The effect of valproic acid in combination with irradiation and temozolomide on primary human glioblastoma cells

Abstract

Glioblastoma multiforme (GBM) has nearly uniformly fatal with a median survival of less than 2 years. While there have not been any novel anti-GBM therapeutics approved for many years, there has been the gradual accumulation of clinical data suggesting that the widely used anti-convulsant agent, valproic acid (VPA) may significantly prolong survival in GBM patients. This pre-clinical study aimed to determine the potential clinical utility of VPA in the treatment of GBM. Primary GBM cells were treated with VPA as a monotherapy and in combination with temozolomide and irradiation. At clinically achievable concentrations, VPA was shown to be effective as a monotherapy agent in the five primary lines tested. VPA was then used as a sensitizing agent to in vitro radiation and showed significant augmentation of in vitro irradiation therapy. In addition, when VPA, radiation and temozolomide were combined an additive, rather than synergistic effect was noted. Gene expression profiling demonstrated close clustering of triple treated cells with VPA mono-treated cells while untreated cells clustered closer with TMZ-irradiation dual treated cells. These microarray data suggest a dominant role of VPA at the gene expression level when combining these different treatment options. Moreover, in an in vivo tumor transplantation model, we were able to demonstrate an increase in animal survival when cells were pre-treated with irradiation-VPA and when triple treated. These findings provide a significant rationale for the investigation of VPA in the treatment of GBM patients.

Cellular Effects of the Antiepileptic Drug Valproic Acid in Glioblastoma

“…In the present study, VPA (750 µM) failed to radiosensitize T98G, U251 and U-87MG in delayed plating colony formation assays (1 h VPA-pre-incubation, VPA co-incubation during irradiation, and 24 h VPA post-incubation). Likewise, in the present experiments on primary glioblastoma spheroid cultures subjected to fractionated irradiation (5 x 0 Gy or 5 x 2 Gy) and temozolomide (5 µM), VPA (750 µM) neither impaired clonogenic survival nor radiosensitized the cells while a previous study using higher VPA concentrations (1-15 mM) in a similar experimental setting (primary glioblastoma cultures, irradiation, temozolomide) reported VPA-mediated inhibition of cell viability by MTT assay [13]. …”

“…These results were unexpected in light of the reported chromatin remodeling action of VPA [18, 48-50] and contradictory to previous in vitro studies [10-15, 17, 48, 51, 52] which clearly show VPA-induced cell death or radio- or chemosensitization in cancer cells including glioma. Unlike the present work, VPA concentrations of 1-5 mM [52], 1.5-2 mM [10], 1-5 mM [51], 2.5-5 mM [11], 1.5-3 mM [12], 1-15 mM [13], 2-16 mM [14], 7.5 mM [18], 2 mM [19], 5-20 mM [53], 1-10 mM [21], or 4.8 mM [22] were applied in these studies.…”

“…Notably, a further in vivo study transplanting primary human WK1 glioblastoma cells pre-treated with radiation and/or temozolomide and/or VPA (1 mM under serum-free conditions) orthotopically in mice observed a prolonged survival as compared to the control group (challenged with vehicle-treated WK1 cells) only in mice inoculated with radiation/VPA co-treated cells or radiation/temozolomide/VPA triple-treated cells [13]. Although suggestive of a VPA-mediated radiosensitization, it has to be noted that this study [13] did not report significant longer mouse survival upon inoculation with VPA/radiation double-treated and VPA/radiation/temozolomide triple-treated cells as compared to the survival of mice challenged with only radiation-treated and radiation/temozolomide co-treated cells, respectively.…”

Does valproic acid affect tumor growth and improve survival in glioblastomas?

“…The combination of VPA, TMZ and RT has been reported to cause a significant radiation enhancement, without antagonizing the cytotoxic effects of TMZ [12]. Recently, an additive, rather than synergistic, effect when VPA, radiation and TMZ are combined has been reported [13].…”

Valproate in Adjuvant Glioblastoma Treatment

“…Dose-response curves for the initial effect can be quite different when used for a new application when repurposing drugs. Our in vitro experimental studies on established cell lines and primary human glioblastoma cells clearly showed an interaction between VPA and chemoradiotherapy 2,3 but only at the upper end of recommended doses for seizure prevention. VPA dose does not directly relate to CSF concentration in humans, and studies using subtherapeutic doses are of limited clinical relevance.…”