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High-content, cell-by-cell assessment of HER2 overexpression and amplification: a tool for intratumoral heterogeneity detection in breast cancer

Laboratory Investigationvolume 99pages722732 (2019) | Download Citation

Abstract

Immunohistochemistry and fluorescence in situ hybridization are the two standard methods for human epidermal growth factor receptor 2 (HER2) assessment. However, they have severe limitations to assess quantitatively intratumoral heterogeneity (ITH) when multiple subclones of tumor cells co-exist. We develop here a high-content, quantitative analysis of breast cancer tissues based on microfluidic experimentation and image processing, to characterize both HER2 protein overexpression and HER2 gene amplification at the cellular level. The technique consists of performing sequential steps on the same tissue slide: an immunofluorescence (IF) assay using a microfluidic protocol, an elution step for removing the IF staining agents, a standard FISH staining protocol, followed by automated quantitative cell-by-cell image processing. Moreover, ITH is accurately detected in both cluster and mosaic form using an analysis of spatial association and a mathematical model that allows discriminating true heterogeneity from artifacts due to the use of thin tissue sections. This study paves the way to evaluate ITH with high accuracy and content while requiring standard staining methods.

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Acknowledgements

We thank the European Union Ideas program for supporting this work (Grant number ERC-2012-AdG-320404).

Funding

ERC-2012-AdG-320404. Work and the open access charge were supported by the European Union Ideas program.

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Affiliations

  1. Laboratory of Microsystems 2, École Polytechnique Fédérale de Lausanne EPFL, Lausanne, Switzerland

    • Huu Tuan Nguyen
    • , Daniel Migliozzi
    •  & Martin A. M. Gijs
  2. Institute of Pathology, Centre Hospitalier Universitaire Vaudois and University of Lausanne, Lausanne, Switzerland

    • Bettina Bisig
    •  & Laurence de Leval

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Conflict of interest

MAMG has a patent application related to the microfluidic immunostaining (Swiss Patent Application 00256/12) and is involved in the startup Lunaphore technologies SA developing this technology. The remaining authors declare that they have no conflict of interest.

Corresponding author

Correspondence to Martin A. M. Gijs.

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https://doi.org/10.1038/s41374-018-0172-y