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Ink4a/Arf−/− and HRAS(G12V) transform mouse mammary cells into triple-negative breast cancer containing tumorigenic CD49f− quiescent cells

Oncogene volume 33, pages 440–448 (2014)Cite this article

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Abstract

Intratumoral heterogeneity within individual breast tumors is a well-known phenomenon that may contribute to drug resistance. This heterogeneity is dependent on several factors, such as types of oncogenic drivers and tumor precursor cells. The purpose of our study was to engineer a mouse mammary tumor model with intratumoral heterogeneity by using defined genetic perturbations. To achieve this, we used mice with knockout (–/–) of Ink4a/Arf, a tumor suppressor locus; these mice are known to be susceptible to non-mammary tumors such as fibrosarcoma. To induce mammary tumors, we retrovirally introduced an oncogene, HRAS(G12V), into Ink4a/Arf−/− mammary cells in vitro, and those cells were inoculated into syngeneic mice mammary fat pads. We observed 100% tumorigenesis. The tumors formed were negative for estrogen receptor, progesterone receptor and HER2. Further, they had pathological features similar to those of human triple-negative breast cancer (TNBC) (for example, pushing borders, central necrosis). The tumors were found to be heterogeneous and included two subpopulations: CD49f− quiescent cells and CD49f+cells. Contrary to our expectation, CD49f− quiescent cells had high tumor-initiating potential and CD49f+cells had relatively low tumor-initiating potential. Gene expression analysis revealed that CD49f− quiescent cells overexpressed epithelial-to-mesenchymal transition-driving genes, reminiscent of tumor-initiating cells and claudin-low breast cancer. Our animal model with intratumoral heterogeneity, derived from defined genetic perturbations, allows us to test novel molecular targeted drugs in a setting that mimics the intratumoral heterogeneity of human TNBC.

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Acknowledgements

We thank N Suzuki, Y Ito and S Hayashi for excellent animal husbandry, I Ishimatsu for expert assistance with histology and K Hashimoto for expert help in the flow cytometry lab. We thank T Suda and MC Hung for invaluable discussions. Thanks are also owed to M Fujiwara for expert help with microarray analysis, T Ishimoto for technical help with confocal microscopy and SC Patterson for editorial assistance. This work was supported by grants from the Japan Society for the Promotion of Science, Japan (to KK and HS). KK was supported by the program ‘Strategic Young Researcher Overseas Visits Program for Accelerating Brain Circulation’ by the Ministry of Education, Culture, Sports, Science and Technology, Japan. This research was also supported in part by the US National Institutes of Health through MD Anderson's Cancer Center Support Grant (5 P30 CA016672-36), grant R01 CA123318-01A1 (to NTU) and by the Nellie B Connally Breast Cancer Research Fund and a donation from Mr and Mrs Sidney J Jansma, Jr.

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Authors and Affiliations

  1. Breast Cancer Translational Research Laboratory, The University of Texas MD Anderson Cancer Center, Houston, TX, USA

    K Kai, C Bartholomeusz, L Pusztai & N T Ueno

  2. Department of Breast Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA

    K Kai, T Iwamoto, C Bartholomeusz, G N Hortobagyi, L Pusztai & N T Ueno

  3. Division of Gene Regulation, Institute for Advanced Medical Research, School of Medicine, Keio University, Tokyo, Japan

    K Kai, T Kobayashi, Y Arima, Y Takamoto, N Ohnishi & H Saya

  4. Division of Pathology, The Cancer Institute of the Japanese Foundation for Cancer Research, Tokyo, Japan

    R Horii & F Akiyama

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Correspondence to H Saya or N T Ueno.

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Kai, K., Iwamoto, T., Kobayashi, T. et al. Ink4a/Arf−/− and HRAS(G12V) transform mouse mammary cells into triple-negative breast cancer containing tumorigenic CD49f− quiescent cells. Oncogene 33, 440–448 (2014). https://doi.org/10.1038/onc.2012.609

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