title: Morphometric analysis of immunoselection against hyperploid cancer cells.
abstract: An at least transient increase of ploidy, usually by whole genome duplication, is a frequent event in oncogenesis, explaining the cytogenetic features of at least 40% of solid cancers. Here, we show that fibrosarcomas induced by the carcinogen methylcholanthrene (MCA) are distinct with respect to their ploidy status when they arise in immunocompetent wild type versus severely immunodeficient Rag2-/-γc-/- mice. MCA-induced fibrosarcomas are particularly hyperploid if they develop in an immunodeficient setting, correlating with higher DNA content, increased nuclear surface, as well as hyperphosphorylation of eukaryotic initiation factor 2α (eIF2α), a biomarker indicating endoplasmic reticulum (ER) stress. Upon transfer of such cells into wild type mice, such hyperploid, ER-stressed cells (that originated in Rag2-/-γc-/- mice) fail to proliferate and actually induce a protective anticancer immune response. In contrast, such cells do form tumors in Rag2-/-γc-/- recipients (which lack T, B and NK cells) as well as in Rag2-/- recipients (which only lack T and B lymphocytes) and conserve their hyperploidy as well as eIF2α hyperphosphorylation. To measure these parameters, we developed a morphometric analysis tool that is applicable to immunohistochemistry of formaldehyde-fixed, paraffin-embedded tissues. This software automatically identifies and quantifies the surface of nuclei and determines the intensity of eIF2α phosphorylation within a perinuclear region of interest. Comparative analyses performed on cultured cells and tissue sections validated the accuracy of this method, which can be used to investigate ploidy and ER stress in cancers in situ.
authors.affiliation: Equipe 11 Labellisée Ligue Contre le Cancer, Centre de Recherche des Cordeliers, INSERM U 1138, 75006 Paris, France.|Université Paris Sud, Faculté de Médecine, 94270 Kremlin Bicêtre, France.|Université Paris Descartes, Sorbonne Paris Cité, 75006 Paris, France.|Université Pierre et Marie Curie, 75006 Paris, France.
authors.forename: Norma
authors.initials: N
authors.lastname: Bloy
authors.affiliation: Equipe 11 Labellisée Ligue Contre le Cancer, Centre de Recherche des Cordeliers, INSERM U 1138, 75006 Paris, France.|Université Paris Descartes, Sorbonne Paris Cité, 75006 Paris, France.|Université Pierre et Marie Curie, 75006 Paris, France.|Metabolomics and Cell Biology Platforms, Gustave Roussy Comprehensive Cancer Institute, 94805 Villejuif, France.
authors.forename: Allan
authors.initials: A
authors.lastname: Sauvat
authors.affiliation: Equipe 11 Labellisée Ligue Contre le Cancer, Centre de Recherche des Cordeliers, INSERM U 1138, 75006 Paris, France.|Université Paris Descartes, Sorbonne Paris Cité, 75006 Paris, France.|Université Pierre et Marie Curie, 75006 Paris, France.
authors.forename: Kariman
authors.initials: K
authors.lastname: Chaba
authors.affiliation: Equipe 11 Labellisée Ligue Contre le Cancer, Centre de Recherche des Cordeliers, INSERM U 1138, 75006 Paris, France.|Université Paris Descartes, Sorbonne Paris Cité, 75006 Paris, France.|Université Pierre et Marie Curie, 75006 Paris, France.
authors.forename: Aitziber
authors.initials: A
authors.lastname: Buqué
authors.affiliation: Equipe 11 Labellisée Ligue Contre le Cancer, Centre de Recherche des Cordeliers, INSERM U 1138, 75006 Paris, France.|Université Paris Sud, Faculté de Médecine, 94270 Kremlin Bicêtre, France.|Université Paris Descartes, Sorbonne Paris Cité, 75006 Paris, France.|Université Pierre et Marie Curie, 75006 Paris, France.
authors.forename: Juliette
authors.initials: J
authors.lastname: Humeau
authors.affiliation: Equipe 11 Labellisée Ligue Contre le Cancer, Centre de Recherche des Cordeliers, INSERM U 1138, 75006 Paris, France.|Université Paris Descartes, Sorbonne Paris Cité, 75006 Paris, France.|Université Pierre et Marie Curie, 75006 Paris, France.
authors.forename: José Manuel
authors.initials: JM
authors.lastname: Bravo-San Pedro
authors.affiliation: Department of Pathology, University of California San Diego, CA 92093, USA.
authors.forename: Jack
authors.initials: J
authors.lastname: Bui
authors.affiliation: Equipe 11 Labellisée Ligue Contre le Cancer, Centre de Recherche des Cordeliers, INSERM U 1138, 75006 Paris, France.|Université Paris Descartes, Sorbonne Paris Cité, 75006 Paris, France.|Université Pierre et Marie Curie, 75006 Paris, France.|Metabolomics and Cell Biology Platforms, Gustave Roussy Comprehensive Cancer Institute, 94805 Villejuif, France.
authors.forename: Oliver
authors.initials: O
authors.lastname: Kepp
authors.affiliation: Equipe 11 Labellisée Ligue Contre le Cancer, Centre de Recherche des Cordeliers, INSERM U 1138, 75006 Paris, France.|Université Pierre et Marie Curie, 75006 Paris, France.|Metabolomics and Cell Biology Platforms, Gustave Roussy Comprehensive Cancer Institute, 94805 Villejuif, France.|Pôle de Biologie, Hôpital Européen Georges Pompidou, AP-HP, 75015 Paris, France.|Karolinska Institute, Department of Women's and Children's Health, Karolinska University Hospital, 17176 Stockholm, Sweden.
authors.forename: Guido
authors.initials: G
authors.lastname: Kroemer
authors.affiliation: Equipe 11 Labellisée Ligue Contre le Cancer, Centre de Recherche des Cordeliers, INSERM U 1138, 75006 Paris, France.|Université Paris Descartes, Sorbonne Paris Cité, 75006 Paris, France.|Université Pierre et Marie Curie, 75006 Paris, France.
authors.forename: Laura
authors.initials: L
authors.lastname: Senovilla
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000235:genetics* / Q000378:metabolism; D004171:Diploidy; D059865:Endoplasmic Reticulum Stress / Q000235:genetics; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism; D005434:Flow Cytometry; D015151:Immunoblotting; D007150:Immunohistochemistry; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008856:Microscopy, Fluorescence; D009374:Neoplasms, Experimental / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D010766:Phosphorylation; D011123:Polyploidy
chemical_list: D015852:Eukaryotic Initiation Factor-2
keywords: ER stress; immunoselection; morphometric analysis; ploidy