High efficiency lentiviral gene delivery in non-dividing cells by deoxynucleoside treatment

2002 ◽  
Vol 4 (2) ◽  
pp. 161-169 ◽  
Author(s):  
Elisabetta Ravot ◽  
Giuditta Comolli ◽  
Franco Lori ◽  
Julianna Lisziewicz
Keyword(s):  
Gene Delivery ◽  
High Efficiency ◽  
Dividing Cells

Recent advances in the analysis full/empty capsid ratio and genome integrity of adeno-associated virus (AAV) gene delivery vectors

2020 ◽  
Vol 20 ◽  
Author(s):  
L. Hajba ◽  
A. Guttman
Keyword(s):  
Gene Delivery ◽  
High Efficiency ◽  
Analytical Techniques ◽  
Viral Gene ◽  
Adeno Associated Virus ◽  
Gene Delivery Vectors ◽  
Empty Capsid ◽  
Purification Methods ◽  
Viral Gene Delivery

: Adeno-associated virus (AAV) is one of the most promising viral gene delivery vectors with long-term gene expression and disease correction featuring high efficiency and excellent safety in human clinical trials. During the production of AAV vectors,there are several quality control (QC)parameters that should be rigorously monitored to comply with clini-cal safety and efficacy. This review gives a short summary of the most frequently used AVV production and purification methods,focusing on the analytical techniques applied to determine the full/empty capsid ratio and the integrity of the encapsidated therapeutic DNA of the products.

Polyethylenimine-grafted polyamidoamine conjugates for gene delivery with high efficiency and low cytotoxicity

2014 ◽  
Vol 22 (7) ◽  
pp. 757-764 ◽  
Author(s):  
Tae-Hun Kim ◽  
Ho Won Seo ◽  
Jin Han ◽  
Kyung Soo Ko ◽  
Joon Sig Choi
Keyword(s):  
Gene Delivery ◽  
High Efficiency ◽  
Low Cytotoxicity

scholarly journals High-efficiency endovascular gene delivery via therapeutic ultrasound

2001 ◽  
Vol 37 (7) ◽  
pp. 1975-1980 ◽  
Author(s):  
Philippe G Amabile ◽  
Jacob M Waugh ◽  
Thomas N Lewis ◽  
Christopher J Elkins ◽  
Wolfgang Janas ◽  
...  
Keyword(s):  
Gene Delivery ◽  
High Efficiency ◽  
Therapeutic Ultrasound

Abstract 17357: Direct Cardiac Reprogramming Using Combinatorial Modified mRNA

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Keerat Kaur ◽  
Asharee Mahmoud ◽  
Hanna Girard ◽  
Ann Anu Kurian ◽  
Magdalena Zak ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Gene Delivery ◽  
High Efficiency ◽  
De Novo ◽  
Heart Function ◽  
Lineage Tracing ◽  
Dominant Negative ◽  
Post Myocardial Infarction ◽  
Transduction Efficiency

Introduction: Despite various clinical modalities, ischemic heart disease remains among the leading causes of mortality and morbidity worldwide. The elemental problem is the immense loss of cardiomyocytes (CMs) post-myocardial infarction (MI). Reprogramming non- cardiomyocytes (non-CMs) into cardiomyocyte (CM)-like cells in vivo is a promising strategy for cardiac regeneration: however, the traditional viral delivery method hampered its application into clinical settings due to low and erratic transduction efficiency. Methods: We used a modified mRNA (modRNA) gene delivery platform to deliver different stoichiometry of cardiac-reprogramming genes (Gata4, Mef2c, Tbx5 and Hand2) together with reprogramming helper genes (Dominant Negative (DN)-TGFβ, DN- Wnt8a and Acid ceramidase (AC)), named 7G, to induce direct cardiac reprogramming post myocardial infarction (MI). Results: Here, we identified 7G modRNA cocktail as an important regulator ofthe cardiac reprogramming. Cardiac transfection with 7G modRNA doubled cardiac reprogramming efficiency (57%) in comparison to Gata4, Mef2C and Tbx5 (GMT) alone (28%) in vitro . By inducing MI in our lineage tracing model, we showed that one-time delivery of the 7G-modRNA cocktail reprogrammed ~25% of the non-CMs in the scar area to CM- like cells. Furthermore, 7G modRNA treated mice showed significantly improved cardiac function, longer survival, reduced scar size and greater capillary density than control mice 28 days post-MI. We attributed the improvement in heart function post modRNA delivery of 7G or 7G with increased Hand2 ratio (7G-GMT Hx2) to significant upregulation of 15 key angiogenic factors without any signs of angioma or edema. Conclusions: 7G or 7G GMT HX2 modRNA cocktails boosts de novo CM-like cells and promotes cardiovascular regeneration post-MI. Thus, we highlight that this gene delivery approach not only has high efficiency but also high margin of safety for translation to clinics.

Early entry and deformation of macropinosomes correlates with high efficiency of decaarginine-polyethylene glycol-lipid-mediated gene delivery

2012 ◽  
Vol 14 (4) ◽  
pp. 262-271 ◽  
Author(s):  
Shouhei Kobayashi ◽  
Yoshiyuki Hattori ◽  
Hiroko Osakada ◽  
Kazunori Toma ◽  
Yoshie Maitani ◽  
...  
Keyword(s):  
Polyethylene Glycol ◽  
Gene Delivery ◽  
High Efficiency ◽  
Early Entry

scholarly journals Long-Circulating Polymeric Nanovectors for Tumor-Selective Gene Delivery

2005 ◽  
Vol 4 (6) ◽  
pp. 615-625 ◽  
Author(s):  
Sushma Kommareddy ◽  
Sandip B. Tiwari ◽  
Mansoor M. Amiji
Keyword(s):  
Gene Therapy ◽  
Gene Delivery ◽  
Viral Vectors ◽  
High Efficiency ◽  
Transfection Efficiency ◽  
The United States ◽  
Medical Intervention ◽  
Hydrophilic Polymers ◽  
Circulation Time ◽  
Viral Gene

Significant advances in the understanding of the genetic abnormalities that lead to the development, progression, and metastasis of neoplastic diseases has raised the promise of gene therapy as an approach to medical intervention. Most of the clinical protocols that have been approved in the United States for gene therapy have used the viral vectors because of the high efficiency of gene transfer. Conventional means of gene delivery using viral vectors, however, has undesirable side effects such as insertion of mutational viral gene into the host genome and development of replication competent viruses. Among non-viral gene delivery methods, polymeric nanoparticles are increasingly becoming popular as vectors of choice. The major limitation of these nanoparticles is poor transfection efficiency at the target site after systemic administration due to uptake by the cells of reticuloendothelial system (RES). In order to reduce the uptake by the cells of the RES and improve blood circulation time, these nanoparticles are coated with hydrophilic polymers such as poly(ethylene glycol) (PEG). This article reviews the use of such hydrophilic polymers employed for improving the circulation time of the nanocarriers. The mechanism of polymer coating and factors affecting the circulation time of these nanocarriers will be discussed. In addition to the long circulating property, modifications to improve the target specificity of the particles and the limitations of steric protection will be analyzed.

scholarly journals Drag-and-drop genome insertion without DNA cleavage with CRISPR-directed integrases

2021 ◽  
Author(s):  
Eleonora I. Ioannidi ◽  
Matthew T. N. Yarnall ◽  
Cian Schmitt-Ulms ◽  
Rohan N. Krajeski ◽  
Justin Lim ◽  
...  
Keyword(s):  
Genome Editing ◽  
Dna Cleavage ◽  
High Efficiency ◽  
End Joining ◽  
Differentiated Cells ◽  
Dividing Cells ◽  
Gene Integration ◽  
Fluorescent Tagging ◽  
Non Homologous End Joining ◽  
Drag And Drop

Programmable and multiplexed genome integration of large, diverse DNA cargo independent of DNA repair remains an unsolved challenge of genome editing. Current gene integration approaches require double-strand breaks that evoke DNA damage responses and rely on repair pathways that are inactive in terminally differentiated cells. Furthermore, CRISPR-based approaches that bypass double stranded breaks, such as Prime editing, are limited to modification or insertion of short sequences. We present Programmable Addition via Site-specific Targeting Elements, or PASTE, which achieves efficient and versatile gene integration at diverse loci by directing insertion with a CRISPR-Cas9 nickase fused to both a reverse transcriptase and serine integrase. Without generating double stranded breaks, we demonstrate integration of sequences as large as ~36 kb with rates between 10-50% at multiple genomic loci across three human cell lines, primary T cells, and quiescent non-dividing primary human hepatocytes. To further improve PASTE, we discover thousands of novel serine integrases and cognate attachment sites from metagenomes and engineer active orthologs for high-efficiency integration using PASTE. We apply PASTE to fluorescent tagging of proteins, integration of therapeutically relevant genes, and production and secretion of transgenes. Leveraging the orthogonality of serine integrases, we engineer PASTE for multiplexed gene integration, simultaneously integrating three different genes at three genomic loci. PASTE has editing efficiencies comparable to or better than those of homology directed repair or non-homologous end joining based integration, with activity in non-dividing cells and fewer detectable off-target events. For therapeutic applications, PASTE can be delivered as mRNA with synthetically modified guides to programmably direct insertion of DNA templates carried by AAV or adenoviral vectors. PASTE expands the capabilities of genome editing via drag-and-drop gene integration, offering a platform with wide applicability for research, cell engineering, and gene therapy.

Understanding why the same gene delivery vector behaves differently in different cell types is essential for developing more adaptable transfection systems

2021 ◽  
Author(s):  
Moataz Dowaidar
Keyword(s):  
Gene Delivery ◽  
High Efficiency ◽  
Transfection Efficiency ◽  
Cell Metabolism ◽  
New Technology ◽  
Cell Types ◽  
Viral Gene ◽  
Proliferation Capacity ◽  
Delivery Vector ◽  
Different Cell Types

Since their origin, non-viral gene delivery reagents have evolved into a variety of effective delivery reagents with a variety of components and designs, and are widely used in gene therapy and gene engineering. A flood of successful commercial gene delivery reagents has also developed, and PEI has emerged as the "gold standard" for the industry. On the other hand, their transfection efficiency must be enhanced and their cell toxicity must be reduced. In recent years, toxicity, efficiency and targeted investigations have progressed. In addition to creating and manufacturing reagents with reduced toxicity and higher efficiency, polypeptides that stimulate cell membrane perforation and tiny molecular compounds that can better compress pDNA, as well as various combinations with liposomes or polymer vectors, have demonstrated improved outcomes. However, most of these freshly created delivery vector reagents are still under investigation, and others require additional refinement to achieve high transfection efficiency and minimum toxicity. The processes behind the effects of various gene delivery reagents, genes, and drugs entering cells, as well as their transit, escape, and cell metabolism, are also unclear. This requires improving relevant research. Understanding why the same reagent reacts differently to different cell types is crucial to creating more adaptive transfection reagents for different cell lines. This is suggested because different cells have different growth cycles. Because of their weak proliferation capacity, primordial cells, for example, are harder to replicate.Artificial intelligence, real-world and virtual-world integration technology, big data, multiomics technology, and signal pathway research have all achieved substantial breakthroughs in recent years, and novel transfection reagents and drug delivery technologies are predicted to continue. It is worth examining how to take advantage of the scientific and high-efficiency benefits that new technology provides for research and how to solve the issues given by the in-depth examination of the selection and mechanism of action of novel composite materials in vector reagent creation.

High efficiency and low toxicity of polyethyleneimine modified Pluronics (PEI–Pluronic) as gene delivery carriers in cell culture and dystrophic mdx mice

2012 ◽  
Vol 22 (13) ◽  
pp. 6038 ◽  
Author(s):  
Mingxing Wang ◽  
Peijuan Lu ◽  
Bo Wu ◽  
Jay D. Tucker ◽  
Caryn Cloer ◽  
...  
Keyword(s):  
Cell Culture ◽  
Gene Delivery ◽  
High Efficiency ◽  
Mdx Mice ◽  
Low Toxicity
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