Relevant bibliographies by topics / Lentiviral vector

Academic literature on the topic 'Lentiviral vector'

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Published: 4 June 2021
Last updated: 10 March 2023

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Journal articles on the topic "Lentiviral vector"

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Yew, Chee-Hong Takahiro, Narmatha Gurumoorthy, Fazlina Nordin, Gee Jun Tye, Wan Safwani Wan Kamarul Zaman, Jun Jie Tan, and Min Hwei Ng. "Integrase deficient lentiviral vector: prospects for safe clinical applications." PeerJ 10 (August 12, 2022): e13704. http://dx.doi.org/10.7717/peerj.13704.

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HIV-1 derived lentiviral vector is an efficient transporter for delivering desired genetic materials into the targeted cells among many viral vectors. Genetic material transduced by lentiviral vector is integrated into the cell genome to introduce new functions, repair defective cell metabolism, and stimulate certain cell functions. Various measures have been administered in different generations of lentiviral vector systems to reduce the vector’s replicating capabilities. Despite numerous demonstrations of an excellent safety profile of integrative lentiviral vectors, the precautionary approach has prompted the development of integrase-deficient versions of these vectors. The generation of integrase-deficient lentiviral vectors by abrogating integrase activity in lentiviral vector systems reduces the rate of transgenes integration into host genomes. With this feature, the integrase-deficient lentiviral vector is advantageous for therapeutic implementation and widens its clinical applications. This short review delineates the biology of HIV-1-erived lentiviral vector, generation of integrase-deficient lentiviral vector, recent studies involving integrase-deficient lentiviral vectors, limitations, and prospects for neoteric clinical use.
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Li, Chen, Biao Qian, Zhao Ni, Qinzhang Wang, Zixiong Wang, Luping Ma, Zhili Liu, Qiang Li, and Xinmin Wang. "Construction of recombinant lentiviral vector containing human stem cell leukemia gene and its expression in interstitial cells of cajal." Open Life Sciences 15, no. 1 (March 25, 2020): 83–91. http://dx.doi.org/10.1515/biol-2020-0010.

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AbstractThis study aims to construct recombinant lentiviral vectors containing the human stem cell leukemia (SCL) gene and investigate their in vitro transfection efficiency in Interstitial Cells of Cajal (ICC) of guinea pig bladders. In this study, the human SCL gene was successfully cloned, and the recombinant lentivirus GV287-SCL was successfully constructed. The titer of the recombinant lentivirus was 5 × 108 TU /mL. After transfecting the ICCs with the lentiviral vector at different MOIs, the optimal MOI was determined to be 10.0, and the optimal transfection time was determined to be 3 days. The amplification product of the lentivirus transfection group was consistent with the target fragment, indicating that the SCL gene had been successfully introduced into ICCs. In conclusion, the recombinant lentiviral vector GV287-SCL was successfully constructed and transfected into the in vitro cultured ICCs. The successful expression of SCL in ICCs may provide an experimental basis for the in vivo transfection of the SCL gene.
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Perry, Christopher, and Andrea C. M. E. Rayat. "Lentiviral Vector Bioprocessing." Viruses 13, no. 2 (February 9, 2021): 268. http://dx.doi.org/10.3390/v13020268.

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Lentiviral vectors (LVs) are potent tools for the delivery of genes of interest into mammalian cells and are now commonly utilised within the growing field of cell and gene therapy for the treatment of monogenic diseases and adoptive therapies such as chimeric antigen T-cell (CAR-T) therapy. This is a comprehensive review of the individual bioprocess operations employed in LV production. We highlight the role of envelope proteins in vector design as well as their impact on the bioprocessing of lentiviral vectors. An overview of the current state of these operations provides opportunities for bioprocess discovery and improvement with emphasis on the considerations for optimal and scalable processing of LV during development and clinical production. Upstream culture for LV generation is described with comparisons on the different transfection methods and various bioreactors for suspension and adherent producer cell cultivation. The purification of LV is examined, evaluating different sequences of downstream process operations for both small- and large-scale production requirements. For scalable operations, a key focus is the development in chromatographic purification in addition to an in-depth examination of the application of tangential flow filtration. A summary of vector quantification and characterisation assays is also presented. Finally, the assessment of the whole bioprocess for LV production is discussed to benefit from the broader understanding of potential interactions of the different process options. This review is aimed to assist in the achievement of high quality, high concentration lentiviral vectors from robust and scalable processes.
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Wang, Nan, Narendiran Rajasekaran, Tieying Hou, and Elizabeth Mellins. "Comparison of protein expression by different lentiviral vectors (51.12)." Journal of Immunology 188, no. 1_Supplement (May 1, 2012): 51.12. http://dx.doi.org/10.4049/jimmunol.188.supp.51.12.

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Abstract HIV1-derived lentiviral vectors have been widely used as gene delivery tools due to their potent transduction capacity and stable expression after chromosomal integration in dividing and non-dividing mammalian cells. Lentiviral vectors were screened for expressing the murine class II chaperone, invariant chain (Ii), in hematopoietic stem and progenitor cells (HSPC). We compared various lentiviral vectors using GFP as a reporter gene in 293T cells and HSPC. We assessed a dual promoter vector (DP) with separate promoters for Ii and GFP, a fusion protein vector (FU) that expresses Ii and GFP together under a single promoter, and T2A vector (T2A) in which Ii and GFP are separated by a self-cleaving 2A peptide. Similar percentages of 293T cells were positive for Ii expression after being directly transfected with DP and T2A. The percentage was slightly higher in FU transfected cells. 293T cells transfected with DP have the highest GFP expression. We packaged each vector into lentivirus and found that both Ii and GFP expression levels dropped in 293T cells. Further reduction in expression was seen in viral transduced HSPC. The trend toward different expression levels of Ii and GFP persisted whether lentiviral transduction or vector transfection was used. Only DP and T2A constructs, but not FU constructs, resulted in functional Ii expression. Based on these results, we hypothesize that the poor expression of Ii may be due to deleterious effects of Ii on assembly of competent virus.
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Nguyen, Tuan Huy, Tatiana Khakhoulina, Andrew Simmons, Philippe Morel, and Didier Trono. "A Simple and Highly Effective Method for the Stable Transduction of Uncultured Porcine Hepatocytes Using Lentiviral Vector." Cell Transplantation 14, no. 7 (August 2005): 489–96. http://dx.doi.org/10.3727/000000005783982828.

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Gene therapy is an attractive approach for the treatment of a wide spectrum of liver diseases. Lentiviral vectors allow the stable integration of transgenes into the genome of nondividing differentiated cells including hepatocytes and could provide long-lasting expression of a therapeutic gene. To develop such approaches, preclinical studies in large animal models such as pigs are necessary to evaluate the feasibility and safety of stable lentiviral integration and long-term vector expression. In addition, effective lentivector-mediated gene transfer onto porcine hepatocytes could advance in cell-based therapies for acute liver failure. To investigate this issue, porcine hepatocytes were transduced in suspension immediately after their isolation in University of Wisconsin (UW) solution containing vitamin E. Up to 80% of hepatocytes stably expressed a GFP transgene after a single exposure to lentiviral vector coding for GFP under the control of either liver-specific or ubiquitous promoters. Moreover, porcine hepatocytes cryopreserved in UW solution containing fetal bovine serum, dimethyl sulfoxide, and vitamin E remained highly transducible with lentiviral vector after thawing. When thawed, transduced in suspension, and immediately transplanted into the spleen of immunodeficient mice, ex vivo lentivirally transgene marked xenogeneic hepatocytes were detected in murine liver. We demonstrated that porcine hepatocytes are highly susceptible to lentiviral vector and describe an easy methodology to efficiently, rapidly, and stably introduce transgenes into uncultured porcine hepatocytes.
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Kubo, Shuji, and Kohnosuke Mitani. "A New Hybrid System Capable of Efficient Lentiviral Vector Production and Stable Gene Transfer Mediated by a Single Helper-Dependent Adenoviral Vector." Journal of Virology 77, no. 5 (March 1, 2003): 2964–71. http://dx.doi.org/10.1128/jvi.77.5.2964-2971.2003.

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ABSTRACT To achieve efficient and sustained gene expression, we developed a new lentivirus/adenovirus hybrid vector (LA vector) that encodes sequences required for production of a human immunodeficiency virus-based lentiviral vector (i.e., a lentiviral vector, a gag/pol/rev expression cassette, a tetracycline-inducible envelope cassette, and the tetracycline-inducible transcriptional activator cassette) in a single helper-dependent adenovirus vector backbone. Via either transfection or infection, human cell lines transduced with the LA vector produced a lentiviral vector in a doxycycline-dependent manner at titers up to 105 to 106 green fluorescent protein transducing units per ml, which are comparable to the titers obtained by conventional multiple plasmid transfection methods. Efficient spread and persistent expression of the transgene were observed in cells maintained in long-term culture that had been infected with the LA vector. Furthermore, when cocultured with adherent cells infected with the LA vector, the human T-cell leukemia cell line was successfully transduced with a marker gene. This LA vector possesses the advantages of efficient gene transfer from an adenoviral vector and stable integration from a lentiviral vector; therefore, it might have potential for a variety of gene therapy applications.
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Sakuma, Toshie, Michael A. Barry, and Yasuhiro Ikeda. "Lentiviral vectors: basic to translational." Biochemical Journal 443, no. 3 (April 16, 2012): 603–18. http://dx.doi.org/10.1042/bj20120146.

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More than two decades have passed since genetically modified HIV was used for gene delivery. Through continuous improvements these early marker gene-carrying HIVs have evolved into safer and more effective lentiviral vectors. Lentiviral vectors offer several attractive properties as gene-delivery vehicles, including: (i) sustained gene delivery through stable vector integration into host genome; (ii) the capability of infecting both dividing and non-dividing cells; (iii) broad tissue tropisms, including important gene- and cell-therapy-target cell types; (iv) no expression of viral proteins after vector transduction; (v) the ability to deliver complex genetic elements, such as polycistronic or intron-containing sequences; (vi) potentially safer integration site profile; and (vii) a relatively easy system for vector manipulation and production. Accordingly, lentivector technologies now have widespread use in basic biology and translational studies for stable transgene overexpression, persistent gene silencing, immunization, in vivo imaging, generating transgenic animals, induction of pluripotent cells, stem cell modification and lineage tracking, or site-directed gene editing. Moreover, in the present high-throughput ‘-omics’ era, the commercial availability of premade lentiviral vectors, which are engineered to express or silence genome-wide genes, accelerates the rapid expansion of this vector technology. In the present review, we assess the advances in lentiviral vector technology, including basic lentivirology, vector designs for improved efficiency and biosafety, protocols for vector production and infection, targeted gene delivery, advanced lentiviral applications and issues associated with the vector system.
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Breckpot, Karine, David Escors, Frederick Arce, Lucienne Lopes, Katarzyna Karwacz, Sandra Van Lint, Marleen Keyaerts, and Mary Collins. "HIV-1 Lentiviral Vector Immunogenicity Is Mediated by Toll-Like Receptor 3 (TLR3) and TLR7." Journal of Virology 84, no. 11 (March 17, 2010): 5627–36. http://dx.doi.org/10.1128/jvi.00014-10.

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ABSTRACT Lentiviral vectors are promising vaccine vector candidates that have been tested extensively in preclinical models of infectious disease and cancer immunotherapy. They are also used in gene therapy clinical trials both for the ex vivo modification of cells and for direct in vivo injection. It is therefore critical to understand the mechanism(s) by which such vectors might stimulate the immune system. We evaluated the effect of lentiviral vectors on myeloid dendritic cells (DC), the main target of lentiviral transduction following subcutaneous immunization. The activation of DC cultures was independent of the lentiviral pseudotype but dependent on cell entry and reverse transcription. In vivo-transduced DC also displayed a mature phenotype, produced tumor necrosis factor alpha (TNF-α), and stimulated naive CD8+ T cells. The lentiviral activation of DC was Toll-like receptor (TLR) dependent, as it was inhibited in TRIF/MyD88 knockout (TRIF/MyD88−/−) DC. TLR3−/− or TLR7−/− DC were less activated, and reverse transcription was important for the activation of TLR7−/− DC. Moreover, lentivirally transduced DC lacking TLR3 or TLR7 had an impaired capacity to induce antigen-specific CD8+ T-cell responses. In conclusion, we demonstrated TLR-dependent DC activation by lentiviral vectors, explaining their immunogenicity. These data allow the rational development of strategies to manipulate the host's immune response to the transgene.
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Park, Frank. "Lentiviral vectors: are they the future of animal transgenesis?" Physiological Genomics 31, no. 2 (October 2007): 159–73. http://dx.doi.org/10.1152/physiolgenomics.00069.2007.

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Lentiviral vectors have become a promising new tool for the establishment of transgenic animals and the manipulation of the mammalian genome. While conventional microinjection-based methods for transgenesis have been successful in generating small and large transgenic animals, their relatively low transgenic efficiency has opened the door for alternative approaches, including lentiviral vectors. Lentiviral vectors are an appealing tool for transgenesis in part because of their ability to incorporate into genomic DNA with high efficiency, especially in cells that are not actively dividing. Lentiviral vector-mediated transgene expression can also be maintained for long periods of time. Recent studies have documented high efficiencies for lentiviral transgenesis, even in animal species and strains, such as NOD/ scid and C57Bl/6 mouse, that are very difficult to manipulate using the standard transgenic techniques. These advantages of the lentiviral vector system have broadened its use as a gene therapy vector to additional applications that include transgenesis and knockdown functional genetics. This review will address the components of the lentiviral vector system and recent successes in lentiviral transgenesis using both male- and female-derived pluripotent cells. The advantages and disadvantages of lentiviral transgenesis vs. other approaches to produce transgenic animals will be compared with regard to efficiency, the ability to promote persistent transgene expression, and the time necessary to generate a sufficient number of animals for phenotyping.
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Lucke, Susann, Thomas Grunwald, and Klaus Überla. "Reduced Mobilization of Rev-Responsive Element-Deficient Lentiviral Vectors." Journal of Virology 79, no. 14 (July 2005): 9359–62. http://dx.doi.org/10.1128/jvi.79.14.9359-9362.2005.

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ABSTRACT Infection of cells transduced with a lentiviral vector by human immunodeficiency virus (HIV) could lead to packaging of the lentiviral vector RNA into HIV particles and unintended transfer of the vector. To prevent this, the Rev-responsive element (RRE) of an HIV-1 vector was functionally replaced by a heterologous RNA element (MS2). Providing Rev fused to an MS2 binding protein allowed efficient vector production. Mobilization of the vector from infected target cells was below the level of detection and at least 103- to 104-fold lower than for the RRE-containing vector. Thus, RRE-deficient lentiviral vectors provide a novel approach to reduce the risk of vector mobilization.
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Dissertations / Theses on the topic "Lentiviral vector"

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Trimby, Christopher Matthew. "STRATEGIES FOR TARGETING LENTIVIRAL VECTORS." UKnowledge, 2011. http://uknowledge.uky.edu/gradschool_diss/157.

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Lentiviral gene therapy has held great promise for treating a wide range of neurological disorders due to its ability to stably integrate into the genome of nondividing cells like neurons, in addition to dividing cells. The nervous system is a complex and highly heterogeneous system, and while a therapeutic intervention may have beneficial effects in one population of cells it may have severe side effects in another. For this reason, specific targeting of lentiviral vectors is crucial for their ultimate utility for research and clinical research use. Two different approaches for focusing the targeting of lentiviral vectors were employed in these studies. The first method involved assessing the effects of vector production strategies on the resulting virus’s tropism both in vivo and in vitro. The changes in vector transduction were determined via flow cytometry on cells in culture and immunohistochemistry following brain injections. Results from these experiments suggest that while the production conditions do impact the vectors efficacy, there is not a distinct effect on their tropism. A unique characteristic of retroviral and lentiviral vectors is their capacity for being pseudotyped, conferring a new tropism on the vector. Native tropisms are generally not specific beyond very broad cell types, which may not be sufficient for all applications. In this case, chimeric targeting molecules can provide an even more refined targeting profile compared to native pseudotypes. The second approach utilizes novel chimeric glycoproteins made from nerve growth factor and the vesicular stomatitis virus glycoprotein. These chimeras are designed to pseudotype lentiviral vectors to target nociceptive sensory neurons for a variety of disorders. While these chimeras were successfully produced as protein, they were misfolded and sequestered in the endoplasmic reticulum and therefore unavailable to produce lentivirus. While neither strategy was completely successful, they do provide interesting information for the design and creation of lentiviral vectors. This research shows that small differences in the steps followed as part of a lentivirus production protocol can greatly impact the resulting vectors efficacy. It also shows that while VSV has been used to create chimeric glycoproteins, not all targeting molecules are suitable for this purpose.
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Ingrao, Dina. "Etude de l'étape d'entrée des vecteurs lentiviraux dérivés du VIH-1 dans les cellules hématopoïétiques humaines." Thesis, Evry-Val d'Essonne, 2013. http://www.theses.fr/2013EVRY0021/document.

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Les vecteurs lentiviraux (LV) sont des outils efficaces de transfert de gène, largement utilisés en thérapie génique, en particulier pour la transduction ex vivo de cellules souches et progénitrices hématopoïétiques (CSPH). Afin d’étudier simultanément la fusion et la transduction dans les CSPH avec les LV, nous avons adapté une méthode basée sur latechnologie du transfert d’énergie entre deux molécules fluorescentes (FRET). Pour mettre en place cette technique, des LV capables d’incorporer spécifiquement une enzyme, la bétalactamase (BLAM-LV) et de coder une forme tronquée du récepteur au facteur de croissance nerveuse (DELTA-NGFR), sont produits. Nos résultats montrent que les LV sont soumis à une restriction post-entrée forte dans les cellules hématopoïétiques, que ce soit dans des lymphocytes T immortalisés ou bien des CSH CD34+. Nous montrons également que cette inhibition post-entrée peut être partiellement saturée après une forte augmentation de la multiplicité d’infection ou en présence d’additifs de culture, comme la Vectofusin-1® ou laRetronectin®. De plus, nous avons montré lors de la transduction de CSPH avec des vecteurs BLAM-LV que la Vectofusin-1® agit sur l’étape d’entrée en augmentant l’adhésion et la fusion entre les membranes virale et cellulaire. Cette technique représente donc un nouvel outil sensible et efficace pour étudier de façon concomitante l’étape de fusion et le niveau de transduction dans les cellules cibles. A terme, ce travail permettra une meilleure compréhension de la biologie des LV mais pourra également conduire à l’élaboration de protocoles de transduction lentivirale plus efficaces
Lentiviral vectors (LV) are used for various gene transfer applications, notably for hematopoietic gene therapy, but methods are lacking to precisely evaluate parameters that control the efficiency of transduction in relation with the entry of vectors into target cells. We adapted a fluorescence resonance energy transfer (FRET)-based HIV-1 fusion assay to measure the entry of non-replicative recombinant LV in various cell types, including primary human hematopoietic stem and progenitor cells, and to quantify the level of transduction of he same initially-infected cells. The assay utilizes recombinant LV containing betalactamase (BLAM)-Vpr chimeric proteins (BLAM-LV) and encoding a truncated form of thelow affinity nerve growth factor receptor (DELTA-NGFR). This LV-based fusion/transduction assay is a dynamic and versatile tool, revealing for instance the extent of lentiviral post-entry restrictions occuring in cells of hematopoietic origin. The assay also shows that transduction enhancers like Vectofusin®-1 or Retronectin® can partially relieve this post-entry block but their effects differ in the way to promote LV entry. Furthermore, our results show that Vectofusin®-1 acts at the entry step by promoting the adhesion and the fusion between lentiviral and cellular membranes. In conclusion, one such assay should be useful to study hematopoietic post-entry restrictions directed against LV and should allow improvements in various LV-based gene therapy protocols
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Thomas, Joan Helen. "Studies in gene transfer using pseudotyped lentiviral vector systems." Thesis, University of Cambridge, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.621818.

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Gelinas, Jean-Francois. "Enhancement of lentiviral vector production through alteration of virus-cell interactions." Thesis, University of Oxford, 2016. https://ora.ox.ac.uk/objects/uuid:9921b8b4-e2b5-4eec-9efc-6036765c8d55.

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Gene therapy is the introduction or alteration of genetic material with the intention to treat disease. To support this aim, viruses have been modified, with elements linked to viral pathogenicity removed from their genome and replaced by the genetic material to be delivered. Gene therapy vectors based on lentiviruses have many advantages, such as the ability to transduce non-dividing cells and to target specific cell types via pseudotyping. They have been successfully used in ex vivo clinical trials for several haematopoietic stem cell disorders. Lentiviral vectors, however, suffer from substantially lower titres than the more popular adeno-associated virus (AAV)-based vectors and therefore have limited applicability for in vivo gene therapy which requires much greater quantities of virus. The main aim of this thesis was to investigate strategies to improve lentiviral vector productivity during manufacture, in order to increase the likelihood of lentiviruses being adopted for disease treatment. Initial experiments were based on the lentiviral vector manufacturing process currently being developed by the United Kingdom Cystic Fibrosis Gene Therapy Consortium for the generation of highly concentrated, purified lentivirus for clinical use. Supplementation of FreeStyle 293 Expression Medium used during upstream processing was attempted, but none of the assessed supplements led to significant increases in lentiviral vector production. Investigation into intrinsic immunity to viral infection indicated that over-expression of the protein kinase RNA-activated (PKR) led to lower production titres, but over-expression of its inhibitors was not successful at increasing titres. The focus then shifted to reducing, or 'knocking-down', inhibitory factors present in the host cells, which could adversely affect viral titres. Investigation of the published HIV-1 literature revealed a possible 152 candidate inhibitory factors described as having a negative impact on HIV-1 replication in the late stages of the life cycle of the virus. A novel siRNA screen was developed to assess the effect of ‘knock-down' of inhibitory factors on lentiviral vector titre. Application of the screen to 89 candidate inhibitory factors identified nine genes which, when knocked-down, resulted in increased lentiviral vector production by more than 40%. Further work will be necessary to understand the role of the inhibitory factors in lentiviral vector production, but novel cell lines in which genes encoding these factors have been permanently deleted from producer cells could lead to higher titres, reducing costs in the manufacture of lentiviral vectors and making in vivo gene therapy more feasible from a health economics perspective.
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Zhang, Bing. "Lentiviral vector-mediated gene transfer in vitro and in vivo /." St. Lucia, Qld, 2004. http://www.library.uq.edu.au/pdfserve.php?image=thesisabs/absthe18024.pdf.

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Booth, C. A. "Lentiviral vector mediated gene therapy for X-linked lymphoproliferative disease." Thesis, University College London (University of London), 2012. http://discovery.ucl.ac.uk/1356299/.

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X-linked lymphoproliferative disease (XLP) is a rare primary immunodeficiency characterised by severe immune dysregulation and is caused by mutations in the SH2D1A gene. Clinical manifestations vary and include haemophagocytic lymphohistiocytosis (HLH), lymphoma and dysgammaglobulinaemia, often triggered by Epstein-Barr virus (EBV) infection. SLAM-associated protein (SAP) is a key regulator of immune function in T, NK, and NKT cells and defects in this protein lead to the cellular and humoral immune defects described in patients. Treatment options for XLP are limited and currently haematopoietic stem cell transplant (HSCT) is the only curative option. Results are variable and dependent on a good donor match and absence of active infection at transplant. Somatic gene therapy is now successfully used to correct certain severe immunodeficiencies and offers a potential cure in XLP. The use of self-inactivating (SIN) lentiviral vectors with transgene expression driven by non-viral promoters has improved the biosafety profile of haematopoietic stem cell gene therapy procedures. In this study we have successfully corrected both cellular and humoral defects in a SAP deficient murine model using a SIN lentiviral vector with a codon optimised SAP transgene under the transcriptional control of the elongation factor 1α short form (EFS) promoter. Initial attempts with a non-codon optimised version of SAP led to insufficient protein expression levels to restore immune function. We also assessed the CD2 locus control region (LCR) to evaluate any lymphoid specificity to permit more regulated SAP expression but were unable to demonstrate any benefit with this regulatory element. The results presented here provide proof of concept for the development of gene therapy for XLP and further work is warranted to improve the efficiency of gene transfer, secure engraftment of long term repopulating haematopoietic stem cell progenitors and additional characterisation of immune reconstitution after gene therapy.
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Macdonald, D. "Lentiviral vector vaccines for T-cell-mediated protection against influenza." Thesis, University College London (University of London), 2014. http://discovery.ucl.ac.uk/1417882/.

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Vaccines that induce T cells which recognize conserved viral proteins could confer cross-strain protection against pathogens with fast-mutating B cell epitopes. Influenza is an example of such a pathogen for which there is a pressing need for a universal vaccine. Lentiviral vectors are a counterintuitive choice as vaccines since they have low inherent immunogenicity. However, their efficient transduction of non-dividing cells and high capacity permits transduction of antigen presenting cells with not only antigen but also molecular adjuvants that directly or indirectly enhance the T cell response. We therefore investigated the potential of two such adjuvants: viral flice-like inhibitor protein, which activates dendritic cells through nuclear factor kappa-B, and 4-1BB ligand, which activates T cells directly through 4-1BB. By co-encoding these with influenza nucleoprotein, we have shown that the influenza-specific T cell response to lentiviral vector vaccination is significantly enhanced in mice. Furthermore, we have demonstrated that intranasally delivered lentiviral vectors transduce alveolar macrophages with high efficiency, recalling and expanding large and sustained populations of nucleoprotein-specific CD8+ T cells in the lung and airway in mice that have been primed subcutaneously or previously exposed to influenza. These lung-resident T cell populations persist for at least 4 months and are sufficiently abundant to rapidly control a mouse-adapted lethal influenza challenge without invocation of a secondary cytokine response, weight loss or lung injury. Furthermore, dendritic cells expressing 4-1BBL potently trans-activate bystander dendritic cells, both in vitro and in vivo, demonstrating an indirect mechanism by which the 4-1BBL:4-1BB signaling axis can enhance T cell responses.
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Oakland, Mayumi. "Improving lentiviral vector-mediated gene transfer by understanding cellular barriers." Diss., University of Iowa, 2013. https://ir.uiowa.edu/etd/4709.

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Cystic fibrosis (CF) is an autosomal recessive genetic disorder of which lung disease is the leading cause of morbidity and mortality. One attractive strategy for the treatment of CF lung disease is to directly deliver CF transmembrane conductance regulator gene to airway epithelia. Although promising results have been reported, barriers present in the lung make successful gene transfer to the respiratory tract difficult. In order to improve gene transfer strategies in the intrapulmonary airways, we need to identify the bottlenecks of transduction for the vector system. A previous study reported that feline immunodeficiency virus (FIV)-mediated gene transfer was more efficient in the nasal airways in mice than the intrapulmonary airways (Sinn, P.L. et al. 2008, J. Viol). Our first goal was to identify barriers to lentiviral gene transfer in the murine airways. We demonstrate that host immune response is not the major barrier preventing efficient FIV-mediated transduction in the intrapulmonary airways. We show that the FIV vector transduces murine primary nasal epithelial cell cultures with greater efficiency than murine primary tracheal epithelial cell cultures. In addition, GP64 pseudotyped vesicular stomatitis virus (VSV) transduces better in nasal epithelia compared to intrapulmonary airways in mice. On the other hand, we observed that VSVG glycoprotein-pseudotyped VSV transduces the intrapulmonary airway as well as nasal epithelia in mice with similar efficiency. Our results suggest that differentially expressed cellular factor(s) specific for GP64 or FIV vector may be the major barrier(s) for FIV vector-mediated gene transfer in the murine intrapulmonary airways. The recent development of CF porcine models prompted us to investigate possible barriers for lentiviral vector-mediated gene transfer in porcine cells. Our preliminary results showed that HIV transduction was restricted in porcine but not human lung-derived cell lines. Porcine TRIM5 has sequences similar to restrictive bovine TRIM5 orthologs. Therefore, our second goal was to investigate the possible restriction of lentiviral vectors by porcine TRIM5. We demonstrate that transient overexpression or knockdown of endogenously expressed porcine TRIM5 does not affect HIV or FIV transduction. Lastly, we characterized a mucin domain-deleted EBOV (EBOVΔO) glycoprotein mutant with increased transduction. This EBOVΔO 5-mer mutant was generated based on mutants with an increased transduction as identified by alanine scanning mutagenesis (Brindlay, M.A. et al. 2007. J. Viol). We show that VSV pseudotyped with the 5-mer mutant increased transduction both in vitro and in mice when compared to the wild-type EBOVΔO. Structural analysis demonstrated that 5 mutations were located proximal to the GP1-GP2 interface. Enhanced transduction likely results from a lower energy metastable state of the glycoprotein. FIV pseudotyped with 5-mer also shows increased transduction in multiple cell lines. Identification of barriers in intrapulmonary airways and improvements of vector systems will help the advancement of gene therapy for CF.
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Mekkaoui, Leila. "Lentiviral vector purification using genetically encoded biotin mimic in packaging cell." Thesis, University College London (University of London), 2018. http://discovery.ucl.ac.uk/10053191/.

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Lentiviral vectors (LVs) are powerful tools in gene therapy that have recently witnessed an increasing demand in both research and clinical applications. Current LVs purification represents the main bottle neck in their application as several methods are employed which are time consuming, cumbersome and yield low recoveries. The aim of this project was to develop a one-step method to specifically and efficiently purify LVs, with high vector yields and reduced levels of impurities, using the biotin-streptavidin system. Herein, packaging 293T cells were genetically engineered with biotin mimicking synthetic peptides and different cell membrane anchoring strategies for optimal streptavidin binding were tested. We have identified a flanked disulphide-constrained peptide, termed Ctag (ECHPQGPPCIEGRK), displayed on a CD8α stalk to be the most promising. LVs were modified with Ctag by its random incorporation onto viral surfaces during budding, without viral protein engineering or hindrance on infectivity. The expression of Ctag on LVs allowed complete capture of infectious particles by streptavidin magnetic beads. As Ctag binds streptavidin in the nanomolar range, we hypothesised that gentle elution from streptavidin matrix should occur by biotin’s competitive binding. Accordingly, addition of micromolar concentrations of biotin to captured LVs resulted in an overall yield of ≥60%. Analysis of eluted LVs revealed high purity levels, with a ≤3-log and 2-log reduction of DNA contamination and host cell proteins, respectively. This one-step purification was also tested for scalable vector processing using streptavidin monolith affinity chromatography and preliminary results were encouraging with 20% overall yield. In conclusion, we developed a single-step affinity chromatography which allows specific purification and concentration of infectious vectors modified with a biotin mimic. Based on intended usage, efficient LV purification can be achieved using both magnetic beads and column chromatography. This method will be of valuable use for both research and clinical applications of LVs.
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10

FIRRITO, CLAUDIA. "Targeted Gene Correction and Reprogramming of SCID-X1 Fibroblasts to Rescue IL2RG Expression in iPSC-derived Hematopoietic Cells." Doctoral thesis, Università degli Studi di Milano-Bicocca, 2015. http://hdl.handle.net/10281/94656.

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La terapia genica basata sull’utilizzo di vettori integranti è stata già applicata con successo per la cura di varie malattie genetiche come le malattie da accumulo lisosomiale (LSD), la beta-talassemia (β-Thal) e le immunodeficienze primarie (PID). L’immunodeficienza combinata grave legata al cromosoma X (SCID-X1) è una malattia monogenica letale causata da mutazioni del gene codificante la catena comune gamma del recettore per l’interleuchina 2 (IL2RG). I primi studi clinici per la SCID-X1 hanno mostrato il potenziale terapeutico della terapia genica basata su vettori integranti, risultando nella ricostituzione del compartimento linfoide grazie al vantaggio selettivo delle cellule geneticamente modificate. D’altra parte, tali studi hanno evidenziato il rischio di mutagenesi inserzionale dovuto all’integrazione casuale del virus nel genoma della cellula ospite e all’espressione non regolata del transgene, sottolineando la necessità di sviluppare nuove strategie di terapia genica più sicure. In questo lavoro, sfruttando la tecnologia delle Zinc-Finger Nucleasi (ZFN) per indurre una rottura del doppio filamento del DNA in maniera sito specifica e dei vettori lentivirali difettivi per l’integrazione (IDLV) per l’introduzione di un templato donatore, abbiamo impiegato il processo di riparazione del DNA guidata dall’omologia per la correzione delle mutazioni che causano la SCID-X1, ripristinando così la funzione genica e l’espressione fisiologica del gene IL2RG. Mediante l’integrazione di un cDNA correttivo del gene IL2RG a valle del promotore endogeno sia in cellule B linfoblastodi, che esprimono costitutivamente la catena gamma comune, sia in linfociti T da donatori sani, che richiedono IL2RG per la loro sopravvivenza, abbiamo dimostrato la funzionalità e l’attività fisiologica del gene modificato. Abbiamo quindi accoppiato la correzione genica con la selezione delle cellule mediante l’inclusione di una cassetta excidibile di espressione della GFP o della resistenza alla puromicina (PuroR) a valle del cDNA correttivo, al fine di correggere fibroblasti, che normalmente non esprimono IL2R, derivati da pazienti SCID-X1. Abbiamo quindi ottenuto una popolazione di fibroblasti corretti che abbiamo “ riprogrammato” mediante un nuovo vettore di reprogramming che esprime i fattori di trascrizione (SOX2, OCT4, KLf4) e il microRNA cluster 367, generando così una fonte illimitata di cellule staminali pluripotenti indotte (iPSC) geneticamente corrette di interesse terapeutico. L’espressione transiente della Cre-ricombinasi mediante IDLV ha inoltre permesso l’excisione del vettore di reprogramming e della cassetta di selezione, permettendo così l’ottenimento di cellule iPSC corrette, prive di vettore e con un normale cariotipo. Infine, attraverso il differenziamento delle cellule iPSC in progenitori T-linfoidi, un tipo cellulare assente nei pazienti SCID-X1, e l’osservazione di un vantaggio selettivo delle cellule linfoidi derivate dalle iPSC corrette, abbiamo dimostrato la correzione funzionale dell’allele IL2RG mutato. In conclusione questi dati dimostrano la validità della nostra strategia di integrazione sito-specifica che, mediante la correzione e la riprogrammazione cellulare, consente di ottenere cellule iPSC geneticamente corrette, aprendo la strada a nuove opportunità terapeutiche più sicure per il trattamento della SCID-X1.
Gene replacement by integrating vectors has been successfully used to treat several inherited diseases, such as Lysosomal Storage Disorders (LSD), Thalassemia and Primary Immunodeficiencies (PIDs). X-linked Combined Immunodeficiency (SCID-X1) is a fatal monogenic disorder, caused by mutation of the Interleukin 2 Receptor common γ-chain (IL2RG) gene. For SCID-X1, the early clinical studies have clearly shown the therapeutic potential of integrating vector based gene replacement therapy, which achieved efficient lymphoid reconstitution thanks to the selective growth advantage of the genetically modified cells. However, these studies also highlighted the potential risk of insertional mutagenesis due to random integration of the vector into the host cell genome and to unregulated transgene expression, thus calling for the development of safer gene therapy approaches. Here, by combining the Zinc Finger Nuclease (ZFNs) technology to induce site-specific DNA double-strand breaks (DSB) and of Integrase-Defective Lentiviral Vector (IDLV) to deliver a corrective donor template, we exploited Homology Driven Repair (HDR) to correct SCID-X1 mutation in situ, restoring both physiological expression and function of the IL2RG gene . By knocking-in a corrective IL2RG cDNA transgene downstream of its endogenous promoter in B-lymphoblastoid cells, which constitutively express IL2RG, and in primary T-lymphocytes, which requires IL2RG for their survival and growth, we provide evidence of physiologic activity of the gene-edited IL2RG gene. By including an excisable GFP- or a Puromycin Resistance (PuroR) expression cassette downstream of the corrective cDNA, we coupled correction with exogenous selection of corrected SCID-X1 primary fibroblasts, which do not physiologically express IL2RG, and obtained an enriched population of gene-corrected cells. We then reverted this population to pluripotency by using a novel reprogramming vector that expresses OCT4, SOX2, KLF4 and microRNA cluster 302-367 to obtain a potentially unlimited source of gene-corrected induced pluripotent stem cells (iPSC). We thus generated several gene-corrected bona-fide iPSCs, as confirmed by molecular analyses for targeted integration, which were characterized for their pluripotent state. IDLV-mediated transient delivery of the Cre-recombinase resulted in the co-excision of the reprogramming vector together with the selector cassette, thus allowing the generation of several gene-corrected, reprogramming-factor free iPSCs with normal karyotypes. Finally, by differentiating corrected iPSC to T-lymphoid progenitor cells, which are lacking in SCID-X1 patients, and showing a selective growth advantage of those derived from corrected iPSCs, we provide evidence of the functional correction of the IL2RG mutant allele. Overall these data demonstrate the feasibility of our targeted gene editing strategy, which couples gene correction with cell reprogramming to generate disease-free IPSC, thus paving the way for the development of novel and safer therapeutic approaches for SCID-X1.
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Books on the topic "Lentiviral vector"

1

Lentiviral vector systems for gene transfer. Georgetown, TX: Eurekah.com, 2003.

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Trono, Didier, ed. Lentiviral Vectors. Berlin, Heidelberg: Springer Berlin Heidelberg, 2002. http://dx.doi.org/10.1007/978-3-642-56114-6.

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Escors, David, Karine Breckpot, Frederick Arce, Grazyna Kochan, and Holly Stephenson. Lentiviral Vectors and Gene Therapy. Basel: Springer Basel, 2012. http://dx.doi.org/10.1007/978-3-0348-0402-8.

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Escors, David. Lentiviral Vectors and Gene Therapy. Basel: Springer Basel, 2012.

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Maurizio, Federico, ed. Lentivirus gene engineering protocols. Totowa, N.J: Humana Press, 2003.

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Federico, Maurizio, ed. Lentiviral Vectors and Exosomes as Gene and Protein Delivery Tools. New York, NY: Springer New York, 2016. http://dx.doi.org/10.1007/978-1-4939-3753-0.

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Maurizio, Federico, ed. Lentivirus gene engineering protocols. 2nd ed. New York: Humana Press, 2010.

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Lentiviral Vector Systems for Gene Transfer (Medical Intelligence Unit, 31). Springer, 2003.

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Lentiviral Vector Systems for Gene Transfer (Medical Intelligence Unit, 31). Eurekah.com, 2001.

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Buchschacher, Gary L. Lentiviral Vector Systems for Gene Transfer (Medical Intelligence Unit, 31). Eurekah.com, 2001.

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Book chapters on the topic "Lentiviral vector"

1

Larochelle, A., K. W. Peng, and S. J. Russell. "Lentiviral Vector Targeting." In Current Topics in Microbiology and Immunology, 143–63. Berlin, Heidelberg: Springer Berlin Heidelberg, 2002. http://dx.doi.org/10.1007/978-3-642-56114-6_7.

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Segura, María Mercedes, Alain Garnier, Yves Durocher, Sven Ansorge, and Amine Kamen. "New Protocol for Lentiviral Vector Mass Production." In Lentivirus Gene Engineering Protocols, 39–52. Totowa, NJ: Humana Press, 2009. http://dx.doi.org/10.1007/978-1-60761-533-0_2.

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Ramezani, Ali, and Robert G. Hawley. "Strategies to Insulate Lentiviral Vector-Expressed Transgenes." In Lentivirus Gene Engineering Protocols, 77–100. Totowa, NJ: Humana Press, 2009. http://dx.doi.org/10.1007/978-1-60761-533-0_5.

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Han, Shuhong, and Lung-Ji Chang. "Immunity of Lentiviral Vector-Modified Dendritic Cells." In Gene Therapy of Cancer, 245–59. Totowa, NJ: Humana Press, 2009. http://dx.doi.org/10.1007/978-1-59745-561-9_13.

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Chong, Mark S. K., and Jerry Chan. "Lentiviral Vector Transduction of Fetal Mesenchymal Stem Cells." In Lentivirus Gene Engineering Protocols, 135–47. Totowa, NJ: Humana Press, 2009. http://dx.doi.org/10.1007/978-1-60761-533-0_9.

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Cui, Yan, and Lung-Ji Chang. "Detection and Selection of Lentiviral Vector-Transduced Cells." In Lentivirus Gene Engineering Protocols, 69–85. Totowa, NJ: Humana Press, 2003. http://dx.doi.org/10.1385/1-59259-393-3:69.

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Scherr, Michaela, Letizia Venturini, and Matthias Eder. "Lentiviral Vector-Mediated Expression of pre-miRNAs and AntagomiRs." In Lentivirus Gene Engineering Protocols, 175–85. Totowa, NJ: Humana Press, 2009. http://dx.doi.org/10.1007/978-1-60761-533-0_12.

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ter Brake, Olivier, Jan-Tinus Westerink, and Ben Berkhout. "Lentiviral Vector Engineering for Anti-HIV RNAi Gene Therapy." In Lentivirus Gene Engineering Protocols, 201–13. Totowa, NJ: Humana Press, 2009. http://dx.doi.org/10.1007/978-1-60761-533-0_14.

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Du, Zhong-Wei, and Su-Chun Zhang. "Lentiviral Vector-Mediated Transgenesis in Human Embryonic Stem Cells." In Lentivirus Gene Engineering Protocols, 127–34. Totowa, NJ: Humana Press, 2009. http://dx.doi.org/10.1007/978-1-60761-533-0_8.

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Verhoeyen, Els, Caroline Costa, and Francois-Loic Cosset. "Lentiviral Vector Gene Transfer into Human T Cells." In Genetic Modification of Hematopoietic Stem Cells, 97–114. Totowa, NJ: Humana Press, 2009. http://dx.doi.org/10.1007/978-1-59745-409-4_8.

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Conference papers on the topic "Lentiviral vector"

1

Ranzani, Marco, Daniela Cesana, Cynthia C. Bartholomä, Francesca Sanvito, Michela Riba, Mauro Pala, Fabrizio Benedicenti, et al. "Abstract 3169: Lentiviral vector-based insertional mutagenesis identifies new clinically relevant liver cancer genes." In Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC. American Association for Cancer Research, 2013. http://dx.doi.org/10.1158/1538-7445.am2013-3169.

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Albershardt, Tina C., David J. Campbell, Andrea J. Parsons, Jan H. ter Meulen, and Peter Berglund. "Abstract 2506: Preclinical characterization of LV305, a lentiviral vector targeting tumors expressing NY-ESO-1." In Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA. American Association for Cancer Research, 2015. http://dx.doi.org/10.1158/1538-7445.am2015-2506.

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Dwivedi, Alka, Ling Su, Justin Mirazee, Mehdi Benzaoui, Christopher Chien, Nirali Shah, Xiaolin Wu, and Naomi Taylor. "327 Clonal expansion of CD22 CAR T-cells following lentiviral vector integration in the PWWP3A gene." In SITC 37th Annual Meeting (SITC 2022) Abstracts. BMJ Publishing Group Ltd, 2022. http://dx.doi.org/10.1136/jitc-2022-sitc2022.0327.

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Qin, Shi, and Jie Wang. "The construction of RGD-TAT-KDR siRNA fusion gene lentiviral vector and the study of its antitumor activityin vitro." In International Conference on Medical Engineering and Bioinformatics. Southampton, UK: WIT Press, 2014. http://dx.doi.org/10.2495/meb140171.

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Bajaj, Anshika, Tsai-Yu Lin, Lisa Y. Ngo, Michele Murphy, Brenna Kelley-Clarke, Wayne R. Gombotz, Jan H. ter Meulen, and Peter Berglund. "Abstract 5919: Component-specific qPCR assays for characterization and identity testing of multigenome ZVex®, a dendritic cell-targeting lentiviral vector platform." In Proceedings: AACR Annual Meeting 2018; April 14-18, 2018; Chicago, IL. American Association for Cancer Research, 2018. http://dx.doi.org/10.1158/1538-7445.am2018-5919.

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Bajaj, Anshika, Lisa Y. Ngo, Peter Berglund, and Jan ter Meulen. "Abstract 5092: ZVex® lentiviral vector strongly activates pro-inflammatory, antigen processing, and anti-viral defense response pathways in monocyte-derived dendritic cells." In Proceedings: AACR Annual Meeting 2017; April 1-5, 2017; Washington, DC. American Association for Cancer Research, 2017. http://dx.doi.org/10.1158/1538-7445.am2017-5092.

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Ranzani, Marco, Daniela Cesana, Cynthia Bartholomae, Francesca Sanvito, Mauro Pala, Fabrizio Benedicenti, Lucia Sergi Sergi, et al. "Abstract 4982: Identification of new human liver cancer genes by a novel lentiviral vector-based insertional mutagenesis approach in three mouse models of hepatocarcinogenesis." In Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL. American Association for Cancer Research, 2011. http://dx.doi.org/10.1158/1538-7445.am2011-4982.

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Ranzani, Marco, Daniela Cesana, Cynthia C. Bartholomae, Francesca Sanvito, Mauro Pala, Fabrizio Benedicenti, Pierangela Gallina, et al. "Abstract 104: New liver cancer genes identified by lentiviral vector-based insertional mutagenesis in mice are associated to differential survival in hepatocellular carcinoma patients." In Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL. American Association for Cancer Research, 2012. http://dx.doi.org/10.1158/1538-7445.am2012-104.

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Bryson, Paul D., Xiaolu Han, Norman Truong, and Pin Wang. "Abstract 2888: Dendritic cell-targeted lentiviral vector vaccines overcome tolerance to generate a protective T-cell immune response to breast cancer antigens ERBB2 and α-lactalbumin." In Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA. American Association for Cancer Research, 2014. http://dx.doi.org/10.1158/1538-7445.am2014-2888.

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Nicolai, Chris, Jim Qin, Way Wu, Mollie McDonnell, Erica Shirazi, Greyson Hamilton, Max Chen, et al. "1230 VivoVec lentiviral vector particles surface-engineered with T cell activating and co-stimulatory ligands enhancein vivoCAR T cell generation and antitumor activity." In SITC 37th Annual Meeting (SITC 2022) Abstracts. BMJ Publishing Group Ltd, 2022. http://dx.doi.org/10.1136/jitc-2022-sitc2022.1230.

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Reports on the topic "Lentiviral vector"

1

Yes, Jiing-Kuan. Anti-Angiogenic Gene Therapy of Prostate Cancer with Lentiviral Vectors. Fort Belvoir, VA: Defense Technical Information Center, July 2004. http://dx.doi.org/10.21236/ada428533.

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Yee, Jiing-Kuan. Anti-Angiogenic Gene Therapy of Prostate Cancer With Lentiviral Vectors. Fort Belvoir, VA: Defense Technical Information Center, July 2002. http://dx.doi.org/10.21236/ada410315.

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Yee, Jiing-Kuan. Anti-Angiogenic Gene Therapy of Prostate Cancer with Lentiviral Vectors. Fort Belvoir, VA: Defense Technical Information Center, July 2003. http://dx.doi.org/10.21236/ada418264.

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