Polyamine biosynthesis inhibition enhances HSV‐1 thymidine kinase/ganciclovir‐mediated cytotoxicity in tumor cells

Pasanen, Tiina; Karppinen, Anne; Alhonen, Leena; Jänne, Juhani; Wahlfors, Jarmo

doi:10.1002/ijc.10942

Cited by 4 publications

(10 citation statements)

References 15 publications

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“…As demonstrated in our earlier study, also the 2-difluoromethyl ornithine (DFMO)-induced polyamine biosynthesis inhibition accompanied with a predominant cell cycle arrest can be used to enhance HSV-TK/GCV gene therapy (5). We showed that DFMO caused a long-lasting disturbance to the cell cycle of different tumor cells and a correct timing of DFMO and GCV treatments yielded synergistic cytotoxic effect.…”

Section: Introductionmentioning

confidence: 75%

“…In theory, this can be accomplished by arresting the cell population at a certain point of the cell cycle, releasing the block and then destroying the enriched S phase fraction of the cells with HSV-TK/GCV gene therapy. Perhaps the simplest way to introduce transient cell cycle arrest is to use serum deprivation, but our results have shown that increase of the S phase cells is too transient to be synergized with the HSV-TK/GCV gene therapy (5) and certainly this type of approach would not be useful in a clinical setting. Fortunately, there are many drugs available today that can halt cell proliferation at certain point of the cell cycle via different mechanisms.…”

Section: Introductionmentioning

confidence: 91%

“…The cell cycle phases of 9L and U-251 MG cells were analyzed after a 12 h treatment with different drugs (concentrations indicated above). Cell samples were collected immediately after drug removal and 2, 4, 6, 9, 12 and 24 h later, stained with propidium iodide and the DNA content was analyzed with flow cytometer as described before (5).…”

Section: Cell Lines and Viral Vectorsmentioning

confidence: 99%

“…Treatment (12 h) with cell cycle altering drugs was started 24 or 12 h after GCV treatment initiation (GCV1 and GCV2), simultaneously with GCV initiation (GCV3) or 12 h before the start of GCV treatment (GCV4). The TK-GFP positive cells (6000 cells/well in a 48-well plate) were incubated with 1 μg/ml GCV for 5 days, followed by cell viability analysis with MTT assay (cell proliferation kit II, Roche) as described before (5). Control cells were treated with different drugs for 12 h, followed by incubation without GCV for 60, 72, 84 and 96 h (GCV1, GCV2, GCV3 and GCV4 controls, respectively) and viability analysis with MTT assay.…”

Section: Cell Lines and Viral Vectorsmentioning

confidence: 99%

“…However, the rapid by-pass of the elevated S phase fraction is not considered a favorable feature for enhancement of the HSV-TK/GCV system. Our earlier study (5) revealed that elevated proportion of S phase cells induced by serum deprivation was very transient (passed in 24 h after release from the arrest) and could not be used to augment HSV-TK/GCV cytotoxicity. On the other hand, reversal of the DFMO-induced polyamine depletion and the accompanying cell cycle disturbance was a significantly longer process (proportion of S phases remained elevated for 2-3 days) and allowed synergy with the HSV-TK/GCV gene therapy.…”

Section: Cell Cycle Arrest Induced By Aphidicolin Hydroxyurea Lovasmentioning

confidence: 99%

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Polyamine depletion and cell cycle manipulation in combination with HSV thymidine kinase/ganciclovir cancer gene therapy

Wahlfors

Karppinen²,

Jänne³

et al. 2006

Int J Oncol

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Abstract. We have shown earlier that polyamine biosynthesis inhibition is accompanied by cell cycle alterations that can be utilized to enhance the efficacy of herpes simplex virus thymidine kinase -ganciclovir (HSV-TK/GCV) cancer gene therapy. In the present study, we asked 1) can the activated polyamine catabolism instead of biosynthesis inhibition be utilized to enhance the efficacy of HSV-TK/GCV gene therapy, and 2) can other known cell cycle inhibitors be used to make tumor cells more sensitive to this form of gene therapy? We show, using rat (9L) and human (U251-MG) glioma cell populations with 15% of HSV-TK-positive cells that DENSPM-induced activation of polyamine catabolism caused a profound polyamine deprivation in U251-MG cells, but there were no associated cell cycle effects in these cells. Consequently, we did not see any enhancement of the HSV-TK/GCV system. Aphidicolin, hydroxyurea, mimosine and resveratrol, but not lovastatin induced an apparent cell cycle arrest, followed by an intense but transient increase of the S phase cells after removal of the drug. This effect was shown to potentiate the HSV-TK/GCV cytotoxicity to some extent, especially in 9L cells and when the GCV treatment was started 0-24 h before the drug treatment. However, the enhancement was weaker than observed earlier with DFMO-induced cell cycle arrest and a considerable degree of the effect appeared to result from the growth-inhibitory actions of the drugs. In summary, we demonstrate that polyamine deprivation via DENSPM action is not associated with cell cycle effects and is not sufficient to cause enhancement of the HSV-TK/GCV system. Also, drugs with a rapid effect to the cell cycle are weak boosters of the HSVTK/GCV gene therapy, thus being less useful than DFMO for enhancement of this gene therapy form in animal studies and clinical trials.

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Section: Introductionmentioning

confidence: 75%

Section: Introductionmentioning

confidence: 91%

Section: Cell Lines and Viral Vectorsmentioning

confidence: 99%

Section: Cell Lines and Viral Vectorsmentioning

confidence: 99%

Section: Cell Cycle Arrest Induced By Aphidicolin Hydroxyurea Lovasmentioning

confidence: 99%

See 3 more Smart Citations

Polyamine depletion and cell cycle manipulation in combination with HSV thymidine kinase/ganciclovir cancer gene therapy

Wahlfors

Karppinen²,

Jänne³

et al. 2006

Int J Oncol

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In vivo enhancement of herpes simplex virus thymidine kinase/ganciclovir cancer gene therapy with polyamine biosynthesis inhibition

Wahlfors

Hakkarainen

Jänne

et al. 2006

Intl Journal of Cancer

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We have earlier demonstrated that inhibition of polyamine biosynthesis with difluoromethylornithine (DFMO) can be used to enhance the cytotoxicity of herpes simplex virus thymidine kinase/ ganciclovir (HSV-TK/GCV) gene therapy in different tumor cell lines. Here, the utility of this treatment combination was tested in vivo in a nude mouse tumor model. First, the effect of DFMO was verified by treating mice bearing subcutaneous 9L rat glioma tumors with 2% DFMO in drinking water. The drug treatment induced almost complete suppression of ornithine decarboxylase activity, and as a result, a strong decrease in intratumoral putrescine and spermidine concentrations, which were normalized 4 days after drug removal. Consequently, the tumors displayed a significant reduction in the proliferation activity that was increased to 20% higher than the normal level at day 4 and returned to normal level 7 days after DFMO removal. Next, 9L tumors with 30% of TK-GFP fusion gene positive cells were induced and the animals were given DFMO and GCV in 2 treatment schemes, with the drug administration periods overlapping either 5 or 2 days. The analysis of tumor size at the end of the treatment revealed that DFMO can enhance HSV-TK/GCV cytotoxicity when the overlap between DFMO and GCV was 5 days, but the result was not significant. However, the 2-day overlap scheme yielded a significantly (p < 0.05, ANOVA) enhanced antitumor effect. In conclusion, the data here confirms that a novel combination of 2 clinically relevant treatment modalities, polyamine deprivation and HSV-TK/ GCV suicide gene therapy, can be used synergistically in vivo. ' 2005 Wiley-Liss, Inc.Key words: polyamines; DFMO; cancer gene therapy; suicide gene; TK-GFP fusion; lentivirus vector; animal model Herpes simplex virus thymidine kinase/ganciclovir (HSV-TK/ GCV) gene therapy 1 is a typical form of suicide gene therapy and one of the most widely used approaches to destroy malignant cells by genetic means. Despite a number of attempts to enhance the efficacy of this tumor therapy form, results from clinical trials have been modest and true clinical breakthroughs have not occurred yet.2 Thus, thymidine kinase-ganciclovir suicide gene therapy still needs improvement and new, innovative means to increase the tumor toxicity of this treatment form are actively being sought. Our approach to enhance the efficacy of HSV-TK/ GCV method is to manipulate the cell cycle of the target tumor. When proliferation of a tumor cell population is blocked and then released, an increased proportion of cells transiently enter S phase. Because HSV-TK/GCV gene therapy affects through DNA replication and thus kills only dividing cells, it can be anticipated that cell population with maximal number of cells in their S phase is most vulnerable for HSV-TK/GCV toxic effect. It has been shown that polyamine deprivation induced with a-difluoromethylornithine (DFMO, an irreversible inhibitor of ornithine decarboxylase) causes cytostasis that is accompanied by cell cycle arrest through inhibition of G...

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Polyamines: metabolism and implications in human diseases

2005

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Polyamine biosynthesis inhibition enhances HSV‐1 thymidine kinase/ganciclovir‐mediated cytotoxicity in tumor cells

Cited by 4 publications

References 15 publications

Polyamine depletion and cell cycle manipulation in combination with HSV thymidine kinase/ganciclovir cancer gene therapy

Polyamine depletion and cell cycle manipulation in combination with HSV thymidine kinase/ganciclovir cancer gene therapy

In vivo enhancement of herpes simplex virus thymidine kinase/ganciclovir cancer gene therapy with polyamine biosynthesis inhibition

Polyamines: metabolism and implications in human diseases

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