Fractionation increasingly reduces FLASH sparing for acute murine skin damage

BACKGROUND AND PURPOSE: Preclinical studies report a favourable normal tissue-sparing FLASH effect. Most studies report single-fraction irradiations. The interplay between fractionation and FLASH sparing is highly relevant for clinical translation but is scarce in the literature. This study compared the tissue-sparing effect of FLASH on acute skin toxicity with single-fraction, four-fraction and eight-fraction schemes. MATERIALS AND METHODS: Acute skin toxicity after conventional dose rate (CONV, 0.16 Gy/s) and FLASH (mean ± sd 251 ± 12 Gy/s) irradiation was assessed in female CDF1 mice. The right hindleg of unanaesthetised mice was irradiated using a single dose (1 fraction), one daily dose for four consecutive days (4 fractions) or two daily doses with a 6-hour interval for four consecutive days (8 fractions). The study had 4-12 mice per dose group. The total dose per group ranged from 24.5 to 89.9 Gy. A FLASH-enabled accelerator (TrueBeam, Varian) delivered irradiation with a 16 MeV electron beam. Acute skin toxicity was quantified daily from 9 to 28 days post-treatment. RESULTS: Single-fraction irradiation had a tissue-sparing FLASH effect, which was halved for four fractions and markedly reduced for eight fractions. The dose modification ratio was 1.41 for a single fraction, 1.18 for four fractions, and nonsignificantly 1.05 for eight fractions. CONCLUSION: Fractionation increasingly reduced the acute skin-sparing effect seen in single-fraction FLASH studies. However, a tissue-sparing effect of 18 % was still present at four fractions, while eight fractions provided a nonsignificant 5 % FLASH skin-sparing. Clinical implementation of FLASH may work best in highly hypofractionated settings.