PURPOSE: This study aimed to analyze the intra-tumoral heterogeneity of γH2AX foci in tumor specimens following ex vivo radiation to evaluate the potential of γH2AX foci as predictors for radiosensitivity.

MATERIAL AND METHODS: γH2AX foci were quantified in tumor specimens of 3hHNSCC tumor models with known differences in radiosensitivity after reoxygenation in culture medium (10h, 24h), single dose exposure (0Gy, 4Gy), and fixation 24h post-irradiation. Multiple, equally treated samples of the same tumor were analyzed for foci, normalized and fitted in a linear mixed-effects model.

RESULTS: The ex vivo reoxygenation time had no significant effect on γH2AX foci counts. A significant intra model heterogeneity could be shown for FaDu (p=0.033) but not for SKX (p=0.167) and UT-SCC-5 (p=0.082) tumors, respectively. All tumor models showed a significant intra-tumoral heterogeneity between specimens of the same tumor (p<0.01) or among microscopic fields of a particular tumor specimen (p<0.0001).

CONCLUSION: Similar results for ex vivo γH2AX foci between 10h and 24h reoxygenation time support the applicability of the assay in a clinical setting. The high intra-tumoral heterogeneity underlines the necessity of multiple analyzable samples per patient and therewith the need for an automated foci analysis.