Long-term inhibition of HIV-1 replication with RNA interference against cellular co-factors

Eekels, Julia J.M.; Geerts, Dirk; Jeeninga, Rienk E.; Berkhout, Ben

doi:10.1016/j.antiviral.2010.11.005

Cited by 53 publications

(46 citation statements)

References 69 publications

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“…Thus, the chance that resistance mutations are selected in host mRNAs is negligible compared to HIV-1 target sequences. Recent screens revealed several cofactor-encoding mRNAs whose knock-down resulted in diminished HIV-1 replication [63][64][65][66][67] . However, knockdown of cellular proteins at the mRNA level might have negative effects on cell viability and anti-host shRNAs must be carefully designed.…”

Section: Resultsmentioning

confidence: 99%

Selection of RNAi-based inhibitors for anti-HIV gene therapy

Knoepfel

Centlivre²,

Liu³

et al. 2012

WJV

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In the last decade, RNA interference (RNAi) advanced to one of the most widely applied techniques in the biomedical research field and several RNAi therapeutic clinical trials have been launched. We focus on RNAibased inhibitors against the chronic infection with human immunodeficiency virus type 1 (HIV-1). A lentiviral gene therapy is proposed for HIV-infected patients that will protect and reconstitute the vital immune cell pool. The RNAi-based inhibitors that have been developed are short hairpin RNA molecules (shRNAs), of which multiple are needed to prevent viral escape. In ten distinct steps, we describe the selection process that started with 135 shRNA candidates, from the initial design criteria, via testing of the in vitro and in vivo antiviral activity and cytotoxicity to the final design of a combinatorial therapy with three shRNAs. These shRNAs satisfied all 10 selection criteria such as targeting conserved regions of the HIV-1 RNA genome, exhibiting robust inhibition of HIV-1 replication and having no impact on cell physiology. This combinatorial shRNA vector will soon move forward to the first clinical studies.

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Section: Resultsmentioning

confidence: 99%

Selection of RNAi-based inhibitors for anti-HIV gene therapy

Knoepfel

Centlivre²,

Liu³

et al. 2012

WJV

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“…2 --6 Both viral RNA and cellular transcripts that encode cofactors necessary for viral replication can be targeted. 7 Especially for the attack on viruses that cause a persistent infection the preferred method is the generation of stable knockdown cells, in which constitutive expression of the antiviral shRNAs is achieved by vector transduction. In particular, lentiviral vectors have been very successful because they are able to transduce many different cell types, both actively dividing, and quiescent cells, in which the viral genome is stably integrated in the host cell DNA.…”

Section: Introductionmentioning

confidence: 99%

A competitive cell growth assay for the detection of subtle effects of gene transduction on cell proliferation

et al. 2011

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RNA interference (RNAi) is a sequence-specific gene silencing mechanism with therapeutic potential against many human pathogens. To obtain a durable therapeutic effect, stable transduction of target cells with for instance a lentiviral vector that expresses a short hairpin (shRNA) inducer of the RNAi pathway is necessary. Apart from the intended therapeutic effect, this treatment can induce negative effects on cell proliferation via off-target effects. A careful evaluation of the transduced cells is required to develop a safe gene therapy approach. Stably transduced cells are usually selected by expression of the enhanced green fluorescent protein (GFP) marker. In this study we show that the mixed transduction culture, containing both transduced GFP þ and untransduced GFP À cells, can simply be passaged to score the GFP þ /GFP À ratio by longitudinal flow cytometric analysis as a measure of the negative impact of the RNAi treatment on the cellular proliferation rate. We show that this assay is sensitive, easy to use and internally controlled for assessing subtle effects on cell proliferation of lentiviral transduction and transgene expression.

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“…This increased HIV-1 expression level is ascribed to the inhibition of PKR (8,25,29) and to a direct activity of TRBP that relieves the translational block caused by the TAR RNA structure (2,36). The transfection of small interfering RNAs (siRNAs) against TRBP decreases the level of HIV-1 production in HeLa cells, and the transduction of short hairpin RNAs (shRNAs) against TRBP leads to the long-term inhibition of HIV-1 replication in human SupT1 lymphocytic cells, indicating that the protein contributes to viral expression and replication (24,39), but these findings have not yet been reproduced in primary cells. This property seems to be mediated mainly by a decreased TRBP-PKR interaction and a consequent enhancement of PKR activation, which prevents viral translation, but could also be due to TRBP's interactions with other molecules (24,109,127).…”

Section: Activity In Hiv-1 Replicationmentioning

confidence: 99%

The Multiple Functions of TRBP, at the Hub of Cell Responses to Viruses, Stress, and Cancer

Daniels

Gatignol

2012

Microbiol Mol Biol Rev

100

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SUMMARY The TAR RNA binding protein (TRBP) has emerged as a key player in many cellular processes. First identified as a cellular protein that facilitates the replication of human immunodeficiency virus, TRBP has since been shown to inhibit the activation of protein kinase R (PKR), a protein involved in innate immune responses and the cellular response to stress. It also binds to the PKR activator PACT and regulates its function. TRBP also contributes to RNA interference as an integral part of the minimal RNA-induced silencing complex with Dicer and Argonaute proteins. Due to its multiple functions in the cell, TRBP is involved in oncogenesis when its sequence is mutated or its expression is deregulated. The depletion or overexpression of TRBP results in malignancy, suggesting that the balance of TRBP expression is key to normal cellular function. These studies show that TRBP is multifunctional and mediates cross talk between different pathways. Its activities at the molecular level impact the cellular function from normal development to cancer and the response to infections.

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Long-term inhibition of HIV-1 replication with RNA interference against cellular co-factors

Cited by 53 publications

References 69 publications

Selection of RNAi-based inhibitors for anti-HIV gene therapy

Selection of RNAi-based inhibitors for anti-HIV gene therapy

A competitive cell growth assay for the detection of subtle effects of gene transduction on cell proliferation

The Multiple Functions of TRBP, at the Hub of Cell Responses to Viruses, Stress, and Cancer

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