Prolonged Integration Site Selection of a Lentiviral Vector in the Genome of Human Keratinocytes

Qian, Wei; Wang, Yong; Li, Ruifu; Zhou, Xian‐Liang; Liu, Jing; Peng, Dai-zhi

doi:10.12659/msm.903094

Cited by 11 publications

(6 citation statements)

References 21 publications

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“…HPKs should be expanded after genetic modification by CRISPR/ Cas9 from single cells, allowing the selection of true knockouts and even knockins and their characterization by sequencing. Lentiviral or retroviral integration can disrupt genetic information, resulting in aberrant transcripts and possibly leading to cancer development (Hirsch et al, 2017;Moiani et al, 2012;Qian et al, 2017). This can be avoided using other currently available approaches, such as transfection of the recombinant Cas9 protein with the sgRNA of interest.…”

Section: Discussionmentioning

confidence: 99%

Genome Editing of Human Primary Keratinocytes by CRISPR/Cas9 Reveals an Essential Role of the NLRP1 Inflammasome in UVB Sensing

Fenini

Grossi

Contassot

et al. 2018

Journal of Investigative Dermatology

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By forming a protective barrier, epidermal keratinocytes represent the first line of defense against environmental insults. UVB radiation of the sun is a major challenge for the skin and can induce inflammation, aging, and eventually skin cancer. UVB induces an immune response in human keratinocytes resulting in activation and secretion of the proinflammatory cytokines proIL-1β and -18. This is mediated by an assembly of protein complexes, termed inflammasomes. However, the mechanisms underlying sensing of UVB by keratinocytes, and particularly the types of inflammasomes required for cytokine secretion, are a matter of debate. To address these questions, we established a protocol that allows the generation of CRISPR/Cas9-targeted human primary keratinocytes. Our experiments showed an essential role of the NLRP1 rather than the NLRP3 inflammasome in UVB sensing and subsequent IL-1β and -18 secretion by keratinocytes. Moreover, NLRP1 but not NLRP3 was required for inflammasome activation in response to nigericin, a potassium ionophore and well-established NLRP3 activator in immune cells. Because the CRISPR/Cas9-targeted cells retained their full differentiation capacity, genome editing of human primary keratinocytes might be useful for numerous research and medical applications.

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Section: Discussionmentioning

confidence: 99%

Genome Editing of Human Primary Keratinocytes by CRISPR/Cas9 Reveals an Essential Role of the NLRP1 Inflammasome in UVB Sensing

Fenini

Grossi

Contassot

et al. 2018

Journal of Investigative Dermatology

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“… 250 – 252 Lentiviral DNA integrates into the host genome in a non-random manner with preference for transcriptionally active sites. 253 – 255 …”

Section: Introductionmentioning

confidence: 99%

Viral vector platforms within the gene therapy landscape

Bulcha

Wang

et al. 2021

Sig Transduct Target Ther

796

572

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Throughout its 40-year history, the field of gene therapy has been marked by many transitions. It has seen great strides in combating human disease, has given hope to patients and families with limited treatment options, but has also been subject to many setbacks. Treatment of patients with this class of investigational drugs has resulted in severe adverse effects and, even in rare cases, death. At the heart of this dichotomous field are the viral-based vectors, the delivery vehicles that have allowed researchers and clinicians to develop powerful drug platforms, and have radically changed the face of medicine. Within the past 5 years, the gene therapy field has seen a wave of drugs based on viral vectors that have gained regulatory approval that come in a variety of designs and purposes. These modalities range from vector-based cancer therapies, to treating monogenic diseases with life-altering outcomes. At present, the three key vector strategies are based on adenoviruses, adeno-associated viruses, and lentiviruses. They have led the way in preclinical and clinical successes in the past two decades. However, despite these successes, many challenges still limit these approaches from attaining their full potential. To review the viral vector-based gene therapy landscape, we focus on these three highly regarded vector platforms and describe mechanisms of action and their roles in treating human disease.

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“…A report for self-inactivating MLV vectors in primary keratinocytes previously reported a reduced 'genotoxic' profile in transduced cells 26 . Integration sites for lentiviral vectors in a spontaneously immortalised keratinocyte cell line (HaCat) were also reported with integration preferences for gene and gene-rich regions as expected 27,28 . In This paper has been peer-reviewed and accepted for publication, but has yet to undergo copyediting and proof correction.…”

Section: Discussionmentioning

confidence: 86%

Generation and Clinical Application of Gene-Modified Autologous Epidermal Sheets in Netherton Syndrome: Lessons Learned from a Phase 1 Trial

Di¹,

Lwin

Petrova³

et al. 2019

Human Gene Therapy

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Prolonged Integration Site Selection of a Lentiviral Vector in the Genome of Human Keratinocytes

Cited by 11 publications

References 21 publications

Genome Editing of Human Primary Keratinocytes by CRISPR/Cas9 Reveals an Essential Role of the NLRP1 Inflammasome in UVB Sensing

Genome Editing of Human Primary Keratinocytes by CRISPR/Cas9 Reveals an Essential Role of the NLRP1 Inflammasome in UVB Sensing

Viral vector platforms within the gene therapy landscape

Generation and Clinical Application of Gene-Modified Autologous Epidermal Sheets in Netherton Syndrome: Lessons Learned from a Phase 1 Trial

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