Circulating tumor cell copy-number heterogeneity in ALK-rearranged non-small-cell lung cancer resistant to ALK inhibitors

Abstract Gatekeeper mutations are identified in only 50% of the cases at resistance to Anaplastic Lymphoma Kinase (ALK)-tyrosine kinase inhibitors (TKIs). Circulating tumor cells (CTCs) are relevant tools to identify additional resistance mechanisms and can be sequenced at the single-cell level. Her...

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Autores principales:	Marianne Oulhen, Patrycja Pawlikowska, Tala Tayoun, Marianna Garonzi, Genny Buson, Claudio Forcato, Nicolò Manaresi, Agathe Aberlenc, Laura Mezquita, Yann Lecluse, Pernelle Lavaud, Charles Naltet, David Planchard, Benjamin Besse, Françoise Farace
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Neoplasms. Tumors. Oncology. Including cancer and carcinogens RC254-282
Acceso en línea:	https://doaj.org/article/17945349780b47ecb1ff35beb8ce4404
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spelling	oai:doaj.org-article:17945349780b47ecb1ff35beb8ce44042021-12-02T18:30:50ZCirculating tumor cell copy-number heterogeneity in ALK-rearranged non-small-cell lung cancer resistant to ALK inhibitors10.1038/s41698-021-00203-12397-768Xhttps://doaj.org/article/17945349780b47ecb1ff35beb8ce44042021-07-01T00:00:00Zhttps://doi.org/10.1038/s41698-021-00203-1https://doaj.org/toc/2397-768XAbstract Gatekeeper mutations are identified in only 50% of the cases at resistance to Anaplastic Lymphoma Kinase (ALK)-tyrosine kinase inhibitors (TKIs). Circulating tumor cells (CTCs) are relevant tools to identify additional resistance mechanisms and can be sequenced at the single-cell level. Here, we provide in-depth investigation of copy number alteration (CNA) heterogeneity in phenotypically characterized CTCs at resistance to ALK-TKIs in ALK-positive non-small cell lung cancer. Single CTC isolation and phenotyping were performed by DEPArray or fluorescence-activated cell sorting following enrichment and immunofluorescence staining (ALK/cytokeratins/CD45/Hoechst). CNA heterogeneity was evaluated in six ALK-rearranged patients harboring ≥ 10 CTCs/20 mL blood at resistance to 1st and 3rd ALK-TKIs and one presented gatekeeper mutations. Out of 82 CTCs isolated by FACS, 30 (37%) were ALK+/cytokeratins-, 46 (56%) ALK-/cytokeratins+ and 4 (5%) ALK+/cytokeratins+. Sequencing of 43 CTCs showed highly altered CNA profiles and high levels of chromosomal instability (CIN). Half of CTCs displayed a ploidy >2n and 32% experienced whole-genome doubling. Hierarchical clustering showed significant intra-patient and wide inter-patient CTC diversity. Classification of 121 oncogenic drivers revealed the predominant activation of cell cycle and DNA repair pathways and of RTK/RAS and PI3K to a lower frequency. CTCs showed wide CNA heterogeneity and elevated CIN at resistance to ALK-TKIs. The emergence of epithelial ALK-negative CTCs may drive resistance through activation of bypass signaling pathways, while ALK-rearranged CTCs showed epithelial-to-mesenchymal transition characteristics potentially contributing to ALK-TKI resistance. Comprehensive analysis of CTCs could be of great help to clinicians for precision medicine and resistance to ALK-targeted therapies.Marianne OulhenPatrycja PawlikowskaTala TayounMarianna GaronziGenny BusonClaudio ForcatoNicolò ManaresiAgathe AberlencLaura MezquitaYann LeclusePernelle LavaudCharles NaltetDavid PlanchardBenjamin BesseFrançoise FaraceNature PortfolioarticleNeoplasms. Tumors. Oncology. Including cancer and carcinogensRC254-282ENnpj Precision Oncology, Vol 5, Iss 1, Pp 1-11 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Neoplasms. Tumors. Oncology. Including cancer and carcinogens RC254-282
spellingShingle	Neoplasms. Tumors. Oncology. Including cancer and carcinogens RC254-282 Marianne Oulhen Patrycja Pawlikowska Tala Tayoun Marianna Garonzi Genny Buson Claudio Forcato Nicolò Manaresi Agathe Aberlenc Laura Mezquita Yann Lecluse Pernelle Lavaud Charles Naltet David Planchard Benjamin Besse Françoise Farace Circulating tumor cell copy-number heterogeneity in ALK-rearranged non-small-cell lung cancer resistant to ALK inhibitors
description	Abstract Gatekeeper mutations are identified in only 50% of the cases at resistance to Anaplastic Lymphoma Kinase (ALK)-tyrosine kinase inhibitors (TKIs). Circulating tumor cells (CTCs) are relevant tools to identify additional resistance mechanisms and can be sequenced at the single-cell level. Here, we provide in-depth investigation of copy number alteration (CNA) heterogeneity in phenotypically characterized CTCs at resistance to ALK-TKIs in ALK-positive non-small cell lung cancer. Single CTC isolation and phenotyping were performed by DEPArray or fluorescence-activated cell sorting following enrichment and immunofluorescence staining (ALK/cytokeratins/CD45/Hoechst). CNA heterogeneity was evaluated in six ALK-rearranged patients harboring ≥ 10 CTCs/20 mL blood at resistance to 1st and 3rd ALK-TKIs and one presented gatekeeper mutations. Out of 82 CTCs isolated by FACS, 30 (37%) were ALK+/cytokeratins-, 46 (56%) ALK-/cytokeratins+ and 4 (5%) ALK+/cytokeratins+. Sequencing of 43 CTCs showed highly altered CNA profiles and high levels of chromosomal instability (CIN). Half of CTCs displayed a ploidy >2n and 32% experienced whole-genome doubling. Hierarchical clustering showed significant intra-patient and wide inter-patient CTC diversity. Classification of 121 oncogenic drivers revealed the predominant activation of cell cycle and DNA repair pathways and of RTK/RAS and PI3K to a lower frequency. CTCs showed wide CNA heterogeneity and elevated CIN at resistance to ALK-TKIs. The emergence of epithelial ALK-negative CTCs may drive resistance through activation of bypass signaling pathways, while ALK-rearranged CTCs showed epithelial-to-mesenchymal transition characteristics potentially contributing to ALK-TKI resistance. Comprehensive analysis of CTCs could be of great help to clinicians for precision medicine and resistance to ALK-targeted therapies.
format	article
author	Marianne Oulhen Patrycja Pawlikowska Tala Tayoun Marianna Garonzi Genny Buson Claudio Forcato Nicolò Manaresi Agathe Aberlenc Laura Mezquita Yann Lecluse Pernelle Lavaud Charles Naltet David Planchard Benjamin Besse Françoise Farace
author_facet	Marianne Oulhen Patrycja Pawlikowska Tala Tayoun Marianna Garonzi Genny Buson Claudio Forcato Nicolò Manaresi Agathe Aberlenc Laura Mezquita Yann Lecluse Pernelle Lavaud Charles Naltet David Planchard Benjamin Besse Françoise Farace
author_sort	Marianne Oulhen
title	Circulating tumor cell copy-number heterogeneity in ALK-rearranged non-small-cell lung cancer resistant to ALK inhibitors
title_short	Circulating tumor cell copy-number heterogeneity in ALK-rearranged non-small-cell lung cancer resistant to ALK inhibitors
title_full	Circulating tumor cell copy-number heterogeneity in ALK-rearranged non-small-cell lung cancer resistant to ALK inhibitors
title_fullStr	Circulating tumor cell copy-number heterogeneity in ALK-rearranged non-small-cell lung cancer resistant to ALK inhibitors
title_full_unstemmed	Circulating tumor cell copy-number heterogeneity in ALK-rearranged non-small-cell lung cancer resistant to ALK inhibitors
title_sort	circulating tumor cell copy-number heterogeneity in alk-rearranged non-small-cell lung cancer resistant to alk inhibitors
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/17945349780b47ecb1ff35beb8ce4404
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Circulating tumor cell copy-number heterogeneity in ALK-rearranged non-small-cell lung cancer resistant to ALK inhibitors

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