Harnessing endogenous miR-181a to segregate transgenic antigen receptor expression in developing versus post-thymic T cells in murine hematopoietic chimeras

Papapetrou, Eirini P.; Kovalovsky, Damián; Beloeil, Laurent; Sant’Angelo, Derek B.; Sadelain, Michel

doi:10.1172/jci37216

Cited by 37 publications

(53 citation statements)

References 52 publications

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“…Overlapping PCR was used to generate bicistronic expression cassettes encoding vexGFP, mCitrine, mCherry, and mCerulean linked by a P2A peptide preceded by a Gly-Ser-Gly linker (35) to the cDNAs of human Oct4, Sox2, Klf4 and c-Myc, respectively, which were cloned in a lentiviral vector under the transcriptional control of the human phosphoglycerate kinase (hPGK) promoter. Vector supernatants were produced as previously described (36).…”

Section: Methodsmentioning

confidence: 99%

Stoichiometric and temporal requirements of Oct4, Sox2, Klf4, and c-Myc expression for efficient human iPSC induction and differentiation

Papapetrou

Tomishima²,

Chambers³

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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266

228

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Human-induced pluripotent stem cells (hiPSCs) are generated from somatic cells by ectopic expression of the 4 reprogramming factors (RFs) Oct-4, Sox2, Klf4, and c-Myc. To better define the stoichiometric requirements and dynamic expression patterns required for successful hiPSC induction, we generated 4 bicistronic lentiviral vectors encoding the 4 RFs co-expressed with discernable fluorescent proteins. Using this system, we define the optimal stoichiometry of RF expression to be highly sensitive to Oct4 dosage, and we demonstrate the impact that variations in the relative ratios of RF expression exert on the efficiency of hiPSC induction. Monitoring of expression of each individual RF in single cells during the course of reprogramming revealed that vector silencing follows acquisition of pluripotent cell markers. Pronounced lentiviral vector silencing was a characteristic of successfully reprogrammed hiPSC clones, but lack of complete silencing did not hinder hiPSC induction, maintenance, or directed differentiation. The vector system described here presents a powerful tool for mechanistic studies of reprogramming and the optimization of hiPSC generation. fluorescent proteins ͉ lentiviral vectors ͉ silencing ͉ stoichiometry

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Section: Methodsmentioning

confidence: 99%

Stoichiometric and temporal requirements of Oct4, Sox2, Klf4, and c-Myc expression for efficient human iPSC induction and differentiation

Papapetrou

Tomishima²,

Chambers³

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

266

228

View full text Add to dashboard Cite

show abstract

“…In both cases, the aim was to prevent the expression of the foreign gene by antigen-presenting cells 126,127 to avoid activation of specific lymphocyte responses to transgene products. In a similar approach, Papapetrou et al 128 demonstrated specific expression of foreign genes only in post-thymic naive or activated T cells by incorporating mirR-181a target sequence (expressed in developing T cells) in the therapeutic vector. Recently the Naldini's group 129 has demonstrated that this strategy can be used to tolerize rodents to a lentiviral-encoded antigen.…”

Section: Box 2 Tissue-specific Vs Physiologically Regulated Vectorsmentioning

confidence: 99%

Physiological and tissue-specific vectors for treatment of inherited diseases

Toscano

Romero

Muñoz³

et al. 2010

Gene Ther

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After more than 1500 gene therapy clinical trials in the past two decades, the overall conclusion is that for gene therapy (GT) to be successful, the vector systems must still be improved in terms of delivery, expression and safety. The recent development of more efficient and stable vector systems has created great expectations for the future of GT. Impressive results were obtained in three primary immunodeficiencies and other inherited diseases such as congenital blindness, adrenoleukodystrophy or junctional epidermolysis bullosa. However, the development of leukemia in five children included in the GT clinical trials for X-linked severe combined immunodeficiency and the silencing of the therapeutic gene in the chronic granulomatous disease clearly showed the importance of improving safety and efficiency. In this review, we focus on the main strategies available to achieve physiological or tissue-specific expression of therapeutic transgenes and discuss the importance of controlling transgene expression to improve safety. We propose that tissue-specific and/or physiological viral vectors offer the best balance between efficiency and safety and will be the tools of choice for future clinical trials in GT of inherited diseases.

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“…Several miR-181 targets were inserted in the LV that encodes tumor antigen receptors. 199 This miRNA is expressed in developing thymocytes, but its expression ceases in mature thymocytes. This property was used to restrict the LV to developing thymocytes in mice upon re-engraftment with modified bone-marrow cells.…”

Section: Controlled Transgene Expression Via Mirnasmentioning

confidence: 99%

Improving miRNA Delivery by Optimizing miRNA Expression Cassettes in Diverse Virus Vectors

Herrera-Carrillo

Liu

Berkhout

2017

Human Gene Therapy Methods

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The RNA interference pathway is an evolutionary conserved post-transcriptional gene regulation mechanism that is exclusively triggered by double-stranded RNA inducers. RNAi-based methods and technologies have facilitated the discovery of many basic science findings and spurred the development of novel RNA therapeutics. Transient induction of RNAi via transfection of synthetic small interfering RNAs can trigger the selective knockdown of a target mRNA. For durable silencing of gene expression, either artificial short hairpin RNA or microRNA encoding transgene constructs were developed. These miRNAs are based on the molecules that induce the natural RNAi pathway in mammals and humans: the endogenously expressed miRNAs. Significant efforts focused on the construction and delivery of miRNA cassettes in order to solve basic biology questions or to design new therapy strategies. Several viral vectors have been developed, which are particularly useful for the delivery of miRNA expression cassettes to specific target cells. Each vector system has its own unique set of distinct properties. Thus, depending on the specific application, a particular vector may be most suitable. This field was previously reviewed for different viral vector systems, and now the recent progress in the field of miRNA-based gene-silencing approaches using lentiviral vectors is reported. The focus is on the unique properties and respective limitations of the available vector systems for miRNA delivery.Keywords: miRNA cassettes, gene therapy, viral vectors, vector tropism INTRODUCTION NON-CODING RNA MOLECULES play an essential role in gene regulation of the cell via a mechanism called RNA interference (RNAi). The RNAi mechanism is evolutionary conserved in eukaryotes and triggers the sequence-specific inhibition or degradation of a single or a unique set of complementary mRNAs. Three small RNA classes have been described to participate in mammalian RNAi mechanisms: microRNAs (miRNAs), 1,2 endogenous small interfering RNAs (endo-siRNAs), 3 and PIWI-associated RNAs (piRNAs). 4 The endo-siRNAs and piRNAs are involved in suppression of transposable elements. The miRNAs are broadly involved in the regulated expression of multiple cellular genes and execute their effect at the posttranscriptional level. 5,6 MiRNA-mediated regulation of gene expression plays a significant role in cell metabolism and cellular developmental and differentiation processes in mammals. More than a thousand human miRNAs have already been identified that are involved in the regulated expression of around 30% of all genes, which is a conservative estimate.7 Because of the miRNA abundance and their important regulatory functions, these molecules have received much attention from the scientific community over the last decade. For instance, efforts have been made to explain the biological function of the natural miRNAs by identifying the target mRNA and the role in cellular physiology. On the other hand, the design of man-made miRNA mimics to impose control over gene expression beca...

show abstract

Harnessing endogenous miR-181a to segregate transgenic antigen receptor expression in developing versus post-thymic T cells in murine hematopoietic chimeras

Cited by 37 publications

References 52 publications

Stoichiometric and temporal requirements of Oct4, Sox2, Klf4, and c-Myc expression for efficient human iPSC induction and differentiation

Stoichiometric and temporal requirements of Oct4, Sox2, Klf4, and c-Myc expression for efficient human iPSC induction and differentiation

Physiological and tissue-specific vectors for treatment of inherited diseases

Improving miRNA Delivery by Optimizing miRNA Expression Cassettes in Diverse Virus Vectors

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