Enhanced inhibition of clonogenic survival of human medulloblastoma cells by multimodal treatment with ionizing irradiation, epigenetic modifiers, and differentiation-inducing drugs.

Ina Patties,Franziska Menzel,Rolf-Dieter Kortmann,Annegret Glasow

doi:10.1186/s13046-016-0376-1

Ina Patties, Franziska Menzel + Show 2 more

Open Access

https://doi.org/10.1186/s13046-016-0376-1

Copy DOI

Abstract

BackgroundMedulloblastoma (MB) is the most common pediatric brain tumor. Current treatment regimes consisting of primary surgery followed by radio- and chemotherapy, achieve 5-year overall survival rates of only about 60 %. Therapy-induced endocrine and neurocognitive deficits are common late adverse effects. Thus, improved antitumor strategies are urgently needed. In this study, we combined irradiation (IR) together with epigenetic modifiers and differentiation inducers in a multimodal approach to enhance the efficiency of tumor therapy in MB and also assessed possible late adverse effects on neurogenesis.MethodsIn three human MB cell lines (DAOY, MEB-Med8a, D283-Med) short-time survival (trypan blue exclusion assay), apoptosis, autophagy, cell cycle distribution, formation of gH2AX foci, and long-term reproductive survival (clonogenic assay) were analyzed after treatment with 5-aza-2′-deoxycytidine (5-azadC), valproic acid (VPA), suberanilohydroxamic acid (SAHA), abacavir (ABC), all-trans retinoic acid (ATRA) and resveratrol (RES) alone or combined with 5-aza-dC and/or IR. Effects of combinatorial treatments on neurogenesis were evaluated in cultured murine hippocampal slices from transgenic nestin-CFPnuc C57BL/J6 mice. Life imaging of nestin-positive neural stem cells was conducted at distinct time points for up to 28 days after treatment start.ResultsAll tested drugs showed a radiosynergistic action on overall clonogenic survival at least in two-outof-three MB cell lines. This effect was pronounced in multimodal treatments combining IR, 5-aza-dC and a second drug. Hereby, ABC and RES induced the strongest reduction of clongenic survival in all three MB cell lines and led to the induction of apoptosis (RES, ABC) and/or autophagy (ABC). Additionally, 5-aza-dC, RES, and ABC increased the S phase cell fraction and induced the formation of gH2AX foci at least in oneout-of-three cell lines. Thereby, the multimodal treatment with 5-aza-dC, IR, and RES or ABC did not change the number of normal neural progenitor cells in murine slice cultures.ConclusionIn conclusion, the radiosensitizing capacities of epigenetic and differentiation-inducing drugs presented here suggest that their adjuvant administration might improve MB therapy. Thereby, the combination of 5-aza-dC/IR with ABC and RES seemed to be the most promising to enhance tumor control without affecting the normal neural precursor cells.

Highlights

Medulloblastoma (MB) is the most common pediatric brain tumor
Based on the positive results of our previous studies investigating the effects of IR and 5-aza-dC [4], and of 5-aza-dC with additional modulators [18], we investigated here for the first time the effect of a triple combination: IR, 5-aza-dC and one additional epigenetic modifier (SAHA, valproic acid (VPA)) or differentiation-inducer (RES, ABC and all-trans retinoic acid (ATRA))
A higher amount of untreated D283-Med cells is located in the radiosensitive G2/M cell cycle phase compared to DAOY cells which exhibited a higher amount of radioresistant G1/G0 cells, more double-strand breaks (DSB) were induced in the DAOY in D283-Med cells

Summary

Introduction

Current treatment regimes consisting of primary surgery followed by radio- and chemotherapy, achieve 5-year overall survival rates of only about 60 %. Therapy-induced endocrine and neurocognitive deficits are common late adverse effects. We combined irradiation (IR) together with epigenetic modifiers and differentiation inducers in a multimodal approach to enhance the efficiency of tumor therapy in MB and assessed possible late adverse effects on neurogenesis. Medulloblastoma (MB) is the most common malignant brain tumor (WHO °IV) in children aged < 15 years [1]. No radiation therapy but a more aggressive chemotherapy is given to children below 4 years to avoid radiation-related adverse late effects, like neuroendocrine and neurocognitive deficits. The 5-year overall survival of approximately 60 % implies the urgent need of improved antitumor therapies to enhance the outcome especially of high-risk patients (infants, metastatic disease: ~ 55 % of all MB patients)

Methods

Results

Discussion

Conclusion