P06.01.A Deconvolution of immunotherapy-treated glioblastoma identifies cellular heterogeneity and plasticity at the single-cell level

Abstract

Abstract Background Glioblastoma is the most aggressive cancer originating in the brain with an average survival of 15 months. One of the characteristics of glioblastoma is the high level of intra-tumour heterogeneity (ITH), but the composition and complexity at the single-cell level is poorly understood. Here, we aimed to assess the effects and consequences of immune checkpoint inhibitor (ICI) on the cellular and molecular heterogeneity of glioblastoma tumours using at the single cell level. Material and Methods In collaboration with the phase I trials unit at Rigshospitalet, we performed paired molecular analysis of glioma cells from primary and relapse surgery after ICI treatment. Samples were analysed using single-cell RNA sequencing (scRNA-seq) as well as bulk RNA sequencing and whole exome DNA sequencing. Results In an effort to trace cellular lineages we developed and refined methods to a identify copy number changes using scRNA-seq. To this end, we identified clonal and subclonal tumour cell populations in each sample. We found high levels of ITH prior to treatment, both with respect to the glioblastoma subtype enrichment and the cell type-specific gene expression. Using expression-based cell-type classification, we found defined recurrent cell-type populations present at both surgery time points. The immune checkpoint treatment had consequences on the cellular phenotypes and proportions of tumour cells, suggesting a level of plasticity in the neoplastic cells. Moreover, we identified examples of clonal dynamics and sweeps following ICI treatment, pointing to potential treatment response and resistance in these population. Conclusion In summary, we pursued single cell-focused analysis of ICI treated glioblastoma patients to study the cellular and molecular heterogeneity within and between glioblastoma patients, which pointed to recurrent patterns of cellular responses following ICI treatment.