Real-time response prediction to GBM CART cell therapy with patient-matched organoids

Glioblastoma (GBM) is the most common primary brain tumor in adults and one of the most aggressive cancers currently without effective treatment. Recent clinical trial results from our group, employing a dual-targeting chimeric antigen receptor (CAR) T cell approach, have shown the treatment potential of harnessing the patient’s own immune system. However, not all responses have been on the same scale or shown the same durability, demonstrating the significant variability found in GBM. This spread of responses highlights the need for clinically validated predictive biomarkers, to help guide subsequent therapeutic decisions. The lack of complete tumor eradication and subsequent regrowth in some patients also confirms tumor escape, suggesting there are resistant tumor populations that need to be characterized. This research program works to develop a biomarker to predict patient response to CAR T cell therapy and identify mechanisms of resistance to the CAR T cell treatment. In order to generate results with a high level of fidelity, the work will make use of the GBM organoid. GBOs preserve the native genetic, transcriptional, and cellular heterogeneity of their parent tumor, and notably, they maintain non-neoplastic cells that play an essential role in the TME. GBOs can be established rapidly after surgery, biobanked, and used in a wide range of experimental settings, both in vitro and in animal models. In these ways, GBOs are an ideal model system to study the effect of CAR T cells on tumor heterogeneity. In addition, we have been taking advantage of ongoing clinical trials to compare responses of GBOs to CAR T treatment in vitro and GBM responses in patients. We have also leveraged recent advancements in genomic technologies to construct a highly multiplexed single-cell multi-omic platform that enables us to dissect more completely the compound complexity of our biological system. Enabled by these latest technologies and based on initial findings, we propose to expand on the effects of activated CAR T cells on GBM cells in order to optimize potential therapeutic strategies. The use of animal models in the project is critical for investigating the disease mechanisms under conditions that are as close as possible to physiological conditions. Mice present a platform that is more representative of human disease than other models used in developmental biology such as zebrafish, frogs or birds. In Aim 1, we will develop a predictive biomarker, utilizing both GBOs and clinical trial samples. We will expand our existing data set, examining GBO cell killing, cytokine production, and CAR T cell population characterization, both in the infusion product and post-infusion patient samples. These data will be compared to patient outcomes from our ongoing clinical trials, focusing on progression-free survival and overall survival. We will then validate this marker prospectively in our follow-up clinical trial. In Aim 2, we will use GBOs to examine the live post-CAR T-treated tumor cells. This population will provide insight into tumor escape mechanisms and lead to the development of the next generation of CAR T cells. Project Number: 1R01CA304417-01A1 | Fiscal Year: 2026 | NIH Institute/Center: National Cancer Institute (NCI) | Principal Investigator: DONALD O'ROURKE (+1 co-PI) | Institution: UNIVERSITY OF PENNSYLVANIA, PHILADELPHIA, PA | Award Amount: $668,864 | Activity Code: R01 | Study Section: Cellular Immunotherapy of Cancer Study Section[CIC] View on NIH RePORTER: https://reporter.nih.gov/project-details/11449429