Glioblastoma (GBM) is highly resistant to treatment and invasion into the surrounding brain is a cancer hallmark that leads to recurrence despite surgical resection. With the emergence of precision medicine, patient-derived 3D systems are considered potentially robust GBM preclinical models. In this study, we screened a library of 22 anti-invasive compounds (i.e., NF-kB, GSK-3-B, COX-2, and tubulin inhibitors) using glioblastoma U-251 MG cell spheroids. We evaluated toxicity and invasion inhibition using a 3D Matrigel invasion assay. We next selected three compounds that inhibited invasion and screened them in patient-derived glioblastoma organoids (GBOs). We developed a platform using available macros for FIJI/ImageJ to quantify invasion from the outer margin of organoids. Our data demonstrated that a high-throughput invasion screening can be done using both an established cell line and patient-derived 3D model systems. Tubulin inhibitor compounds had the best efficacy with U-251 MG cells, however, in ex vivo patient organoids the results were highly variable. Our results indicate that the efficacy of compounds is highly related to patient intra and inter-tumor heterogeneity. These results indicate that such models can be used to evaluate personal oncology therapeutic strategies.
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A Functional Precision Medicine Pipeline Combines Comparative Transcriptomics and Tumor Organoid Modeling to Identify Bespoke Treatment Strategies for Glioblastoma
Li Fraumeni syndrome (LFS) is a hereditary cancer predisposition syndrome caused by germline mutations in TP53. TP53 is the most common mutated gene in human cancer, occurring in 30–50% of glioblastomas (GBM). Here, we highlight a precision medicine platform to identify potential targets for a GBM patient with LFS. We used a comparative transcriptomics approach to identify genes that are uniquely overexpressed in the LFS GBM patient relative to a cancer compendium of 12,747 tumor RNA sequencing data sets, including 200 GBMs. STAT1 and STAT2 were identified as being significantly overexpressed in the LFS patient, indicating ruxolitinib, a Janus kinase 1 and 2 inhibitors, as a potential therapy. The LFS patient had the highest level of STAT1 and STAT2 expression in an institutional high-grade glioma cohort of 45 patients, further supporting the cancer compendium results. To empirically validate the comparative transcriptomics pipeline, we used a combination of adherent and organoid cell culture techniques, including ex vivo patient-derived organoids (PDOs) from four patient-derived cell lines, including the LFS patient. STAT1 and STAT2 expression levels in the four patient-derived cells correlated with levels identified in the respective parent tumors. In both adherent and organoid cultures, cells from the LFS patient were among the most sensitive to ruxolitinib compared to patient-derived cells with lower STAT1 and STAT2 expression levels. A spheroid-based drug screening assay (3D-PREDICT) was performed and used to identify further therapeutic targets. Two targeted therapies were selected for the patient of interest and resulted in radiographic disease stability. This manuscript supports the use of comparative transcriptomics to identify personalized therapeutic targets in a functional precision medicine platform for malignant brain tumors.
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- Award ID(s):
- 1903357
- NSF-PAR ID:
- 10329135
- Date Published:
- Journal Name:
- Cells
- Volume:
- 10
- Issue:
- 12
- ISSN:
- 2073-4409
- Page Range / eLocation ID:
- 3400
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Abstract Mutations in the
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Abstract Following treatment with androgen receptor (AR) pathway inhibitors, ≈20% of prostate cancer patients progress by shedding their AR‐dependence. These tumors undergo epigenetic reprogramming turning castration‐resistant prostate cancer adenocarcinoma (CRPC‐Adeno) into neuroendocrine prostate cancer (CRPC‐NEPC). No targeted therapies are available for CRPC‐NEPCs, and there are minimal organoid models to discover new therapeutic targets against these aggressive tumors. Here, using a combination of patient tumor proteomics, RNA sequencing, spatial‐omics, and a synthetic hydrogel‐based organoid, putative extracellular matrix (ECM) cues that regulate the phenotypic, transcriptomic, and epigenetic underpinnings of CRPC‐NEPCs are defined. Short‐term culture in tumor‐expressed ECM differentially regulated DNA methylation and mobilized genes in CRPC‐NEPCs. The ECM type distinctly regulates the response to small‐molecule inhibitors of epigenetic targets and Dopamine Receptor D2 (DRD2), the latter being an understudied target in neuroendocrine tumors. In vivo patient‐derived xenograft in immunocompromised mice showed strong anti‐tumor response when treated with a DRD2 inhibitor. Finally, we demonstrate that therapeutic response in CRPC‐NEPCs under drug‐resistant ECM conditions can be overcome by first cellular reprogramming with epigenetic inhibitors, followed by DRD2 treatment. The synthetic organoids suggest the regulatory role of ECM in therapeutic response to targeted therapies in CRPC‐NEPCs and enable the discovery of therapies to overcome resistance.
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null (Ed.)Type-II diabetes (T2D) patients affected by underlying hyperglycemic (high glucose/blood sugar) conditions often suffer from cardiac atrophy, resulting in tissue mass reduction and debilitating cardiac health. To understand pathophysiological mechanisms during progression of cardiac atrophy, a 3D bioprinted organoid platform was developed from a mixture of hydrogels containing human cardiac cells, including cardiomyocytes (CM), fibroblasts (CF) and endothelial cells (EC), to mimic the functionality of the in-vivo tissue. The organoids were cultured using normoglycemic- or hyperglycemic-conditions. The expression of essential biomarkers in these organoids, for myocardin (Myocd), troponin-I (TRP-I), fibroblast protein-1 (FSP-1) and endothelin-1 (ET-1) was confirmed. To assess the physiological cellular connections during hyperglycemia, the presence of Connexin-43 (CX-43) was assessed in the presence of a CX-43 blocker, gap26. Epigenomic tools were used to simultaneously interrogate histone-modifications by histone 3 lysine 9 mono-methylation (H3K9me1) along with the co-regulation of inflammatory mediators, such as the high mobility group box 1 (HMGB1) and toll like receptor 4 (TLR4) in the cardiac organoids cultured using normal versus hyperglycemic conditions. Organoids exposed to high glucose showed an increased expression of H3K9me1 as well as inflammatory mediators HMGB1 and TLR4. Hyperglycemia also exhibited alterations in expression of Myocd and FSP-1 in the organoids, compared to normoglycemic conditions. Treatment with gap26 affected the CX-43 expression significantly, in organoids cultured under hyperglycemia suggesting that high glucose conditions associated with prolonged diabetes may lead to compromised CM-CF coupling, essential for maintenance of cardiac functionality. Increased levels of H3K9me1 suggest decreased expression of Myocd, which may lead to CM degeneration. Epigenetic modifications including alterations in histone methylation in regulation of the myocardial genes and gap junction proteins under hyperglycemic conditions, may lead to cardiac atrophy. We expect to establish an actual T2D patient iPSC cell derived cardiac platform, to offer new therapeutic opportunities within the field.more » « less
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Abstract Temozolomide (TMZ) is the standard‐of‐care chemotherapy drug for treating glioblastomas (GBMs), the most aggressive cancer that affects people of all ages. However, its therapeutic efficacy is limited by the drug‐resistance mediated by a DNA repair protein, O6‐methylguanine‐DNA methyltransferase (MGMT), which eliminates the TMZ‐induced DNA lesions. Here, the development of an iron oxide nanoparticle (NP) system for targeted delivery of small interfering RNAs to suppress the TMZ‐resistance gene (MGMT) is reported. The NPs are able to overcome biological barriers, bind specifically to tumor cells, and reduce MGMT expression in tumors of mice bearing orthotopic GBM serially passaged patient‐derived xenografts. The treatment with sequential administration of this NP and TMZ result in increased apoptosis of GBM stem‐like cells, reduced tumor growth, and significantly prolonged survival as compared to mice treated with TMZ alone. This study introduces an approach that holds great promise to improve the outcomes of GBM patients.
- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.3390/cells10123400
