Risks Associated With Lentiviral Vector Exposures and Prevention Strategies

Schlimgen, Ryan; Howard, John; Wooley, Dawn P.; Thompson, Maureen; Baden, Lindsey R.; Yang, Otto O.; Christiani, David C.; Mostoslavsky, Gustavo; Diamond, David V.; Duane, Elizabeth Gilman; Byers, Karen B.; Winters, Thomas; Gelfand, Jeffrey A.; Fujimoto, Gary; Hudson, T; Vyas, Jatin M.

doi:10.1097/jom.0000000000000879

Cited by 113 publications

(88 citation statements)

References 27 publications

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“…While LV-mChABC gene therapy has undeniable benefit, there remains concern about the safety of the use of the therapy in humans. Lentiviral vectors risk insertional mutagenesis, transgene oncogenesis, generation of replication-competent retrovirus's (Schlimgen et al, 2016), as well as the possibility of the modified prokaryote ChABC enzyme still being capable of eliciting an immune response. Is there a mammalian endogenous alternative?…”

Section: Introductionmentioning

confidence: 99%

Astrocyte-selective AAV-ADAMTS4 gene therapy combined with hindlimb rehabilitation promotes functional recovery after spinal cord injury

Griffin

Fackelmeier

Clemett

et al. 2019

Preprint

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Chondroitin sulphate proteoglycans (CSPGs) are inhibitors to axon regeneration and plasticity. Bacterial chondroitinase ABC degrades CSPGs and has been extensively reported to be therapeutic after SCI but there remain concerns for its clinical translation. A disintegrin and metalloproteinase with thrombospondin motifs-4 (ADAMTS4) is a human enzyme that catalyses the proteolysis of CSPG protein cores. Infusion of ADAMTS4 into the damaged spinal cord was previously shown to improve functional recovery after SCI, however, this therapy is limited in its enzyme form. Adeno-associated viral (AAV) vector gene therapy has emerged as the vector of choice for safe, robust and long-term transgene expression in the central nervous system. Here, an AAV expression cassette containing ADAMTS4 under the control of the astrocytic GfaABC 1 D promoter was packaged into an AAV5 vector. Sustained expression of ADAMTS4 was achieved in vitro and in vivo, leading to widespread degradation of CSPGs. AAV-ADAMTS4 resulted in significantly decreased lesion size, increased sprouting of hindlimb corticospinal tract axons, increased serotonergic fiber density caudal to the injury, and improved functional recovery after moderate contusive SCI.Hindlimb-specific exercise rehabilitation was used to drive neuroplasticity towards improving functional connections. The combination of hindlimb rehabilitation with AAV-ADAMTS4 led to enhanced functional recovery after SCI. Thus, widespread and long-term degradation of CSPGs through AAV-ADAMTS4 gene therapy in a combinational approach with rehabilitation represents a promising candidate for further preclinical development. AbbreviationsAAV Adeno-associated virus ADAMTS A disintegrin and metalloproteinase with thrombospondin motifs ChABC Chondroitinase ABC CS-GAG Chondroitin sulphate glycosaminoglycan CSPG Chondroitin sulphate proteoglycan ECM Extracellular matrix GAG Glycosaminoglycan LV Lentiviral vector

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Section: Introductionmentioning

confidence: 99%

Astrocyte-selective AAV-ADAMTS4 gene therapy combined with hindlimb rehabilitation promotes functional recovery after spinal cord injury

Griffin

Fackelmeier

Clemett

et al. 2019

Preprint

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“…Noteworthy, lentiviral transduction requires ongoing cellular division [46], which is incompatible with understanding the impacts of miRNA on resting NK cells. These infections and long culture times can have unintended consequences, convey insertional mutagenesis or alter the distribution or function of NK cell subsets inadvertently, and require strict safety precautions [45,47]. Hence, for short-term study or alteration of miRNA in primary NK cells, non-viral transfections are desirable.…”

Section: Discussionmentioning

confidence: 99%

Highly efficient serum-free manipulation of miRNA in human NK cells without loss of viability or phenotypic alterations is accomplished with TransIT-TKO

2020

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Natural killer (NK) cells are innate lymphocytes with functions that include target cell killing, inflammation and regulation. NK cells integrate incoming activating and inhibitory signals through an array of germline-encoded receptors to gauge the health of neighbouring cells. The reactive potential of NK cells is influenced by microRNA (miRNA), small non-coding sequences that interfere with mRNA expression. miRNAs are highly conserved between species, and a single miRNA can have hundreds to thousands of targets and influence entire cellular programs. Two miRNA species, miR-155-5p and miR-146a-5p are known to be important in controlling NK cell function, but research to best understand the impacts of miRNA species within NK cells has been bottlenecked by a lack of techniques for altering miRNA concentrations efficiently and without off-target effects. Here, we describe a nonviral and straightforward approach for increasing or decreasing expression of miRNA in primary human NK cells. We achieve >90% transfection efficiency without off-target impacts on NK cell viability, education, phenotype or function. This opens the opportunity to study and manipulate NK cell miRNA profiles and their impacts on NK cellular programs which may influence outcomes of cancer, inflammation and autoimmunity.

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“…Lentiviral vectors are powerful tools for genetic engineering and their use in clinical trials is rising. Nevertheless, the use of lentiviral vectors might be associated with safety concerns such as an increased risk for tumorigenesis (56). In contrast to the in vivo application of lentiviral vectors, ex vivo organ perfusion allows for the selective genetic modification of the target organ thereby reducing the possibility for off-target and systemic adverse effects.…”

Section: Safetymentioning

confidence: 99%

Genetic Engineering of the Kidney to Permanently Silence MHC Transcripts During ex vivo Organ Perfusion

et al. 2020

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Organ gene therapy represents a promising tool to correct diseases or improve graft survival after transplantation. Polymorphic variation of the major histocompatibility complex (MHC) antigens remains a major obstacle to long-term graft survival after transplantation. Previously, we demonstrated that MHC-silenced cells are protected against allogeneic immune responses. We also showed the feasibility to silence MHC in the lung. Here, we aimed at the genetic engineering of the kidney toward permanent silencing of MHC antigens in a rat model. We constructed a sub-normothermic ex vivo perfusion system to deliver lentiviral vectors encoding shRNAs targeting β2-microglobulin and the class II transactivator to the kidney. In addition, the vector contained the sequence for a secreted nanoluciferase. After kidney transplantation (ktx), we detected bioluminescence in the plasma and urine of recipients of an engineered kidney during the 6 weeks of post-transplant monitoring, indicating a stable transgene expression. Remarkably, transcript levels of β2-microglobulin and the class II transactivator were decreased by 70% in kidneys expressing specific shRNAs. Kidney genetic modification did not cause additional cell death compared to control kidneys after machine perfusion. Nevertheless, cytokine secretion signatures were altered during perfusion with lentiviral vectors as revealed by an increase in the secretion of IL-10, MIP-1α, MIP-2, IP-10, and EGF and a decrease in the levels of IL-12, IL-17, MCP-1, and IFN-γ. Biodistribution assays indicate that the localization of the vector was restricted to the graft. This study shows the potential to generate immunologically invisible kidneys showing great promise to support graft survival after transplantation and may contribute to reduce the burden of immunosuppression.

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Risks Associated With Lentiviral Vector Exposures and Prevention Strategies

Cited by 113 publications

References 27 publications

Astrocyte-selective AAV-ADAMTS4 gene therapy combined with hindlimb rehabilitation promotes functional recovery after spinal cord injury

Astrocyte-selective AAV-ADAMTS4 gene therapy combined with hindlimb rehabilitation promotes functional recovery after spinal cord injury

Highly efficient serum-free manipulation of miRNA in human NK cells without loss of viability or phenotypic alterations is accomplished with TransIT-TKO

Genetic Engineering of the Kidney to Permanently Silence MHC Transcripts During ex vivo Organ Perfusion

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