Metabolic analysis of HPV positive and negative HNSCC cell lines after irradiation with carbon-12 (12C) particles
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Cancer cells are highly heterogeneous regarding their metabolic activity and phenotype. Proliferation of these cells is a rapid and dynamic process that needs significant energy. Generally, compared to normal cells, cancer cells to a large extent use the so-called Warburg effect, or aerobe glycolysis, for energy production. Metabolic parameters of six (three HPV-ve and three HPV+ve) head and neck squamous cell carcinoma (HNSCC) cancer cell lines with diverse anatomic origins and with different genetic backgrounds (UM-SCC-3, UM-SCC-29, UT-SCC-26A, UM-SCC-47, UM-SCC-104, UPCI-SCC-154) were analyzed under standard culture conditions and after radiation with 12C particles.
Metabolic studies were performed with the Agilent Seahorse XF96 analyzer using the XF Real-Time ATP rate assay. With this assay, the oxygen consumption rate (OCR) in pmol/min and extracellular acidification rate (ECAR) in mpH/min, were measured and converted to mitochondrial ATP production rate (oxidative phosphorylation) and glycolytic ATP production rate (glycolysis) respectively.
Cells to be irradiated were seeded together with control cells in the same Seahorse tissue culture microplate, so that metabolic analysis could be performed for irradiated as well as non-irradiated control cells at the same time.
Different doses (1, 2, 4, 8 and 12 Gy) of 12C particle irradiation were used and the energy metabolism of the cancer cell lines was evaluated at different time points (2, 6, 10 and 24 hours) after irradiation.
A decrease of glycolytic activity and mitochondrial respiration in the tested HNSCC cell lines was observed after exposure to irradiation, which was reflected by a reduction of the glycolytic as well as mitochondrial ATP production rate in irradiated cells compared with the control cells in most of our assays. This effect was more pronounced after application of higher irradiation doses.
The reduction in glycolysis was more obvious and statistically significant in HPV ve HNSCC cell lines compared with HPV+ve HNSCC cell lines. Moreover, there was some significant increase of glycolytic ATP production rate in irradiated HPV+ve HNSCC cell lines cells compared to control cells with low irradiation dose. At the same time, a significant reduction of mitochondrial ATP production rate in irradiated cells compared with the control cells was observed in most of the assays for HPV+ve HNSCC cell lines. On the other hand, mitochondrial ATP production rate for HPV-ve HNSCC cell lines tend to increase in irradiated cells compared to control cells with low irradiation dose at the early time points. With higher irradiation doses it tends to decrease significantly at the late time points.
Based on previous results when comparing HPV-ve and HPV+ve HNSCC cell lines, ATP production in HPV+ve HNSCC cell lines was significantly more derived from oxidative phosphorylation and significantly less from glycolysis compared with HPV-ve HNSCC cell lines. which means that 12C particle irradiation could be more selective in affecting the dominant energy production mechanism in cancer cells. however, there is limited understanding of the exact mechanism.
There are only few reports regarding the effect of radiation therapy, especially particle therapy, on cancer cell metabolism, wherefore there is an increasing demand for such studies. The present study could contribute to a better understanding of metabolic changes in HPV-ve and HPV+ve HNSCC after 12C particle irradiation thereby helping to exploit potential metabolic weak points of HNSCC cells after 12C irradiation, which could be used for therapeutic targeting. Future studies will test inhibitors of the glycolytic pathway for their potential use as radiosensitizers that could be implemented in the therapy scheme of head & neck cancer patients.
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Issued: 2025-11-18
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Has part: Agilent Seahorse XF Real-Time ATP Rate Assay Kit User Guide Notices (2024) Agilent Seahorse XF Real-Time ATP Rate Assay Kit For use with Seahorse XF/XFe96 and XFe24 Extracellular Flux Analyzers with the corresponding V3 or V7 PS cell culture microplates. Not compatible with other cell culture plates. Agilent Seahorse XF Real-Time ATP Rate Assay Kit User Guide Notices.Has part: Fleming, J.C. et al. (2019) ‘HPV, tumor metabolism and novel target identification in head and neck squamous cell carcinoma’, British Journal of Cancer, 120(3), pp. 356–367. Available at: https://doi.org/10.1038/s41416-018-0364-7.Has part: Ikawa, H. et al. (2019) ‘Multicenter study of carbon-ion radiation therapy for nonsquamous cell carcinomas of the oral cavity’, Cancer Medicine, 8(12), pp. 5482–5491. Available at: https://doi.org/10.1002/cam4.2408.
Faculty
FB20:Medizin
Language
en
Keywords
HNSCCPARTICLE THERAPYMito ATPGlyco ATP
DFG-subjects
2.22-28 - Zahnheilkunde; Mund-, Kiefer- und Gesichtschirurgie2.22-29 - Hals-Nasen-Ohrenheilkunde, Phoniatrie und Audiologie2.22-30 - Radiologie
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Al Rabadi, Hytham: Metabolic analysis of HPV positive and negative HNSCC cell lines after irradiation with carbon-12 (12C) particles. : 2025-11-18.
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Except where otherwised noted, this item's license is described as Attribution 3.0 Germany