Organotypic culture assays for murine and human primary and metastatic-site tumors

Veena Padmanaban; Eloise M. Grasset; Neil M. Neumann; Andrew K. Fraser; Elodie Henriet; William Matsui; Phuoc T. Tran; Kevin J. Cheung; Dan Georgess; Andrew J. Ewald

doi:10.1038/s41596-020-0335-3

Organotypic culture assays for murine and human primary and metastatic-site tumors

Veena Padmanaban, Eloise M. Grasset, Neil M. Neumann, Andrew K. Fraser, Elodie Henriet, William Matsui, Phuoc T. Tran, Kevin J. Cheung, Dan Georgess, Andrew J. Ewald

School of Medicine

Research output: Contribution to journal › Article › peer-review

9 Scopus citations

Abstract

Cancer invasion and metastasis are challenging to study in vivo since they occur deep inside the body over extended time periods. Organotypic 3D culture of fresh tumor tissue enables convenient real-time imaging, genetic and microenvironmental manipulation and molecular analysis. Here, we provide detailed protocols to isolate and culture heterogenous organoids from murine and human primary and metastatic site tumors. The time required to isolate organoids can vary based on the tissue and organ type but typically takes <7 h. We describe a suite of assays that model specific aspects of metastasis, including proliferation, survival, invasion, dissemination and colony formation. We also specify comprehensive protocols for downstream applications of organotypic cultures that will allow users to (i) test the role of specific genes in regulating various cellular processes, (ii) distinguish the contributions of several microenvironmental factors and (iii) test the effects of novel therapeutics.

Original language	English (US)
Pages (from-to)	2413-2442
Number of pages	30
Journal	Nature protocols
Volume	15
Issue number	8
DOIs	https://doi.org/10.1038/s41596-020-0335-3
State	Published - Aug 1 2020

ASJC Scopus subject areas

Biochemistry, Genetics and Molecular Biology(all)

Access to Document

10.1038/s41596-020-0335-3

Cite this

@article{23b77401ab934713ac51a210705e88f8,

title = "Organotypic culture assays for murine and human primary and metastatic-site tumors",

abstract = "Cancer invasion and metastasis are challenging to study in vivo since they occur deep inside the body over extended time periods. Organotypic 3D culture of fresh tumor tissue enables convenient real-time imaging, genetic and microenvironmental manipulation and molecular analysis. Here, we provide detailed protocols to isolate and culture heterogenous organoids from murine and human primary and metastatic site tumors. The time required to isolate organoids can vary based on the tissue and organ type but typically takes <7 h. We describe a suite of assays that model specific aspects of metastasis, including proliferation, survival, invasion, dissemination and colony formation. We also specify comprehensive protocols for downstream applications of organotypic cultures that will allow users to (i) test the role of specific genes in regulating various cellular processes, (ii) distinguish the contributions of several microenvironmental factors and (iii) test the effects of novel therapeutics.",

author = "Veena Padmanaban and Grasset, {Eloise M.} and Neumann, {Neil M.} and Fraser, {Andrew K.} and Elodie Henriet and William Matsui and Tran, {Phuoc T.} and Cheung, {Kevin J.} and Dan Georgess and Ewald, {Andrew J.}",

note = "Funding Information: We thank Jin Zhu for assistance with FACS experiments. K.J.C. was supported by the Burroughs Welcome Fund Career Award for Medical Scientists 1013355.01. D.G. was supported by a Postdoctoral Fellowship Grant from the Susan G. Komen Foundation (PDF15332336). A.J.E. received support for this project through grants from The Breast Cancer Research Foundation (BCRF-18-048), the Metastatic Breast Cancer Network, Twisted Pink, Hope Scarves, Theresa{\textquoteright}s Research Foundation and the National Institutes of Health/ National Cancer Institute (U01CA217846, U01CA212007, U54CA2101732 and 3P30CA006973). Publisher Copyright: {\textcopyright} 2020, The Author(s), under exclusive licence to Springer Nature Limited.",

year = "2020",

month = aug,

day = "1",

doi = "10.1038/s41596-020-0335-3",

language = "English (US)",

volume = "15",

pages = "2413--2442",

journal = "Nature Protocols",

issn = "1754-2189",

publisher = "Nature Publishing Group",

number = "8",

}

TY - JOUR

T1 - Organotypic culture assays for murine and human primary and metastatic-site tumors

AU - Padmanaban, Veena

AU - Grasset, Eloise M.

AU - Neumann, Neil M.

AU - Fraser, Andrew K.

AU - Henriet, Elodie

AU - Matsui, William

AU - Tran, Phuoc T.

AU - Cheung, Kevin J.

AU - Georgess, Dan

AU - Ewald, Andrew J.

N1 - Funding Information: We thank Jin Zhu for assistance with FACS experiments. K.J.C. was supported by the Burroughs Welcome Fund Career Award for Medical Scientists 1013355.01. D.G. was supported by a Postdoctoral Fellowship Grant from the Susan G. Komen Foundation (PDF15332336). A.J.E. received support for this project through grants from The Breast Cancer Research Foundation (BCRF-18-048), the Metastatic Breast Cancer Network, Twisted Pink, Hope Scarves, Theresa’s Research Foundation and the National Institutes of Health/ National Cancer Institute (U01CA217846, U01CA212007, U54CA2101732 and 3P30CA006973). Publisher Copyright: © 2020, The Author(s), under exclusive licence to Springer Nature Limited.

PY - 2020/8/1

Y1 - 2020/8/1

N2 - Cancer invasion and metastasis are challenging to study in vivo since they occur deep inside the body over extended time periods. Organotypic 3D culture of fresh tumor tissue enables convenient real-time imaging, genetic and microenvironmental manipulation and molecular analysis. Here, we provide detailed protocols to isolate and culture heterogenous organoids from murine and human primary and metastatic site tumors. The time required to isolate organoids can vary based on the tissue and organ type but typically takes <7 h. We describe a suite of assays that model specific aspects of metastasis, including proliferation, survival, invasion, dissemination and colony formation. We also specify comprehensive protocols for downstream applications of organotypic cultures that will allow users to (i) test the role of specific genes in regulating various cellular processes, (ii) distinguish the contributions of several microenvironmental factors and (iii) test the effects of novel therapeutics.

AB - Cancer invasion and metastasis are challenging to study in vivo since they occur deep inside the body over extended time periods. Organotypic 3D culture of fresh tumor tissue enables convenient real-time imaging, genetic and microenvironmental manipulation and molecular analysis. Here, we provide detailed protocols to isolate and culture heterogenous organoids from murine and human primary and metastatic site tumors. The time required to isolate organoids can vary based on the tissue and organ type but typically takes <7 h. We describe a suite of assays that model specific aspects of metastasis, including proliferation, survival, invasion, dissemination and colony formation. We also specify comprehensive protocols for downstream applications of organotypic cultures that will allow users to (i) test the role of specific genes in regulating various cellular processes, (ii) distinguish the contributions of several microenvironmental factors and (iii) test the effects of novel therapeutics.

UR - http://www.scopus.com/inward/record.url?scp=85088277356&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85088277356&partnerID=8YFLogxK

U2 - 10.1038/s41596-020-0335-3

DO - 10.1038/s41596-020-0335-3

M3 - Article

C2 - 32690957

AN - SCOPUS:85088277356

SN - 1754-2189

VL - 15

SP - 2413

EP - 2442

JO - Nature Protocols

JF - Nature Protocols

IS - 8

ER -

Organotypic culture assays for murine and human primary and metastatic-site tumors

Abstract

ASJC Scopus subject areas

Access to Document

Other files and links

Fingerprint

Cite this