Highly efficient cell-mediated gene transfer using non-viral vectors and FuGene™6: in vitro and in vivo studies

2000 ◽  
Vol 57 (8) ◽  
pp. 1326-1333 ◽  
Author(s):  
I. Hellgren* ◽  
V. Drvota ◽  
R. Pieper ◽  
S. Enoksson ◽  
P. Blomberg ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Viral Vectors ◽  
In Vivo Studies ◽  
Highly Efficient

Liver-directed gene transfer: a linear polyethylenimine derivative mediates highly efficient DNA delivery to primary hepatocytes in vitro and in vivo

1998 ◽  
Vol 5 (6) ◽  
pp. 369-375 ◽  
Author(s):  
I. Chemin ◽  
D. Moradpour ◽  
S. Wieland ◽  
W. -B. Offensperger ◽  
E. Walter ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Dna Delivery ◽  
Primary Hepatocytes ◽  
Highly Efficient ◽  
Linear Polyethylenimine

Functional Amphiphiles for Gene Delivery

MRS Bulletin ◽  
2005 ◽  
Vol 30 (9) ◽  
pp. 647-653 ◽  
Author(s):  
Philippe Barthélémy ◽  
Michel Camplo
Keyword(s):  
Gene Transfer ◽  
Gene Delivery ◽  
Viral Vectors ◽  
Transfection Efficiency ◽  
Gene Transfection ◽  
Research Activity ◽  
Safety Issues ◽  
Significant Research

AbstractThe design of safe and efficient gene transfer vectors remains one of the key challenges in gene therapy. Despite their remarkable transfection efficiency, viral vectors suffer from known safety issues. Consequently, significant research activity has been undertaken to develop nonviral approaches to gene transfer during the last decade. Numerous academic and industrial research groups are investigating synthetic cationic vectors, such as cationic amphiphiles, with the objective of increasing the gene transfection activity. Within this area, the development of functional synthetic vectors that respond to local environmental effects have met with success. These synthetic vectors are based on mechanistic principles and represent a significant departure from earlier systems. Many of these systems for gene delivery in vitro and in vivo are discussed in this article.

scholarly journals Canine Adenovirus Vectors for Lung-Directed Gene Transfer: Efficacy, Immune Response, and Duration of Transgene Expression Using Helper-Dependent Vectors

2006 ◽  
Vol 80 (3) ◽  
pp. 1487-1496 ◽  
Author(s):  
Anne Keriel ◽  
Céline René ◽  
Chad Galer ◽  
Joseph Zabner ◽  
Eric J. Kremer
Keyword(s):  
Gene Transfer ◽  
Respiratory Tract ◽  
Neutralizing Antibodies ◽  
Transgene Expression ◽  
Viral Vectors ◽  
Ex Vivo ◽  
Adenovirus Type ◽  
Transduction Efficiency

ABSTRACT A major hurdle to the successful clinical use of some viral vectors relates to the innate, adaptive, and memory immune responses that limit the efficiency and duration of transgene expression. Some of these drawbacks may be circumvented by using vectors derived from nonhuman viruses such as canine adenovirus type 2 (CAV-2). Here, we evaluated the potential of CAV-2 vectors for gene transfer to the respiratory tract. We found that CAV-2 transduction was efficient in vivo in the mouse respiratory tract, and ex vivo in well-differentiated human pulmonary epithelia. Notably, the in vivo and ex vivo efficiency was poorly inhibited by sera from mice immunized with a human adenovirus type 5 (HAd5, a ubiquitous human pathogen) vector or by human sera containing HAd5 neutralizing antibodies. Following intranasal instillation in mice, CAV-2 vectors also led to a lower level of inflammatory cytokine secretion and cellular infiltration compared to HAd5 vectors. Moreover, CAV-2 transduction efficiency was increased in vitro in human pulmonary cells and in vivo in the mouse respiratory tract by FK228, a histone deacetylase inhibitor. Finally, by using a helper-dependent CAV-2 vector, we increased the in vivo duration of transgene expression to at least 3 months in immunocompetent mice without immunosuppression. Our data suggest that CAV-2 vectors may be efficient and safe tools for long-term clinical gene transfer to the respiratory tract.

scholarly journals Critical Aspects of Viral Vectors for Gene Transfer into the Kidney

2000 ◽  
Vol 11 (suppl 2) ◽  
pp. S149-S153
Author(s):  
DAVID FAVRE ◽  
NICOLAS FERRY ◽  
PHILIPPE MOULLIER
Keyword(s):  
Skeletal Muscle ◽  
Gene Transfer ◽  
Viral Vectors ◽  
Viral Vector ◽  
Accurate Determination ◽  
Coagulation Factor ◽  
Factor Ix ◽  
Adeno Associated Virus

Abstract.Viral vectors have been usedin vitroandin vivofor more than a decade, with some significant results in specific situations,e.g., when recombinant adeno-associated virus is used for the long-term transduction of skeletal muscle in coagulation factor IX-deficient patients. However, the kidney has been quite difficult to transduce with any viral vector currently available. When viral transduction occurs, it is often heterogeneous, transient, and eventually associated with immune and toxic side effects. However, recombinant adeno-associated virus and lentiviral vectors remain to be fully evaluated in the kidney; the former is small enough to be filtered through the glomerular basement membrane. This may be critical, because glomerular filtration is required for DNA complex-mediated transduction of tubular cells. An alternative toin siturenal gene transfer is secretion of a therapeutic protein from a distant site, such as skeletal muscle. Several examples provide evidence that this could be a clinically relevant approach. It also may allow accurate determination of the pathophysiologic mechanisms involved in the establishment and maintenance of experimental glomerulonephritis.

Highly Efficient mRNA- and cDNA-Based Transient Gene Delivery into Human Progenitor Cells.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 5471-5471
Author(s):  
Jochen Greiner ◽  
Jan Torzewski ◽  
Peter Ponsaerts ◽  
Markus T. Rojewski ◽  
Desiree Kronawitter ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Gene Therapy ◽  
Stem Cell ◽  
Gene Transfer ◽  
Progenitor Cells ◽  
Transfection Efficiency ◽  
Highly Efficient ◽  
Human Cd34

Abstract The method of gene transfer into progenitor cells is critical as viral vector transduction involves the risk of tumor induction by non-specific genomic integration. Non-viral transfection systems often fail due to low transfection efficiency. However, gene transfer into human CD34+ hematopoietic progenitor (HPC) and mesenchymal stem cells (MSC) is an essential tool for in vitro- and in vivo-applications and therapeutic strategies such as tissue engineering and gene therapy. We recently reported an transient genetic labelling of human CD34+ HPC with deltaLNGFR-plasmid-DNA for in vivo application: Transient transfection was efficient for both, CD34+ HSC (41% ± 2%) and leukemia cell lines (55% ± 4.9%) using the method of nucleofection. Moreover, mature myeloid cells (CD66b+) derived from transfected human CD34+ HPC and leukemia cells maintained deltaLNGFR expression at a high percentage (70% ± 1.6% and 58% ± 2% respectively). In this work, we investigated labelling of CD34+ HPC with mRNA. Human CD34+ HPC and human MSC were transfected with in vitro-transcribed mRNA for deltaLNGFR, a marker gene approved for human in vivo-application, using nucleofection. EGFP was used as a control. 24h after nucleofection, FACS-analysis showed a higher transfection efficiency compared to plasmid transfected CD34+ HPC and MSC: A high transfection frequency was found for mRNA-transfected HPCs using deltaLNGFR (82.4±9.7%) and EGFP (88.7±2.6%). We found also a high transfection rate for MSC using the marker genes deltaLNGFR (92.4±3.6%) and EGFP (83.3±4.1%). Cell viability was not affected by mRNA-transfection. Moreover, differentiation assays of deltaLNGFR-selected MSC after transfection, showed that differentiation of MSC into mesenchymal cells like chondrocytes, adipocytes and osteoblasts was not affected by mRNA nucleofection. Taken together, mRNA based nucleofection is a powerful, highly efficient and non-toxic approach for transient labelling of human progenitor cells or, via transfection of selective proteins, for transient manipulation of stem cell function. It may thus be useful to transiently manipulate stem cell characteristics and combine principles of gene therapy and tissue engineering.

scholarly journals Biochemical and functional analysis of an adenovirus-based ligand complex for gene transfer

1994 ◽  
Vol 299 (1) ◽  
pp. 49-58 ◽  
Author(s):  
K J Fisher ◽  
J M Wilson
Keyword(s):  
Gene Transfer ◽  
Plasmid Dna ◽  
Viral Vectors ◽  
Recombinant Adenovirus ◽  
Particle Structure ◽  
Ligand Complex ◽  
Lysine Modification ◽  
Transgene Delivery

Ligand-mediated approaches to gene transfer offer an alternative to viral vectors for both in vivo and in vitro applications. Although a significant percentage of the plasmid-based DNA complex is lost to lysosomal degradation following receptor-mediated endocytosis, simultaneous infection with adenovirus has been shown to increase the level of transgene expression [Curiel, Agarwal, Wagner and Cotten (1991) Proc. Natl. Acad. Sci. U.S.A. 88, 8850-8854; Wagner, Zatloukal, Cotten, Kirlappos, Mechtler, Curiel and Birnstiel (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 6099-6103]. In this study we describe an adenovirus-based ligand complex where the plasmid DNA, polycation-ligand conjugate and adenovirus are contained within a single particle structure. At the core of the transfection particle is a replication-defective recombinant adenovirus encoding a cDNA minigene for human placenta alkaline phosphatase that was chemically modified with poly(L-lysine) (Ad-pLys). Electron microscopy of an adenovirus-based ligand complex formed by successively adding plasmid DNA and an asialo-orosomucoid-poly(L-lysine) conjugate to Ad-pLys revealed structures that appeared as intact viral particles coated with a dense biomolecular layer. Adenovirus-based ligand complexes containing either a luciferase or beta-galactosidase reporter plasmid were shown to efficiently deliver the plasmid transgene to cells that express the hepatic asialoglycoprotein receptor. Furthermore, the poly(L-lysine) modification greatly reduced the infectivity potential of the virus without causing a concomitant loss of augmented gene transfer. As an alternative to infectious virions, incomplete products of viral assembly were also considered as a source for endosomalytic activity. However, these defective virions were unable to significantly enhance plasmid transgene delivery.

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