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Combining Cell Type-Restricted Adenoviral Targeting with Immunostaining and Flow Cytometry to Identify Cells-of-Origin of Lung Cancer

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Inflammation and Cancer

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1725))

Abstract

Lung cancers display considerable intertumoral heterogeneity, leading to the classification of distinct tumor subtypes. Our understanding of the genetic aberrations that underlie tumor subtypes has been greatly enhanced by recent genomic sequencing studies and state-of-the-art gene targeting technologies, highlighting evidence that distinct lung cancer subtypes may be derived from different “cells-of-origin”. Here, we describe the intra-tracheal delivery of cell type-restricted Ad5-Cre viruses into the lungs of adult mice, combined with immunohistochemical and flow cytometry strategies for the detection of lung cancer-initiating cells in vivo.

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References

  1. Sutherland KD, Berns A (2010) Cell of origin of lung cancer. Mol Oncol 4:397–403

    Article  PubMed  PubMed Central  Google Scholar 

  2. Weeden CE, Chen Y, Ma SB et al (2017) Lung basal stem cells rapidly repair DNA damage using the error-prone nonhomologous end-joining pathway. PLoS Biol 15:e2000731

    Article  PubMed  PubMed Central  Google Scholar 

  3. Sutherland KD, Visvader JE (2015) Cellular mechanisms underlying intertumoral heterogeneity. Trends Cancer 1:15–23

    Article  PubMed  Google Scholar 

  4. Rock JR, Onaitis MW, Rawlins EL et al (2009) Basal cells as stem cells of the mouse trachea and human airway epithelium. Proc Natl Acad Sci U S A 106:12771–12775

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Rawlins EL, Okubo T, Xue Y et al (2009) The role of Scgb1a1+ Clara cells in the long-term maintenance and repair of lung airway, but not alveolar, epithelium. Cell Stem Cell 4:525–534

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Song H, Yao E, Lin C et al (2012) Functional characterization of pulmonary neuroendocrine cells in lung development, injury, and tumorigenesis. Proc Natl Acad Sci U S A 109:17531–17536

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Barkauskas CE, Cronce MJ, Rackley CR et al (2013) Type 2 alveolar cells are stem cells in adult lung. J Clin Invest 123:3025–3036

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. McQualter JL, Yuen K, Williams B et al (2010) Evidence of an epithelial stem/progenitor cell hierarchy in the adult mouse lung. Proc Natl Acad Sci U S A 107:1414–1419

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Ferone G, Song JY, Sutherland KD et al (2016) SOX2 is the determining oncogenic switch in promoting lung squamous cell carcinoma from different cells of origin. Cancer Cell 30:519–532

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Sutherland KD, Proost N, Brouns I et al (2011) Cell of origin of small cell lung cancer: inactivation of Trp53 and Rb1 in distinct cell types of adult mouse lung. Cancer Cell 19:754–764

    Article  CAS  PubMed  Google Scholar 

  11. Sutherland KD, Song JY, Kwon MC et al (2014) Multiple cells-of-origin of mutant K-Ras-induced mouse lung adenocarcinoma. Proc Natl Acad Sci U S A 111:4952–4957

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Muzumdar MD, Tasic B, Miyamichi K et al (2007) A global double-fluorescent Cre reporter mouse. Genesis 45:593–605

    Article  CAS  PubMed  Google Scholar 

  13. Jackson EL, Olive KP, Tuveson DA et al (2005) The differential effects of mutant p53 alleles on advanced murine lung cancer. Cancer Res 65:10280–10288

    Article  CAS  PubMed  Google Scholar 

  14. Mainardi S, Mijimolle N, Francoz S et al (2014) Identification of cancer initiating cells in K-Ras driven lung adenocarcinoma. Proc Natl Acad Sci U S A 111:255–260

    Article  CAS  PubMed  Google Scholar 

  15. Xu X, Rock JR, Lu Y et al (2012) Evidence for type II cells as cells of origin of K-Ras-induced distal lung adenocarcinoma. Proc Natl Acad Sci U S A 109:4910–4915

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Soriano P (1999) Generalized lacZ expression with the ROSA26 Cre reporter strain. Nat Genet 21:70–71

    Article  CAS  PubMed  Google Scholar 

  17. Jackson EL, Willis N, Mercer K et al (2001) Analysis of lung tumor initiation and progression using conditional expression of oncogenic K-ras. Genes Dev 15:3243–3248

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Suvarna KS, Layton C, Bancroft JD (eds) (2013) Bancroft’s theory and practice of histological techniques, 7th edn. London, Churchill Livingstone

    Google Scholar 

Download references

Acknowledgement

We thank Ellen Tsui, Cary Tsui and Ji-Ying Song for their expert histological assistance. The intra-tracheal procedure is performed by highly-trained staff following protocols approved by the WEHI Animal Ethics Committee. K.D.S. is supported by the Peter and Julie Alston Centenary Fellowship, M.L.A.-L. is the recipient of The Viertel Charitable Foundation Senior Medical Research Fellowship, and S.A.B is supported by the Victorian Cancer Agency Early-Career Fellowship. This work was made possible through Victorian State Government Operational Infrastructure Support and Australian Government NHMRC IRIISS.

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Authors and Affiliations

  1. ACRF Stem Cells and Cancer Division, The Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia

    Sarah A. Best, Ariena Kersbergen, Marie-Liesse Asselin-Labat & Kate D. Sutherland

  2. Department of Medical Biology, The University of Melbourne, Parkville, VIC, Australia

    Sarah A. Best, Marie-Liesse Asselin-Labat & Kate D. Sutherland

Authors
  1. Sarah A. Best
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  2. Ariena Kersbergen
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  3. Marie-Liesse Asselin-Labat
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  4. Kate D. Sutherland
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Corresponding author

Correspondence to Kate D. Sutherland .

Editor information

Editors and Affiliations

  1. Centre for Innate Immunity and Infectious Diseases, Hudson Institute of Medical Research, Clayton, Victoria, Australia

    Brendan J. Jenkins

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Best, S.A., Kersbergen, A., Asselin-Labat, ML., Sutherland, K.D. (2018). Combining Cell Type-Restricted Adenoviral Targeting with Immunostaining and Flow Cytometry to Identify Cells-of-Origin of Lung Cancer. In: Jenkins, B. (eds) Inflammation and Cancer. Methods in Molecular Biology, vol 1725. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7568-6_2

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  • DOI: https://doi.org/10.1007/978-1-4939-7568-6_2

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7567-9

  • Online ISBN: 978-1-4939-7568-6

  • eBook Packages: Springer Protocols

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