Sodium phosphate enhances plasmid DNA expression in vivo

Hartikka, Jukka; Bozoukova, Vesselina; Jones, D. Brian; Mahajan, Rohit; Wloch, MK; Sawdey, M.; Buchner, Carol; Sukhu, Loretta; Km, Barnhart; Abai, AM; Meek, J; Shen, Nancy L. L.; Manthorpe, Marston

doi:10.1038/sj.gt.3301226

Cited by 47 publications

(35 citation statements)

References 52 publications

(66 reference statements)

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“…Recently it was reported that sodium phosphate enhances plasmid DNA expression in vivo possibly by inhibiting DNA degradation (Hartikka et al, 2000). When an electric pulse was not applied ( Figure 1A), luciferase activity with distilled water vehicle was about 3 times lower than with normal saline [87 ± 36 versus 269 ± 115 relative luminescence unit (RLU) per muscle, P > 0.05] and about 10 times lower than with sodium phosphate (87 ± 36 versus 904 ± 351 RLU per muscle, P < 0.05).…”

Section: Comparison Of the Effect Of The Three Vehiclesmentioning

confidence: 99%

Optimal salt concentration of vehicle for plasmid DNA enhances gene transfer mediated by electroporation

Lee

Cho²,

Jang³

et al. 2002

Exp Mol Med

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In vivo electroporation has emerged as a leading technology for developing nonviral gene therapies, and the various technical parameters governing electroporation efficiency have been optimized by both theoretical and experimental analysis. However, most electroporation parameters focused on the electric conditions and the preferred vehicle for plasmid DNA injections has been normal saline. We hypothesized that salts in vehicle for plasmid DNA must affect the efficiency of DNA transfer because cations would alter ionic atmosphere, ionic strength, and conductivity of their medium. Here, we show that half saline (71 mM) is an optimal vehicle for in vivo electroporation of naked DNA in skeletal muscle. With various salt concentrations, two reporter genes, luciferase and β β β β-galactosidase were injected intramuscularly under our optimal electric condition (125 V/cm, 4 pulses x 2 times, 50 ms, 1 Hz). Exact salt concentrations of DNA vehicle were measured by the inductively coupled plasma-atomic emission spectrometer (ICP-AES) and the conductivity change in the tissue induced by the salt in the medium was measured by Low-Frequency (LF) Impedance Analyzer. Luciferase expression increased as cation concentration of vehicle decreased and this result can be visualized by X-Gal staining. However, at lower salt concentration, transfection efficiency was diminished because the hypoosmotic stress and electrical injury by low conductivity induced myofiber damage. At optimal salt concentration (71 mM), we observed a 3-fold average increase in luciferase expression in comparison with the normal saline condition (p < 0.01). These results provide a valuable experimental parameter for in vivo gene therapy mediated by electroporation.

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Section: Comparison Of the Effect Of The Three Vehiclesmentioning

confidence: 99%

Optimal salt concentration of vehicle for plasmid DNA enhances gene transfer mediated by electroporation

Lee

Cho²,

Jang³

et al. 2002

Exp Mol Med

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“…Our results indicated that noncarrier naked CRT/E7 DNA in ddH2O or TE buffer can generate better E7-specific immune responses and antitumor effects than in PBS buffer (Figures 4b and d). Hartikka et al 37 observed that the concentration, osmolarity and pH of sodium phosphate can influence plasmid DNA expression in vivo. The optimum sodium phosphate concentration was 150 mM and the luciferase expression was 4.3-fold compared with saline.…”

Section: Discussionmentioning

confidence: 99%

“…The luciferase expression decreased significantly, when solved in sodium phosphate solution less than 40 mM, compared to that of saline. 37 The sodium concentration of PBS buffer was only 6.7 mM in this study. This might be the reason why the PBS group did not generate as potent antigen-specific immune responses and antitumor effects as the other two groups in this study.…”

Section: Discussionmentioning

confidence: 99%

Noncarrier naked antigen-specific DNA vaccine generates potent antigen-specific immunologic responses and antitumor effects

Sun

et al. 2009

Gene Ther

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Genetic immunization strategies have largely focused on the use of plasmid DNA with a gene gun. However, there remains a clear need to further improve the efficiency, safety, and cost of potential DNA vaccines. The gold particle-coated DNA format delivered through a gene gun is expensive, time and process consuming, and raises aseptic safety concerns. This study aims to determine whether a low-pressured gene gun can deliver noncarrier naked DNA vaccine without any particle coating, and generate similarly strong antigen-specific immunologic responses and potent antitumor effects compared with gold particle-coated DNA vaccine. Our results show that mice vaccinated with noncarrier naked chimeric CRT/E7 DNA lead to dramatic increases in the numbers of E7-specific CD8 + T-cell precursors and markedly raised titers of E7-specific antibodies. Furthermore, noncarrier naked CRT/E7 DNA vaccine generated potent antitumor effects against subcutaneous E7-expressing tumors and pre-established E7-expressing metastatic pulmonary tumors. In addition, mice immunized with noncarrier naked CRT/E7 DNA vaccine had significantly less burning effects on the skin compared with those vaccinated with gold particle-coated CRT/E7 DNA vaccine. We conclude that noncarrier naked CRT/E7 DNA vaccine delivered with a low-pressured gene gun can generate similarly potent immunologic responses and effective antitumor effects has fewer side effects, and is more convenient than conventional gold particle-coated DNA vaccine.

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“…Serum anti-NP IgG titers were measured in an indirect ELISA using 96-well plates (Corning Incorporated, Corning, NY, USA) coated with 71 ng/well of influenza A/PR/8/34 nucleoprotein (NP) purified from recombinant baculoviral extracts as previously described [19,21]. The anti-gB ELISA was performed as previously described [7].…”

Section: Antibody Elisa Assaysmentioning

confidence: 99%

“…Plasmid VR4700 was constructed by inserting the A/PR/8/34 influenza NP coding sequence into an expression plasmid containing the CMV immediate early 1 promoter/enhancer and intron A, a modified rabbit β-globin transcriptional terminator and an open reading frame encoding the kanamycin resistance gene [19,20]. Plasmids VCL6368 encoding a detoxified CMV antigen pp65, VCL6365 encoding a secreted form of the CMV gB antigen lacking the transmembrane and cytoplasmic domains, VR1255 encoding firefly luciferase, and VR2956 encoding feline erythropoietin (EPO) were constructed as previously described [7,21].…”

Section: Materials and Plasmid Constructsmentioning

confidence: 99%

Physical characterization and in vivo evaluation of poloxamer‐based DNA vaccine formulations

Hartikka

Geall

Bozoukova

et al. 2008

The Journal of Gene Medicine

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Sodium phosphate enhances plasmid DNA expression in vivo

Cited by 47 publications

References 52 publications

Optimal salt concentration of vehicle for plasmid DNA enhances gene transfer mediated by electroporation

Optimal salt concentration of vehicle for plasmid DNA enhances gene transfer mediated by electroporation

Noncarrier naked antigen-specific DNA vaccine generates potent antigen-specific immunologic responses and antitumor effects

Physical characterization and in vivo evaluation of poloxamer‐based DNA vaccine formulations

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