DOCSLIB.ORG

Tat-Independent Lentivirus Genomes for Vaccination and Host/Pathogen Interaction Studies

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Tat-Independent Lentivirus Genomes for Vaccination and Host/Pathogen Interaction Studies THÈSE Pour obtenir le grade de DOCTEUR DE LA COMMUNAUTÉ UNIVERSITÉ GRENOBLE ALPES Spécialité : Virologie/Microbiologie/Immunologie Arrêté ministériel : 7 août 2006 Présentée par Deepanwita BOSE Thèse dirigée par Dr. Yahia Chebloune préparée au sein du Laboratoire Pathogénèse et Vaccination Lentivirales (PAVAL Lab) dans l'École Doctorale Chimie & Sciences du Vivant (ED218) Tat-independent lentivirus genomes for vaccination and host/pathogen interaction studies. Thèse soutenue publiquement le 26 Janvier 2017, devant le jury composé de : Dr. Giuseppe Bertoni Maitre de conférences, IVI, Bern, Suisse Rapporteur Pr. Francois Villinger Professeur, Director of New Iberia Research Center, USA, Rapporteur Pr. Philippe Sabatier Professeur à l’Université Grenoble Alpes, Président Dr.Yahia Chebloune Research Director, à l’Université Grenoble Alpes, Examinateur Dr. Corinne Ronfort Chargé de recherche, Université Lyon I, Examinateur i “Dedicated to my Mom, Dad and Brother” i Acknowledgements Acknowledgements This thesis in its current form is a result of support, assistance, collaboration, critical inputs and guidance of several people and thus, I would sincerely thank each one of them. Firstly, I would like to express my sincere gratitude to my mentor Dr. Yahia Chebloune for having his faith and confidence in me and accepting me in his research lab for doing a PhD. thesis. I am thankful to him for his support, encouragement and extraordinary guidance all through my graduate work. His insightful discussions and critical comments were an invaluable part in preparation of this thesis. I am grateful for his patience throughout the tribulations, errors, repeats and also the successes in the lab, the corrections in the redaction of the thesis. His mentorship greatly improved and enhanced my scientific grasp and outlook. I would like to thank the members of my Committee: Our collaborator Pr. François Villinger, Director of the New Iberia Research Center, for sharing his expertise in the field and providing invaluable time and suggestions with a lot of encouragement. I am also thankful to him for providing a platform for adapting the newly developed viruses that I developed in Grenoble. For performing these experiments and adapting the viruses, I would like to thank members of his team, specially Dr. Chunxia Zhao at the Yerkes National Primate Center, Margot Lavy at the New Iberia Research Center and other members of the lab working to adapt the viruses in vitro and in vivo. Dr. Giuseppe Bertoni, for his invaluable time and expertise in the field and for being part of jury and reporter of the thesis. Dr. Corinne Ronfort, for all her scientific wisdom providing valuable inputs during the thesis and for her patience, critical review and guidance throughout the correction of the thesis Pr. Phillipe Sabatier, BioHealth Computing Director, for his professional excellence in providing constructive criticism and valuable inputs into the thesis. Dr. Catherine Lemaire, for being part of my thesis committee and providing significant constructive criticisms, I would also like to thank her for allowing me free use of laboratory equipment in emergency situations. I also thank Salima, for her technical help during these visits to the LAPM, Jean Roget Institute. I am thankful to Dr. Jean Gagnon, for his expertise and unwavering kindness, with valuable suggestions throughout my graduate studies and for his precious input for the redaction of my paper and thesis. I am deeply grateful to my lab mates and colleagues Maha, Simaa and Dimitri for their help, motivation, support and encouragement during good and less good times. They were always around to discuss, and gave invaluable suggestions and technical help for which, I am more than grateful. They were a part of my day-to-day life in lab and I cannot imagine my three i Acknowledgements years of lab life without them. I want also to acknowledge the help of Takfarinas, who joined the lab recently for his help in English-French translation and his savviness in MS Word software. I am also thankful and appreciate all other current and past members of the PAVAL team including previous students Honorine, Mikael, Sara, Avika, Ivan, Amna, Nadia, Amandine, Margot, Vanessa, Kathia, and technician Abderrahim. Special thanks to Camille for repeating some experiments of this thesis during her internship. I also want to warmly thank Gour, Monalisa, Ila, Sudha, Vikas, Vinodh, Lakshman, Bharat from the Indian community in Grenoble for their homely, joyful gatherings with loads of food, fun and all their support and encouragements along with valuable inputs for living in the French system. I am thankful for the financial support that I benefited from the Presidential Scholarship, University Grenoble Alpes for the years 2013-2016. And I am thankful to my Parents and Brother in India for always being encouraging and supporting me in all my endeavors in life, and this thesis is dedicated to them. ii Abstract in English Tat-independent lentivirus genomes for vaccination and host/pathogen interaction studies. Abstract in English Our lab has previously described the generation of a unique vaccine prototype against HIV- 1/AIDS. It is a non-integrative lentivector DNA vaccine, tested in pilot studies in animal models of HIV vaccine. The non-human primate study showed protection of all 6/6 macaques and immune response correlates were composed of a variety of effector (EM) and central memory (CM) T cells. More importantly, they also contained high proliferating antigen specific cells containing a type of stem cell-like memory T cells (TSCM). Here in my thesis work, the specificity of the vaccine was enhanced further by switching the CXCR4 envelope of the vaccine to CCR5 tropic envelopes such as the clade B WARO obtained from a chronically infected patient and three transmitted/founder (T/F) HIV Clade C strains from Zambia. To improve the vaccine further, we developed new strategies to incorporate cytokine genes as molecular adjuvants able to enhance and sustain the newly elicited immune responses. Since HIV-1 has developed multiple complex strategies to persist, the focus of the next part of my thesis was to develop tools to ease and better understand the underlying mechanisms of latency in infected memory CD4+ T cells. Latently infected cells have integrated in their genomes the complete viral DNA sequences but viral genes are not expressed. One of the main mechanisms of this latency is the absence of Tat transactivation of the LTR promoter. A recent focus in the era of efficient highly active antiviral therapy (HAART) is the cure of infection by targeting the reservoir of latently infected cells. One of the obstacles faced, is a lack of primate lentivirus prototypes incapable of undergoing latency as an extreme infection model to allow for teasing apart the various mechanisms contributing to latency. We hypothesized that a replication-competent SHIV genome driven by Tat-independent constitutive-expression LTRs such as the one of the caprine arthritis encephalitis virus, CAEV, the natural lentivirus of goats, will be a valuable tool for such studies. We designed chimeric CAEV LTRs bearing terminal sequences of SIV at their extremities, and used them to drive the transcription of the complete genome of SHIV-KU2. The reverse transcription of such a genome generates double stranded viral DNA bearing SIV attachment sequences (att) used efficiently by SIV IN for provirus integration. The resulting construct termed SHIV- YCC is expected to generate a viral infection that will not undergo latency due to uncoupling from Tat activation. We found that cells transfected with the SHIV-YCC DNA produce SHIV proteins that assemble into infectious particles released from the producer cells. These virions are able to infect new target CD4+ T cells both in primary PBMCs and in cell lines. SHIV- YCC passaged in macaques PBMCs increased virus replication and infectivity. SHIV-YCC is the first chimeric primate replication-competent lentivirus that constitutively expresses all viral proteins. This new model offers the possibility of studying the early events by which provirus undergoes latency and will serve as backbone for the construction of chimeric SHIVs with T/F CCR5 tropic HIV-1 envelopes more representative of early infecting isolates. Another major focus of this thesis was the development of replication competent (Tat driven) SHIVs that also more closely represent new HIV infections in humans. There is a need for CCR5 tropic SHIVs that not only infect monkeys but also induce sustained viral load set- points and pathogenesis, as many of the current R5-tropic SHIVs tend to undergo spontaneous control in vivo. In an attempt to palliate this issue in this thesis, SHIV-AD-T/F-31 and SHIV- AD-T/F-18 were developed using the SHIV-AD8EO backbone, which is a recent clade B SHIV with promising replication kinetics in vivo. The tat-env region of SHIV-AD8EO was iii Abstract in English exchanged with those of two HIV-1 clade C isolates: Z331T/F and Z3618T/F. These CCR5- tropic SHIVs will also provide tools to determine the mechanisms of latency/persistence when used in parallel with CAEV LTR chimeric constructs that are independent of the Tat/TAR transactivation. In addition, the replicative SHIV may provide invaluable reagents for testing HIV vaccines and microbicide efficacy in the macaque models. Keywords : HIV/SIV ; Tat ; CAEV ; CCR5 SHIV iv Abstract in French Génomes de lentivirus Tat indépendants pour des études de vaccination et les interactions hôte / pathogène Abstract in French Notre laboratoire a développé un prototype vaccinal unique contre le VIH-1 responsable du SIDA chez l’homme. C'est un lentivecteur ADN non-intégratif qui a été testé dans des études pilotes utilisant des modèles animaux. Une desq études a montré la protection de tous les macaques (6/6) vaccinés et la réponse immunitaire qui corrèle avec cette protection était composée de cellules effectrices (EM) et des cellules T centrales mémoires (CM).
Load more

Recommended publications
  • Non-Primate Lentiviral Vectors and Their Applications in Gene Therapy for Ocular Disorders
    viruses Review Non-Primate Lentiviral Vectors and Their Applications in Gene Therapy for Ocular Disorders Vincenzo Cavalieri 1,2,* ID , Elena Baiamonte 3 and Melania Lo Iacono 3 1 Department of Biological, Chemical and Pharmaceutical Sciences and Technologies (STEBICEF), University of Palermo, Viale delle Scienze Edificio 16, 90128 Palermo, Italy 2 Advanced Technologies Network (ATeN) Center, University of Palermo, Viale delle Scienze Edificio 18, 90128 Palermo, Italy 3 Campus of Haematology Franco e Piera Cutino, Villa Sofia-Cervello Hospital, 90146 Palermo, Italy; [email protected] (E.B.); [email protected] (M.L.I.) * Correspondence: [email protected] Received: 30 April 2018; Accepted: 7 June 2018; Published: 9 June 2018 Abstract: Lentiviruses have a number of molecular features in common, starting with the ability to integrate their genetic material into the genome of non-dividing infected cells. A peculiar property of non-primate lentiviruses consists in their incapability to infect and induce diseases in humans, thus providing the main rationale for deriving biologically safe lentiviral vectors for gene therapy applications. In this review, we first give an overview of non-primate lentiviruses, highlighting their common and distinctive molecular characteristics together with key concepts in the molecular biology of lentiviruses. We next examine the bioengineering strategies leading to the conversion of lentiviruses into recombinant lentiviral vectors, discussing their potential clinical applications in ophthalmological research. Finally, we highlight the invaluable role of animal organisms, including the emerging zebrafish model, in ocular gene therapy based on non-primate lentiviral vectors and in ophthalmology research and vision science in general. Keywords: FIV; EIAV; BIV; JDV; VMV; CAEV; lentiviral vector; gene therapy; ophthalmology; zebrafish 1.
    [Show full text]
  • LENTIVIRUS and LENTIVIRAL VECTORS
    LENTIVIRUS and LENTIVIRAL VECTORS Risk Group: 3 I. Background and Health Hazards Lentiviruses are a subset of retroviruses that have the ability to integrate into host chromosomes and to infect non-dividing cells, and include human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) which can infect humans. Another commonly used lentivirus that is infectious to animals, but not humans, is feline immunodeficiency virus (FIV). Lentiviral vectors consist of recombinant or synthetic nucleic acid sequences and HIV or other lentivirus- based viral packaging and regulatory sequences flanked by either wild-type or chimeric long terminal repeat (LTR) regions. Use of these vector systems is particularly desirable because of their ability to integrate transgenes into dividing, as well as, non-dividing cells. However, there are risks associated with working with lentiviral vectors and these must be carefully evaluated. The two major risks are: 1) the potential generation of replication competent lentivirus (RCL); and 2) the potential for oncogenesis. These risks are largely based upon the vector system used and the transgene insert encoded by the vector. Therefore, it is imperative that prior to utilizing a lentiviral vector system, a risk assessment must be reviewed and documented. Also, because construction and/or use of lentiviral vectors falls under the definition of r/sNA research as outlined in the NIH Guidelines, possession must be reported in the workplace’s biological agent inventory and experiments must be submitted for review and approval by the site IBC prior to initiation. II. MODES OF TRANSMISSION Lentivirus is transmissible through injection, ingestion, exposure to broken skin or contact with mucous membranes of the eyes, nose and mouth.
    [Show full text]
  • Advances in the Study of Transmissible Respiratory Tumours in Small Ruminants Veterinary Microbiology
    Veterinary Microbiology 181 (2015) 170–177 Contents lists available at ScienceDirect Veterinary Microbiology journa l homepage: www.elsevier.com/locate/vetmic Advances in the study of transmissible respiratory tumours in small ruminants a a a a,b a, M. Monot , F. Archer , M. Gomes , J.-F. Mornex , C. Leroux * a INRA UMR754-Université Lyon 1, Retrovirus and Comparative Pathology, France; Université de Lyon, France b Hospices Civils de Lyon, France A R T I C L E I N F O A B S T R A C T Sheep and goats are widely infected by oncogenic retroviruses, namely Jaagsiekte Sheep RetroVirus (JSRV) Keywords: and Enzootic Nasal Tumour Virus (ENTV). Under field conditions, these viruses induce transformation of Cancer differentiated epithelial cells in the lungs for Jaagsiekte Sheep RetroVirus or the nasal cavities for Enzootic ENTV Nasal Tumour Virus. As in other vertebrates, a family of endogenous retroviruses named endogenous Goat JSRV Jaagsiekte Sheep RetroVirus (enJSRV) and closely related to exogenous Jaagsiekte Sheep RetroVirus is present Lepidic in domestic and wild small ruminants. Interestingly, Jaagsiekte Sheep RetroVirus and Enzootic Nasal Respiratory infection Tumour Virus are able to promote cell transformation, leading to cancer through their envelope Retrovirus glycoproteins. In vitro, it has been demonstrated that the envelope is able to deregulate some of the Sheep important signaling pathways that control cell proliferation. The role of the retroviral envelope in cell transformation has attracted considerable attention in the past years, but it appears to be highly dependent of the nature and origin of the cells used. Aside from its health impact in animals, it has been reported for many years that the Jaagsiekte Sheep RetroVirus-induced lung cancer is analogous to a rare, peculiar form of lung adenocarcinoma in humans, namely lepidic pulmonary adenocarcinoma.
    [Show full text]
  • The Expression of Human Endogenous Retroviruses Is Modulated by the Tat Protein of HIV‐1
    The Expression of Human Endogenous Retroviruses is modulated by the Tat protein of HIV‐1 by Marta Jeannette Gonzalez‐Hernandez A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy (Immunology) in The University of Michigan 2012 Doctoral Committee Professor David M. Markovitz, Chair Professor Gary Huffnagle Professor Michael J. Imperiale Associate Professor David J. Miller Assistant Professor Akira Ono Assistant Professor Christiane E. Wobus © Marta Jeannette Gonzalez‐Hernandez 2012 For my family and friends, the most fantastic teachers I have ever had. ii Acknowledgements First, and foremost, I would like to thank David Markovitz for his patience and his scientific and mentoring endeavor. My time in the laboratory has been an honor and a pleasure. Special thanks are also due to all the members of the Markovitz laboratory, past and present. It has been a privilege, and a lot of fun, to work near such excellent scientists and friends. You all have a special place in my heart. I would like to thank all the members of my thesis committee for all the valuable advice, help and jokes whenever needed. Our collaborators from the Bioinformatics Core, particularly James Cavalcoli, Fan Meng, Manhong Dai, Maureen Sartor and Gil Omenn gave generous support, technical expertise and scientific insight to a very important part of this project. Thank you. Thanks also go to Mariana Kaplan’s and Akira Ono’s laboratory for help with experimental designs and for being especially generous with time and reagents. iii Table of Contents Dedication ............................................................................................................................ ii Acknowledgements ............................................................................................................. iii List of Figures ...................................................................................................................
    [Show full text]
  • HIV 101: Pathogenesis and Treatment
    HIV 101: Pathogenesis and Treatment Stephen Raffanti MD MPH Professor of Medicine Vanderbilt University School of Medicine . Dr. Raffanti has no financial disclosures to make. Objectives . After this presentation the attendee should be able to: . Describe current issues in the epidemiology of the HIV epidemic. Describe the life cycle of HIV; . Describe the way that HIV interacts with its host, producing disease. Describe the principles of treatment. Describe Acute HIV infection . Describe the initial evaluation of an HIV infected patient Testing, linkage to care, effective treatment and effective PrEP could stop the epidemic today. Percentages of Diagnoses of HIV Infection among Adults and Adolescents, by Region and Population of Area of Residence, 2015—United States Note. Data include persons with a diagnosis of HIV infection regardless of stage of disease at diagnosis. Data for the year 2015 are preliminary and based on 6 months reporting delay. Data exclude persons whose county of residence is unknown. Percentages of Diagnoses of HIV Infection among Adults and Adolescents, by Population of Area of Residence and Age at Diagnosis, 2015—United States Note. Data include persons with a diagnosis of HIV infection regardless of stage of disease at diagnosis. Data for the year 2015 are preliminary and based on 6 months reporting delay. Data exclude persons whose county of residence is unknown. Diagnoses of HIV Infection among Men Who Have Sex with Men, by Age Group, 2010–2014—United States and 6 Dependent Areas Note. Data include persons with a diagnosis of HIV infection regardless of stage of disease at diagnosis. All displayed data have been statistically adjusted to account for reporting delays and missing transmission category, but not for incomplete reporting.
    [Show full text]
  • Lentivirus Vector Fact Sheet
    LENTIVIRUS VECTOR FACT SHEET LENTIVIRUSES: Lentiviral vector constructs are derived from HIV and are therefore highly efficient vehicles for in vivo gene delivery. Use of these vector systems is particularly desirable because of their ability to integrate transgenes into dividing, as well as, non-dividing cells. However, there are risks associated with working with lentiviral vectors and these must be carefully evaluated. The two major risks are: 1) the potential generation of replication competent virus; and 2) the potential for oncogenesis through insertional mutagenesis. These risks are largely based upon the vector system used and the transgene insert encoded by the vector. Therefore, it is imperative that prior to utilizing a lentiviral vector system, a risk assessment must be completed and documented. Also, because construction and/or use of lentiviral vectors falls under the definition of rDNA research as outlined in the NIH Recombinant DNA Guidelines, possession must be reported in the workplace’s biological agent inventory and experiments must be submitted for review and approval by the site IBC prior to initiation. Typically, lentiviral vectors may be safely handled using either BSL-2 or BSL-2 enhanced controls depending upon the risk assessment. LENTIVIRAL VECTORS: Lentiviruses can infect not only dividing, but also non-dividing host cells, because their pre-integration complex (virus “shell”) can get through the intact membrane of the nucleus of the target cell. Lentiviral vectors can be used to provide highly effective gene therapy as they can provide long-term expression of the vectored transgene in target cells. First-generation lentiviral vectors were manufactured using a single vector packaging system that contained all HIV genes, except the envelope (env) gene, in one plasmid.
    [Show full text]
  • VMC 321: Systematic Veterinary Virology Retroviridae Retro: from Latin Retro,"Backwards”
    VMC 321: Systematic Veterinary Virology Retroviridae Retro: from Latin retro,"backwards” - refers to the activity of reverse RETROVIRIDAE transcriptase and the transfer of genetic information from RNA to DNA. Retroviruses Viral RNA Viral DNA Viral mRNA, genome (integrated into host genome) Reverse (retro) transfer of genetic information Usually, well adapted to their hosts Endogenous retroviruses • RNA viruses • single stranded, positive sense, enveloped, icosahedral. • Distinguished from all other RNA viruses by presence of an unusual enzyme, reverse transcriptase. Retroviruses • Retro = reversal • RNA is serving as a template for DNA synthesis. • One genera of veterinary interest • Alpharetrovirus • • Family - Retroviridae • Subfamily - Orthoretrovirinae [Ortho: from Greek orthos"straight" • Genus -. Alpharetrovirus • Genus - Betaretrovirus Family- • Genus - Gammaretrovirus • Genus - Deltaretrovirus Retroviridae • Genus - Lentivirus [ Lenti: from Latin lentus, "slow“ ]. • Genus - Epsilonretrovirus • Subfamily - Spumaretrovirinae • Genus - Spumavirus Retroviridae • Subfamily • Orthoretrovirinae • Genus • Alpharetrovirus Alpharetrovirus • Species • Avian leukosis virus(ALV) • Rous sarcoma virus (RSV) • Avian myeloblastosis virus (AMV) • Fujinami sarcoma virus (FuSV) • ALVs have been divided into 10 envelope subgroups - A , B, C, D, E, F, G, H, I & J based on • host range Avian • receptor interference patterns • neutralization by antibodies leukosis- • subgroup A to E viruses have been divided into two groups sarcoma • Noncytopathic (A, C, and E) • Cytopathic (B and D) virus (ALV) • Cytopathic ALVs can cause a transient cytotoxicity in 30- 40% of the infected cells 1. The viral envelope formed from host cell membrane; contains 72 spiked knobs. 2. These consist of a transmembrane protein TM (gp 41), which is linked to surface protein SU (gp 120) that binds to a cell receptor during infection. 3. The virion has cone-shaped, icosahedral core, Structure containing the major capsid protein 4.
    [Show full text]
  • Effects of Retroviruses on Host Genome Function
    ANRV361-GE42-20 ARI 1 August 2008 18:2 V I E E W R S I E N C N A D V A Effects of Retroviruses on Host Genome Function Patric Jern and John M. Coffin Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts 02111; email: [email protected], John.Coffi[email protected] Annu. Rev. Genet. 2008. 42:20.1–20.23 Key Words The Annual Review of Genetics is online at Human Endogenous Retrovirus, LTR, transcription, recombination, genet.annualreviews.org methylation This article’s doi: 10.1146/annurev.genet.42.110807.091501 Abstract Copyright c 2008 by Annual Reviews. For millions of years, retroviral infections have challenged vertebrates, All rights reserved occasionally leading to germline integration and inheritance as ERVs, 0066-4197/08/1201-0001$20.00 genetic parasites whose remnants today constitute some 7% to 8% of the human genome. Although they have had significant evolutionary side effects, it is useful to view ERVs as fossil representatives of retro- viruses extant at the time of their insertion into the germline, not as direct players in the evolutionary process itself. Expression of particu- lar ERVs is associated with several positive physiological functions as well as certain diseases, although their roles in human disease as etio- logical agents, possible contributing factors, or disease markers—well demonstrated in animal models—remain to be established. Here we discuss ERV contributions to host genome structure and function, in- cluding their ability to mediate recombination, and physiological effects on the host transcriptome resulting from their integration, expression, and other events.
    [Show full text]
  • Protocol and Reagents for Pseudotyping Lentiviral Particles with SARS-Cov-2 Spike Protein for Neutralization Assays
    bioRxiv preprint doi: https://doi.org/10.1101/2020.04.20.051219; this version posted April 20, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY 4.0 International license. 1 of 15 Protocol and reagents for pseudotyping lentiviral particles with SARS-CoV-2 Spike protein for neutralization assays Katharine H.D. Crawford 1,2,3, Rachel Eguia 1, Adam S. Dingens 1, Andrea N. Loes 1, Keara D. Malone 1, Caitlin R. Wolf 4, Helen Y. Chu 4, M. Alejandra Tortorici 5,6, David Veesler 5, Michael Murphy 7, Deleah Pettie 7, Neil P. King 5,7, Alejandro B. Balazs 8, and Jesse D. Bloom 1,2,9,* 1 Division of Basic Sciences and Computational Biology Program, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA; [email protected] (K.D.C.), [email protected] (R.E.), [email protected] (A.S.D.), [email protected] (K.M.), [email protected] (A.N.L.) 2 Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA 3 Medical Scientist Training Program, University of Washington, Seattle, WA 98195, USA 4 Division of Allergy and Infectious Diseases, University of Washington, Seattle, WA 98195, USA; [email protected] (C.R.W.), [email protected] (H.Y.C.) 5 Department of Biochemistry, University of Washington, Seattle, WA 98109, USA; [email protected] (M.A.T.), [email protected] (D.V.), [email protected] (M.M.), [email protected] (D.P.), [email protected] (N.P.K.) 6 Institute
    [Show full text]
  • Lentivirus-Mediated Gene Transfer to the Respiratory Epithelium: a Promising Approach to Gene Therapy of Cystic fibrosis
    Gene Therapy (2004) 11, S67–S75 & 2004 Nature Publishing Group All rights reserved 0969-7128/04 $30.00 www.nature.com/gt REVIEW Lentivirus-mediated gene transfer to the respiratory epithelium: a promising approach to gene therapy of cystic fibrosis E Copreni, M Penzo, S Carrabino and M Conese Institute for Experimental Treatment of Cystic Fibrosis, HS Raffaele, Milano, Italy Gene therapy of cystic fibrosis (CF) lung disease needs logous envelopes are the strategies currently used to highly efficient delivery and long-lasting complementation overcome the paucity of specific viral receptors on the apical of the CFTR (cystic fibrosis transmembrane conductance surface of airway epithelial cells and to reach the basolateral regulator) gene into the respiratory epithelium. The develop- surface receptors. Preclinical studies on CF mice, demon- ment of lentiviral vectors has been a recent advance in the strating complementation of the CF defect, offer hope that field of gene transfer and therapy. These integrating vectors lentivirus gene therapy can be translated into an effective appear to be promising vehicles for gene delivery into treatment of CF lung disease. Besides a direct targeting of respiratory epithelial cells by virtue of their ability to infect the stem/progenitor niche(s) in the CF airways, an alternative nondividing cells and mediate long-term persistence of approach may envision homing of hematopoietic stem cells transgene expression. Studies in human airway tissues and engineered to express the CFTR gene by lentiviral vectors. animal models have highlighted the possibility of achieving In the context of lentivirus-mediated CFTR gene transfer gene expression by lentiviral vectors, which outlasted the to the CF airways, biosafety aspects should be of primary normal lifespan of the respiratory epithelium, indicating concern.
    [Show full text]
  • Lentiviral Integration Site Targeting: Host Determinants and Consequences
    University of Pennsylvania ScholarlyCommons Publicly Accessible Penn Dissertations Spring 2010 Lentiviral Integration Site Targeting: Host Determinants and Consequences Keshet Ronen University of Pennsylvania, [email protected] Follow this and additional works at: https://repository.upenn.edu/edissertations Part of the Virology Commons Recommended Citation Ronen, Keshet, "Lentiviral Integration Site Targeting: Host Determinants and Consequences" (2010). Publicly Accessible Penn Dissertations. 174. https://repository.upenn.edu/edissertations/174 This paper is posted at ScholarlyCommons. https://repository.upenn.edu/edissertations/174 For more information, please contact [email protected]. Lentiviral Integration Site Targeting: Host Determinants and Consequences Abstract A necessary step in the retroviral lifecycle is integration, the covalent insertion of the viral cDNA into the genome of the infected cell. This means that retroviruses, for example HIV, establish life-long infection. It also means that retroviruses are used as gene-delivery vectors to treat genetic diseases. Integration events are distributed non-randomly in the genome of the infected cell, with characteristic genus-specific preferences. In this dissertation, we focus on the lentiviral class of retroviruses, and explore two aspects of their integration: the means by which integration is targeted to its favored sites, and the consequences of integration at these sites for the host cell. The host protein LEDGF/p75 has been shown to interact with lentiviral integrases and contribute to their preference for integration in genes. We sought to establish the extent to which integration site selection is determined by LEDGF/p75 tethering. We first asked whether LEDGF/p75 was an essential integration tether, by analyzing integration site distribution in cells stringently depleted for LEDGF/p75.
    [Show full text]
  • Preliminary PCR-Based Screening Indicates a Higher Incidence of Porcine Endogenous Retrovirus Subtype C (PERV-C) in Feral Versus Domestic Swine
    Fort Hays State University FHSU Scholars Repository Biology Faculty Publications Biology 2019 Preliminary PCR-based screening indicates a higher incidence of Porcine Endogenous Retrovirus subtype C (PERV-C) in feral versus domestic swine Rashmi Acharya Zoey K. Wallis Robert Keener Eric T. Gillock Follow this and additional works at: https://scholars.fhsu.edu/biology_facpubs Part of the Biology Commons TRANSACTIONS OF THE KANSAS Vol. 122, no. 3-4 ACADEMY OF SCIENCE p. 257-263 (2019) Preliminary PCR-based screening indicates a higher incidence of Porcine Endogenous Retrovirus subtype C (PERV-C) in feral versus domestic swine RASHMI ACHARYA1, ZOEY K. WALLIS1, ROBERT J. KEENER2 AND ERIC T. GILLOCK1 1. Department of Biological Sciences, Fort Hays State University, Hays, Kansas [email protected] 2. Department of Agriculture, Fort Hays State University, Hays, Kansas Xenotransplantation is considered a potential alternative to allotransplantation to relieve the current shortage of human organs. Due to their similar size and physiology, the organs of pigs are of particular interest for this purpose. Endogenous retroviruses are a result of integration of retroviral genomes into the genome of infected germ cells as DNA proviruses, which are then carried in all cells of the offspring of the organism. Porcine endogenous retroviruses (PERVs) are of special concern because they are found in pig organs and tissues that might otherwise be used for xenotransplantation. PERV proviruses can be induced to replicate and recombine in pigs, and have been shown to infect human cells in vitro. There are three subtypes of PERVs based on differences in the receptor binding domain of the env protein; PERV-A, PERV-B, and PERV-C.
    [Show full text]