Senescence Induction by Combined Ionizing Radiation and DNA Damage Response Inhibitors in Head and Neck Squamous Cell Carcinoma Cells
Abstract
:1. Introduction
2. Materials and Methods
2.1. Cell Culture
2.2. Homologous Recombination by Immunofluorescence Microscopy
2.3. Combined Assay for Senescence, Cell Cycle, Apoptosis, and Necrosis by Flow Cytometry
2.4. Gating Strategy for Flow Cytometry
2.5. Senescence by β-Galactosidase and p21/α Tubulin Staining
2.6. Clonogenic Survival by Colony Formation Assay
2.7. Statistics
3. Results
3.1. Ability to Perform Homologous Recombination
3.2. Induction of Senescence by IR, DDRi, and Combined Treatment
3.3. Apoptotic and Necrotic Cell Death and Senescence
3.4. G2 Arrest
3.5. Background Levels of Senescence and Necrosis
3.6. Senescence Validation by β-Galactosidase
3.7. Senescence Validation by p21 and α Tubulin
4. Discussion
4.1. ATR Inhibitor VE-822 and ATM Inhibitor AZD0156 were More Effectively Inducing Senescence and Necrosis for Combinatorial Treatment with IR
4.2. DNA-PK/mTor Inhibitor CC-115 did not Interact with IR for Senescence and Necrosis Induction
4.3. Induction of G2 Arrest in the Six Cell Lines
4.4. Radiosensitization in the Context of Abrogation of G2 Arrest
4.5. HR Status Did Not Have Major Influence on Senescence Induction upon IR and DDRi Treatment
4.6. Individual Reactions and Sensitivity towards DDRi Treatment in HPV+ and HPV-Cell Lines
4.7. Molecular Heterogeneity as the Cause for Varying Efficacy of DDRi in HNSCC Cell Lines
4.8. Decreased Senescence Levels upon DDRi Treatment can be Explained by Involvement of DDRi in Senescence Induction
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Dobler, C.; Jost, T.; Hecht, M.; Fietkau, R.; Distel, L. Senescence Induction by Combined Ionizing Radiation and DNA Damage Response Inhibitors in Head and Neck Squamous Cell Carcinoma Cells. Cells 2020, 9, 2012. https://doi.org/10.3390/cells9092012
Dobler C, Jost T, Hecht M, Fietkau R, Distel L. Senescence Induction by Combined Ionizing Radiation and DNA Damage Response Inhibitors in Head and Neck Squamous Cell Carcinoma Cells. Cells. 2020; 9(9):2012. https://doi.org/10.3390/cells9092012
Chicago/Turabian StyleDobler, Clara, Tina Jost, Markus Hecht, Rainer Fietkau, and Luitpold Distel. 2020. "Senescence Induction by Combined Ionizing Radiation and DNA Damage Response Inhibitors in Head and Neck Squamous Cell Carcinoma Cells" Cells 9, no. 9: 2012. https://doi.org/10.3390/cells9092012