Sendai virus, an RNA virus with no risk of genomic integration, delivers CRISPR/Cas9 for efficient gene editing

Park, Arnold; Hong, Patrick; Won, Sohui T.; Thibault, Patricia A.; Vigant, Frédéric; Oguntuyo, Kasopefoluwa Y.; Taft, Justin; Lee, Benhur

doi:10.1038/mtm.2016.57

Cited by 41 publications

(18 citation statements)

References 57 publications

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“…For example, the use of non‐integrating vectors, transient expression systems and small molecules to deliver transcription factors to induced pluripotent stem (iPS) cells are addressing safety concerns by eliminating unwanted long‐term expression of the encoded transcription factors and the possibility for insertional mutagenesis . Transient expression of Cas9 and other DNA endonuclease enzymes for genome editing might also be advantageous by limiting the number of potential off‐target effects and this is being extensively investigated …”

Section: Evolving Technologies and New Gene Therapy Platformsmentioning

confidence: 99%

Gene therapy clinical trials worldwide to 2017: An update

Ginn

Amaya

Alexander

et al. 2018

The Journal of Gene Medicine

647

489

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To date, almost 2600 gene therapy clinical trials have been completed, are ongoing or have been approved worldwide. Our database brings together global information on gene therapy clinical activity from trial databases, official agency sources, published literature, conference presentations and posters kindly provided to us by individual investigators or trial sponsors. This review presents our analysis of clinical trials that, to the best of our knowledge, have been or are being performed worldwide. As of our November 2017 update, we have entries on 2597 trials undertaken in 38 countries. We have analysed the geographical distribution of trials, the disease indications (or other reasons) for trials, the proportions to which different vector types are used, and the genes that have been transferred. Details of the analyses presented, and our searchable database are available via The Journal of Gene Medicine Gene Therapy Clinical Trials Worldwide website at: http://www.wiley.co.uk/genmed/clinical. We also provide an overview of the progress being made in gene therapy clinical trials around the world, and discuss key trends since the previous review, namely the use of chimeric antigen receptor T cells for the treatment of cancer and advancements in genome editing technologies, which have the potential to transform the field moving forward.

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Section: Evolving Technologies and New Gene Therapy Platformsmentioning

confidence: 99%

Gene therapy clinical trials worldwide to 2017: An update

Ginn

Amaya

Alexander

et al. 2018

The Journal of Gene Medicine

647

489

View full text Add to dashboard Cite

show abstract

“…While this strategy has achieved some noteworthy successes, use of DNA as a vector introduces other unwanted issues including the need for entry into the nucleus and the possibility of genomic integration. RNA-based vectors mitigate this risk by having no DNA phase and performing all of their function in the cytoplasm 32 . Given these attributes, we chose to adapt miRNA-based targeting as a means of instilling cell-specific activity.…”

Section: Discussionmentioning

confidence: 99%

“…By exploiting the biology of RNA viruses, Cas12a, and miRNAs, here we demonstrate that one can design a single RNA that could be systemically delivered with high efficiency but only function for a set amount of time and only within a desired cell lineage. The basic principle underlying this biology could be used with the replicon based strategy outlined here or with existing RNA virus based genetic editors 32 . Taken together, with the rapid advancements of synthetic biology, new CRISPR-Cas systems, and our understanding of virus-host interactions, our progress towards in vivo editing may allow us to view genetic-based errors as something that can be seamlessly overwritten.…”

Section: Discussionmentioning

confidence: 99%

Engineering an RNA-based tissue-specific platform for genetic editing through use of a miRNA-enabled Cas12a

Møeller

Oishi

tenOever

2020

Preprint

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9The capacity to edit genomes using CRISPR-Cas systems holds immense potential for 10 countless genetic-based diseases. However, one significant impediment preventing broad 11 therapeutic utilization is in vivo delivery. While genetic editing at a single cell level in vitro can be 12 achieved with high efficiency, the capacity to utilize these same biologic tools in a desired tissue 13 in vivo remains challenging. Non-integrating RNA virus-based vectors constitute efficient 14 platforms for transgene expression and surpass several barriers to in vivo delivery. However, 15 the broad tissue tropism of viral vectors raises the concern for off-target effects. Moreover, 16 prolonged expression of the Cas proteins, regardless of delivery method, can accumulate 17 aberrant RNAs leading to unwanted immunological responses. In an effort to circumvent these 18 shortcomings, here we describe a versatile RNA virus-based technology that can achieve cell-19 specific activity and self-inactivation by combining host microRNA (miRNA) biology with the 20 CRISPR-Cas12a RNA-guided nuclease. Exploiting the RNase activity of Cas12a, we generated 21 a vector that self-inactivates upon delivery of Cas12a and an accompanying CRISPR RNA 22 (crRNA). Furthermore, we show that maturation of the crRNA can be made dependent on cell-23 specific miRNAs, which confers cell-specificity. We demonstrate that this genetic editing circuit 24delivers diminished yet sufficient levels of Cas12a to achieve effective genome editing whilst 25 inducing a minimal immunological response. It can also function in a cell-specific manner 26 thereby facilitating in vivo editing and mitigating the risk of unwanted, off-target effects. 27

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“…New viral vectors accepted for clinical use are presently being tested to enhance viability and to increase the efficiency of nucleic acid delivery into primary cells (adenovirus, lentivirus; Goswami et al, 2019). Sendai virus, a cytoplasmic RNA virus with no risk of genomic integration, is also been tested (Park et al, 2016).…”

Section: Transfection Efficiencymentioning

confidence: 99%

New Bicistronic TALENs Greatly Improve Genome Editing

Martín-Fernández¹,

Fleischer

Vallejo-Díez

et al. 2020

CP Stem Cell Biology

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Genome editing has become one of the most powerful tools in present‐day stem cell and regenerative medicine research, but despite its rapid acceptance and widespread use, some elements of the technology still need improvement. In this unit, we present data regarding the use of a new, more efficient type of transcription activator‐like effector nuclease (TALEN) for gene editing. Our group has generated bicistronic genes in which classical TALEN coding sequences are linked by 2A elements to different reporter molecules, such as fluorochromes (TALEN‐F) or membrane receptors (TALEN‐M). This structure results in two proteins transcribed from the same transcript, of which the second (the reporter) can be used as the target for selection by fluorescence‐assisted cell sorting (FACS) or magnetic‐activated cell sorting (MACS). The application of these new TALEN genes allows a rapid enrichment of cells in which both members of the TALEN pair are active, thus eliminating the need for lengthy selection in culture and laborious characterization of a large number of clones. © 2020 by John Wiley & Sons, Inc. Basic Protocol 1: Generation of new TALENs Basic Protocol 2: Genome editing using TALEN‐F Alternate Protocol 1: Generation of TALEN‐M Support Protocol 1: mRNA in vitro transcription (IVT) of TALEN‐T2A‐reporter expression vector Alternate Protocol 2: Editing of primary T cells using TALEN‐M Basic Protocol 3: Verifying gene editing Support Protocol 2: Rapid expansion protocol for edited T‐cells

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Sendai virus, an RNA virus with no risk of genomic integration, delivers CRISPR/Cas9 for efficient gene editing

Cited by 41 publications

References 57 publications

Gene therapy clinical trials worldwide to 2017: An update

Gene therapy clinical trials worldwide to 2017: An update

Engineering an RNA-based tissue-specific platform for genetic editing through use of a miRNA-enabled Cas12a

New Bicistronic TALENs Greatly Improve Genome Editing

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