Visualization of targeted transduction by engineered lentiviral vectors

Joo, Kye-Il; Wang, Pin

doi:10.1038/gt.2008.87

Cited by 35 publications

(47 citation statements)

References 47 publications

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“…These data suggest that the interaction between vector CD4 and cell surface Env play a pivotal role in initiating the targeting. We further postulated that the binding should induce receptor-mediated endocytosis to uptake LVs into endosomal compartments, where the low pH environment can prompt the conformation change of the vector-carrying FM molecule and induce fusion between endosome and vector membranes (Joo and Wang, 2008). Using an endosomal neutralization assay, we found that the transduction efficiencies of the targeting vectors decreased with increasing concentrations of NH 4 Cl.…”

Section: Discussionmentioning

confidence: 99%

“…We further incorporated several different fusogenic molecules (FMs) that were engineered based on Kielian and co-workers’ study (Lu et al, 1999) and showed that these FMs could significantly improve the transduction efficiency of targeting vectors (Yang et al, 2008). An entry study revealed that the engineered vector particles can be internalized through clathrin-dependent endocytosis upon binding to target cells and further transported into the endosomal compartment, where the FMs on the vector surface sense the low pH and undergo a conformation change to trigger fusion, releasing the viral core into the cytosol (Joo and Wang, 2008). …”

Section: Introductionmentioning

confidence: 99%

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Engineered lentiviral vectors pseudotyped with a CD4 receptor and a fusogenic protein can target cells expressing HIV-1 envelope proteins

et al. 2011

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Lentiviral vectors (LVs) derived from human immunodeficiency virus type 1 (HIV-1) are promising vehicles for gene delivery because they not only efficiently transduce both dividing and non-dividing cells, but also maintain long-term transgene expression. Development of an LV system capable of transducing cells in a cell type-specific manner can be beneficial for certain applications that rely on targeted gene delivery. Previously it was shown that an inverse fusion strategy that incorporated an HIV-1 receptor (CD4) and its co-receptor (CXCR4 or CCR5) onto vector surfaces could confer to LVs the ability to selectively deliver genes to HIV-1 envelope-expressing cells. To build upon this work, we aim to improve its relatively low transduction efficiency and circumvent its inability to target multiple tropisms of HIV-1 by a single vector. We investigated a method to create LVs co-enveloped with the HIV-1 cellular receptor CD4 and a fusogenic protein derived from the Sindbis virus glycoprotein and tested its efficiency to selectively deliver genes into cells expressing HIV-1 envelope proteins. The engineered LV system yields a higher level of transduction efficiency and a broader tropism towards cells displaying the HIV-1 envelope protein (Env) than the previously developed system. Furthermore, we demonstrated in vitro that this engineered LV can preferentially deliver suicide gene therapy to HIV-1 envelope-expressing cells. We conclude that it is potentially feasible to target LVs towards HIV-1-infected cells by functional co-incorporation of the CD4 and fusogenic proteins, and provide preliminary evidence for further investigation on a potential alternative treatment for eradicating HIV-1-infected cells that produce drug-resistant viruses after highly active antiretroviral therapy (HAART).

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Engineered lentiviral vectors pseudotyped with a CD4 receptor and a fusogenic protein can target cells expressing HIV-1 envelope proteins

et al. 2011

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“…GFP-Vpr was cloned previously in our lab [19]. DsRed-hDC-SIGN was cloned by PCR-amplifying hDC-SIGN with HindIII and EcoR1 cut sites.…”

Section: Methodsmentioning

confidence: 99%

Visualization of DC-SIGN-Mediated Entry Pathway of Engineered Lentiviral Vectors in Target Cells

et al. 2013

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Dendritic cells (DCs) are potent antigen-presenting cells and therefore have enormous potential as vaccine targets. We have previously developed an engineered lentiviral vector (LV) that is pseudotyped with a mutated Sindbis virus glycoprotein (SVGmu), which is capable of targeting DCs through Dendritic Cell-specific ICAM3-grabbing Nonintegrin (DC-SIGN), a receptor that is predominantly expressed by DCs. In this study, we aimed to elucidate the internalization and trafficking mechanisms of this viral vector system through direct visualization of GFP-Vpr-tagged viral particles in target DCs, which was further corroborated by drug inhibition and dominant-negative mutants of cellular proteins that regulate the endocytic traffic. We demonstrated that our engineered LVs enter the cell via receptor-mediated clathrin- and dynamin-dependent endocytosis. Microtubule networks were also involved in a productive infection. Viral vector fusion was low-pH-dependent and occurred in the early endosomal stage of the intracellular transport. Autophagy was also examined for its effect on transduction efficiency, and we observed that enhanced autophage activity reduced vector infectivity, while suppressed autophagy boosted transduction efficiency. This study shed some light on the internalization and trafficking mechanisms of DC-directed LVs and offers some strategies to further improve the efficiency of LV-mediated gene therapy.

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“…야생형 녹색형광 단백질(wtGFP)의 아미노산을 점 돌연변이 시켜 만들어진 EGFP (enhanced green fluorescent protein) 단백질은 야생형에 비해 높은 강도를 나타내는 녹색형광 단백질로 (Cinelli et al, 2000), 세포 및 분자생물학 분야에서 녹색형광 발현 형질전환체를 제작하거나 형 질전환체 선별을 위한 선발마커 등에 흔히 이용되고 있으며 (Arun et al, 2005, Joo andWang, 2008), 우리나라에 서도 GFP 발현 쥐와 닭 모델이 개발 되어 있다 (Sim et al, 2009, Kwon et al, 2004 (Lee and Kim, 2000). 최초의 형질전환누에가 개발된 이래 형질전환 누에에서 기능성 단백질을 생산하기 위한 연구가 활발히 진행되고 있으며, 이 중 인공 실크 생산을 위한 곤충으로 서의 이용가능성이 대두되고 있다 (Imamura et al, 2003, Prudhomme and Couble, 2002, Satoshi et al 2007, Tamura et al, 2000.…”

Section: Introductionunclassified

Single-dose oral toxicity study of genetically modified silkworm expressing EGFP protein in ICR mouse

Jang¹,

Kim²,

Park³

et al. 2016

Korean Journal of Agricultural Science

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Silk has had a reputation as a luxurious and sensuous fabric but it is not popular due to the expensive price and poor durability. To develop the silk materials that apply the various industries, the artificially synthesized gene can be introduced into the silkworm and expressed in the silk gland. Transgenic silkworms for the mass production of green fluorescent silks are generated using a fibroin H-chain expression system. For commercial use, safety assessment of the transgenic silkworms is essential. The purpose of this study was to examine the potential acute oral toxicity of EGFP protein expressed in genetically modified (GM) fluorescence silkworm and to obtain the approximative lethal dose in the male and female at 6-weeks ICR mice. EGFP protein was fed at a dose of 2,000 mg/kg body weight in five male or five female mice. Mortalities, clinical findings and body weight changes were monitored for 1, 3, 7, 14 days after dosing. At the end of 14 day observation period, all mice were sacrificed, and the postmortem necropsy were performed. The test group was not observed death case. Also the effect was not admitted by test substance administration in common symptoms, the body weight and postmortem. The results of single-dose oral toxicity test showed that approximative lethal dose of EGFP protein expressed in fluorescence silkworm was considered to exceed the 2,000 mg/kg body weight in both sexes.

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Visualization of targeted transduction by engineered lentiviral vectors

Cited by 35 publications

References 47 publications

Engineered lentiviral vectors pseudotyped with a CD4 receptor and a fusogenic protein can target cells expressing HIV-1 envelope proteins

Engineered lentiviral vectors pseudotyped with a CD4 receptor and a fusogenic protein can target cells expressing HIV-1 envelope proteins

Visualization of DC-SIGN-Mediated Entry Pathway of Engineered Lentiviral Vectors in Target Cells

Single-dose oral toxicity study of genetically modified silkworm expressing EGFP protein in ICR mouse

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