Abstract 4705: Single cell derived 3D organoids recapitulate the tumorigenic features of glioblastoma
Authors: Chadwick Michelle , Mfon Rachael , Jara Kelly , Danish Shabbar , Aiken Robert , Sabaawy Hatem E. 
Published: August 2019

In recent years, three-dimensional (3D) models of human brain derived from pluripotent embryonic and iPS cells have emerged as brain organoids, providing established models for brain development and genetic engineering. However, tumor organoids derived from adult glioblastoma multiforme (GBM) patients were seldomly formed. WHO grade IV GBM is a devastating locally invasive brain cancer with a median survival of 14.6 months. Standard of care for GBM includes surgical resection followed by radiotherapy plus concomitant and maintenance temozolomide (TMZ). Resistance to TMZ develops rapidly and neither dose-intensified TMZ nor anti-angiogenic approaches could improve survival. Genomic, transcriptomic and epigenetic profiling allowed classifying adult GBM into three groups: isocitrate dehydrogenase (IDH)-mutant (mut), 1p/19q co-deleted oligodendroglial GBM with best prognosis; IDH-mut, 1p/19q non-co-deleted astrocytic GBM with intermediate outcome; and IDH wild-type (WT) GBM with poor prognosis. Preclinical models that reflect these GBM profiles and heterogeneity are urgently needed to examine new therapies. Patient derived orthotopic xenografts (PDOXs) are thought to better mimic the GBM environment. Genetically engineered mouse models (GEMMs) make gliomas in mice with competent immune systems, but are laborious and expensive requiring compound genetics to mimic human GBM. Here, we describe a 3D serum-free organoid system in low-adherence plates derived from primary and sphere cultures that supports the long-term growth and expansion of GBM organoids for several months. We generated organoids from GBM with IDH-WT or IDH-Mut, EGFR amplification, +7q/-10q genotype, PTEN mutation, and EGFRvIII expression. GBM organoids are comprised of a heterogenous cell populations, thus mimicking the original tumor. GBM organoids could be generated from a single cell, therefore allowing to track intratumor heterogeneity. GBM organoids could be utilized for investigating aspects of GBM biology such as 3D cellular self-renewal showing expression of stem cell and differentiation targets, 3D cell cycle regulation including synchronized growth and phase cycling, 3D cell metabolism including intracellular adenosine triphosphate (ATP) level and protein synthesis rate, 3D cell invasiveness validated with IHC assays, and for evaluation of drug effects in the context of IDH, PI3K and EGFR aberrations. We have demonstrated that GBM sphere cells exhibit a solid growth pattern when implanted orthotopically into the mouse PDOXs. Since GBM organoids form tighter cell-cell contact, oxygen and nutrient gradients, we implanted GBM organoids in mouse orthotopic xenografts to demonstrate their tumor growth and invasive phenotypes. This human GBM organoid platform allows for novel preclinical therapeutic approaches to be assessed and provides personalized therapeutic options for individual GBM patients.

Citation Format: Michelle Chadwick, Rachael Mfon, Kelly Jara, Shabbar Danish, Robert Aiken, Hatem E. Sabaawy. Single cell derived 3D organoids recapitulate the tumorigenic features of glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4705.