Cellular and molecular characterization of IDH1-mutated diffuse low grade gliomas reveals tumor heterogeneity and absence of EGFR/PDGFRα activation

Diffuse low grade gliomas (DLGG, grade II gliomas) are slowly-growing brain tumors that often progress into high grade gliomas. Most tumors have a missense mutation for IDH1 combined with 1p19q codeletion in oligodendrogliomas or ATRX/TP53 mutations in astrocytomas. The phenotype of tumoral cells, their environment and the pathways activated in these tumors are still ill-defined and are mainly based on genomics and transcriptomics analysis. Here we used freshly-resected tumors to accurately characterize the tumoral cell population and their environment. In oligodendrogliomas, cells express the transcription factors MYT1, Nkx2.2, Olig1, Olig2, Sox8, four receptors (EGFR, PDGFRα, LIFR, PTPRZ1) but not the co-receptor NG2 known to be expressed by oligodendrocyte progenitor cells. A variable fraction of cells also express the more mature oligodendrocytic markers NOGO-A and MAG. DLGG cells are also stained for the young-neuron marker doublecortin (Dcx) which is also observed in oligodendrocytic cells in nontumoral human brain. In astrocytomas, MYT1, PDGFRα, PTPRZ1 were less expressed whereas Sox9 was prominent over Sox8. The phenotype of DLGG cells is overall maintained in culture. Phospho-array screening showed the absence of EGFR and PDGFRα phosphorylation in DLGG but revealed the strong activation of p44/42 MAPK/ERK which was present in a fraction of tumoral cells but also in nontumoral cells. These results provide evidence for the existence of close relationships between the cellular phenotype and the mutations found in DLGG. The slow proliferation of these tumors may be associated with the absence of activation of PDGFRα/EGFR receptors

MANOTA: a promising bifunctional chelating agent for copper-64 immunoPET

International audienceImproved bifunctional chelating agents (BFC) are required for copper-64 radiolabelling of monoclonal antibodies (mAbs) under mild conditions to yield stable, target-specific imaging agents. Four different bifunctional chelating agents (BFC) were evaluated for Fab (Fragment antigen binding) conjugation and radiolabelling with copper-64. Two DOTA- (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) and two NOTA- (1,4,7-triazacyclononane-1,4,7-triacetic acid) derivatives bearing a p-benzyl-isothiocyanate group were conjugated to Fab-trastuzumab - which targets the HER2/neu receptor - and the average number of chelators attached ranged from 2.4 to 4.3 macrocycles per Fab. Labelling of the immunoconjugate with copper-64 was achieved in high radiochemical yields after 45 min at 37 °C, and the radiochemical purity of each (64)Cu-BFC-Fab-trastuzumab reached 97% after purification. The affinity of each (64)Cu-BFC-Fab-trastuzumab ranged between 10 and 50 nM as evaluated by in vitro saturation assays using the HCC1954 breast cancer cell line. PET-MR imaging and biodistribution studies were performed in mice bearing breast cancer BT-474 xenografts. BT-474 tumours were clearly visualized on PET images at 4 and 24 hours post-injection. The tumour uptake of (64)Cu-BFC-Fab-trastuzumab reached 8.9 to 12.8% ID g(-1) 24 hours post-injection and significant differences in non-specific liver uptake were observed depending on the BFC conjugated, the lowest being observed with MANOTA. These results show that MANOTA is a valuable tool for copper-64 radiolabelling