Comparative response of head and neck cancer cells to photons and protons: Optimising radiosensitivity through targeting the DNA damage response


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Jason Parsons1, Maria Rita Fabbrizi1, Rachel Carter1, Catherine Nickson1, Eirini-Terpsi Vitti1, Mark Hill2
1University of Liverpool, 2University of Oxford

Abstract

Background

Radiotherapy is a mainstay for treatment of head and neck squamous cell carcinoma (HNSCC), and where proton beam therapy (PBT) is increasingly being utilised as a precision targeted approach. However, PBT displays increases in linear energy transfer (LET) at and around the Bragg peak leading to complex DNA damage (CDD), containing several lesions in close proximity within DNA, which contributes significantly to enhanced biological effectiveness. Consequently, there are still uncertainties regarding the optimal treatment strategies using PBT and the comparisons to photon radiotherapy in HNSCC.

Method

The 60 MeV cyclotron at the Clatterbridge Cancer Centre was used for proton irradiations at low-LET (58 MeV;1 keV/μm) generated at the Bragg peak, versus high-LET (11 MeV;12 keV/μm) protons at the distal end. Irradiations were also performed using low-LET photons/x-rays (100 kV). HeLa or HNSCC cells from the oropharynx (UMSCC74A, UMSCC6), hypopharynx (FaDu) or oral cavity (A253) were analysed for cell survival (clonogenic assay) or DNA damage repair (comet assays) post-irradiation. HNSCC cells were also cultured as 3D spheroids and growth monitored using microscopy.

Results

High-LET protons induce significant decreases in cell survival post-irradiation versus low-LET protons or photons due to elevated levels of CDD. siRNA screening has subsequently identified specific enzymes that when targeted using siRNA/inhibitors can further exacerbate the cell killing effects under the different irradiation conditions, with a specific focus on those responsive to high-LET protons. For example, inhibition of the DNA strand break sensor PARP-1 or the DNA glycosylase enzyme OGG1 can significantly enhance radiosensitivity of cells to high-LET protons, but not to low-LET protons or photons. Conversely using a more targeted approach, specific inhibitors against the DNA double strand break protein kinases ATM, ATR and DNA-Pkcs were effective in enhancing radiosensitivity of 2D and 3D models of HNSCC to both low-LET protons and photons.

Conclusion

We demonstrate that targeting key enzymes involved in the cellular DNA damage response is an effective strategy to optimise the impact of protons (including of increasing LET) and/or photons on HNSCC cell models.

Impact statement

Optimising radiosensitivity of preclinical HNSCC cell models to photons and protons will inform on future clinical trials and effective treatments for HNSCC patients.