R. Counsell Guillaume De Brabandere Rajvinder Karda Marc Moore Antonio Greco Alysha Bray Juan Antinao Diaz Dany P. Perocheau Ulrike Mock Simon N. Waddington
spellingShingle	John R. Counsell Guillaume De Brabandere Rajvinder Karda Marc Moore Antonio Greco Alysha Bray Juan Antinao Diaz Dany P. Perocheau Ulrike Mock Simon N. Waddington Re-structuring lentiviral vectors to express genomic RNA via cap-dependent translation Molecular Therapy: Methods & Clinical Development
author_facet	John R. Counsell Guillaume De Brabandere Rajvinder Karda Marc Moore Antonio Greco Alysha Bray Juan Antinao Diaz Dany P. Perocheau Ulrike Mock Simon N. Waddington
author_sort	John R. Counsell
title	Re-structuring lentiviral vectors to express genomic RNA via cap-dependent translation
title_short	Re-structuring lentiviral vectors to express genomic RNA via cap-dependent translation
title_full	Re-structuring lentiviral vectors to express genomic RNA via cap-dependent translation
title_fullStr	Re-structuring lentiviral vectors to express genomic RNA via cap-dependent translation
title_full_unstemmed	Re-structuring lentiviral vectors to express genomic RNA via cap-dependent translation
title_sort	re-structuring lentiviral vectors to express genomic rna via cap-dependent translation
publisher	Elsevier
series	Molecular Therapy: Methods & Clinical Development
issn	2329-0501
publishDate	2021-03-01
description	Lentiviral (LV) vectors based on human immunodeficiency virus type I (HIV-1) package two copies of their single-stranded RNA into vector particles. Normally, this RNA genome is reverse transcribed into a double-stranded DNA provirus that integrates into the cell genome, providing permanent gene transfer and long-term expression. Integration-deficient LV vectors have been developed to reduce the frequency of genomic integration and thereby limit their persistence in dividing cells. Here, we describe optimization of a reverse-transcriptase-deficient LV vector, which enables direct translation of LV RNA genomes upon cell entry, for transient expression of vector payloads as mRNA without a DNA intermediate. We have engineered a novel LV genome arrangement in which HIV-1 sequences are removed from the 5′ end, to enable ribosomal entry from the 5′ 7-methylguanylate cap for efficient translation of the vector payload. We have shown that this LV-mediated mRNA delivery platform provides transient transgene expression in vitro and in vivo. This has a potential application in gene and cell therapy scenarios requiring temporary payload expression in cells and tissues that can be targeted with pseudotyped LV vectors.
url	http://www.sciencedirect.com/science/article/pii/S2329050120302539
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Re-structuring lentiviral vectors to express genomic RNA via cap-dependent translation

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