Patient-derived Xenograft Cultures Research Articles

Pan-Cancer Pharmacogenomic Analysis of Patient-Derived Tumor Cells Using Clinically Relevant Drug Exposures.

As a result of tumor heterogeneity and solid cancers harboring multiple molecular defects, precision medicine platforms in oncology are most effective when both genetic and pharmacologic determinants of a tumor are evaluated. Expandable patient-derived xenograft (PDX) mouse tumor and corresponding PDX culture (PDXC) models recapitulate many of the biological and genetic characteristics of the original patient tumor, allowing for a comprehensive pharmacogenomic analysis. Here, the somatic mutations of 23 matched patient tumor and PDX samples encompassing four cancers were first evaluated using next-generation sequencing (NGS). 19 antitumor agents were evaluated across 78 patient-derived tumor cultures using clinically relevant drug exposures. A binarization threshold sensitivity classification determined in culture (PDXC) was used to identify tumors that best respond to drug in vivo (PDX). Using this sensitivity classification, logic models of DNA mutations were developed for 19 antitumor agents to predict drug response. We determined that the concordance of somatic mutations across patient and corresponding PDX samples increased as variant allele frequency increased. Notable individual PDXC responses to specific drugs, as well as lineage-specific drug responses were identified. Robust responses identified in PDXC were recapitulated in vivo in PDX-bearing mice and logic modeling determined somatic gene mutation(s) defining response to specific antitumor agents. In conclusion, combining NGS of primary patient tumors, high-throughput drug screen using clinically relevant doses, and logic modeling, can provide a platform for understanding response to therapeutic drugs targeting cancer.

Abstract 4106: High throughput chemical profiling of chemoresistant triple negative breast cancer

Abstract Triple-negative breast cancer (TNBC) makes up approximately 10-20% of all breast cancers, is characterized by a lack of targeted therapies, and in approximately 40-50% of cases a significant residual cancer burden remains after neoadjuvant chemotherapy, thus presenting a need to identify actionable targets in both treatment naive and resistant settings. To address the unmet treatment needs of chemo-resistant patients, our multi-institutional collaborative of clinical oncologists and basic researchers are assembling a panel of patient-derived xenograft (PDX) models of TNBC as part of an ongoing IRB-approved clinical trial at MD Anderson Cancer Center (ARTEMIS; NCT02276443). PDX models are generated from serial fine needle aspirates of primary TNBC from patients (1) prior to treatment, and in non-responding tumors (2) after four cycles of Adriamycin plus cyclophosphamide (AC), and (3) after a 3-month course of targeted therapy. We first developed a standardized intake procedure that simultaneously optimized cell seeding densities in 384-well plates, tested multiple cell culturing medias, determined baseline phenotypic characteristics, and established robust assay controls. After optimization, we performed high-throughput drug screens with a panel of 634 mechanistically annotated chemical probes that are FDA approved, late stage investigational or mechanistic probes. A bioluminescent reporter quantifying the amount of ATP (CellTiter Glo, Promega) was used as a surrogate readout of cell viability, which is well suited for the majority of PDX cultures that grew as non-adherent multi-cellular aggregates as well as being integrable into larger datasets run in a similar format. Screening assays are performed with 3-4 technical replicates over a three log concentration range (10.0, 1.0, 0.1 uM). The effectiveness of each drug is summarized as an area under the fitted concentration-response curve, simultaneously capturing alterations in potency and efficacy. In total, we have tested 21 PDX models isolated from 14 unique patients using this strategy. From these data, we show that PDX models generated from individual patients have unique pharmacologic profiles with little correspondence to molecularly defined sub-classifications, which may suggest the need for the development of an actionable sub-classification strategy to better stratify patients. Despite the significant heterogeneity across pharmacologic profiles, we identified a consolidated set of drugs targeting specific epigenetic processes and protein homeostasis that have increased activity in a subset of chemo-resistant samples. We are currently collecting orthogonal data including next-generation sequencing and transcriptomics to further elucidate the mechanism of these targets and identify potential biomarkers. Citation Format: Reid Trenton Powell, Abena Redwood, Lei Guo, Shirong Cai, Yizheng Tu, Yan Jiang, Xuan Liu, Xinhui Zhou, Clifford Stephan, Peter Davies, Jeffrey T. Chang, Stacey Moulder, William F. Symmans, Helen Piwnica-Worms. High throughput chemical profiling of chemoresistant triple negative breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4106.

Abstract LB-036: Targeting the WNT-pathway in 3D-cultures of pancreatic cancer biopsies

Abstract Creating patient-derived xenograft (PDX) models from human cancers allows bio-banking, propagation, characterization and drug testing on a large panel of tumor variants. We developed 3D in vitro culture methods for PDX model-derived cell isolates. Each individual model was subjected to a wide panel of culture medium formulations, amongst which some media contain WNT-pathway stimulating components. We identified an absolute dependency of a pancreatic in vitro PDX tumor to the WNT-signaling potentiating factor R-Spondin, while other pancreas tumors showed little or no dependency. To further study the dependency of pancreatic PDX tumors on the functionality and signaling of the WNT-pathway, we exposed in vitro PDX cultures to increasing doses of anti-FZD7, anti-FZD8 and to the Porcupine inhibitor LGK974, which blocks WNT palmitoylation and therefore secretion. We also applied the same approach to establish cultures from patient-derived pancreatic cancer biopsies. Our findings are that the WNT-pathway is functional and contributing to pancreas cancer outgrowth and morphological differentiation to different extents in different patient-models. Our in vitro culture system in combination with 3D phenotypic analysis allows in-detail study of the WNT-pathway, as well as by supplementation or omission of secreted factors, addition of small molecule inhibitors and by exposure to antibodies. These in vitro models can be used to test novel compounds and antibodies targeting the WNT pathway and underscore the WNT pathway as a potential target in pancreatic cancer. Citation Format: Bram Herpers, Sander Basten, Kuan Yan, Thomas Dijkmans, Julia Schueler, Torsten Giesemann, Leo S. Price. Targeting the WNT-pathway in 3D-cultures of pancreatic cancer biopsies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-036.

Abstract LB-A16: Targeting the WNT-pathway in 3D-cultured in vitro PDX models for pancreas cancer

Abstract Creating patient-derived xenograft (PDX) models from human cancers allows bio-banking, propagation, characterization and drug testing on a large panel of tumor variants. To scale up the number of compound tests, identify relevant compound doses, increase the throughput, and study relevant cancer-driving pathways we developed 3D in vitro culture methods for PDX model-derived cell isolates. In this effort, each individual model is subjected to a wide panel of culture medium formulations, amongst which some media contain WNT-pathway stimulating components. In one of our campaigns we identified an absolute dependency of one of the tested Pancreatic in vitro PDX models to the WNT-signaling potentiating factor R-Spondin, while other Pancreas models showed little or no dependency. To further study the dependency of Pancreatic PDX models on the functionality and signaling of the WNT-pathway, we exposed these in vitro PDX models to increasing doses of bio-similars of 18R5 (anti-FZD7), 54F28 (FZD8-Fc decoy receptor) and to the Porcupine inhibitor LGK974, which blocks WNT palmitoylation and therefore secretion. Our findings are that the WNT-pathway is functional and contributing to Pancreas cancer outgrowth and morphological differentiation at different extents and levels in different patient-models. Our in vitro PDX culture system in combination with 3D phenotypic analysis allows in-detail study of the WNT-pathway, as well as by supplementation or omission of secreted factors, addition of small molecule inhibitors and by exposure to antibodies. These in vitro PDX models can be used to test novel compounds and antibodies targeting the WNT pathway. Citation Format: Thomas Dijkmans. Targeting the WNT-pathway in 3D-cultured in vitro PDX models for pancreas cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-A16.

Abstract LB-A18: Targeting the WNT-pathway in 3D-cultured in vitro PDX models for pancreas cancer

Abstract Creating patient-derived xenograft (PDX) models from human cancers allows bio-banking, propagation, characterization and drug testing on a large panel of tumor variants. To scale up the number of compound tests, identify relevant compound doses, increase the throughput, and study relevant cancer-driving pathways we developed 3D in vitro culture methods for PDX model-derived cell isolates. In this effort, each individual model is subjected to a wide panel of culture medium formulations, amongst which some media contain WNT-pathway stimulating components. We identified an absolute dependency of a pancreatic in vitro PDX tumor to the WNT-signaling potentiating factor R-Spondin, while other pancreas tumors showed little or no dependency. To further study the dependency of pancreatic PDX tumors on the functionality and signaling of the WNT-pathway, we exposed in vitro PDX cultures to increasing doses of anti-FZD7, anti-FZD8 and to the Porcupine inhibitor LGK974, which blocks WNT palmitoylation and therefore secretion. Our findings are that the WNT-pathway is functional and contributing to pancreas cancer outgrowth and morphological differentiation to different extents in different patient-models. Our in vitro PDX culture system in combination with 3D phenotypic analysis allows in-detail study of the WNT-pathway, as well as by supplementation or omission of secreted factors, addition of small molecule inhibitors and by exposure to antibodies. These in vitro PDX models can be used to test novel compounds and antibodies targeting the WNT-pathway and underscore the WNT-pathway as a potential target in pancreatic cancer. Citation Format: Thomas Dijkmans, Bram Herpers, Sander Basten, Kuan Yan, Torsten Giesemann, Julia Schueler, Leo Price. Targeting the WNT-pathway in 3D-cultured in vitro PDX models for pancreas cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-A18.

Abstract LB-044: A 384-well in vitro PDX culture platform for compound screening and PDX model selection

Abstract Background. There is very high attrition of compounds in early stage anti-cancer drug discovery. This is partly due to the failure of early stage drug testing platforms to represent human biology. 2D cultures of human cells lack the realistic complexity of an organism, leading to overestimation of compound potency. Animal models do retain complexity, but are limited in throughput and frequently fail to reflect human tumor biology. Patient-derived xenograft (PDX) models in immune-compromised mice allow propagation of and compound testing in human-derived tumors. To expand the potential of these human-relevant PDX models, we sought to develop 3D in vitro culture methods for PDX-derived tumor cells that show in vivo-like growth characteristics, invasion and responses to therapeutics. In combination with advanced 3D image analysis methods, we created a unique high throughput in vitro PDX screening platform that not only allows efficient identification of active and selective molecules but also enables selection of the optimal PDX tumor models for subsequent validation of candidates in vivo. Results. Each PDX model has its own unique growth characteristics. Hydrogel and growth media composition were optimized to support growth of tumor tissues in vitro from cells derived from bladder, gastric, breast, colon and lung cancer PDX tumors. Tumor tissues were cultured in a 384-well format and used to screen chemotherapeutics, small molecules, antibodies and antibody-drug-conjugates (ADCs). Using OcellO's 3D image analysis platform, Ominer, tumoroid growth, cell proliferation, apoptosis, invasion, cell polarity, differentiation and other aspects of cell and tissue architecture were analyzed and the effects of compound exposure on tumoroids was determined. By performing feature training based on reference compounds, we selected ±10 morphological features (out of more than 500) to generate a phenotypic signature that described the unique phenotypic change induced by each compound. Different compounds that target the same molecule were found to induce a similar morphological change whereas compounds with off-target effects could be discriminated. This approach enabled a high resolution evaluation and comparison of compound activity in an automated manner. Conclusions. We established several bladder, gastric, breast, colon and lung cancer PDX model-derived 3D tumor cultures in which standard-of-care and novel therapeutic agents (small molecules, antibodies and ADCs) can efficiently be screened, based on therapeutically relevant parameters and their changing morphological profile. This method enables both the in vitro selection of promising compounds in a pre-clinically relevant setting and the selection of optimum PDX tumor models for follow-up in vivo studies. This highly translational in vitro-in vivo PDX pipeline is expected to reduce attrition and increase efficiency in early drug-discovery. Citation Format: Bram Herpers, Lidia Daszkiewicz, Torsten Giesemann, Leo Price. A 384-well in vitro PDX culture platform for compound screening and PDX model selection. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-044.