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Spatial transcriptomics of tumor microenvironment in formalin-fixed paraffin-embedded breast cancer

Lou Romanens, Prasad Chaskar, Jean-Christophe Tille, Stephan Ryser, Nicolas Liaudet, Ketty Hu-Heimgartner, Killian Heimgartner, Gurkan Kaya, Petros Tsantoulis, S. Intidhar Labidi-Galy
doi: https://doi.org/10.1101/2020.01.31.928143
Lou Romanens
Department of Medicine, Faculty of Medicine, University of Geneva, Genève, Switzerland
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Prasad Chaskar
Department of Medicine, Faculty of Medicine, University of Geneva, Genève, SwitzerlandDepartment of Oncology, Hôpitaux Universitaires de Genève, Genève, Switzerland
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Jean-Christophe Tille
Department of Diagnosis, Division of Clinical Pathology, Hôpitaux Universitaires de Genève, Genève, Switzerland
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Stephan Ryser
Department of Medicine, Faculty of Medicine, University of Geneva, Genève, Switzerland
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Nicolas Liaudet
Bioimaging Core Facility, Faculty of Medicine, University of Geneva, Genève, Switzerland
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Ketty Hu-Heimgartner
Department of Medicine, Faculty of Medicine, University of Geneva, Genève, Switzerland
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Killian Heimgartner
Department of Medicine, Faculty of Medicine, University of Geneva, Genève, Switzerland
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Gurkan Kaya
Department of Medicine, Division of Dermatology, Hôpitaux Universitaires de Genève, Genève, Switzerland
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Petros Tsantoulis
Department of Medicine, Faculty of Medicine, University of Geneva, Genève, SwitzerlandDepartment of Oncology, Hôpitaux Universitaires de Genève, Genève, Switzerland
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S. Intidhar Labidi-Galy
Department of Medicine, Faculty of Medicine, University of Geneva, Genève, SwitzerlandDepartment of Oncology, Hôpitaux Universitaires de Genève, Genève, Switzerland
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  • For correspondence: sana.labidi-galy@unige.ch
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Abstract

Tumor samples are conserved in clinical practice in formalin-fixed paraffin-embedded (FFPE) blocks. Formalin fixation chemically alters nucleic acids, rendering transcriptomic analysis challenging. RNA-sequencing is usually performed on tumor bulk, without distinction of cell subtypes or location. Here we describe the development of a robust method for RNA extraction and exome-capture RNA-sequencing of laser-capture microdissected tumor cells (TC) and stromal immune cells (TIL) based on their morphology. We applied this method on 7 tumor samples (surgical or core needle biopsy) of triple-negative breast cancer (TNBC) stored in FFPE blocks over 3-10 years. Unsupervised clustering and principal component analysis showed a clear separation between gene-expression profile of TIL and TC. TIL were enriched in markers of B cells (CD79B, PAX5 and BLNK) and T cells (CD2, CD3D and CD8B) whereas tumor cells expressed epithelial markers (EPCAM, MUC1 and KRT8). Microenvironment cell populations-counter (MCP)-counter deconvolution showed an enrichment in adaptive immune cell signatures in microdissected TIL. Transcripts of immune checkpoints were differentially expressed in TIL and TC. We further validated our results by qRT-PCR and multispectral immunohistochemistry. In conclusion, we showed that combining laser-capture microdissection and RNA-sequencing on archived FFPE blocks is feasible and allows spatial transcriptional characterization of tumor microenvironment.

Footnotes

  • Conflict of interest: All the authors disclose no potential conflict of interest

  • Financial support: ILG is supported by funding from the Swiss National Science Foundation (Ambizione n° PZ00P3_173959), la Ligue Genevoise Contre le Cancer (grant n°1712), la Fondation pour la lutte contre le cancer et pour des recherches médico-biologiques and The Novartis Biomedical Research Foundation (#17C150). PT and PC are supported by funding from La Ligue Genevoise Contre le Cancer.

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted February 02, 2020.
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Spatial transcriptomics of tumor microenvironment in formalin-fixed paraffin-embedded breast cancer
Lou Romanens, Prasad Chaskar, Jean-Christophe Tille, Stephan Ryser, Nicolas Liaudet, Ketty Hu-Heimgartner, Killian Heimgartner, Gurkan Kaya, Petros Tsantoulis, S. Intidhar Labidi-Galy
bioRxiv 2020.01.31.928143; doi: https://doi.org/10.1101/2020.01.31.928143
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Spatial transcriptomics of tumor microenvironment in formalin-fixed paraffin-embedded breast cancer
Lou Romanens, Prasad Chaskar, Jean-Christophe Tille, Stephan Ryser, Nicolas Liaudet, Ketty Hu-Heimgartner, Killian Heimgartner, Gurkan Kaya, Petros Tsantoulis, S. Intidhar Labidi-Galy
bioRxiv 2020.01.31.928143; doi: https://doi.org/10.1101/2020.01.31.928143

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