Article
Standardised generation of tumour-organoids as novel drug screening platform in meningioma
Einsatz von standardisiert hergestellten Tumororganoiden aus Meningeomen als neuartige Medikamentenscreeningplattform
Search Medline for
Authors
Published: | May 25, 2022 |
---|
Outline
Text
Objective: Tumor-organoids (TOs) are novel, complex three-dimensional ex vivo tissue cultures that under optimal conditions accurately reflect the genotype and phenotype of the original tissue with preserved cellular heterogeneity and morphology. They offer a new and exciting platform for studying cancer biology and directing personalized therapies. Our study aimed to establish TOs from meningioma (MGM) and to test their usability for large-scale drug screenings.
Methods: TOs were established by controlled reaggregation of freshly prepared single cell suspensions of MGM tissues in 96-well plates. Size and shape were continuously observed by light microscopy. Structural tissue architecture was assessed by H&E stainings. Live/dead (Invitrogen) staining assessed integrity and viability of TOs. TOs were treated with drug concentrations ranging from 10 nM to 30 µM. Viability was assessed with CellTiterGlo3D (Promega). Resulting dose curves were converted to the z-transformed area under curve (z-AUC) values.
Results: We were capable of forming several hundred TO equal in size with a success rate of 98%. In total, standardized TOs from 60 patients were formed, including eight grade II and three grade III MGMs. TOs reaggregated within 3 days seen by a reduction of the initial diameter by 50%. Thereafter, diameter remained stable throughout a 14 days observation period. TOs consisted of largely viable cells, whereas dead cells were predominantly found outside of the organoid. H&E stainings confirmed the successful establishment of dense tissue-like structures. As a next step, we queried the suitability and reliability of TOs for robust large-scale drug testing by employing nine highly potent compounds, derived from a drug screening performed on MGM cell lines. First, we assessed if drug responses depend on the TO size. Interestingly, drug responses to these drugs remained identical independent of their sizes. Based on a sufficient representation of low abundance cell types such as T-cells and macrophages an overall number of 25.000 cells/TO was selected for further experiments revealing FDA-approved HDAC inhibitors as highly effective drugs in most of the TOs with a mean z-AUC score of -1.33.
Conclusion: Taken together, we developed a protocol to generate standardized TO from MGM containing low abundant cell types of the tumor microenvironment in a representative manner. Robust and reliable drug responses suggest patient-derived TOs as a novel drug testing platform.