Patient's Derived Organoids for Drug Screening in Glioblastoma

An amount of tissue of approximately 2-3 cm3, if available, will be allocated to the study. The specimen will be divided in three parts (depending on the volume of the biopsy) and used to: a) obtain PDOs according to established procedures (Chadwick, et al., 2020; Gamboa, et al., 2021); b) flash-frozen for molecular analysis of original tissue; c) used to isolate GSCs by flow-cytometry cell sorting. Only PDOs characterized by histological and molecular conformity with primary tumors will be used. The mutational status of genes frequently associated with GBM onset and progression will be analyzed in PDOs, and compared with data derived from tumor DNA, in order to assess their representation of the genetic heterogeneity of original tumors. These studies will allow us the set up a reliable procedure for the ex-vivo establishment of pre-clinical models of GBM.

PDOs and GSCs representing different GBM molecular subtypes will be treated with epigenetic modulators , with spliceosome inhibitors or with drugs that indirectly target the splicing machinery, such as PRMT5 inhibitors. These drugs will be tested for their ability to suppress growth and/or induce cell death, when administered either alone or in combination with standard chemotherapy. Furthermore, the investigators will perform RNA sequencing experiments to identify TE-derived transcripts and splice variants induced by the treatments. By employing a computational pipeline developed in our laboratory (Pieraccioli and Sette, unpublished), the investigators will also characterize the affinity for MHC-I and immunogenicity of neoepitopes encoded by the treatment-induced TE-derived transcripts and splice variants. The results of these analysis will allow to identify neoepitopes to be used for designing immunotherapy approaches.