With recent advances in therapeutics exploiting the growing number of potential nucleic acid molecules used as drugs, in vivo-jetPEI™ has become the non-viral delivery reagent of choice for efficient and reproducible nucleic acid delivery in animal models and in the clinic.

in vivo-jetPEI™ was initially developed to deliver DNA and oligonucleotides in order to mediate gene expression in various tissues upon in vivo administration. For example In vivo imaging shows high level of gene expression in the lung upon intravenous injection of a Luciferase encoding plasmid in mice (Fig.1 and Table 1). Depending on the route of administration, in vivo-jetPEI™-mediated gene expression was also observed in the brain, liver, pancreas, spleen, kidney, heart, bladder, skin, retina, artery, etc.

Figure 1. Systemic delivery using in vivo-jetPEI™. Bioluminescent imaging of luciferase expression in living Balb/C mouse using a cooled camera 24 h after gene delivery. pCMVLuc (50 µg) was complexed with in vivo-jetPEI™ in 400 µl of 5% glucose solution and injected into the tail vein. Courtesy J.L. Coll.

The stability of in vivo-jetPEI™DNA complexes allows the use of numerous routes of administration shown in Figure 2. The targeted organs however depend mainly on the injection route.

Routes and target organ:

Upon intravenous injection, in vivo-jetPEI™mediated DNA delivery leads to gene expression in the lung (Fig 1 and Fig 3) liver, pancreas, spleen, kidney, heart, bladder and artery. In addition, in vivo-jetPEI™ is also well-adapted for local delivery such as application onto the skin, intratumoral, intracerebral or intra-articular injections (see Table 1). For experimental conditions using in vivo-jetPEI™, see:

Technical Note: Guidelines to set up your gene delivery experiments in mice

Literature references are available in the Product citations online database.

Figure 2. Successful delivery routes in mouse using in vivo-jetPEI™.

Animal models:
in vivo-jetPEI™ was successfully used to deliver nucleic acids to a wide range of species including mouse, rat, guinea pig, duck, rabbit, monkey, goat, sheep, chicken, quail, hamster, cow, tadpole, shrimp and fish. As a result, the experimental set up can be adapted to most species. Our technical support team will be pleased to assist you in order to adjust the reagent to nucleic acid ratio according to your model.

Technical Note: In vivo publications with Polyplus reagents using the most common administration routes

Technical Note: In vivo publications with Polyplus reagents by target organ/tissue

Being versatile and robust, in vivo-jetPEI™ is a reagent of choice to inhibit gene expression and to perform RNA interference in selected organs using synthetic siRNA including siRNA with chemically modified nucleobases and Sticky siRNA, a novel design of siRNA leading to higher gene silencing. Being very potent for DNA delivery, in vivo-jetPEI™ is suitable for shRNA and miRNA delivery in vivo. Specific inhibition of protein synthesis can be achieved by delivering siRNA, antisenses oligonucleotides or ribozymes with in vivo-jetPEI™ in animal models (Fig.3). Recently, in vivo-jetPEI™ and derivatives were successfully used as delivery reagents for anti-cancer therapy by RNA interference (Table 1).

Figure 3. RNA interference in the lung with in vivo-jetPEI™. pCMVLuc (40µg) was co-transfected with a control siRNA (upper) or with 10 µg of specific anti-Luc siRNA (lower). The complexes were injected into the tail vein of nude mice. Luciferase gene expression was monitored in living mice 24 h later by bioluminescence imaging using a cooled CCD camera.

shRNA
Since in vivo-jetPEI™ was developed for DNA delivery, it has been used successfully for shRNA approaches for RNA interference. Interestingly, it was used for gene silencing in the liver (George and Tsutsumi (2007), Gene Ther 14:890; Parajpe et al. (2007),Hepatology 45:147). Another interesting application in keeping with recent RNAi therapeutic developments in the clinic is the use of shRNA for RNAi in the retina. The method has been optimized by Liao and collaborators (Liao et al. (2007), BioTechniques 42: 285). Furthermore, in vivo-jetPEI™ is also suitable for shRNA approaches for RNAi in the brain (Hassani et al. (2007), NAR 35:e65).

siRNA
For siRNA delivery in vivo, several groups have also used in vivo-jetPEI™ successfully. This reagent is suitable for any siRNAs whether they include chemically modified nucleobases or not. Current successfully targeted organs in mice include vacsular endothelial cells (Choi et al., (2008) J Biol Chem 283; 20186 ), skin (Murase D., et al., (2009) J Biol Chem 284; 4343), subcutaneous xenograft tumors (Storci G., (2008) J Pathol 214; 25; Urban-Klein B. et al., (2005) Gene Ther. 12; 461, lung (Lively et al., (2008) J Allergy Clin immunol, 121;88) and dendritic cells (Poeck H., (2008) , Nature Med 14;1256; Cubillos-Ruiz (2009) J. Clin. Invest. in press).To consult all references see Table 1 or browse the product citation database.

STICKY SIRNA™ (ssiRNA): a novel approach
In order to improve siRNA delivery in vivo, Polyplus transfection has recently developed a novel type of siRNA. By including longer overhangs within the siRNA, we have generated STICKY siRNA (ssiRNA) that are able to form concatemers in the presence of in vivo-jetPEI™, thereby mimicking the structure of DNA and thus enhancing siRNA transfer. This design was shown to improve in vitro siRNA delivery and gene silencing efficiency (Bolcato-Bellemin (2007), PNAS 104:16050). More recently we have shown that intraperitoneal injection of STICKY siRNA™ cyclinB1 in mice using in vivo-jetPEI™ in a PC-3 tumor model, lead to an inhibition of tumor metastasis compared to mismatch and control. Moreover, mice survival is increased dramatically with STICKY siRNA™ delivered by in vivo-jetPEI™.

For more information see STICKY SIRNA™

Linear PEI such as in vivo-jetPEI™ does not induce any significant pro-inflammatory response after systemic injection of DNA or siRNA, especially when compared to other highly immunogenic reagents such as branched PEI or other lipid transfection reagents (Kawakami et al. (2006), J Pharmacol Exp Ther 317:1382).

Bonnet and collaborators also showed that there was no induction of major pro-inflammatory cytokines such as  TNF-alpha, IL6, IL12/IL23 and IL-1beta upon DNA and siRNA IV delivery using in vivo-jetPEI. Furthermore no increase in sera levels of hepatic enzyme were detected, suggesting that in vivo-jetPEI does not induce hepatotoxicity (Fig. 4, Bonnet et al. (2008), Pharm Res, 25:2972). In addition, linear PEI such as in vivo-jetPEI fails to generate neutralizing antibodies, thus permitting repeated administrations (Garzon et al. (2005), Vaccine 23:138).

Figure 4 : Serum concentration of TNF-α, IL12/IL23, IFNγ and IL6 following intravenous nucleic acid delivery using in vivo-jetPEI™ (N/P=8) respectively 1h, 6h, 12h and 6 h after delivery. The negative control consists of an IV injection of 5% glucose; the positive control an IP injection of 50 μg E.Coli LPS. (A) 40 μg DNA and (B) 40 ug siRNA were delivered with or without in vivo-jetPEI™.

Gene delivery using in vivo-jetPEI™ is reliable providing reproducible data (Fig 5.) with limited toxicity in contrast to other non-viral delivery methods. The reproducibility is linked to the unique properties of in vivo-jetPEI™. In 5% glucose, in vivo-jetPEI™ condenses nucleic acids into stable nanoparticles of ca. 50 nm in diameter (Fig 6). As a result, aggregation of blood cells is reduced compared to other reagents (Kircheis et al. (2001), Gene Ther 8: 28), thereby preventing a restriction in diffusion within tissues, erythrocyte aggregation and microembolia. These nanoparticles are sufficiently small to diffuse within the tissues and enter the cells by endocytosis. At the cellular level, in vivo-jetPEI™ possesses the unique feature of facilitating escape from the endosome using a proton sponge mechanism (Akinc et al. (2005), J Gene Med 7: 657). It also favors crossing of the nuclear membrane (Brunner et al. (2002), Mol Ther 5: 80).

Fig. 5. Systemic delivery using in vivo-jetPEI™. pCMVLuc (50 µg) was complexed with in vivo-jetPEI™ in 5% glucose and injected retro-orbitally. After 24 h, luciferase gene expression was assessed in the lung (n=8). Viability of mice was 100%.

Figure 6. in vivo-jetPEI™ forms small spherical particles with plasmid DNA. in vivo-jetPEI™/DNA complexes are prepared in 5% glucose solution at N/P ratio of 10. Complexes were added on a carbon covered grid and stained with uranyl acetate. Observation was carried out under a TEM. Bar is 100 nm. Complexes produced in glucose solution are discrete spheres having a mean size of 50 +/- 30 nm (Courtesy J-S Remy, Laboratoire Chimie Génétique, CNRS UMR 7514, Illkirch, France).