Human glioblastoma stem-like cells accumulate protoporphyrin IX when subjected to exogenous 5-aminolaevulinic acid, rendering them sensitive to photodynamic treatment.

Glioblastoma (GBM) is the most frequent and lethal primary brain tumor in adults. Despite multimodal therapy combining resection, radio- and alkylating chemotherapy, disease recurrence is universal and prognosis of patients is poor. Glioblastoma stem-like cells (GSC), which can be grown as neurospheres from primary tumors in vitro, appear to be resistant to the established therapies and are suspected to be the driving force for disease recurrence. Thus, efficacy of emerging therapies may depend on targeting GSC. 5-aminolaevulinic acid-mediated photodynamic therapy (5-ALA/PDT) is a promising therapeutic approach in GBM. It utilizes the selective accumulation of the photosensitizer protoporphyrin IX (PPIX) in GBM cells after application of 5-ALA. When exposed to laser light of 635nm wavelength, PPIX initiates a photochemical reaction resulting in the generation of reactive oxygen species, which kill the tumor cells. Whether GSC accumulate PPIX and are sensitive to 5-ALA/PDT is currently unknown. Therefore, human GSC were derived from primary tumors and grown as neurospheres under serum free conditions. When subjected to exogenous 5-ALA, a dose- and time-dependent accumulation of PPIX in GSC was observed by flow cytometry, which varied between individual GSC preparations. Subsequent exposure to laser light of 635nm wavelength substantially killed GSC, whereas treatment with 5-ALA or exposure to laser light only had no effect. LD50 values differed between GSC preparations, but were negatively correlated with PPIX accumulation in GSC. In summary, we report for the first time that glioblastoma stem-like cells accumulate PPIX when subjected to 5-aminolaevulinic acid and are sensitive to 5-aminolaevulinc acid based photodynamic therapy.

Migration of mesenchymal stem cells towards glioblastoma cells depends on hepatocyte-growth factor and is enhanced by aminolaevulinic acid-mediated photodynamic treatment.

Hepatocyte-growth factor (HGF) is expressed by glioblastomas and contributes to their growth, migration and invasion. HGF also mediates migration of mesenchymal stem cells (MSC) to sites of apoptotic cell death. Moreover, MSC show tropism for glioblastomas, which is exploited in gene therapy to deliver the therapeutics to the tumor cells. Here, we have studied whether HGF contributes to the recruitment of MSC by glioblastoma cells and whether aminolaevulinic acid-mediated photodynamic therapy (ALA/PDT), a novel therapeutic approach that induces apoptosis in glioblastoma cells, affects HGF release and this migratory response. MSC expressed the HGF receptor MET and migrated towards U87 and U251 glioblastoma spheroids. Migration increased significantly when spheroids were subjected to ALA/PDT, which was associated with induction of apoptosis and up-regulation of HGF. Neutralizing HGF resulted in significant inhibition of MSC migration towards untreated as well as ALA/PDT-treated spheroids. Thus, glioblastoma cells express HGF, which contributes to the attraction of MSC. ALA/PDT induces apoptosis and augments HGF release causing enhanced MSC migration towards the tumor cells. ALA/PDT may therefore be exploited to improve targeting of MSC delivered gene therapy, but it may also constitute a risk in terms of beneficial effects for the tumor.