430. EGFRvIII Retargeted Oncolytic Measles Virus Strains Have Significant Antitumor Activity Against Gliomas

Top of pageAbstract Gliomas are the most common primary CNS tumor and have a dismal prognosis despite the use of multimodality treatment. A very common genetic alteration in gliomas is the amplification of the epidermal growth factor receptor (EGFR) gene, present in 40-60% of glioblastoma multiforme (GBM). Amplification of the EGFR gene in GBM is associated with a tumor specific EGFR mutation, EGFRvIII, thus providing a target for tumor specific entry. We have previously shown that attenuated vaccine strains of measles virus have antitumor activity against gliomas (Phuong, 2003). Mutations have been identified which ablate recognition of the natural receptors CD46 and SLAM (Vongpunsawad, 2004, Nakamura, 2004). We therefore tested the hypothesis that engineering the virus to bind through a glioma specific receptor, while eliminating natural receptor binding, could result in improved specificity comparable therapeutic efficacy in tumors expressing EGFRvIII. METHODS/RESULTS: Recombinant viruses were generated from the Edmonston-NSe vaccine strain displaying an EGFRvIII single chain antibody at the c-terminus of the H protein. All viruses contained the GFP gene at position 1. Two different H mutants were used: HSNS (V451S, Y481N, A527S) (CD46 knockout) and HAA (Y481A, R533A) (CD46 and SLAM knockout). MV-GFP was employed as a positive control. These viruses were tested against the EGFRvIII expressing glioma lines GBM6, GBM39 and U118-EGFRvIII and the EGFRvIII negative lines GBM14, U118, normal human astrocytes, and fibroblasts. Specificity of retargeted strains was verified in CHO cell lines stably expressing CD46, SLAM or EGFRvIII and in U118 and U118-EGFRvIII glioma lines. Both retargeted viruses had significant antitumor activity against EGFRvIII expressing primary glioma lines. MV-GFP-HAA-scEGFRvIII-H6 had comparable antitumor activity to MV-GFP in vitro; while MV-GFP-HSNS-scEGFRvIII resulted in later onset of cytopathic effect. In contrast to MV-GFP, infection with retargeted strains resulted in lack of cytotoxicity against untransformed cells such as normal human astrocytes and fibroblasts even at high MOIs, indicating an improved therapeutic index. In vivo testing in orthotopically implanted GBM39 xenografts that maintain the EGFRvIII amplification, demonstrated statistically significant prolongation of survival of the animals treated with the retargeted strains as compared to UV-inactivated virus treated animals (p=0.02 and 0.002 respectively). There was no difference in the in vivo efficacy between retargeted strains and the unmodified MV-GFP strain. Viral propagation was verified in tumor samples from animals treated in vivo with the retargeted strains by examining GFP expression in frozen sections and by in situ hybridization for measles virus N-mRNA in paraffin imbedded samples. Extensive macrophage and NK cell infiltration was observed surrounding tumor induced syncytia. CONCLUSIONS: Measles virus strains retargeted against EGFRvIII maintain oncolytic activity and have comparable therapeutic efficacy with improved therapeutic index as compared to the unmodified MV-GFP strain in vitro and in vivo.