DDDR-19. An optimized explant organoid method for preclinical brain tumour research

Abstract Tumour heterogeneity presents a significant obstacle to identifying effective treatments for brain cancer, explaining the ineffectiveness of standard treatments. Patient-derived brain tumour explant organoids (GBOs) replicate this heterogeneity and can predict a patient’s response to therapy, constituting a critical precision Neuro-oncology tool. However, current methods to generate GBOs are time-consuming, costly, and technically challenging, precluding the development of larger diverse organoid biobanks that capture the true spectrum of the disease and impeding rapid patient-personalised preclinical testing. By introducing semi-automated tissue processing, size exclusion chromatography and immediate cryopreservation, we optimized current methods for the generation and biobanking of tumour pieces (from low-grade glioma, high-grade glioma and primary/recurrent glioblastoma) suitable for GBO culture, reducing processing time from 6 hours to under 60 minutes. We used this biobank to compare effects of novel idronoxil-conjugated benzopyran compounds (NX786, NX904) on GBO growth, viability, cell death, invasion and senescence utilizing a combination of live and fixed imaging and multi-omics. We generated 24 GBOs from 33 samples of diverse brain tumour types (low-grade glioma, high-grade glioma and primary/recurrent glioblastoma) and tested the effect of NX786 and NX904 on 9 of these. NX786 and NX904 significantly reduced GBO growth (≥50%) and invasion. Low-grade gliomas were more resistant. NX904 induced significant cell death (>60%), while NX786 increased cellular senescence and induction of the mesenchymal-like state. By developing an efficient method for processing and cryopreserving brain tumour tissues suitable for GBO growth, we captured more patient samples and established an extensive biobank collection, which we successfully utilised for targeted drug screening, highlighting its potential for patient-personalised preclinical testing. We found that through the inhibition of crucial brain cancer hallmarks, including proliferation and invasion, together with the induction of a less aggressive senescence phenotype, NX786 and NX904 have potential for development as new therapeutic agents for brain cancer.