Generation and Culture of Human Pancreatic Ductal Adenocarcinoma Organoids from Resected Tumor Specimens

Baker, Lindsey A.; Tiriac, Hervé; Tuveson, David A.

doi:10.1007/978-1-4939-8879-2_9

Cited by 29 publications

(26 citation statements)

References 8 publications

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“…These limitations hamper their use in studies of pancreas duct cell biology and genetics as well as for disease modelling of the exocrine compartment and potential cell therapy approaches. While healthy, non-transformed, human pancreas tissue had proven difficult to maintain ex vivo, human pancreas organoids derived from tumour tissue [21,32] or tumour biopsies [33,34] have already been established and utilised for modelling pancreas cancer in vitro and for identifying drug sensitivities, as these faithfully recapitulate the architecture, transcriptome and mutational landscape of the tumour of origin in a patient-specific manner. However, other exocrine pancreas diseases, such as cystic fibrosis or pancreatitis, have not yet been modelled in vitro due to the lack of a culture system for manipulating primary human ductal epithelium ex vivo.…”

Section: Discussionmentioning

confidence: 99%

Long-term expansion, genomic stability and in vivo safety of adult human pancreas organoids

Georgakopoulos

Prior

Angres³

et al. 2020

BMC Dev Biol

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Background: Pancreatic organoid systems have recently been described for the in vitro culture of pancreatic ductal cells from mouse and human. Mouse pancreatic organoids exhibit unlimited expansion potential, while previously reported human pancreas organoid (hPO) cultures do not expand efficiently long-term in a chemically defined, serum-free medium. We sought to generate a 3D culture system for long-term expansion of human pancreas ductal cells as hPOs to serve as the basis for studies of human pancreas ductal epithelium, exocrine pancreatic diseases and the development of a genomically stable replacement cell therapy for diabetes mellitus. Results: Our chemically defined, serum-free, human pancreas organoid culture medium supports the generation and expansion of hPOs with high efficiency from both fresh and cryopreserved primary tissue. hPOs can be expanded from a single cell, enabling their genetic manipulation and generation of clonal cultures. hPOs expanded for months in vitro maintain their ductal morphology, biomarker expression and chromosomal integrity. Xenografts of hPOs survive long-term in vivo when transplanted into the pancreas of immunodeficient mice. Notably, mouse orthotopic transplants show no signs of tumorigenicity. Crucially, our medium also supports the establishment and expansion of hPOs in a chemically defined, modifiable and scalable, biomimetic hydrogel. Conclusions: hPOs can be expanded long-term, from both fresh and cryopreserved human pancreas tissue in a chemically defined, serum-free medium with no detectable tumorigenicity. hPOs can be clonally expanded, genetically manipulated and are amenable to culture in a chemically defined hydrogel. hPOs therefore represent an abundant source of pancreas ductal cells that retain the characteristics of the tissue-of-origin, which opens up avenues for modelling diseases of the ductal epithelium and increasing understanding of human pancreas exocrine biology as well as for potentially producing insulin-secreting cells for the treatment of diabetes.

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Section: Discussionmentioning

confidence: 99%

Long-term expansion, genomic stability and in vivo safety of adult human pancreas organoids

Georgakopoulos

Prior

Angres³

et al. 2020

BMC Dev Biol

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“…Previous studies showed different approaches defining the "right" organoid morphology and characteristics, which resemble traits of the corresponding tumor [8,9,19,25,26,43]. Baker et al described organoid passaging for five or more times as sufficient organoid propagation, which was achieved in 66% of initiated cultures.…”

Section: Discussionmentioning

confidence: 99%

“…Baker et al described organoid passaging for five or more times as sufficient organoid propagation, which was achieved in 66% of initiated cultures. No correlation between traits of the originating tumor and corresponding organoids has been made regarding cellular architecture or histopathological features [43]. Boj et al described murine and human PDAC tumor models.…”

Section: Discussionmentioning

confidence: 99%

Novel methods for in vitro modeling of pancreatic cancer reveal important aspects for successful primary cell culture

et al. 2020

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Background: Pancreatic cancer remains a fatal disease. Experimental systems are needed for personalized treatment strategies, drug testing and to further understand tumor biology. Cell cultures can serve as an excellent preclinical platform, but their generation remains challenging. Methods: Tumor cells from surgically removed pancreatic ductal adenocarcinoma (PDAC) specimens were cultured under novel protocols. Cellular growth and composition were analyzed and culture conditions were continuously optimized. Characterization of cell cultures and primary tumors was performed via hematoxylin and eosin (HE) and immunofluorescence (IF) staining. Results: Protocols for two-and three-dimensional PDAC primary cell cultures could successfully be established. Primary cell culture depended on dissociation techniques, growth factor supplementation and extracellular matrix components containing Matrigel being crucial for the transformation to three-dimensional PDAC organoids. The generated cultures showed to be highly resemblant to established PDAC primary cell cultures. HE and IF staining for cell culture and corresponding primary tumor characterization could successfully be performed. Conclusions: The work presented herein shows novel and effective methods to successfully establish primary PDAC cell cultures in a distinct time frame. Factors contributing to cell growth and differentiation could be identified with important implications for further primary cell culture protocols. The established protocols might serve as novel tools in personalized tumor therapy.

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“…Nevertheless, these models carry numerous constraints, especially in poorly resectable, low-cellularity cancers such as PDAC, where the issue of representativity, both cellular (intratumoural subpopulations) and at the population level (interpatient tumour heterogeneity) plays a limiting role. Organoid culture methods have been recently established from clinical specimens [43][44][45][46][47] and represent an incredible advantage over monolayer cell lines: they can be generated from a resected PDAC in about 2-4 weeks [43], are amenable to therapeutic screening as well as genetic and biochemical perturbation [48], and are able to recapitulate interactions between tumour and stromal compartment. Because organoids can be generated with high efficiency and speed from fine-needle aspirations, biopsies or resection specimens [43][44][45][46], they can serve as a personal cancer model (Fig.…”

Section: Organoids (Patient-derived Organoids)mentioning

confidence: 99%

“…Hence, the efficacy may be increased by personalizing the type of therapy given to any patient with PDAC. Per-sonalized treatment could soon become a more standard practice by using these cell cultures for extensive molecular diagnosis and drug screening [43,47,49]. Drug sensitivity assays can give a clinically actionable sensitivity profile of a patient's cancer.…”

Section: Organoids (Patient-derived Organoids)mentioning

confidence: 99%

Molecular biology in pancreatic ductal adenocarcinoma: implications for future diagnostics and therapy

et al. 2019

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Background Novel technology has enabled researchers to better characterize pancreatic cancers at the molecular level. We wanted to explore some of the emerging discoveries, such as molecular subclassification, use of liquid biopsy and use of organoids in cancer assessment. Methods A literature review with a search specific to the topic, with recent reviews in major journals and a focus on the last 5 years (until December 2018), was done. Results Pancreatic ductal adenocarcinoma (PDAC) may now be classified into clinical subgroups based on the predominant genomic profiles, but consensus on one classification system is lacking. Several subtypes have been suggested, including categories such as basal-like, stroma-activated, desmoplastic, pure classical and immune classical types. Further refinement may translate into clinically meaningful groups for therapeutic or prognostic purposes. Liquid

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Generation and Culture of Human Pancreatic Ductal Adenocarcinoma Organoids from Resected Tumor Specimens

Cited by 29 publications

References 8 publications

Long-term expansion, genomic stability and in vivo safety of adult human pancreas organoids

Long-term expansion, genomic stability and in vivo safety of adult human pancreas organoids

Novel methods for in vitro modeling of pancreatic cancer reveal important aspects for successful primary cell culture

Molecular biology in pancreatic ductal adenocarcinoma: implications for future diagnostics and therapy

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