Characterizing the DNA damage response in putative stem cells of resected normal lung and matched NSCLC patient samples

Marti, Thomas; Tièche, Colin; Peng, Ren-Wang; Hall, Sean; Froment, Laurène; Dorn, Patrick; Berezowska, Sabina Anna; Schmid, Ralph (7 May 2017). Characterizing the DNA damage response in putative stem cells of resected normal lung and matched NSCLC patient samples. Annals of oncology, 28(suppl_2). Oxford University Press 10.1093/annonc/mdx090.013

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Background More than 80% of lung tumors are non-small cell lung cancers (NSCLC). Tumor initiation and propagation are mediated by tumor initiating cells (TICs). In NSCLC and glioblastoma, TICs are characterized by high glycine decarboxylase (GLDC) expression. The DNA damage response (DDR) network evolved to maintain genome integrity and its dysregulation is associated with TICs and therapy resistance. Our aim was to establish a protocol to characterize the DDR status in NSCLC TICs. Methods We evaluated a small cohort of eleven consenting patients who underwent elective surgery for lung cancer. Dissection of the tumor and non-adjacent normal lung tissue was performed by a pathologist. Tissue was dissociated to a single cell suspension, which was stained and analyzed by multicolor flow cytometry (FACS). Results Based on EpCAM and GLDC expression, we identified four distinct subpopulations within the lineage negative compartment (live cells/CD31-/CD45-). In normal lung samples, a rare subpopulation was characterized by increased expression of the epithelial marker EpCAM and GLDC. Relative GLDC expression was increased in matched tumor samples. In normal samples, GLDC expression was associated with proliferation as indicated by Ki-67 staining, which was increased in matched tumor samples. Basal activation of the DDR as indicated by H2AX phosphorylation was increased in tumor versus normal samples. Upon DNA damage induction in both normal and cancer samples, H2AX was phosphorylated in the majority of the EpCAM+/GLDC+/- cells, in a smaller fraction of the EpCAM-/GLDC+ cells and absent in EpCAM-/GLDC- cells. Interestingly, the relative increase in basal versus DNA damage-induced H2AX phosphorylation was significantly higher in normal compared to matched cancer subpopulations. Further, variation in H2AX phosphorylation levels was higher in cancer samples. A larger cohort will be required to correlate DDR activation with the mutational status of the NSCLC samples. Conclusions We established a FACS-based protocol allowing us to analyze DDR activation in normal versus tumor tissue and bulk versus cancer stem cells. This protocol can easily be adapted for the analysis of other types of patient tissue samples.

Item Type:

Conference or Workshop Item (Abstract)

Division/Institute:

04 Faculty of Medicine > Department of Gastro-intestinal, Liver and Lung Disorders (DMLL) > Clinic of Thoracic Surgery
04 Faculty of Medicine > Service Sector > Institute of Pathology > Clinical Pathology
04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > Forschungsbereich Mu50 > Forschungsgruppe Thoraxchirurgie
04 Faculty of Medicine > Service Sector > Institute of Pathology

Graduate School:

Graduate School for Cellular and Biomedical Sciences (GCB)

UniBE Contributor:

Marti, Thomas; Tièche, Colin; Peng, Ren-Wang; Hall, Sean; Froment, Laurène; Dorn, Patrick; Berezowska, Sabina Anna and Schmid, Ralph

Subjects:

600 Technology > 610 Medicine & health
500 Science > 570 Life sciences; biology

ISSN:

0923-7534

Publisher:

Oxford University Press

Language:

English

Submitter:

Thomas Michael Marti

Date Deposited:

01 Feb 2018 16:22

Last Modified:

01 Feb 2018 16:22

Publisher DOI:

10.1093/annonc/mdx090.013

BORIS DOI:

10.7892/boris.110191

URI:

https://boris.unibe.ch/id/eprint/110191

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