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The HDAC inhibitor FK228 enhances adenoviral transgene expression by a transduction-independent mechanism but does not increase adenovirus replication.

Danielsson A, Dzojic H, Rashkova V, Cheng WS, Essand M - PLoS ONE (2011)

Bottom Line: In the present study, we found that the highest increase in transgene expression was achieved when non-toxic concentrations of FK228 were added immediately after transduction, demonstrating that the main effect by which FK228 enhances transgene expression is transduction-independent.In some cases, the effects were dramatic, as demonstrated by an increase in CD40L expression by FK228 from 0.3% to 62% when the murine prostate cancer cell line TRAMP-C2 was transduced with Ad[CD40L].One unexpected finding was that FK228 decreased the transgene expression of an adenoviral vector with the prostate cell-specific PPT promoter in the human prostate adenocarcinoma cell lines LNCaP and PC-346C.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, Genetics and Pathology, Science for Life Laboratory, Uppsala University, Uppsala, Sweden. angelika.danielsson@igp.uu.se

ABSTRACT
The histone deacetylase inhibitor FK228 has previously been shown to enhance adenoviral transgene expression when cells are pre-incubated with the drug. Upregulation of the coxsackie adenovirus receptor (CAR), leading to increased viral transduction, has been proposed as the main mechanism. In the present study, we found that the highest increase in transgene expression was achieved when non-toxic concentrations of FK228 were added immediately after transduction, demonstrating that the main effect by which FK228 enhances transgene expression is transduction-independent. FK228 had positive effects both on Ad5 and Ad5/f35 vectors with a variety of transgenes and promoters, indicating that FK228 works mainly by increasing transgene expression at the transcriptional level. In some cases, the effects were dramatic, as demonstrated by an increase in CD40L expression by FK228 from 0.3% to 62% when the murine prostate cancer cell line TRAMP-C2 was transduced with Ad[CD40L]. One unexpected finding was that FK228 decreased the transgene expression of an adenoviral vector with the prostate cell-specific PPT promoter in the human prostate adenocarcinoma cell lines LNCaP and PC-346C. This is probably a consequence of alteration of the adenocarcinoma cell lines towards a neuroendocrine differentiation after FK228 treatment. The observations in this study indicate that FK228 enhances adenoviral therapy by a transduction-independent mechanism. Furthermore, since histone deacetylase inhibitors may affect the differentiation of cells, it is important to keep in mind that the activity and specificity of tissue- and tumor-specific promoters may also be affected.

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FK228 treatment enhances adenoviral-mediated CD40L expression.Prostate cancer cell lines were transduced with various MOI of Ad[CD40L] and cultured in medium with various concentrations of FK228 for 24 h. Cell surface expression of CD40L was analyzed by flow cytometry. Percentages of CD40L positive cells (upper right) and MFI values (lower right) are given. FK228 increased CD40L expression in a dose-dependent manner. One representative experiment out of three is shown.
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pone-0014700-g004: FK228 treatment enhances adenoviral-mediated CD40L expression.Prostate cancer cell lines were transduced with various MOI of Ad[CD40L] and cultured in medium with various concentrations of FK228 for 24 h. Cell surface expression of CD40L was analyzed by flow cytometry. Percentages of CD40L positive cells (upper right) and MFI values (lower right) are given. FK228 increased CD40L expression in a dose-dependent manner. One representative experiment out of three is shown.

Mentions: Since FK228 had an effect on the GFP transgene we next wanted to investigate if it can increase also therapeutic transgene expression. An Ad5-based vector with the therapeutic CD40 ligand transgene driven by the CMV promoter, Ad[CD40L], was studied in prostate cancer cell lines. Cells were transduced with the vector for 2 h, washed and then incubated with or without various concentrations of FK228. After 24 h of incubation, the cells were stained for CD40L expression and analyzed by flow cytometry. FK228 treatment increased the percentage of CD40L-expressing cells as well as MFI in a dose-dependent manner (Figure 4). The most pronounced effect was observed in TRAMP-C2 cells, which are normally difficult to transduce with Ad5-based vectors. At an MOI as low as 3 FFU/cell, the percentage of CD40L-expressing cells increased from 0.3% for untreated cells to 62% for FK228-treated (3 ng/ml) cells. At MOI 0.3, FK228 treatment increased CD40L expression about 3 times in LNCaP and 2 times in PC-346C.


The HDAC inhibitor FK228 enhances adenoviral transgene expression by a transduction-independent mechanism but does not increase adenovirus replication.

Danielsson A, Dzojic H, Rashkova V, Cheng WS, Essand M - PLoS ONE (2011)

FK228 treatment enhances adenoviral-mediated CD40L expression.Prostate cancer cell lines were transduced with various MOI of Ad[CD40L] and cultured in medium with various concentrations of FK228 for 24 h. Cell surface expression of CD40L was analyzed by flow cytometry. Percentages of CD40L positive cells (upper right) and MFI values (lower right) are given. FK228 increased CD40L expression in a dose-dependent manner. One representative experiment out of three is shown.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3040751&req=5

pone-0014700-g004: FK228 treatment enhances adenoviral-mediated CD40L expression.Prostate cancer cell lines were transduced with various MOI of Ad[CD40L] and cultured in medium with various concentrations of FK228 for 24 h. Cell surface expression of CD40L was analyzed by flow cytometry. Percentages of CD40L positive cells (upper right) and MFI values (lower right) are given. FK228 increased CD40L expression in a dose-dependent manner. One representative experiment out of three is shown.
Mentions: Since FK228 had an effect on the GFP transgene we next wanted to investigate if it can increase also therapeutic transgene expression. An Ad5-based vector with the therapeutic CD40 ligand transgene driven by the CMV promoter, Ad[CD40L], was studied in prostate cancer cell lines. Cells were transduced with the vector for 2 h, washed and then incubated with or without various concentrations of FK228. After 24 h of incubation, the cells were stained for CD40L expression and analyzed by flow cytometry. FK228 treatment increased the percentage of CD40L-expressing cells as well as MFI in a dose-dependent manner (Figure 4). The most pronounced effect was observed in TRAMP-C2 cells, which are normally difficult to transduce with Ad5-based vectors. At an MOI as low as 3 FFU/cell, the percentage of CD40L-expressing cells increased from 0.3% for untreated cells to 62% for FK228-treated (3 ng/ml) cells. At MOI 0.3, FK228 treatment increased CD40L expression about 3 times in LNCaP and 2 times in PC-346C.

Bottom Line: In the present study, we found that the highest increase in transgene expression was achieved when non-toxic concentrations of FK228 were added immediately after transduction, demonstrating that the main effect by which FK228 enhances transgene expression is transduction-independent.In some cases, the effects were dramatic, as demonstrated by an increase in CD40L expression by FK228 from 0.3% to 62% when the murine prostate cancer cell line TRAMP-C2 was transduced with Ad[CD40L].One unexpected finding was that FK228 decreased the transgene expression of an adenoviral vector with the prostate cell-specific PPT promoter in the human prostate adenocarcinoma cell lines LNCaP and PC-346C.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, Genetics and Pathology, Science for Life Laboratory, Uppsala University, Uppsala, Sweden. angelika.danielsson@igp.uu.se

ABSTRACT
The histone deacetylase inhibitor FK228 has previously been shown to enhance adenoviral transgene expression when cells are pre-incubated with the drug. Upregulation of the coxsackie adenovirus receptor (CAR), leading to increased viral transduction, has been proposed as the main mechanism. In the present study, we found that the highest increase in transgene expression was achieved when non-toxic concentrations of FK228 were added immediately after transduction, demonstrating that the main effect by which FK228 enhances transgene expression is transduction-independent. FK228 had positive effects both on Ad5 and Ad5/f35 vectors with a variety of transgenes and promoters, indicating that FK228 works mainly by increasing transgene expression at the transcriptional level. In some cases, the effects were dramatic, as demonstrated by an increase in CD40L expression by FK228 from 0.3% to 62% when the murine prostate cancer cell line TRAMP-C2 was transduced with Ad[CD40L]. One unexpected finding was that FK228 decreased the transgene expression of an adenoviral vector with the prostate cell-specific PPT promoter in the human prostate adenocarcinoma cell lines LNCaP and PC-346C. This is probably a consequence of alteration of the adenocarcinoma cell lines towards a neuroendocrine differentiation after FK228 treatment. The observations in this study indicate that FK228 enhances adenoviral therapy by a transduction-independent mechanism. Furthermore, since histone deacetylase inhibitors may affect the differentiation of cells, it is important to keep in mind that the activity and specificity of tissue- and tumor-specific promoters may also be affected.

Show MeSH
Related in: MedlinePlus