Abstract Post-mortem pediatric brain tumor tissue provides a uniquely valuable yet rarely utilizable research resource, particularly for recurrent or end-stage disease where surgical access is impossible. Through the Gift from a Child program, we’ve received postmortem pediatric tumor specimens to characterize and evaluate their functional viability using our Screening Live Cancer Explants (SLiCE) platform, which supports uncultured, zero-passage patient tissue, including hard-to-maintain tumor subtypes. Because few preclinical models exist that can meaningfully utilize autopsy-derived tumors, where postmortem interval, ischemia, and tissue degradation pose major barriers, we first assessed whether these specimens retain viable cells upon arrival and evaluated how variables such as time-to-autopsy and shipping conditions influenced viability. We confirmed the presence of live tumor cells in received specimens and demonstrated that SLiCE supports the survival of the post-mortem tissue through our standard four-day assay endpoint comparably to non-post-mortem tissue. Simultaneously, we found no survival of samples plated in standard in vitro culture in several different high-quality medias. Following confirmation of tumor persistence, we evaluated therapeutic response to agents the patients had and had not previously received, observing differential tumor killing. We next characterized the cellular composition of each specimen prior to SLiCE using flow cytometry and identified both tumor and tumor-associated cell populations which persist through post-mortem tissue collection. Finally, we evaluated whether SLiCE could support active tumor–microenvironment signaling by quantifying soluble factors in transwell media over a four-day assay window, including baseline and dynamic changes in human IFN-γ, TGF-β, IL-10, and other cytokines. Together, these studies are signaling that post-mortem donations can be leveraged not only for genomic and transcriptomic profiling, but also for functional viability and TME biology within SLiCE, expanding the scientific impact of these extraordinary gifts from families and establishing a framework for incorporating post-mortem pediatric tumor tissue into precision modeling pipelines.

Abstract CAR T therapy has revolutionized treatment for pediatric hematologic malignancies; however, widespread adoption in pediatric brain tumors remains elusive, largely due to preclinical models failing to capture patient tumor heterogeneity and the immunosuppressive tumor microenvironment (TME). Conventional systems, including cell lines, xenografts, and organoids, often lack critical TME features, leading to overestimation of therapeutic potency. Progress in next-generation CAR T therapies requires more biologically representative models. We have therefore developed Screening Live Cancer Explants (SLiCE), a New Approach Methodology platform that engrafts living, passage-zero patient tumors onto substrates of living tissue. This design preserves tumor heterogeneity better than in vitro culture and enables investigation of how patient tumor-associated cells influence T cell activity. We tested B7-H3–targeted CAR T cells from three healthy donors against cell lines and passage-zero high-grade pediatric brain tumors (n = 2 medulloblastoma, 1 ATRT). B7-H3 expression was quantified by IHC (H-score) and flow cytometry. CAR T or control T cells were added to SLiCE-engrafted tumors at effector-to-target ratios from 1:3 to 3:1. Tumor kill was measured by live tumor cell bioluminescence within four days of seeding. Killing of B7H3+ tumors occurred only in the presence of CAR+ T cells and not non-transduced T cells; killing among multiple tumor specimens collected from the same patient positively correlated with B7H3 expression (assessed by flow cytometry). Notably, the ATRT tumor showed minimal killing despite high B7-H3 expression (H-score = 152). Real-time cytokine secretion profiling and Flow/CyTOF analyses revealed active, inhibitory human myeloid cells, regulatory T cells and PD-L1+ tumor cells, features likely suppressing CAR T activity. These findings indicate SLiCE maintains key immunosuppressive TME components in pediatric brain tumors and can dissect mechanisms of immune evasion. Ongoing studies aim to overcome these barriers. These findings underscore SLiCE’s potential to accelerate development of CAR T therapies tailored for pediatric brain tumors.

Abstract Functional precision medicine (FPM) offers a promising path forward in neuro-oncology, where genomic profiling alone often fails to predict therapeutic response. To bridge this gap, we developed the Screening Live Cancer Explants (SLiCE) platform, a rapid ex vivo drug screening assay that functionally tests passage-zero patient brain tumor tissues engrafted atop living organotypic brain slice cultures (OBSCs). With an assay time of just four days, SLiCE preserves key tumor characteristics not maintained in vitro, including genomic fidelity, growth, invasion, and treatment response, with higher engraftment rates and faster assay speeds than in vivo models. Our standard cryopreservation workflow enables reproducible, iterative, and on-demand testing of a single zero-passage specimen banked in multiple replicate aliquots, setting SLiCE apart from organoid and precision-cut tumor explant models. Here, we describe results from our actively accruing clinical feasibility study (NCT05978557), where we successfully engrafted and tested 35 of 36 diverse brain tumor specimens on SLiCE, achieving our study’s primary endpoint ahead of schedule. SLiCE produced multi-parametric drug sensitivity scores (DSSs), each normalized to off-target toxicity, for all samples within a clinically actionable 28-day window. Across 530 experiments, we generated 142 DSSs from unique drug-tumor combinations, forming a reference library for future benchmarking. We then further analyzed a subset of IDH-WT glioblastoma tumor specimens in which SLiCE DSSs correlated with patient response to temozolomide (AUC = 0.875, p = 0.0175) and overall survival (R2 = 0.73). Additionally, this study validated surgically aspirated tumor tissue as a genomically, transcriptomically, and functionally similar tumor source compared to the standard, manually excised remnant tumor sample approved by clinical pathology. Collecting this often-discarded tumor source increased the mass of tumors accrued by nearly 5-fold and enabled collection from 11 additional patients, significantly increasing tumor tissue for downstream testing on SLiCE. These findings establish SLiCE as a scalable, clinically relevant platform for FPM in brain cancer, with potential to guide individualized treatment decisions and accelerate preclinical drug development. Citation Format: Breanna Mann, Nichole Artz, Adebimpe Adefolaju, Alain Valdivia, Xiaopei Zhang, Rajaneekar Dasari, Caroline Stockwell, Morrent Thang, Allison Murray, Noah Bell, Andrew Buckley, Rami Darawsheh, Kerry Fitzgerald, Jonathan Williams, Hardik Parikh, Shuang Gao, Jie An, Yvette Rodriguez, Daniel Metzger, Collin Parrow, Dylan Riley, Robert Seager, Stephanie B. Hastings, Taylor Jensen, Shakti Ramkissoon, Dominique Higgins, Yasmeen Rauf, Scott Elton, Kimberly Hamilton, Jeremy Wang, Albert Baldwin, Shawn Hingtgen, David Kram, Andrew Satterlee. Rapid drug sensitivity testing of passage-zero patient brain tumor tissues to inform preclinical and clinical decisions: results from a clinical feasibility study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 653.

Abstract Access to living patient tumor tissue remains a major barrier to accelerating preclinical drug development and informing clinical care. Over the last decade, we have developed a “New Approach Methodology” platform called Screening Live Cancer Explants (SLiCE), which reproducibly engrafts living, passage-zero patient tumors atop substrates of living tissue. Our streamlined four-day drug screening workflow enables tumor engraftment within one day of resection, drug addition one day post-engraftment, and quantification of tumor survival three days later. This procedure is fast enough to accelerate preclinical workflows and inform clinical decisions while minimizing genetic drift. As we expand use of SLiCE in the preclinical setting, our established protocols for collecting and cryopreserving passage-zero tumor tissue have thus far enabled us to cryopreserve over 150 grams of viable brain tumor tissue in over 700 cryovials from 41 patients for personalized drug sensitivity profiling and preclinical testing. Through an active collaboration between UNC and Labcorp, we have shown that replicate cryopreserved aliquots of tumor samples post-thaw are equivalent by whole-exome sequencing (WES), RNA sequencing, and functional drug response. Still, availability of living, passage-zero patient tumor tissue remains our limiting component to further scaling this assay for broad adoption and impact. To overcome this hurdle, our SLiCE Team at UNC is expanding collection of viable tumor tissue to other groups and institutions. A new collaboration with the Mayfield Clinic has successfully yielded shipment of tumor tissues freshly resected from patients at partnering hospitals to UNC. These tumor specimens were viably maintained throughout overnight shipping at 4C, quantified by live tumor cell metabolism via Presto Blue assay, and have been successfully engrafted on SLiCE, quantified by live tumor cell bioluminescence on Day 4 after seeding, the length of our standard drug screening assay. These data provide our first proof-of-concept evidence that tumors resected from patients at distant institutions can be shipped fresh to UNC for testing on SLiCE. Our goal is to continue receiving 3-4 tumors per month from Mayfield to optimize and refine this process. These efforts position SLiCE as a leading New Approach Methodology to test promising experimental agents and one day guide clinical care using passage-zero patient tumor tissues, including fresh, living tumors shipped from other sites. Citation Format: Adebimpe Adefolaju, Breanna Mann, Rajaneekar Dasari, Emma Livne, Caroline Stockwell, Alain Valdivia, Sonia Lipp, Angel Augst, Kerry Fitzgerald, Jonathan Williams, Hardik Parikh, Shuang Gao, Jie An, Taylor Jensen, Shakti Ramkissoon, Dominique Higgins, Yasmeen Rauf, Scott Elton, Kimberly Hamilton, Yair Gozal, Vincent DiNapoli, Albert Baldwin, Shawn Hingtgen, David Kram, Andrew Satterlee. Integrating passage-zero patient tumor tissues from distant institutions into a rapid, ex vivo drug screening platform [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 660.

Abstract Adoptive T cell therapies, including chimeric antigen receptor (CAR) T cells, have shown limited success in solid tumors, largely due to widespread preclinical models that fail to capture patient tumor heterogeneity and the immunosuppressive tumor microenvironment (TME). Conventional systems, including cell lines, xenografts, and organoids, often lack critical TME features, leading to overestimation of therapeutic potency. Progress in next-generation CAR T therapies requires more biologically representative models. We have therefore developed Screening Live Cancer Explants (SLiCE), a New Approach Methodology platform that engrafts living, passage-zero patient tumors onto substrates of living tissue. This design preserves tumor heterogeneity better than in vitro culture and enables investigation of how patient tumor-associated cells influence T cell activity. As proof of concept, we tested B7-H3-targeted CAR T cells from three healthy donors against cell lines and passage-zero adult and pediatric brain tumors (n = 3 GBM, 2 medulloblastoma, 1 ATRT). B7-H3 expression was quantified by IHC (H-score) and flow cytometry. CAR T or control T cells were added to SLiCE-engrafted tumors at effector-to-target ratios from 1:3 to 3:1. Tumor kill was measured by live tumor cell bioluminescence within four days of seeding. Killing of B7H3+ tumors occurred only in the presence of CAR+ T cells and not non-transduced T cells; killing among multiple tumor specimens collected from the same patient positively correlated with B7H3 expression (assessed by flow cytometry). Notably, the ATRT tumor showed minimal killing despite high B7-H3 expression (H-score = 152). Flow/CyTOF analyses revealed inhibitory myeloid cells, regulatory T cells (∼30% CD4+CD25+FoxP3+), and >60% PD-L1+ tumor cells, features likely suppressing CAR T activity. These findings indicate SLiCE maintains key immunosuppressive TME elements and can dissect mechanisms of immune evasion. Ongoing studies aim to overcome these barriers. Together, these data highlight SLiCE as a model that can uniquely leverage passage-zero patient tumor and tumor-associated cells to rapidly predict outcomes and potential challenges for CAR T cells and other cell-based therapies. Citation Format: Xiaopei Zhang, Breanna Mann, Adebimpe Adefolaju, Alain Valdivia, Rajaneekar Dasari, Caroline Stockwell, Noah Bell, Ashley Geiger, Tracy A. Withers, Xin Zhou, Alexa Rodriguez, Kerry Fitzgerald, Jon Williams, Hardik Parikh, Shuang Gao, Jie An, Taylor Jensen, Shakti Ramkissoon, Dominique Higgins, Yasmeen Rauf, Scott Elton, Kimberly Hamilton, Albert Baldwin, Barbara Savoldo, Shawn Hingtgen, David Kram, Andrew Satterlee. Rapid CAR T testing in an ex vivo platform that maintains passage-zero patient tumor tissues and their native immunosuppressive microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 661.

The lack of functional precision models that recapitulate the structural and microenvironmental features of advanced ovarian cancer remains a major barrier to improving therapeutic selection. Here, we present an organotypic mesentery membrane culture (OMMC) model, an ex vivo platform that supports rapid engraftment of freshly resected human ovarian cancer tissue and established cell lines onto intact rat mesenteric membranes. This system enables functional assessment of responses to standard-of-care therapies within five days. We demonstrate sustained viability of mesenteric tissue, successful engraftment and expansion of tumor cells and patient-derived tumor tissue, and measurable tumor-mesentery interactions. Using a multi-parametric drug sensitivity score (DSS), we quantified tumor response relative to off-target mesenteric toxicity across a panel of FDA-approved therapies. Application of the platform to patient-derived tumor samples revealed inter-patient variability in drug sensitivity profiles. Comparison of OMMC-derived responses with matched clinical outcomes suggests that this approach may help inform treatment response in a subset of cases. Together, these findings support the potential of the OMMC platform as a functional assay to complement existing preclinical and molecular approaches for evaluating therapeutic responses in ovarian cancer.

Here, we review a growing paradigm shift from genomics-based precision medicine toward functional precision medicine, which evaluates therapeutic efficacy by directly treating living patient tumors ex vivo to better predict patient-specific responses to treatment. We discuss several classes of patient-derived models of central nervous system tumors, highlighting unique features of each. Each class of models holds promise to improve treatment selection, prolong survival, and enhance patient outcomes.

Functional precision medicine (FPM) offers a promising path forward in neuro-oncology, where genomic profiling alone often fails to predict therapeutic response. To bridge this gap, we developed the Screening Live Cancer Explants (SLiCE) platform, a rapid ex vivo drug screening assay that functionally tests passage-zero patient brain tumor tissues engrafted atop living organotypic brain slice cultures (OBSCs). With an assay time of just four days, SLiCE preserves key tumor characteristics not maintained in vitro, including genomic fidelity, growth, invasion, and treatment response, with higher engraftment rates and faster assay speeds than in vivo models. Our standard cryopreservation workflow enables reproducible, iterative, and on-demand testing of a single zero-passage specimen banked in multiple replicate aliquots, setting SLiCE apart from organoid and precision-cut tumor explant models. Here, we describe results from our actively accruing clinical feasibility study (NCT05978557), where we successfully engrafted and tested 35 of 36 diverse brain tumor specimens on SLiCE, achieving our study's primary endpoint ahead of schedule. SLiCE produced multi-parametric drug sensitivity scores (DSSs), each normalized to off-target toxicity, for all samples within a clinically actionable 28-day window. Across 530 experiments, we generated 142 DSSs from unique drug-tumor combinations, forming a reference library for future benchmarking. We then further analyzed a subset of IDH-WT glioblastoma tumor specimens in which SLiCE DSSs correlated with patient response to temozolomide (AUC = 0.875, p = 0.0175) and overall survival (R2 = 0.73). Additionally, this study validated surgically aspirated tumor tissue as a genomically, transcriptomically, and functionally similar tumor source compared to the standard, manually excised remnant tumor sample approved by clinical pathology. Collecting this often-discarded tumor source increased the mass of tumors accrued by nearly 5-fold and enabled collection from 11 additional patients, significantly increasing tumor tissue for downstream testing on SLiCE. These findings establish SLiCE as a scalable, clinically relevant platform for FPM in brain cancer, with potential to guide individualized treatment decisions and accelerate preclinical drug development.

Glioblastoma (GBM) remains one of the most lethal and treatment-resistant cancers, underscoring the urgent need for novel therapeutic strategies. Marine-derived nutraceuticals offer a promising avenue due to their unique and myriad bioactive compounds. Immune-12 is an orally available marine organism-derived product that has shown preliminary evidence of anticancer activity in the published literature. Now, our team is conducting a comprehensive investigation of Immune-12 to characterize its molecular makeup, mechanisms of action, and in vitro/in vivo antitumor potencies. We have already conducted proteomic, peptidomic, and metabolomic profiling of this complex compound alongside several in vitro and in vivo efficacy and safety studies. In vitro cytotoxicity assays across 18 tumor cell lines, including six GBM lines, revealed potent activity, with IC₅₀ values ranging from 0.1% to 1.4% Immune-12 in media (volume/volume). In vivo toxicity studies, in which healthy mice were given Immune-12 for 28 days via oral gavage, showed no signs of systemic toxicity or weight loss. Furthermore, in orthotopic tumor-bearing mouse models, including GBM, Immune-12 inhibited tumor growth as a monotherapy and enhanced the antitumor efficacy of standard therapeutics in combination regimens. These findings suggest that Immune-12 exerts broad-spectrum cytotoxic effects against tumor cells while maintaining a favorable safety profile in vivo, highlighting its potential as a marine-derived therapeutic candidate for GBM and other malignancies. Based on these promising early data, we are rapidly conducting further experiments to maximize its potential for clinical application.

Of the roughly 1% of brain cancer therapeutics that show promise in preclinical testing, about 85% fail in clinical trials, indicating that drugs should be further scrutinized in preclinical settings. This failure is partially attributed to the inability of many preclinical models to recapitulate relevant aspects of disease and response to treatment, thereby misrepresenting a drug's therapeutic potential. Hence, we developed an Organotypic Brain Slice Culture (OBSC)-based platform that allows the direct testing of drugs on cell lines and living, uncultured patient brain tumor tissues to obtain functional readouts that capture both anti-tumor efficacy and off-target toxicity in a summarized drug sensitivity score (DSS). In this rapid, four-day assay, cells or tumor tissues are seeded atop coronal sections of postnatal day 8 rat pup brains just after slicing, therapeutics are added the next day, and live tumor bioluminescence is quantified three days after treatment initiation. In a parallel assay, off-target toxicity to non-tumor-bearing OBSCs after treatment is assessed using a propidium iodide assay that quantifies OBSC-associated cell death. Here, we investigated the antitumor activity of two experimental compounds, TR-107, a potent ONC201 derivative, and EdU, a newly repurposed DNA labeling agent, against an array of cell lines and brain tumor tissues. We then compared tumor survival on OBSCs to the standard of care and/or other relevant drugs tested in our system. The potency and selectivity of TR-107 in our assay is unsurprising as its parent compound, ONC 201, is the first imipridone in Phase 3 trials for treating H3- K27M gliomas. When investigating EdU against patient tumor tissues on OBSCs, we observed that intertumoral differences in Ki-67 abundance, a proliferation marker measured by clinical pathology, correlated with the effective doses necessary to eliminate up to 50% of these heterogeneous tumors. Importantly, while many of our results correlate with expected outcomes, we also identified non-responding tumor tissues whose “static” biomarkers predicted otherwise, reinforcing the need for functional approaches to drug development. In summary, this data supports the use of our OBSC platform as a functional biomarker for drug performance against cell lines and heterogeneous patient tumor tissues, and as an assay that can provide insight into discordance between genomic predictions and functional outcomes long before a drug fails in the clinic. Citation Format: Adebimpe Adefolaju, Humeyra Kaanoglu, Breanna Mann, Xiaopei Zhang, Noah Bell, Morrent Thang, Rajaneekar Dasari, Nichole Artz, Andrew Buckley, Lee Graves, Shawn Hingtgen, Aziz Sancar, Andrew Satterlee. The organotypic brain slice culture platform as a functional biomarker for preclinical drug performance testing [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Functional and Genomic Precision Medicine in Cancer: Different Perspectives, Common Goals; 2025 Mar 11-13; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2025;85(5 Suppl):Abstract nr B007.

Brain and nervous system cancers account for only ∼1.3% of new cancer diagnoses but rank ninth in US cancer mortality, a disparity partly driven by limited therapeutic options and inadequate preclinical models that misrepresent a drug's therapeutic potential. Considering that about 90% of drugs validated with these models fail in late-phase clinical trials, it is imperative to further scrutinize drugs in preclinical settings that better model relevant aspects of disease and treatment response. New paradigms must account for challenges unique to brain cancers such as lack of relevant biomarkers and both intra-disease and patient to patient heterogeneity, which cause treatments to be effective in a suboptimal proportion of the population. In this review, we investigate the current brain cancer drug development landscape, introduce a growing field of functional precision medicine, and propose the inclusion of "preclinical trials" that interrogate the effects of new therapies and drug delivery mechanisms on living patient tumors ex vivo. These preclinical trials respond to the FDA's recent announcement to phase out and replace live animal testing with human-based lab models. Functional models can address heterogeneity and biomarker identification through accrual of living patient tumor tissue, preclinical drug sensitivity testing, identification of non-responders and resistance mechanisms, and development of functional predictive biomarkers and companion diagnostics. Because functional precision medicine stratification of clinical trials candidates has shown improved clinical trials outcome, using this paradigm earlier in drug development could enhance clinical trial success, leading to more FDA-approved drugs and therapeutic options for brain cancer patients.

Functional Precision Medicine platforms, which investigate the dynamic behavior of a patient’s tumor ex vivo to inform personalized treatment, face unique obstacles to clinical translation. These include limited access to patient tissue and stringent demands for intra-platform accuracy and consistency. In this study, an automated data analysis pipeline addresses these concerns for an organotypic brain slice culture-based functional assay by combining computer vision and dose-response modeling approaches. A 99% reduction in analysis time increases the amount of patient tissue that can be processed on the platform. Comparing automated measurements to previously published manual results revealed that automation increased consistency both within experiments and across replicate experiments. This pipeline also explores implementing complex CV with limited resources, modeling a unique and diverse dataset, and validating automated analysis when no gold standard measurements exist, obstacles that hinder automation efforts across scientific disciplines.

Glioblastoma remains profoundly challenging. Moreover, the unique genetic, epigenetic, and microenvironmental features of glioblastoma contribute to its resistance to current therapies, complicating the development of effective therapeutic strategies. Recurrence represents a significant obstacle, as residual cancer stem cells can survive initial therapies and lead to tumor regrowth, often in the same or more aggressive form. Herein, we aim to repurpose three bioactive compounds—fenbendazole, papain, and sodium selenite—as novel therapeutic agents for brain cancer. Fenbendazole, a benzimidazole anthelmintic, disrupts microtubule dynamics by binding to β-tubulin, inhibiting mitosis, reducing glucose uptake, and inducing apoptosis in rapidly dividing tumor cells. Papain, a cysteine protease derived from papaya, exhibits antitumor potential through the degradation of extracellular matrix components, modulation of immune responses, and promotion of apoptotic pathways. Sodium selenite, an inorganic selenium compound, selectively induces oxidative stress in cancer cells, triggering apoptosis and potentially enhancing sensitivity to conventional therapies. Our in vitro studies have demonstrated significant antitumor activity for each compound, as well as combinatorial synergies against several brain tumor cell lines in vitro. To translate these findings into a clinically actionable approach, we are developing an implantable, biodegradable scaffold capable of localized controlled release of these agents within the brain tumor resection cavity. This polyvinyl alcohol-based delivery system is designed to maximize therapeutic efficacy while minimizing systemic toxicity. By integrating these repurposed compounds within our novel biomaterial platform, our strategy offers a promising avenue for improving outcomes in patients with malignant brain tumors. It represents a potentially impactful advancement in neuro-oncology care.

Glioblastoma (GBM) is the most common and aggressive type of malignant central nervous system tumors, with over 300, 000 new cases diagnosed globally each year. Despite intensive efforts to improve survival with different treatment regimens, GBM is still deemed largely incurable. We previously found that 5-ethynyl-2’-deoxyuridine (EdU), which is commonly used in laboratories to study DNA replication, is recognized by mammalian cells as DNA damage, removed from the genome by nucleotide excision repair, and causes death of proliferating cells. Here, we evaluate the therapeutic potential of EdU for treatment of GBM, as EdU has been shown to cross the blood-brain barrier and incorporate into proliferating GBM cells but not into post-mitotic neurons. We tested the efficacy of EdU against several different model systems, including not only GBM cell lines in in vitro cell culture and in vivo orthotopic mouse models of GBM, but also living, uncultured GBM patient tumor tissues grown within our Organotypic Brain Slice Culture (OBSC) ex vivo platform. In all three model systems, EdU was effective in killing GBM cells. When compared to the standard of care drug Temozolomide (TMZ) in in vitro GBM cell survival assays, EdU displayed ED50 values orders of magnitude lower than TMZ in all five GBM tumor cell lines tested. Against two in vivo orthotopic brain tumor models, EdU significantly extended survival relative to controls. Using our OBSC ex vivo platform, we were able to quantify the capability of EdU to potently kill rapidly dividing cells within heterogeneous GBM patient tumor tissues without harming the healthy OBSC-associated brain tissue. EdU efficacy against a panel of patient GBMs largely correlated with the clinical Ki-67 status of each tumor, save for one tumor that remained unresponsive to treatment with both EdU and TMZ. Overall, these data suggest that (1) EdU has potential to be repurposed as an anticancer therapeutic and is especially adept at killing rapidly proliferating cells with low off-target toxicity; (2) the OBSC platform can measure nuanced differences in efficacy of experimental therapeutics on heterogeneous patient tumor tissues; and (3) OBSCs can continue to help identify potential responders and non-responders to EdU treatment via functional precision testing of patient tumors ex vivo. [HK and AA contributed equally to this work.] Humeyra Kaanoglu, Adebimpe Adefolaju, Casey Fraley, Mariah Shobande, Rajaneekar Dasari, Breanna Mann, Noah Bell, Stephen T. Keir, Shawn Hingtgen, Aziz Sancar, Andrew B. Satterlee. Repurposing the DNA labeling agent EdU for glioblastoma treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6872.

Transdifferentiation (TD), a somatic cell reprogramming process that eliminates pluripotent intermediates, creates cells that are ideal for personalized anti-cancer therapy. Here, we provide the first evidence that extracellular vesicles (EVs) from TD-derived induced neural stem cells (Exo-iNSCs) are an efficacious treatment strategy for brain cancer. We found that genetically engineered iNSCs generated EVs loaded with the tumoricidal gene product TRAIL at nearly twice the rate of their parental fibroblasts, and TRAIL produced by iNSCs was naturally loaded into the lumen of EVs and arrayed across their outer membrane (Exo-iNSC-TRAIL). Uptake studies in ex vivo organotypic brain slice cultures showed that Exo-iNSC-TRAIL selectively accumulates within tumor foci, and co-culture assays demonstrated that Exo-iNSC-TRAIL killed metastatic and primary brain cancer cells more effectively than free TRAIL. In an orthotopic mouse model of brain cancer, Exo-iNSC-TRAIL reduced breast-to-brain tumor xenografts by approximately 3000-fold compared to treatment with free TRAIL, with all Exo-iNSC-TRAIL treated animals surviving through 90 days post-treatment. In additional in vivo testing against aggressive U87 and invasive GBM8 glioblastoma tumors, Exo-iNSC-TRAIL also induced a statistically significant increase in survival. These studies establish a novel, easily generated, stable, tumor-targeted EV to efficaciously treat multiple forms of brain cancer.

The lack of functional precision models that recapitulate the pathology and structure/function relationship of advanced ovarian cancer (OC) within an appropriate anatomic setting constitutes a hurdle on the path to developing more reliable therapies and matching those therapies with the right patients. Here, we developed and characterized an Organotypic Mesentery Membrane Culture (OMMC) model as a novel ex-vivo platform where freshly resected human patient OC tumor tissue or established cell lines are seeded directly atop living intact rat mesenteric membranes, rapidly engraft, and enable functional assessment of treatment response to FDA-approved standard care of treatment as single and combination drug therapies within just five days. This study showed successful survival of dissected mesentery tissue, survival and engraftment of tumor cells and patient tumor tissue seeded on OMMCs, mesentery-tumor cell interaction, and quantification of tumor response to treatment and off-target toxicity. Summarized “drug sensitivity scores”, using a multi-parametric algorithm, were also calculated for each patient’s treatment response, enabling us to suggest the most effective therapeutic option. Finally, we compared drug sensitivity results from patient tumor tissue on OMMCs to matched outcomes of individual patients in the clinic and identified positive correlations in drug sensitivity, beginning to validate the functionality of OMMCs as a functional predictor of treatment response.Summary sentence We have successfully developed and characterized a novel ex-vivo platform for personalized treatment of metastatic ovarian cancer.### Competing Interest StatementThe authors have declared no competing interest.

Pediatric low grade glioma (pLGG), the most common subgroup of central nervous system tumors in children, suffers from a paucity of models that support pLGG biological discovery, preclinical target identification, and drug screening studies. We propose the use of an organotypic brain slice culture (OBSC) platform as a novel model in which we are able to reliably support fresh, zero-passage pLGG patient tumor tissue for preclinical testing. We performed individualized drug screening with the frequently used up-front agents: carboplatin, vincristine, trametinib, and dabrafenib. We have collected and established on OBSCs 100% (3/3) pLGGs since 2023. With these three patient samples, we have applied our quantitative drug screening algorithm, which combines tumor killing efficacy with healthy tissue toxicity, and yielded resultant drug sensitivity scores (DSS) to quantify patient-specific treatment efficacies. Overall, these early results suggest that the OBSC platform may serve as a novel and reliable model for pLGG, filling a critical void in representative models for this common and often highly morbid pediatric tumor.

Abstract A challenge in cancer research is developing reproducible, reliable, and practical models which can capture the complexity of cancer development and treatment. The development of functional precision medicine platforms is emerging as a promising strategy for improving pre-clinical drug testing and guiding clinical decisions. We have developed an organotypic brain slice culture (OBSC) technology composed of intact multicellular tissue which can be rapidly used for spatiotemporal drug response testing. OBSCs are reproducibly generated from Sprague-Dawley rat pups and used as living tissue substrates to culture treat different tumor cell lines and uncultured patient brain tumor resection tissue. We evaluated OBSC quality and reproducibility throughout the study by using a propidium iodide nuclear permeability assay. This technique enabled a broad dynamic range to distinguish between healthy and unhealthy slices. In these studies, the rat pup age at the time of generation had an impact on OBSC quality and quantity, indicating that OBSCs should be generated from eight-day-old pups. We also found that optimal OBSCs were generated using improved methods for brain dissection and OBSC culture conditions, establishing our robust, standardized procedure for OBSC generation. Following this optimization period, we continued to conduct quality control analysis with 6 OBSCs per batch for reproducibility. We also conducted immunohistochemistry analysis immediately after slicing and concluded that morphology of neurons in OBSCs remained unchanged between day 0 and 4. In addition, the activation of astrocytes attenuates by day 4 but persists in macrophages/microglia, suggesting that the myeloid cells can phagocytose dead cells and debris in OBSCs. In summary, these results indicate that the OBSC platform may be an effective model that accelerates preclinical drug testing and directs drug development towards clinical evaluation.