OS3.2 Delta-aminolevulinic acid can decrease the radioresistance of glioma stem cells with mesenchymal phenotypes in vitro and in vivo

Abstract

INTRODUCTION: High-grade gliomas (HGGs) is divided into 3 major subtypes based on core gene signatures; mesenchymal (MES), proneural (PN) and proliferative. Among them, HGGs profiled in the MES subtype tend to show worse prognosis than others. Recently, also as for glioma stem cells, two mutually exclusive subtypes with distinct dysregulated signaling and metabolic pathways has been identified; MES-GSC and PN-GSC. It is known that MES-GSC displays more aggressive phenotypes and is markedly resistant to radiation as compared with PN-GSC. Delta-aminolevurinic acid (ALA) is the first metabolite in the porphyrin synthesis and the exogenous administration of ALA increased the endogenous production of photosensitizer protoporphyrin-IX (PpIX) in HGG tumor cells. Recently, several reports have revealed that ALA has an ability to enhance the therapeutic efficacy of X-ray radiotherapy (XRT) for various malignant tumors in vitro and in vivo. The aim of the present study is to investigate whether ALA can decrease the radioresistance of GSCs. MATERIAL & METHODS: We used 3 MES-GSCs and 2 PN-GSCs established from surgical specimens of human malignant gliomas. We defined the above GSCs cultured in an adherent condition with FBS and without glutamine for more than 2 weeks as glioma «differentiated» cells (GDCs). Firstly, we quantified the intracellular PpIX in the GSCs and their corresponding GDCs by FACS and LC. Then, we assessed the in vitro therapeutic effect of ALA-XRT on them; MTT-assay, sphere- or colony- forming assay, and apoptosis assay were performed. We made an intracerebral glioma model using one MES-GSC to evaluate in vivo therapeutic effect of ALA-XRT; MES-GSC transplanted mice were randomly divided into 4 groups, ALA-XRT, XRT only, ALA only and control. Total 30 Gy in 3 Gy fractions of X-ray was irradiated to the mice in 1st and 2nd group. The intraperitoneal injection of 240 mg/kg ALA was done 6 hours prior to each irradiation for the mice in 1st and 3rd group. RESULTS: 1) All MES-GSCs showed significantly higher PpIX accumulation than their corresponding MES-GDCs. Meanwhile, two PN-GSCs showed lower intracellular PpIX level than their corresponding PN-GDCs. 2) All MES-GSCs showed higher resistance to X-ray than their corresponding MES-GDCs, but the radioresistance of MES-GSCs significantly decreased by ALA in vitro. 3) ALA significantly increased the ROS production during XRT in a dose dependent manner both in MES-GSCs and MES-GDCs in vitro. 4) In the in vivo therapeutic experiment, median survivals of control, ALA-only, X-ray only, ALA-XRT group were 19, 22, 30, 38 days, respectively. (ALA-XRT vs. XRT only: p=0.02, Control vs. XRT only: p>0.001, Control vs. ALA only: p=0.09) CONCLUSIONS: MES-GSCs had a higher activity of porphyrin synthesis from exogenous ALA than PN-GSCs. In fact, the redioresistance of MES-GSCs can be effectively decreased by the concomitant use of ALA both in vitro and in vivo.