DIFFERENTIAL CELL SURFACE PROTEOMICS OF FLOTILLIN-1, HMGB1 AND AMINOPEPTIDASE IN H3 MUTATED PAEDIATRIC HIGH-GRADE GLIOMA

Abstract

Abstract AIMS High-grade gliomas (HGG) are the deadliest central nervous system tumour in children. Despite this, treatment options remain elusive, leaving children with a dismal prognosis. Histone H3.3 mutations, K27M and G34V/R have been identified as the two most common recurrent somatic mutations in paediatric HGG (pHGG). These mutations are present in 50% of pHGG compared to only 1% in adults. Our previous research on cell membrane protein analyses have identified different signature of unique and common cell membrane proteins in H3-G34R/V mutated pHGG, in comparison to H3 wild type tumour. Among these we have identified flotillin-1, HMGB1 and aminopeptidase N. Flotillin-1 promotes numerous malignancies, High mobility group box 1(HMGB1) functions as an oncogene and aminopeptidase N is a cell surface metallo- protease involved in tumour progression. We have now verified the expression and localization of these cell surface proteins in H3 mutated and wildtype tumour, in order to target them for developing novel therapeutics. METHOD Cell membrane proteomics were further verified though mass-spectrometric data analyses. Expression and localization of flotillin-1, HMGB1 and Aminopeptidase in H3.3 mutated pHGG cell lines in comparison to H3 wildtype glioma were analysed, using different protein expression assays including immunofluorescence microscopy and western blot assays. RESULTS Experimental analyses showed differential protein expression and localization of Flotillin-1, HMGB1 and Aminopeptidase and verified the mass-spectrometric proteomic data in H3-G34V pHGG, in comparison to wild- type pHGG. CONCLUSIONS This new knowledge will help us utilise Flotillin-1, HMGB1 and Aminopeptidase N, as potential therapeutic targets for these unique H3 mutated children’s brain tumours.