Indicators of late normal tissue response after radiotherapy for head and neck cancer: fibroblasts, lymphocytes, genetics, DNA repair, and chromosome aberrations

Abstract

Purpose: To investigate the relationship between late tissue response after radiotherapy, cellular sensitivity and DNA repair capacity measured in dermal fibroblasts and chromosomal aberrations measured in lymphocytes. The study was in particular designed to compare cellular parameters of patients with maximum differences in late tissue reactions. Materials and methods: The study was performed with 16 pair-wise matched head and neck cancer patients 2–7 years after curative therapy exhibiting maximum differences (grade 1 vs. grade 3) in late normal tissue reactions. Clinical endpoints were fibrosis, telangiectasia, mucositis and xerostomia using the radiation therapy oncology group score. Patients with grade 3 reactions were tested for mutations in ataxia telangiectasia (AT), Nijmegen Breakage Syndrome (NBS), MRE11, RAD50 and DNA ligase IV genes by means of polymerase chain reaction-single-strand conformation polymorphism and sequencing analysis. Skin fibroblasts obtained from biopsies were used to determine the cellular sensitivity by colony formation and the induction and repair of DNA double-strand breaks (dsb) using constant-field gel electrophoresis. Lymphocytes were taken to measure chromosomal damage either in metaphase using conventional chromosome analysis or in G 0 using premature chromosome condensation (PCC)-technique. Results: Patients with extreme late reactions (grade 3) showed no evidence for an AT, NBS, MRE11 or RAD50 mutation. Studies with fibroblasts revealed that extreme late reactions were associated neither with a pronounced cellular radiosensitivity nor with a difference in dsb repair capacity. In contrast, there was a significant difference in chromosomal damage measured in lymphocytes. After in vitro irradiation with 6 Gy, lymphocytes taken from overreacting patients showed on average a significantly higher number of lethal aberrations than lymphocytes isolated from patients with mild reactions (7.2±0.8 vs. 5.0±0.3). Similar differences were found for PCC fragments. Conclusion: This study suggests that lymphocytes are more promising than fibroblasts to predict patient's normal tissue response after radiotherapy.