783. Transcriptional Profiling of HIV-1 Infection and Lentiviral Vector Transduction: Impact of HIV-1 Infection on Cell Transcriptome and Attenuated Response to Advanced Generation Lentiviral Vectors

ABSTRACT AIDS vaccination has a pressing need for more potent vaccination vectors capable of eliciting strong, diversified, and long-lasting cellular immune responses against human immunodeficiency virus (HIV). Lentiviral vectors have demonstrated efficiency not only as gene delivery vehicles for gene therapy applications but also as vaccination tools. This is likely due to their ability to transduce nondividing cells, including dendritic cells, enabling sustained endogenous antigen presentation and thus the induction of high proportions of specific cytotoxic T cells and long-lasting memory T cells. We show in a first proof-of-concept pilot study that a prime/boost vaccination strategy using lentiviral vectors pseudotyped with a glycoprotein G from two non-cross-reactive vesicular stomatitis virus serotypes elicited robust and broad cellular immune responses against the vector-encoded antigen, simian immunodeficiency virus (SIV) GAG, in cynomolgus macaques. Vaccination conferred strong protection against a massive intrarectal challenge with SIVmac251, as evidenced both by the reduction of viremia at the peak of acute infection (a mean of over 2 log10 fold reduction) and by the full preservation of the CD28+ CD95+ memory CD4+ T cells during the acute phase, a strong correlate of protection against pathogenesis. Although vaccinees continued to display lower viremia than control macaques during the early chronic phase, these differences were not statistically significant by day 50 postchallenge. A not-optimized SIV GAG antigen was chosen to show the strong potential of the lentiviral vector system for vaccination. Given that a stronger protection can be anticipated from a modern HIV-1 antigen design, gene transfer vectors derived from HIV-1 appear as promising candidates for vaccination against HIV-1 infection.

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Restriction of Feline Immunodeficiency Virus by Ref1, Lv1, and Primate TRIM5α Proteins

Journal of Virology ◽

10.1128/jvi.79.24.15175-15188.2005 ◽

2005 ◽

Vol 79 (24) ◽

pp. 15175-15188 ◽

Cited By ~ 70

Author(s):

Dyana T. Saenz ◽

Wulin Teo ◽

John C. Olsen ◽

Eric M. Poeschla

Keyword(s):

Cell Lines ◽

Feline Immunodeficiency Virus ◽

Equine Infectious Anemia Virus ◽

Lentiviral Vector ◽

Leukemia Virus ◽

Human T Cell ◽

Immunodeficiency Virus ◽

T Cell Lines ◽

Hiv 1 ◽

Vector Transduction

ABSTRACT The Ref1 and Lv1 postentry restrictions in human and monkey cells have been analyzed for lentiviruses in the primate and ungulate groups, but no data exist for the third (feline) group. We compared feline immunodeficiency virus (FIV) to other restricted (human immunodeficiency virus type 1 [HIV-1], equine infectious anemia virus [EIAV]) and unrestricted (NB-tropic murine leukemia virus [NB-MLV]) retroviruses across wide ranges of viral inputs in cells from multiple primate and nonprimate species. We also characterized restrictions conferred to permissive feline and canine cells engineered to express rhesus and human TRIM5α proteins and performed RNA interference (RNAi) against endogenous TRIM5α. We find that expression of rhesus or human TRIM5α proteins in feline cells restricts FIV, impairing pseudotyped vector transduction and viral replication, but rhesus TRIM5α is more restricting than human TRIM5α. Notably, however, canine cells did not support restriction by human TRIM5α and supported minimal restriction by rhesus TRIM5α, suggesting that these proteins may not function autonomously or that a canine factor interferes. Stable RNAi knockdown of endogenous rhesus TRIM5α resulted in marked increases in FIV and HIV-1 infectivities while having no effect on NB-MLV. A panel of nonprimate cell lines varied widely in susceptibility to lentiviral vector transduction, but normalized FIV and HIV-1 vectors varied concordantly. In contrast, in human and monkey cells, relative restriction of FIV compared to HIV-1 varied from none to substantial, with the greatest relative infectivity deficit for FIV vectors observed in human T-cell lines. Endogenous and introduced TRIM5α restrictions of FIV could be titrated by coinfections with FIV, HIV-1, or EIAV virus-like particles. Arsenic trioxide had complex and TRIM5α-independent enhancing effects on lentiviral but not NB-MLV infection. Implications for human gene therapy are discussed.

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A single administration of lentiviral vectors expressing either full-length human immunodeficiency virus 1 (HIV-1)HXB2 Rev/Env or codon-optimized HIV-1JR-FL gp120 generates durable immune responses in mice

Journal of General Virology ◽

10.1099/vir.0.81706-0 ◽

2006 ◽

Vol 87 (6) ◽

pp. 1625-1634 ◽

Cited By ~ 21

Author(s):

Viviana Buffa ◽

Donatella R. M. Negri ◽

Pasqualina Leone ◽

Roberta Bona ◽

Martina Borghi ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Immune Responses ◽

Viral Vectors ◽

Lentiviral Vector ◽

Lentiviral Vectors ◽

Full Length ◽

Immunodeficiency Virus ◽

Hiv 1 ◽

Human Immunodeficiency Virus 1

Genetic immunization using viral vectors provides an effective means to elicit antigen-specific cellular immune responses. Several viral vectors have proven efficacious in inducing immune responses after direct injection in vivo. Among them, recombinant, self-inactivating lentiviral vectors are very attractive delivery systems, as they are able to efficiently transduce into and express foreign genes in a wide variety of mammalian cells. A self-inactivating lentiviral vector was evaluated for the delivery of human immunodeficiency virus 1 (HIV-1) envelope sequences in mice in order to elicit specific immune responses. With this aim, BALB/c mice were immunized with a single injection of self-inactivating lentiviral vectors carrying either the full-length HIV-1HXB2 Rev/Env (TY2-IIIBEnv) or the codon-optimized HIV-1JR-FL gp120 (TY2-JREnv) coding sequence. Both vectors were able to elicit specific cellular responses efficiently, as measured by gamma interferon ELISPOT and chromium-release assays, upon in vitro stimulation of splenocytes from BALB/c immunized mice. However, only the TY2-JREnv-immunized mice were able to elicit specific humoral responses, measured as anti-gp120 antibody production. These data provide the first evidence that a single, direct, in vivo administration of a lentiviral vector encoding a viral gene might represent a useful strategy for vaccine development.

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HIV-1 sequences in lentiviral vector genomes can be substantially reduced without compromising transduction efficiency

Scientific Reports ◽

10.1038/s41598-021-91309-w ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Helin Sertkaya ◽

Mattia Ficarelli ◽

Nathan P. Sweeney ◽

Hannah Parker ◽

Conrad A. Vink ◽

...

Keyword(s):

Lentiviral Vector ◽

Lentiviral Vectors ◽

Large Deletion ◽

Transduction Efficiency ◽

Rev Response Element ◽

Vector Genome ◽

Long Read ◽

Viral Sequences ◽

Hiv 1 ◽

Splice Site Usage

AbstractMany lentiviral vectors used for gene therapy are derived from HIV-1. An optimal vector genome would include only the viral sequences required for transduction efficiency and gene expression to minimize the amount of foreign sequence inserted into a patient’s genome. However, it remains unclear whether all of the HIV-1 sequence in vector genomes is essential. To determine which viral sequences are required, we performed a systematic deletion analysis, which showed that most of the gag region and over 50% of the env region could be deleted. Because the splicing profile for lentiviral vectors is poorly characterized, we used long-read sequencing to determine canonical and cryptic splice site usage. Deleting specific regions of env sequence reduced the number of splicing events per transcript and increased the proportion of unspliced genomes. Finally, combining a large deletion in gag with repositioning the Rev-response element downstream of the 3’ R to prevent its reverse transcription showed that 1201 nucleotides of HIV-1 sequence can be removed from the integrated vector genome without substantially compromising transduction efficiency. Overall, this allows the creation of lentiviral vector genomes that contain minimal HIV-1 sequence, which could improve safety and transfer less viral sequence into a patient’s DNA.

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Reduced Mobilization of Rev-Responsive Element-Deficient Lentiviral Vectors

Journal of Virology ◽

10.1128/jvi.79.14.9359-9362.2005 ◽

2005 ◽

Vol 79 (14) ◽

pp. 9359-9362 ◽

Cited By ~ 16

Author(s):

Susann Lucke ◽

Thomas Grunwald ◽

Klaus Überla

Keyword(s):

Human Immunodeficiency Virus ◽

Lentiviral Vector ◽

Binding Protein ◽

Lentiviral Vectors ◽

Target Cells ◽

Novel Approach ◽

Immunodeficiency Virus ◽

Efficient Vector ◽

Responsive Element ◽

Hiv 1

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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563. In Vivo Derivation of HIV-1 Resistant Transgenic T Cells by Lentiviral Vector Transduction of a Triple Combination of Anti-HIV Genes Anti-CCR5 Ribozyme, Tat-Rev siRNA and a TAR Decoy

Molecular Therapy ◽

10.1016/s1525-0016(16)44769-6 ◽

2007 ◽

Vol 15 ◽

pp. S217

Keyword(s):

T Cells ◽

Lentiviral Vector ◽

Triple Combination ◽

Anti Hiv ◽

Hiv 1 ◽

Vector Transduction

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384. Abrogation of Post-Entry Restriction of HIV-1 Based Lentiviral Vector Transduction in Simian Cells. Implications for Gene Therapy

Molecular Therapy ◽

10.1016/s1525-0016(16)40826-9 ◽

2003 ◽

Vol 7 (5) ◽

pp. S150

Keyword(s):

Gene Therapy ◽

Lentiviral Vector ◽

Post Entry ◽

Hiv 1 ◽

Vector Transduction

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Construction and identification of recombinant lentiviral vector containing HIV-1 Tat gene and its expression in 293T cells

Journal of Biomedical Research ◽

10.1016/s1674-8301(10)60009-7 ◽

2010 ◽

Vol 24 (1) ◽

pp. 58-63 ◽

Cited By ~ 6

Author(s):

Bingbing Wei ◽

Ninghan Feng ◽

Feng Zhou ◽

Chun Lu ◽

Jiantang Su ◽

...

Keyword(s):

Lentiviral Vector ◽

Hiv 1

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Myocyte-specific enhancer factor 2c triggers transdifferentiation of adipose tissue-derived stromal cells into spontaneously beating cardiomyocyte-like cells

Scientific Reports ◽

10.1038/s41598-020-80848-3 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Shinichiro Takashima ◽

Soichiro Usui ◽

Oto Inoue ◽

Chiaki Goten ◽

Kosei Yamaguchi ◽

...

Keyword(s):

Adipose Tissue ◽

Lentiviral Vector ◽

Troponin T ◽

Stromal Vascular Fraction ◽

Cardiac Regeneration ◽

Cardiac Lineage ◽

Cell Transcriptome ◽

Single Cell Transcriptome ◽

Culture Protocol ◽

Enhancer Factor

AbstractCardiomyocyte regeneration is limited in adults. The adipose tissue-derived stromal vascular fraction (Ad-SVF) contains pluripotent stem cells that rarely transdifferentiate into spontaneously beating cardiomyocyte-like cells (beating CMs). However, the characteristics of beating CMs and the factors that regulate the differentiation of Ad-SVF toward the cardiac lineage are unknown. We developed a simple culture protocol under which the adult murine inguinal Ad-SVF reproducibly transdifferentiates into beating CMs without induction. The beating CMs showed the striated ventricular phenotype of cardiomyocytes and synchronised oscillation of the intracellular calcium concentration among cells on day 28 of Ad-SVF primary culture. We also identified beating CM-fated progenitors (CFPs) and performed single-cell transcriptome analysis of these CFPs. Among 491 transcription factors that were differentially expressed (≥ 1.75-fold) in CFPs and the beating CMs, myocyte-specific enhancer 2c (Mef2c) was key. Transduction of Ad-SVF cells with Mef2c using a lentiviral vector yielded CFPs and beating CMs with ~ tenfold higher cardiac troponin T expression, which was abolished by silencing of Mef2c. Thus, we identified the master gene required for transdifferentiation of Ad-SVF into beating CMs. These findings will facilitate the development of novel cardiac regeneration therapies based on gene-modified, cardiac lineage-directed Ad-SVF cells.

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