Limits...
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.

Show MeSH

Related in: MedlinePlus

Differential gene expression in virus-transduced prostate cancer cells after 48 h of treatment with HDAC inhibitors.LNCaP and PC-346C were transduced with Ad[i/PPT-LUC] and then incubated with 3 ng/ml FK228 or 5 mM VPA for 48 h. RNA was extracted, cDNA was produced and gene expression was evaluated by quantitative PCR (triplicate samples). Gene expression in FK228 and VPA-treated cells was related to untreated cells. A two-fold difference in gene expression was set as a limit for true difference. The expression of prostate-associated genes (AR, ARA24, PSA, PSMA, SRD5A1, SRD5A2 and TARP) was not considerably affected by the HDACi. The expression of neuroendocrine-associated genes (β-tubIII, CgA, NSE and SYP) was increased in HDACi-treated cells.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3040751&req=5

pone-0014700-g007: Differential gene expression in virus-transduced prostate cancer cells after 48 h of treatment with HDAC inhibitors.LNCaP and PC-346C were transduced with Ad[i/PPT-LUC] and then incubated with 3 ng/ml FK228 or 5 mM VPA for 48 h. RNA was extracted, cDNA was produced and gene expression was evaluated by quantitative PCR (triplicate samples). Gene expression in FK228 and VPA-treated cells was related to untreated cells. A two-fold difference in gene expression was set as a limit for true difference. The expression of prostate-associated genes (AR, ARA24, PSA, PSMA, SRD5A1, SRD5A2 and TARP) was not considerably affected by the HDACi. The expression of neuroendocrine-associated genes (β-tubIII, CgA, NSE and SYP) was increased in HDACi-treated cells.

Mentions: The increased transgene expression from Ad[CgA-LUC] in the prostate cancer cell lines LNCaP and PC346-C and the decreased expression of Ad[i/PPT-LUC] upon HDACi treatment led us to further investigate the effect of FK228 or VPA on these cell lines. RNA expression of the genes normally controlled by the regulatory elements of PPT: PSA, PSMA and TARP were therefore evaluated along with genes involved in the androgen receptor pathway: AR, ARA24, SRD5A1 and SRD5A2, since the PPT promoter is partly regulated by testosterone. The prostate epithelial cell-associated genes: AR, ARA24, PSA, PSMA, SRD5A1, SRD5A2 and TARP were generally not affected (less than 2-fold up- or down-regulation) by the FK228 and VPA treatment. However, in LNCaP, the expression of AR was slightly decreased to just below a 2-fold down-regulation for both HDACi. In PC-346C, the expression of SRD5A1 was increased after VPA treatment and SRD5A2 expression was increased after both FK228 and VPA treatment while PSMA expression (VPA treatment) and TARP expression (FK228 and VPA treatment) were decreased (Figure 7).


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)

Differential gene expression in virus-transduced prostate cancer cells after 48 h of treatment with HDAC inhibitors.LNCaP and PC-346C were transduced with Ad[i/PPT-LUC] and then incubated with 3 ng/ml FK228 or 5 mM VPA for 48 h. RNA was extracted, cDNA was produced and gene expression was evaluated by quantitative PCR (triplicate samples). Gene expression in FK228 and VPA-treated cells was related to untreated cells. A two-fold difference in gene expression was set as a limit for true difference. The expression of prostate-associated genes (AR, ARA24, PSA, PSMA, SRD5A1, SRD5A2 and TARP) was not considerably affected by the HDACi. The expression of neuroendocrine-associated genes (β-tubIII, CgA, NSE and SYP) was increased in HDACi-treated cells.
© Copyright Policy
Related In: Results  -  Collection

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

pone-0014700-g007: Differential gene expression in virus-transduced prostate cancer cells after 48 h of treatment with HDAC inhibitors.LNCaP and PC-346C were transduced with Ad[i/PPT-LUC] and then incubated with 3 ng/ml FK228 or 5 mM VPA for 48 h. RNA was extracted, cDNA was produced and gene expression was evaluated by quantitative PCR (triplicate samples). Gene expression in FK228 and VPA-treated cells was related to untreated cells. A two-fold difference in gene expression was set as a limit for true difference. The expression of prostate-associated genes (AR, ARA24, PSA, PSMA, SRD5A1, SRD5A2 and TARP) was not considerably affected by the HDACi. The expression of neuroendocrine-associated genes (β-tubIII, CgA, NSE and SYP) was increased in HDACi-treated cells.
Mentions: The increased transgene expression from Ad[CgA-LUC] in the prostate cancer cell lines LNCaP and PC346-C and the decreased expression of Ad[i/PPT-LUC] upon HDACi treatment led us to further investigate the effect of FK228 or VPA on these cell lines. RNA expression of the genes normally controlled by the regulatory elements of PPT: PSA, PSMA and TARP were therefore evaluated along with genes involved in the androgen receptor pathway: AR, ARA24, SRD5A1 and SRD5A2, since the PPT promoter is partly regulated by testosterone. The prostate epithelial cell-associated genes: AR, ARA24, PSA, PSMA, SRD5A1, SRD5A2 and TARP were generally not affected (less than 2-fold up- or down-regulation) by the FK228 and VPA treatment. However, in LNCaP, the expression of AR was slightly decreased to just below a 2-fold down-regulation for both HDACi. In PC-346C, the expression of SRD5A1 was increased after VPA treatment and SRD5A2 expression was increased after both FK228 and VPA treatment while PSMA expression (VPA treatment) and TARP expression (FK228 and VPA treatment) were decreased (Figure 7).

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