Evaluation of different biomarkers to predict individual radiosensitivity in an inter-laboratory comparison--lessons for future studies

Abstract

Radiotherapy is a powerful cure for several types of solid tumours, but its application is often limited because of severe side effects in individual patients. With the aim to find biomarkers capable of predicting normal tissue side reactions we analysed the radiation responses of cells from individual head and neck tumour and breast cancer patients of different clinical radiosensitivity in a multicentric study. Multiple parameters of cellular radiosensitivity were analysed in coded samples of peripheral blood lymphocytes (PBLs) and derived lymphoblastoid cell lines (LCLs) from 15 clinical radio-hypersensitive tumour patients and compared to age- and sex-matched non-radiosensitive patient controls and 15 lymphoblastoid cell lines from age- and sex- matched healthy controls of the KORA study. Experimental parameters included ionizing radiation (IR)-induced cell death (AnnexinV), induction and repair of DNA strand breaks (Comet assay), induction of yH2AX foci (as a result of DNA double strand breaks), and whole genome expression analyses. Considerable inter-individual differences in IR-induced DNA strand breaks and their repair and/or cell death could be detected in primary and immortalised cells with the applied assays. The group of clinically radiosensitive patients was not unequivocally distinguishable from normal responding patients nor were individual overreacting patients in the test system unambiguously identified by two different laboratories. Thus, the in vitro test systems investigated here seem not to be appropriate for a general prediction of clinical reactions during or after radiotherapy due to the experimental variability compared to the small effect of radiation sensitivity. Genome-wide expression analysis however revealed a set of 67 marker genes which were differentially induced 6 h after in vitro-irradiation in lymphocytes from radio-hypersensitive and non-radiosensitive patients. These results warrant future validation in larger cohorts in order to determine parameters potentially predictive for clinical radiosensitivity.

Figure 1. Dose-response relationship of Olive tail moment (OTM) in PBLs at different time points.

Data from two different centres E (white boxes) and D (hatched boxes) are presented. Comet assay of both laboratories revealed similar results by investigating OTM in PBLs directly after irradiation with 5 Gy and at different time points. Non-radiosensitive and radiosensitive individuals were not distinguishable.

Figure 2. Dose-response relationship of Gamma-H2AX at different time points from two different centres (white boxes – centre C, hatched boxes – centre B).

Each measured value corresponds to the n-fold change to the time point at 0 Gy after 24 hours. (a) Data for the PBLs: Box plots only include samples with an initial cell viability of higher than 80%. (b) Data for LCLs over all samples.