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Skin electroporation: effects on transgene expression, DNA persistence and local tissue environment.

Roos AK, Eriksson F, Timmons JA, Gerhardt J, Nyman U, Gudmundsdotter L, Bråve A, Wahren B, Pisa P - PLoS ONE (2009)

Bottom Line: Electrical pulses have been used to enhance uptake of molecules into living cells for decades.In vivo imaging of luciferase activity after electrovaccination demonstrated a rapid onset (minutes) and a long duration (months) of transgene expression.This study provides important insights to how DNA delivery by intradermal electrovaccination affects the local immunological responses of the skin, transgene expression and clearance of the plasmid.

View Article: PubMed Central - PubMed

Affiliation: Department of Oncology and Pathology, Cancer Center Karolinska R8:01, Immune and Gene Therapy Laboratory, Karolinska Institute, Stockholm, Sweden. roos.anki@gmail.com

ABSTRACT

Background: Electrical pulses have been used to enhance uptake of molecules into living cells for decades. This technique, often referred to as electroporation, has become an increasingly popular method to enhance in vivo DNA delivery for both gene therapy applications as well as for delivery of vaccines against both infectious diseases and cancer. In vivo electrovaccination (gene delivery followed by electroporation) is currently being investigated in several clinical trials, including DNA delivery to healthy volunteers. However, the mode of action at molecular level is not yet fully understood.

Methodology/principal findings: This study investigates intradermal DNA electrovaccination in detail and describes the effects on expression of the vaccine antigen, plasmid persistence and the local tissue environment. Gene profiling of the vaccination site showed that the combination of DNA and electroporation induced a significant up-regulation of pro-inflammatory genes. In vivo imaging of luciferase activity after electrovaccination demonstrated a rapid onset (minutes) and a long duration (months) of transgene expression. However, when the more immunogenic prostate specific antigen (PSA) was co-administered, PSA-specific T cells were induced and concurrently the luciferase expression became undetectable. Electroporation did not affect the long-term persistence of the PSA-expressing plasmid.

Conclusions/significance: This study provides important insights to how DNA delivery by intradermal electrovaccination affects the local immunological responses of the skin, transgene expression and clearance of the plasmid. As the described vaccination approach is currently being evaluated in clinical trials, the data provided will be of high significance.

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Related in: MedlinePlus

Plasmid persistence in skin over time.Histogram showing numbers of persisting plasmid copies at the indicated time-points after intradermal DNA injection of the specified doses. Error bars represent mean±SD (n = 6 individual samples). Each individual sample was analyzed in triplicate for each QPCR. The QPCR assay was run three times. EP, electroporation.
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pone-0007226-g004: Plasmid persistence in skin over time.Histogram showing numbers of persisting plasmid copies at the indicated time-points after intradermal DNA injection of the specified doses. Error bars represent mean±SD (n = 6 individual samples). Each individual sample was analyzed in triplicate for each QPCR. The QPCR assay was run three times. EP, electroporation.

Mentions: To investigate whether electrovaccination affects not only gene expression, but also the persistence of plasmid DNA at the injection site, mice were injected either with 2, 10 or 50 µg pVax-PSA plasmid followed by electroporation or with 50 µg pVax-PSA alone. DNA injection sites were surgically removed 7, 30 or 60 days after DNA administration and the number of persisting plasmid copies in the skin biopsies was determined using qPCR. The group that was not subjected to electroporation had a higher average number of plasmid copies at day 7 compared to mice receiving the same dose followed by electroporation, though the difference was not significant (Fig. 4). Plasmid persistence at day 30 or 60 after DNA administration did not differ between mice that received 50 µg DNA by needle injection without electroporation and mice receiving 50 µg DNA together with electroporation (Fig. 4). Taken together, the results demonstrate that the number of plasmids persisting in skin decreases over time and that adding electroporation to intradermal DNA administration does not significantly affect the number of persisting plasmid copies in skin.


Skin electroporation: effects on transgene expression, DNA persistence and local tissue environment.

Roos AK, Eriksson F, Timmons JA, Gerhardt J, Nyman U, Gudmundsdotter L, Bråve A, Wahren B, Pisa P - PLoS ONE (2009)

Plasmid persistence in skin over time.Histogram showing numbers of persisting plasmid copies at the indicated time-points after intradermal DNA injection of the specified doses. Error bars represent mean±SD (n = 6 individual samples). Each individual sample was analyzed in triplicate for each QPCR. The QPCR assay was run three times. EP, electroporation.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2748717&req=5

pone-0007226-g004: Plasmid persistence in skin over time.Histogram showing numbers of persisting plasmid copies at the indicated time-points after intradermal DNA injection of the specified doses. Error bars represent mean±SD (n = 6 individual samples). Each individual sample was analyzed in triplicate for each QPCR. The QPCR assay was run three times. EP, electroporation.
Mentions: To investigate whether electrovaccination affects not only gene expression, but also the persistence of plasmid DNA at the injection site, mice were injected either with 2, 10 or 50 µg pVax-PSA plasmid followed by electroporation or with 50 µg pVax-PSA alone. DNA injection sites were surgically removed 7, 30 or 60 days after DNA administration and the number of persisting plasmid copies in the skin biopsies was determined using qPCR. The group that was not subjected to electroporation had a higher average number of plasmid copies at day 7 compared to mice receiving the same dose followed by electroporation, though the difference was not significant (Fig. 4). Plasmid persistence at day 30 or 60 after DNA administration did not differ between mice that received 50 µg DNA by needle injection without electroporation and mice receiving 50 µg DNA together with electroporation (Fig. 4). Taken together, the results demonstrate that the number of plasmids persisting in skin decreases over time and that adding electroporation to intradermal DNA administration does not significantly affect the number of persisting plasmid copies in skin.

Bottom Line: Electrical pulses have been used to enhance uptake of molecules into living cells for decades.In vivo imaging of luciferase activity after electrovaccination demonstrated a rapid onset (minutes) and a long duration (months) of transgene expression.This study provides important insights to how DNA delivery by intradermal electrovaccination affects the local immunological responses of the skin, transgene expression and clearance of the plasmid.

View Article: PubMed Central - PubMed

Affiliation: Department of Oncology and Pathology, Cancer Center Karolinska R8:01, Immune and Gene Therapy Laboratory, Karolinska Institute, Stockholm, Sweden. roos.anki@gmail.com

ABSTRACT

Background: Electrical pulses have been used to enhance uptake of molecules into living cells for decades. This technique, often referred to as electroporation, has become an increasingly popular method to enhance in vivo DNA delivery for both gene therapy applications as well as for delivery of vaccines against both infectious diseases and cancer. In vivo electrovaccination (gene delivery followed by electroporation) is currently being investigated in several clinical trials, including DNA delivery to healthy volunteers. However, the mode of action at molecular level is not yet fully understood.

Methodology/principal findings: This study investigates intradermal DNA electrovaccination in detail and describes the effects on expression of the vaccine antigen, plasmid persistence and the local tissue environment. Gene profiling of the vaccination site showed that the combination of DNA and electroporation induced a significant up-regulation of pro-inflammatory genes. In vivo imaging of luciferase activity after electrovaccination demonstrated a rapid onset (minutes) and a long duration (months) of transgene expression. However, when the more immunogenic prostate specific antigen (PSA) was co-administered, PSA-specific T cells were induced and concurrently the luciferase expression became undetectable. Electroporation did not affect the long-term persistence of the PSA-expressing plasmid.

Conclusions/significance: This study provides important insights to how DNA delivery by intradermal electrovaccination affects the local immunological responses of the skin, transgene expression and clearance of the plasmid. As the described vaccination approach is currently being evaluated in clinical trials, the data provided will be of high significance.

Show MeSH
Related in: MedlinePlus