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A peptide-based vector for efficient gene transfer in vitro and in vivo.

ABSTRACT: Finding suitable nonviral delivery vehicles for nucleic acid-based therapeutics is a landmark goal in gene therapy. Cell-penetrating peptides (CPPs) are one class of delivery vectors that has been exploited for this purpose. However, since CPPs use endocytosis to enter cells, a large fraction of peptides remain trapped in endosomes. We have previously reported that stearylation of amphipathic CPPs, such as transportan 10 (TP10), dramatically increases transfection of oligonucleotides in vitro partially by promoting endosomal escape. Therefore, we aimed to evaluate whether stearyl-TP10 could be used for the delivery of plasmids as well. Our results demonstrate that stearyl-TP10 forms stable nanoparticles with plasmids that efficiently enter different cell-types in a ubiquitous manner, including primary cells, resulting in significantly higher gene expression levels than when using stearyl-Arg9 or unmodified CPPs. In fact, the transfection efficacy of stearyl-TP10 almost reached the levels of Lipofectamine 2000 (LF2000), however, without any of the observed lipofection-associated toxicities. Most importantly, stearyl-TP10/plasmid nanoparticles are nonimmunogenic, mediate efficient gene delivery in vivo, when administrated intramuscularly (i.m.) or intradermally (i.d.) without any associated toxicity in mice.

SUBMITTER: Lehto T

PROVIDER: S-EPMC3149163 | biostudies-literature | 2011 Aug

REPOSITORIES: biostudies-literature

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A peptide-based vector for efficient gene transfer in vitro and in vivo.

Lehto Taavi T Simonson Oscar E OE Mäger Imre I Ezzat Kariem K Sork Helena H Copolovici Dana-Maria DM Viola Joana R JR Zaghloul Eman M EM Lundin Per P Moreno Pedro M D PM Mäe Maarja M Oskolkov Nikita N Suhorutšenko Julia J Smith C I Edvard CI Andaloussi Samir E L SE

Molecular therapy : the journal of the American Society of Gene Therapy 20110222 8

Finding suitable nonviral delivery vehicles for nucleic acid-based therapeutics is a landmark goal in gene therapy. Cell-penetrating peptides (CPPs) are one class of delivery vectors that has been exploited for this purpose. However, since CPPs use endocytosis to enter cells, a large fraction of peptides remain trapped in endosomes. We have previously reported that stearylation of amphipathic CPPs, such as transportan 10 (TP10), dramatically increases transfection of oligonucleotides in vitro pa ...[more]

PMID: 21343913

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Project description:Efficient infection with adenovirus (Ad) vectors based on serotype 5 (Ad5) requires the presence of coxsackievirus-adenovirus receptors (CAR) and alpha(v) integrins on cells. The paucity of these cellular receptors is thought to be a limiting factor for Ad gene transfer into hematopoietic stem cells. In a systematic approach, we screened different Ad serotypes for interaction with noncycling human CD34(+) cells and K562 cells on the level of virus attachment, internalization, and replication. From these studies, serotype 35 emerged as the variant with the highest tropism for CD34(+) cells. A chimeric vector (Ad5GFP/F35) was generated which contained the short-shafted Ad35 fiber incorporated into an Ad5 capsid. This substitution was sufficient to transplant all infection properties from Ad35 to the chimeric vector. The retargeted, chimeric vector attached to a receptor different from CAR and entered cells by an alpha(v) integrin-independent pathway. In transduction studies, Ad5GFP/F35 expressed green fluorescent protein (GFP) in 54% of CD34(+) cells. In comparison, the standard Ad5GFP vector conferred GFP expression to only 25% of CD34(+) cells. Importantly, Ad5GFP transduction, but not Ad5GFP/F35, was restricted to a specific subset of CD34(+) cells expressing alpha(v) integrins. The actual transduction efficiency was even higher than 50% because Ad5GFP/F35 viral genomes were found in GFP-negative CD34(+) cell fractions, indicating that the cytomegalovirus promoter used for transgene expression was not active in all transduced cells. The chimeric vector allowed for gene transfer into a broader spectrum of CD34(+) cells, including subsets with potential stem cell capacity. Fifty-five percent of CD34(+) c-Kit(+) cells expressed GFP after infection with Ad5GFP/F35, whereas only 13% of CD34(+) c-Kit(+) cells were GFP positive after infection with Ad5GFP. These findings represent the basis for studies aimed toward stable gene transfer into hematopoietic stem cells.

Dataset Information

A peptide-based vector for efficient gene transfer in vitro and in vivo.

Publications

A peptide-based vector for efficient gene transfer in vitro and in vivo.

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