2437 - Insights Into the Mechanism of Circulating Lymphocyte Sparing by Ultra-High Dose Rate Proton Irradiation Through Simulation of Lymphocyte Kinetics

Purpose/Objective(s): There is a desire to preserve the adaptive immune system following radiation therapy (RT). Ultra-high dose rate irradiation has the potential to spare circulating lymphocytes as the volume of blood traversing the irradiated field diminishes. Whether a biological FLASH effect exists in addition to lymphocyte sparing from the physical circulation effect is not yet known.

Materials/Methods: 96 C57BL/6 mice received radiation to the brain (16Gy/1fx) or liver (8Gy/1fx) from passively scattered protons. Mice were split into three cohorts receiving radiation at 0.5 Gy/s (conventional), 10 Gy/s, and 100 Gy/s. The irradiated field size was 1x1cm². Cheek bleed was used to retrieve 50 µl of blood prior to irradiation, on day 1, and day 8 following irradiation. Circulating CD4/8+ lymphocyte populations were determined using flow cytometry. Dose distributions to circulating blood were calculated using the HEDOS (HEmatological DOSe) approach, extended to simulate lymphocyte infiltration into damaged tissue. Expected lymphocyte survival from the dose to circulating blood was calculated with a=0.46. Linear mixed-effects models were developed to describe lymphocyte survival following irradiation. Significance testing was performed using pairwise Wald tests on estimated marginal means with a Tukey-Kramer correction to adjust for multiple comparisons.

Results: Acute CD8+ lymphocyte reduction following brain irradiation at conventional, 10 Gy/s, and 100 Gy/s dose-rates was 45.1%, 31.5%, and 23.7% (p = 0.02, compared to conventional) respectively. Acute CD8+ lymphocyte reduction following liver irradiation at increasing dose-rate was 34.2%, 19.3% (p = 0.02), and 11.3% (p < 0.01) respectively. Expected lymphocyte reduction from dose to circulating blood volume was 19.1%, 9.4%, 8.0% for brain irradiation and 37.7%, 28.4%, 27.8% for liver irradiation respectively. Computational modeling indicates that a CD8+ lymphocyte uptake of 400 per mm³ into damaged brain tissue following irradiation would be required to explain the experimental results.

Conclusion: Increasing dose rate results in significant sparing of circulating lymphocytes. The marginal difference in circulating blood volume irradiated between 10 Gy/s and 100 Gy/s irradiations can not explain the magnitude of sparing observed; this suggests the presence of a potential FLASH sparing effect beyond the sparing caused by a reduction in dose to circulating blood alone.

After brain irradiation, the acute reduction of circulating lymphocytes is disproportionately larger than the volume of blood irradiated. This effect could be explained by recruitment of lymphocytes into damaged tissue, experimental confirmation of this hypothesis is ongoing.