A surgical orthotopic organoid transplantation approach in mice to visualize and study colorectal cancer progression

Abstract

Most currently available colorectal cancer (CRC) mouse models are not suitable for studying progression toward the metastatic stage. Recently, establishment of tumor organoid lines, either from murine CRC models or patients, and the possibility of engineering them with genome-editing technologies, have provided a large collection of tumor material faithfully recapitulating phenotypic and genetic heterogeneity of native tumors. To study tumor progression in the natural in vivo environment, we developed an orthotopic approach based on transplantation of CRC organoids into the cecal epithelium. The 20-min procedure is described in detail here and enables growth of transplanted organoids into a single tumor mass within the intestinal tract. Due to long latency, tumor cells are capable of spreading through the blood circulation and forming metastases at distant sites. This method is designed to generate tumors suitable for studying CRC progression, thereby providing the opportunity to visualize tumor cell dynamics in vivo in real time by intravital microscopy.

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Figure 1: Isolation and preparation of organoids for orthotopic transplantation.
Figure 2: Orthotopic transplantation of intestinal tumor organoids.
Figure 3: Analysis of the phenotypic outcome of orthotopically transplanted intestinal tumor organoids.
Figure 4: Intravital imaging of inter- and intra-tumor heterogeneity.

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Acknowledgements

We thank A. de Graaff and the Hubrecht Imaging Centre for imaging support. We thank O. Sansom and E. Hong Tan (Beatson Institute, Glasgow, UK) for providing the murine tumor organoid line. This work was financially supported by a Dutch Cancer Society Fellowship (BUIT-2013-5847 to S.J.E.S.), by the Dutch Cancer Society (KWF)/Alpe d′HuZes Bas Mulder Award (KWF/Alpe d′HuZes 10218, to J.D.), by European Research Council Grant CANCER-RECURRENCE 648804 (to J.v.R.), by the CancerGenomics.nl (Netherlands Organisation for Scientific Research) program (to J.v.R.), by the Doctor Josef Steiner Foundation (to J.v.R) and by the European Union′s Horizon 2020 research and innovation program under Marie Sklodowska-Curie grant agreement no. 642866 (to J.v.R).

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A.F. developed the orthotopic transplantation technique; A.F. and J.D. performed the experiments; H.B., E.B. and S.J.E.S. helped with data analysis and preparation of the figures; S.J.E.S edited the video; K.C.O. and H.J.S. provided the patient-derived CRC organoids; J.D. and J.v.R. supervised the study; A.F., J.D. and J.v.R. wrote the manuscript.

Corresponding authors

Correspondence to Jacco van Rheenen or Jarno Drost.

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The authors declare no competing financial interests.

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Fumagalli, A., Suijkerbuijk, S., Begthel, H. et al. A surgical orthotopic organoid transplantation approach in mice to visualize and study colorectal cancer progression. Nat Protoc 13, 235–247 (2018). https://doi.org/10.1038/nprot.2017.137

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