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In Rspondin-based 3D cultures, Lgr5 stem cells from multiple organs form ever-expanding epithelial organoids that retain their tissue identity. We report the establishment of tumor organoid cultures from 20 consecutive colorectal carcinoma (CRC) patients. For most, organoids were also generated from adjacent normal tissue. Organoids closely recapitulate several properties of the original tumor. The spectrum of genetic changes within the "living biobank" agrees well with previous large-scale mutational analyses of CRC. Gene expression analysis indicates that the major CRC molecular subtypes are represented. Tumor organoids are amenable to high-throughput drug screens allowing detection of gene-drug associations. As an example, a single organoid culture was exquisitely sensitive to Wnt secretion (porcupine) inhibitors and carried a mutation in the negative Wnt feedback regulator RNF43, rather than in APC. Organoid technology may fill the gap between cancer genetics and patient trials, complement cell-line- and xenograft-based drug studies, and allow personalized therapy design. PAPERCLIP.
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Bancos de Muestras Biológicas , Neoplasias Colorrectales/patología , Ensayos de Selección de Medicamentos Antitumorales/métodos , Organoides , Neoplasias Colorrectales/tratamiento farmacológico , Proteínas de Unión al ADN/metabolismo , Humanos , Proteínas Oncogénicas/metabolismo , Técnicas de Cultivo de Órganos , Organoides/efectos de los fármacos , Medicina de Precisión , Ubiquitina-Proteína LigasasRESUMEN
The pleural lining of the thorax regulates local immunity, inflammation and repair. A variety of conditions, both benign and malignant, including pleural mesothelioma, can affect this tissue. A lack of knowledge concerning the mesothelial and stromal cells comprising the pleura has hampered the development of targeted therapies. Here, we present the first comprehensive single-cell transcriptomic atlas of the human parietal pleura and demonstrate its utility in elucidating pleural biology. We confirm the presence of known universal fibroblasts and describe novel, potentially pleural-specific, fibroblast subtypes. We also present transcriptomic characterisation of multiple in vitro models of benign and malignant mesothelial cells, and characterise these through comparison with in vivo transcriptomic data. While bulk pleural transcriptomes have been reported previously, this is the first study to provide resolution at the single-cell level. We expect our pleural cell atlas will prove invaluable to those studying pleural biology and disease. It has already enabled us to shed light on the transdifferentiation of mesothelial cells, allowing us to develop a simple method for prolonging mesothelial cell differentiation in vitro.
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Mesotelioma Maligno , Mesotelioma , Neoplasias Pleurales , Humanos , Pleura/patología , Mesotelioma/genética , Mesotelioma/patología , Mesotelioma Maligno/patología , Neoplasias Pleurales/genética , Neoplasias Pleurales/patología , Perfilación de la Expresión GénicaRESUMEN
Human tissue three-dimensional (3D) organoid cultures have the potential to reproduce in vitro the physiological properties and cellular architecture of the organs from which they are derived. The ability of organoid cultures derived from human stomach, liver, kidney, and colon to metabolically activate three dietary carcinogens, aflatoxin B1 (AFB1), aristolochic acid I (AAI), and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), was investigated. In each case, the response of a target tissue (liver for AFB1; kidney for AAI; colon for PhIP) was compared with that of a nontarget tissue (gastric). After treatment cell viabilities were measured, DNA damage response (DDR) was determined by Western blotting for p-p53, p21, p-CHK2, and γ-H2AX, and DNA adduct formation was quantified by mass spectrometry. Induction of the key xenobiotic-metabolizing enzymes (XMEs) CYP1A1, CYP1A2, CYP3A4, and NQO1 was assessed by qRT-PCR. We found that organoids from different tissues can activate AAI, AFB1, and PhIP. In some cases, this metabolic potential varied between tissues and between different cultures of the same tissue. Similarly, variations in the levels of expression of XMEs were observed. At comparable levels of cytotoxicity, organoids derived from tissues that are considered targets for these carcinogens had higher levels of adduct formation than a nontarget tissue.
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Aductos de ADN , Neoplasias , Humanos , Carcinógenos/toxicidad , Carcinógenos/metabolismo , Hígado/metabolismo , Organoides/metabolismoAsunto(s)
Investigación Biomédica/organización & administración , Investigación Biomédica/tendencias , Objetivos , Cooperación Internacional , Terapia Molecular Dirigida/tendencias , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Antineoplásicos Inmunológicos/farmacología , Antineoplásicos Inmunológicos/uso terapéutico , Investigación Biomédica/economía , Sistemas CRISPR-Cas/genética , Supervivencia Celular/efectos de los fármacos , Análisis Mutacional de ADN , Conjuntos de Datos como Asunto , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Difusión de la Información , Aprendizaje Automático , Mutación , Neoplasias/metabolismo , Neoplasias/patología , Proyectos Piloto , Medicina de Precisión , Reproducibilidad de los ResultadosRESUMEN
We report the derivation of 30 patient-derived organoid lines (PDOs) from tumors arising in the pancreas and distal bile duct. PDOs recapitulate tumor histology and contain genetic alterations typical of pancreatic cancer. In vitro testing of a panel of 76 therapeutic agents revealed sensitivities currently not exploited in the clinic, and underscores the importance of personalized approaches for effective cancer treatment. The PRMT5 inhibitor EZP015556, shown to target MTAP (a gene commonly lost in pancreatic cancer)-negative tumors, was validated as such, but also appeared to constitute an effective therapy for a subset of MTAP-positive tumors. Taken together, the work presented here provides a platform to identify novel therapeutics to target pancreatic tumor cells using PDOs.
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Organoids are 3D cultures that to some extent reproduce the structure, composition and function of the mammalian tissues from which they derive, thereby creating in vitro systems with more in vivo-like characteristics than 2D monocultures. Here, the ability of human organoids derived from normal gastric, pancreas, liver, colon and kidney tissues to metabolise the environmental carcinogen benzo[a]pyrene (BaP) was investigated. While organoids from the different tissues showed varied cytotoxic responses to BaP, with gastric and colon organoids being the most susceptible, the xenobiotic-metabolising enzyme (XME) genes, CYP1A1 and NQO1, were highly upregulated in all organoid types, with kidney organoids having the highest levels. Furthermore, the presence of two key metabolites, BaP-t-7,8-dihydrodiol and BaP-tetrol-l-1, was detected in all organoid types, confirming their ability to metabolise BaP. BaP bioactivation was confirmed both by the activation of the DNA damage response pathway (induction of p-p53, pCHK2, p21 and γ-H2AX) and by DNA adduct formation. Overall, pancreatic and undifferentiated liver organoids formed the highest levels of DNA adducts. Colon organoids had the lowest responses in DNA adduct and metabolite formation, as well as XME expression. Additionally, high-throughput RT-qPCR explored differences in gene expression between organoid types after BaP treatment. The results demonstrate the potential usefulness of organoids for studying environmental carcinogenesis and genetic toxicology.
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Benzo(a)pireno , Aductos de ADN , Organoides , Humanos , Activación Metabólica , Benzo(a)pireno/toxicidad , Citocromo P-450 CYP1A1/metabolismo , Aductos de ADN/metabolismo , Hígado/metabolismo , Organoides/efectos de los fármacos , Organoides/metabolismoRESUMEN
Malignant pleural mesothelioma (MPM) is an aggressive cancer most commonly caused by prior exposure to asbestos. Median survival is 12-18 months, since surgery is ineffective and chemotherapy offers minimal benefit. Preclinical models that faithfully recapitulate the genomic and histopathological features of cancer are critical for the development of new treatments. The most commonly used models of MPM are two-dimensional cell lines established from primary tumours or pleural fluid. While these have provided some important insights into MPM biology, these cell models have significant limitations. In order to address some of these limitations, spheroids and microfluidic chips have more recently been used to investigate the role of the three-dimensional environment in MPM. Efforts have also been made to develop animal models of MPM, including asbestos-induced murine tumour models, MPM-prone genetically modified mice and patient-derived xenografts. Here, we discuss the available in vitro and in vivo models of MPM and highlight their strengths and limitations. We discuss how newer technologies, such as the tumour-derived organoids, might allow us to address the limitations of existing models and aid in the identification of effective treatments for this challenging-to-treat disease.
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Amianto , Neoplasias Pulmonares , Mesotelioma Maligno , Mesotelioma , Neoplasias Pleurales , Animales , RatonesRESUMEN
In vitro cancer cell cultures are facile experimental models used widely for research and drug development. Many cancer cell lines are available and efforts are ongoing to derive new models representing the histopathological and molecular diversity of tumours. Cell models have been generated by multiple laboratories over decades and consequently their annotation is incomplete and inconsistent. Furthermore, the relationships between many patient-matched and derivative cell lines have been lost, and accessing information and datasets is time-consuming and difficult. Here, we describe the Cell Model Passports database; cellmodelpassports.sanger.ac.uk, which provides details of cell model relationships, patient and clinical information, as well as access to associated genetic and functional datasets. The Passports database currently contains curated details and standardized annotation for >1200 cell models, including cancer organoid cultures. The Passports will be updated with newly derived cell models and datasets as they are generated. Users can navigate the database via tissue, cancer-type, genetic feature and data availability to select a model most suitable for specific applications. A flexible REST-API provides programmatic data access and exploration. The Cell Model Passports are a valuable tool enabling access to high-dimensional genomic and phenotypic cancer cell model datasets empowering diverse research applications.
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Línea Celular Tumoral , Bases de Datos Factuales , Antineoplásicos , Conjuntos de Datos como Asunto , Desarrollo de Medicamentos , Genómica , Humanos , Modelos Biológicos , OrganoidesRESUMEN
PURPOSE: Patients with locally advanced rectal cancer often require neoadjuvant chemoradiation therapy to downstage the disease, but the response is variable with no predictive biomarkers. We have previously revealed through proteomic profiling that myoferlin is associated with response to radiation therapy. The aims of this study were to further validate this finding and explore the potential for myoferlin to act as a prognostic and/or therapeutic target. METHODS AND MATERIALS: Immunohistochemical analysis of a tissue microarray (TMA) for 111 patients was used to validate the initial proteomic findings. Manipulation of myoferlin was achieved using small interfering RNA, a small molecular inhibitor (wj460), and a CRISPR-Cas9 knockout cell line. Radiosensitization after treatment was assessed using 2-dimensional clonogenic assays, 3-dimensional spheroid models, and patient-derived organoids. Underlying mechanisms were investigated using electrophoresis, immunofluorescence, and immunoblotting. RESULTS: Analysis of both the diagnostic biopsy and tumor resection samples confirmed that low myoferlin expression correlated with a good response to neoadjuvant long-course chemoradiation therapy. High myoferlin expression was associated with spread to local lymph nodes and worse 5-year survival (P = .01; hazard ratio, 3.5; 95% CI, 1.27-10.04). This was externally validated using the Stratification in Colorectal Cancer database. Quantification of myoferlin using immunoblotting in immortalized colorectal cancer cell lines and organoids demonstrated that high myoferlin expression was associated with increased radioresistance. Biological and pharmacologic manipulation of myoferlin resulted in significantly increased radiosensitivity across all cell lines in 2-dimensional and 3-dimensional models. After irradiation, myoferlin knockdown cells had a significantly impaired ability to repair DNA double-strand breaks. This appeared to be mediated via nonhomologous end-joining. CONCLUSIONS: We have confirmed that high expression of myoferlin in rectal cancer is associated with poor response to neoadjuvant therapy and worse long-term survival. Furthermore, the manipulation of myoferlin led to increased radiosensitivity in vitro. This suggests that myoferlin could be targeted to enhance the sensitivity of patients with rectal cancer to radiation therapy, and further work is required.
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The epigenetic landscape of cancer is regulated by many factors, but primarily it derives from the underlying genome sequence. Chromothripsis is a catastrophic localized genome shattering event that drives, and often initiates, cancer evolution. We characterized five esophageal adenocarcinoma organoids with chromothripsis using long-read sequencing and transcriptome and epigenome profiling. Complex structural variation and subclonal variants meant that haplotype-aware de novo methods were required to generate contiguous cancer genome assemblies. Chromosomes were assembled separately and scaffolded using haplotype-resolved Hi-C reads, producing accurate assemblies even with up to 900 structural rearrangements. There were widespread differences between the chromothriptic and wild-type copies of chromosomes in topologically associated domains, chromatin accessibility, histone modifications, and gene expression. Differential epigenome peaks were most enriched within 10 kb of chromothriptic structural variants. Alterations in transcriptome and higher-order chromosome organization frequently occurred near differential epigenetic marks. Overall, chromothripsis reshapes gene regulation, causing coordinated changes in epigenetic landscape, transcription, and chromosome conformation.
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Adenocarcinoma , Cromotripsis , Neoplasias Esofágicas , Humanos , Haplotipos , Cromatina , Genoma , Adenocarcinoma/genéticaRESUMEN
Genetic screens in cancer cell lines inform gene function and drug discovery. More comprehensive screen datasets with multi-omics data are needed to enhance opportunities to functionally map genetic vulnerabilities. Here, we construct a second-generation map of cancer dependencies by annotating 930 cancer cell lines with multi-omic data and analyze relationships between molecular markers and cancer dependencies derived from CRISPR-Cas9 screens. We identify dependency-associated gene expression markers beyond driver genes, and observe many gene addiction relationships driven by gain of function rather than synthetic lethal effects. By combining clinically informed dependency-marker associations with protein-protein interaction networks, we identify 370 anti-cancer priority targets for 27 cancer types, many of which have network-based evidence of a functional link with a marker in a cancer type. Mapping these targets to sequenced tumor cohorts identifies tractable targets in different cancer types. This target prioritization map enhances understanding of gene dependencies and identifies candidate anti-cancer targets for drug development.
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Pruebas Genéticas , Neoplasias , Humanos , Fenotipo , Descubrimiento de Drogas , Neoplasias/genética , Neoplasias/patología , Línea Celular Tumoral , Sistemas CRISPR-CasRESUMEN
Organoid cell culture methodologies are enabling the generation of cell models from healthy and diseased tissue. Patient-derived cancer organoids that recapitulate the genetic and histopathological diversity of patient tumours are being systematically generated, providing an opportunity to investigate new cancer biology and therapeutic approaches. The use of organoid cultures for many applications, including genetic and chemical perturbation screens, is limited due to the technical demands and cost associated with their handling and propagation. Here we report and benchmark a suspension culture technique for cancer organoids which allows for the expansion of models to tens of millions of cells with increased efficiency in comparison to standard organoid culturing protocols. Using whole-genome DNA and RNA sequencing analyses, as well as medium-throughput drug sensitivity testing and genome-wide CRISPR-Cas9 screening, we demonstrate that cancer organoids grown as a suspension culture are genetically and phenotypically similar to their counterparts grown in standard conditions. This culture technique simplifies organoid cell culture and extends the range of organoid applications, including for routine use in large-scale perturbation screens.
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Neoplasias , Organoides , Técnicas de Cultivo de Célula , ADN , Humanos , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Neoplasias/patología , Organoides/patologíaRESUMEN
INTRODUCTION: We have previously reported that induction of epidermal growth factor receptor and ErbB2 in response to antihormonal agents may provide an early mechanism to allow breast cancer cells to evade the growth-inhibitory action of such therapies and ultimately drive resistant cell growth. More recently, the other two members of the ErbB receptor family, ErbB3 and ErbB4, have been implicated in antihormone resistance in breast cancer. In the present study, we have investigated whether induction of ErbB3 and/or ErbB4 may provide an alternative resistance mechanism to antihormonal action in a panel of four oestrogen receptor (ER)-positive breast cancer cell lines. METHODS: MCF-7, T47D, BT474 and MDAMB361 cell lines were exposed to fulvestrant (100 nM) for seven days, and effects on ErbB3/4 expression and signalling, as well as on cell growth, were assessed. Effects of heregulin ß1 (HRGß1) were also examined in the absence and presence of fulvestrant to determine the impact of ER blockade on the capacity of this ErbB3/4 ligand to promote signalling and cell proliferation. RESULTS: Fulvestrant potently reduced ER expression and transcriptional activity and significantly inhibited growth in MCF-7, T47D, BT474 and MDAMB361 cells. However, alongside this inhibitory activity, fulvestrant also consistently induced protein expression and activity of ErbB3 in MCF-7 and T47D cells and ErbB4 in BT474 and MDAMB361 cell lines. Consequently, fulvestrant treatment sensitised all cell lines to the actions of the ErbB3/4 ligand HRGß1 with enhanced ErbB3/4-driven signalling activity, reexpression of cyclin D1 and significant increases in cell proliferation being observed when compared to untreated cells. Indeed, in T47D and MDAMB361 HRGß1 was converted from a ligand having negligible or suppressive growth activity into one that potently promoted cell proliferation. Consequently, fulvestrant-mediated growth inhibition was completely overridden by HRGß1 in all four cell lines. CONCLUSIONS: These findings suggest that although antihormones such as fulvestrant may have potent acute growth-inhibitory activity in ER-positive breast cancer cells, their ability to induce and sensitise cells to growth factors may serve to reduce and ultimately limit their inhibitory activity.
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Receptores ErbB/metabolismo , Estradiol/análogos & derivados , Neurregulina-1/metabolismo , Receptor ErbB-3/metabolismo , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Ciclina D1/biosíntesis , Resistencia a Antineoplásicos , Receptores ErbB/biosíntesis , Estradiol/farmacología , Moduladores de los Receptores de Estrógeno/farmacología , Femenino , Fulvestrant , Humanos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Receptor ErbB-4 , Receptores de Estrógenos/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Since the approval of trastuzumab for the treatment of breast cancers more than two decades ago, many clinically effective targeted anti-cancer therapies have been developed. Here we consider the evidence that supports genomics-guided drug development and review the concept of oncogene addiction, including recent findings that inform this therapeutic approach. We consider non-oncogene addiction and how this synthetic-lethal paradigm could expand the range of new therapies, particularly for currently undruggable cancers. We discuss how CRISPR-based genetic screening is enhancing the ability to identify new targets. We conclude by considering opportunities for expanding the scope and refining the use of precision cancer medicines.
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Neoplasias de la Mama , Genómica , Neoplasias de la Mama/tratamiento farmacológico , Femenino , Humanos , Medicina de Precisión , TrastuzumabRESUMEN
Selecting appropriate cancer models is a key prerequisite for maximizing translational potential and clinical relevance of in vitro oncology studies. We developed CELLector: an R package and R Shiny application allowing researchers to select the most relevant cancer cell lines in a patient-genomic-guided fashion. CELLector leverages tumor genomics to identify recurrent subtypes with associated genomic signatures. It then evaluates these signatures in cancer cell lines to prioritize their selection. This enables users to choose appropriate in vitro models for inclusion or exclusion in retrospective analyses and future studies. Moreover, this allows bridging outcomes from cancer cell line screens to precisely defined sub-cohorts of primary tumors. Here, we demonstrate the usefulness and applicability of CELLector, showing how it can aid prioritization of in vitro models for future development and unveil patient-derived multivariate prognostic and therapeutic markers. CELLector is freely available at https://ot-cellector.shinyapps.io/CELLector_App/ (code at https://github.com/francescojm/CELLector and https://github.com/francescojm/CELLector_App).
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Línea Celular Tumoral/clasificación , Proyectos de Investigación , Animales , Línea Celular Tumoral/metabolismo , Genoma , Genómica/métodos , Humanos , Modelos Biológicos , Neoplasias/genética , Programas InformáticosRESUMEN
Drug sensitivity testing utilizing preclinical disease models such as cancer cell lines is an important and widely used tool for drug development. Importantly, when combined with molecular data such as gene copy number variation or somatic coding mutations, associations between drug sensitivity and molecular data can be used to develop markers to guide patient therapies. The use of organoids as a preclinical cancer model has become possible following recent work demonstrating that organoid cultures can be derived from patient tumors with a high rate of success. A genetic analysis of colon cancer organoids found that these models encompassed the majority of the somatic variants present within the tumor from which it was derived, and capture much of the genetic diversity of colon cancer observed in patients. Importantly, the systematic sensitivity testing of organoid cultures to anticancer drugs identified clinical gene-drug interactions, suggestive of their potential as preclinical models for testing anticancer drug sensitivity. In this chapter, we describe how to perform medium/high-throughput drug sensitivity screens using 3D organoid cell cultures.
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Antineoplásicos/farmacología , Proliferación Celular/efectos de los fármacos , Neoplasias del Colon/patología , Ensayos de Selección de Medicamentos Antitumorales/métodos , Neoplasias Esofágicas/patología , Técnicas de Cultivo de Órganos/métodos , Organoides/patología , Neoplasias del Colon/tratamiento farmacológico , Neoplasias Esofágicas/tratamiento farmacológico , Humanos , Organoides/efectos de los fármacosRESUMEN
The protocol Drug Sensitivity Assays of Human Cancer Organoid Cultures has now been made available open access under a CC BY 4.0 license.
RESUMEN
Cellular dormancy and heterogeneity in cell cycle length provide important explanations for treatment failure after adjuvant therapy with S-phase cytotoxics in colorectal cancer (CRC), yet the molecular control of the dormant versus cycling state remains unknown. We sought to understand the molecular features of dormant CRC cells to facilitate rationale identification of compounds to target both dormant and cycling tumor cells. Unexpectedly, we demonstrate that dormant CRC cells are differentiated, yet retain clonogenic capacity. Mouse organoid drug screening identifies that itraconazole generates spheroid collapse and loss of dormancy. Human CRC cell dormancy and tumor growth can also be perturbed by itraconazole, which is found to inhibit Wnt signaling through noncanonical hedgehog signaling. Preclinical validation shows itraconazole to be effective in multiple assays through Wnt inhibition, causing both cycling and dormant cells to switch to global senescence. These data provide preclinical evidence to support an early phase trial of itraconazole in CRC.
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Ciclo Celular/efectos de los fármacos , Neoplasias Colorrectales/patología , Itraconazol/farmacología , Animales , Puntos de Control del Ciclo Celular/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Separación Celular , Senescencia Celular/efectos de los fármacos , Neoplasias Colorrectales/metabolismo , Humanos , Ratones , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Organoides/efectos de los fármacos , Organoides/patología , Fenotipo , Receptores Acoplados a Proteínas G/metabolismo , Esferoides Celulares/efectos de los fármacos , Esferoides Celulares/metabolismo , Esferoides Celulares/patología , Coloración y Etiquetado , Vía de Señalización Wnt/efectos de los fármacosRESUMEN
Esophageal adenocarcinoma (EAC) incidence is increasing while 5-year survival rates remain less than 15%. A lack of experimental models has hampered progress. We have generated clinically annotated EAC organoid cultures that recapitulate the morphology, genomic, and transcriptomic landscape of the primary tumor including point mutations, copy number alterations, and mutational signatures. Karyotyping of organoid cultures has confirmed polyclonality reflecting the clonal architecture of the primary tumor. Furthermore, subclones underwent clonal selection associated with driver gene status. Medium throughput drug sensitivity testing demonstrates the potential of targeting receptor tyrosine kinases and downstream mediators. EAC organoid cultures provide a pre-clinical tool for studies of clonal evolution and precision therapeutics.
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Adenocarcinoma/tratamiento farmacológico , Evolución Clonal , Neoplasias Esofágicas/tratamiento farmacológico , Organoides/química , Proteínas Tirosina Quinasas Receptoras/genética , Adenocarcinoma/metabolismo , Anciano , Anciano de 80 o más Años , Variaciones en el Número de Copia de ADN , Análisis Mutacional de ADN , Ensayos de Selección de Medicamentos Antitumorales , Neoplasias Esofágicas/metabolismo , Femenino , Humanos , Concentración 50 Inhibidora , Cariotipificación , Masculino , Persona de Mediana Edad , Mutación , Medicina de Precisión , Análisis de Secuencia de ARN , TranscriptomaRESUMEN
Human liver cancer research currently lacks in vitro models that can faithfully recapitulate the pathophysiology of the original tumor. We recently described a novel, near-physiological organoid culture system, wherein primary human healthy liver cells form long-term expanding organoids that retain liver tissue function and genetic stability. Here we extend this culture system to the propagation of primary liver cancer (PLC) organoids from three of the most common PLC subtypes: hepatocellular carcinoma (HCC), cholangiocarcinoma (CC) and combined HCC/CC (CHC) tumors. PLC-derived organoid cultures preserve the histological architecture, gene expression and genomic landscape of the original tumor, allowing for discrimination between different tumor tissues and subtypes, even after long-term expansion in culture in the same medium conditions. Xenograft studies demonstrate that the tumorogenic potential, histological features and metastatic properties of PLC-derived organoids are preserved in vivo. PLC-derived organoids are amenable for biomarker identification and drug-screening testing and led to the identification of the ERK inhibitor SCH772984 as a potential therapeutic agent for primary liver cancer. We thus demonstrate the wide-ranging biomedical utilities of PLC-derived organoid models in furthering the understanding of liver cancer biology and in developing personalized-medicine approaches for the disease.