Ionizing radiation inhibition of distant untreated tumors (abscopal effect) is immune mediated
Presented in part at the 44th Annual Meeting of the American Society for Therapeutic Radiology and Oncology, New Orleans, LA, October 6–10, 2002.
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Fig. 1
Expression of MHC and costimulatory molecules on 67NR carcinoma cells. Histograms show staining of 67NR cells with mAb against MHC class I molecule H2-Kd, MHC class II molecule Iad, and costimulatory molecules CD80 and CD86. The broken line indicates the fluorescence of unstained control cells.
Fig. 2
Systemic anticancer effects are triggered by local RT and Flt3-L. Mice were injected s.c. with 105 (“primary” tumor) and 5 × 104 (“secondary” tumor) 67NR cells at Day 0. Mice were either left untreated (empty diamonds), were treated with RT (filled diamonds) at Day 20 (arrow) at a single dose of 2 Gy exclusively to primary tumors, or were given Flt3-L alone (empty circles) i.p. for 10 days starting at Day 21 or Flt3-L in combination with RT (filled circles). Data are the mean tumor weight ± SD of each treatment group and are representative of two experiments. Whereas Flt3-L alone had no effect, RT led to a significant growth delay of the primary (p < 0.01 from Day 34) but not secondary tumor. In contrast, the combination of Flt3L and RT to the primary tumor resulted in significant growth delay of secondary tumors also (p < 0.05 from Day 34).
Fig. 3
The abscopal effect triggered by local RT and Flt3-L is tumor specific. Mice were injected s.c. in the left flank with 105 67NR cells at Day 0 (“primary” tumor). Five days later, 105 67NR cells were injected in the right flank (“secondary” 67NR), and 106 A20 cells were injected in the back (“secondary” A20). Mice were either left untreated (empty diamonds), were treated with RT (filled diamonds) at Day 14 (arrow) at a single dose of 2 Gy exclusively to primary tumors, or were given Flt3-L alone i.p. (empty circles) for 10 days starting at Day 15 or Flt3-L in combination with RT (filled circles). Mice in the control group and mice treated with Flt3-L alone were killed at Day 38 because of an overall too-large tumor burden. Data are the mean tumor weight of each treatment group. Growth of 67NR and A20 tumors was not affected by Flt3-L alone. In the absence of Flt3-L, RT led to a significant growth delay of the primary (p < 0.05 from Day 31) but not secondary tumors. In the presence of Flt3-L, RT to the primary tumor resulted in growth delay also of secondary antigenically identical 67NR tumors (p < 0.05 from Day 28). In contrast, growth of the secondary antigenically unrelated A20 tumors was not affected.
Fig. 4
67NR-specific CTL precursors are increased in the spleen of mice treated with local RT and Flt3-L. Spleen T cells were isolated from treated and untreated 67NR tumor–bearing mice, as indicated, at the end of Flt3-L administration and tested after one round of in vitro restimulation in a standard 51Cr-release assay against 67NR (solid bars) and A20 (shaded bars) tumor cells at an E:T ratio of 30:1. Increased CTL activity against 67NR but not A20 cells is detected in the spleen of tumor-bearing mice treated with RT + Flt3-L compared to untreated mice and mice receiving RT or Flt3-L alone. Data are representative of two experiments.
Fig. 5
The abscopal effect triggered by local RT and Flt3-L is T cell dependent. Nude mice were injected s.c. in the left flank with 105 67NR cells at Day 0 (“primary” tumor). Five days later, 105 67NR cells were injected in the right flank (“secondary” tumor). Mice were either left untreated (empty diamonds), were treated with RT (filled diamonds) at Day 14 (arrow) at a single dose of 2 Gy exclusively to primary tumors, or were given Flt3-L alone (empty circles) i.p. for 10 days starting at Day 15 or Flt3-L in combination with RT (filled circles). Data are the mean tumor weight ± SD of each treatment group and are representative of two experiments. RT led to a significant growth delay of the primary but not secondary tumors. In contrast to immunocompetent mice, in nude mice the addition of Flt3-L did not result in any detectable abscopal effect of RT on the growth of the secondary tumors.
Abstract
Purpose
Ionizing radiation can reduce tumor growth outside the field of radiation, known as the abscopal effect. Although it has been reported in multiple malignancies, the abscopal effect remains a rare and poorly understood event. Ionizing radiation generates inflammatory signals and, in principle, could provide both tumor-specific antigens from dying cells and maturation stimuli that are necessary for dendritic cells' activation of tumor-specific T cells. We therefore tested the hypothesis that the abscopal effect elicited by radiation is immune mediated. This was directly tested by enhancing the number of available dendritic cells using the growth factor Flt3-Ligand (Flt3-L).
Methods and materials
Mice bearing a syngeneic mammary carcinoma, 67NR, in both flanks were treated with Flt3-L daily for 10 days after local radiation therapy (RT) to only 1 of the 2 tumors at a single dose of 2 or 6 Gy. The second nonirradiated tumor was used as indicator of the abscopal effect. Data were analyzed using repeated measures regression.
Results
RT alone led to growth delay exclusively of the irradiated 67NR tumor, as expected. Surprisingly, growth of the nonirradiated tumor was also impaired by the combination of RT and Flt3-L. As control, Flt3-L had no effect without RT. Importantly, the abscopal effect was shown to be tumor specific, because growth of a nonirradiated A20 lymphoma in the same mice containing a treated 67NR tumor was not affected. Moreover, no growth delay of nonirradiated 67NR tumors was observed when T cell deficient (nude) mice were treated with RT plus Flt3-L.
Conclusions
These results demonstrate that the abscopal effect is in part immune mediated and that T cells are required to mediate distant tumor inhibition induced by radiation.
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☆The authors thank Elaine Thomas for Flt3-Ligand. S.D. was supported by Grant K08 CA89336 from NIH/NCI and by a grant from the Speaker's Fund for Biomedical Research: Toward the Science of Patient Care, awarded by the city of New York. S.C.F. was supported by Grants DAMD17-01-1-0345 from the Department of Defense, TURSG CCE 103174 from the American Cancer Society, and by a grant from the Breast Cancer Research Foundation.
