Induction of in situ DNA double-strand breaks and apoptosis by 200 MeV protons and 10 MV X-rays in human tumour cell lines

Abstract

Purpose: To clarify the properties of clinical high-energy protons by comparing with clinical high-energy X-rays.

Materials and methods: Human tumor cell lines, ONS76 and MOLT4, were irradiated with 200 MeV protons or 10 MV X-rays. In situ DNA double-strand breaks (DDSB) induction was evaluated by immunocytochemical staining of phosphorylated histone H2AX (γ-H2AX). Apoptosis was measured by flow-cytometry after staining with Annexin V. The relative biological effectiveness (RBE) was obtained by clonogenic survival assay.

Results: DDSB induction was significantly higher for protons than X-rays with average ratios of 1.28 (ONS76) and 1.59 (MOLT4) at 30 min after irradiation. However the differences became insignificant at 6 h. Also, apoptosis induction in MOLT4 cells was significantly higher for protons than X-rays with an average ratio of 2.13 at 12 h. However, the difference became insignificant at 20 h. RBE values of protons to X-rays at 10% survival were 1.06 ± 0.04 and 1.02 ± 0.15 for ONS76 and MOLT4, respectively.

Conclusions: Cell inactivation may differ according to different timings and/or endpoints. Proton beams demonstrated higher cell inactivation than X-rays in the early phases. These data may facilitate the understanding of the biological properties of clinical proton beams.

Keywords:

• proton
• RBE
• DDSB
• apoptosis

Acknowledgements

We are indebted to Ms Azusa Morikawa, Mr Masashi Seki, Ms Masako Nishio, Mr Masaru Sato, and Mr Masaya Ishida for their competent technical assistance. In addition, we thank Ms Junko Zenkoh for preparation of this manuscript. We also thank to Dr Tadao Ohno for his valuable technical advice.

Declaration of interest: This work was partly supported by a Grant-in-Aid (20390323) from the Ministry of Education, Culture, Sports, Science & Technology of Japan.