Background Focal radiotherapy (RT) promotes tumor infiltration by conventional dendritic cells type 1 (cDC1), an effect dependent on radiation’s ability to induce type I interferon (IFN-I) secretion. We recently demonstrated that peritumoral s.c. IL-15, while ineffective by itself, synergized with RT, inducing complete regression of the irradiated tumor and long-term protective memory in two murine carcinomas models (TSA, MCA-38) (1). These responses were abrogated in the absence of CD8 T cells or cDC1. Detailed investigations in the TSA model showed that, whereas IL-15 alone had no effects on cDC1, it did significantly increase intratumoral cDC1 numbers and expression of costimulatory molecules CD80 and CD86 induced by RT (1). In addition to CD8 T cells, IL-15 activates NK cells, which have also been implicated in cDC1 tumor recruitment (2). Thus, we hypothesized that NK cells may play a role in the synergy between radiation therapy and IL-15.

Methods To test this hypothesis, BALB/c mice were injected with TSA mammary carcinoma cells and treated with RT (8Gy X3) and daily subcutaneous injections of IL-15 (5µg). Tumors were excised at day 18 and analyzed by immunostaining for NKp46+ cells on tumor sections and flow cytometry after tumor dissociation.

Results The number of intra-tumoral NKp46+ NK cells was significantly higher (p<0.005) in mice treated with IL-15 as compared to control. Whereas RT itself had no effect, it further increased NK cell numbers above what was achieved by IL-15 alone (p<0.05). In addition, tumor infiltrating NK cells expressed higher levels of CD137/4-1 BB, an effect largely driven by IL-15. Finally, NK cells depletion by anti-asialo GM1 before initiation of the treatment abrogated the enhanced cDC1 infiltration in tumors of mice treated with RT + IL-15, and the therapeutic effect of the combination.

Conclusions Our results strongly suggest a role for NK cells in the anti-tumor immune response induced by the combination of RT and IL-15. We are currently working to confirm the role of NK cells by using a complementary approach of engineering TSA cells to overexpress CLEC2D/Clr-b, the ligand for the inhibitory NKR-P1 NK receptor (3,4). Data obtained will improve current knowledge about the interaction of RT with IL-15 and support a rationale strategy for translation to the clinic.

References

1. Pilones KA, Charpentier M, Garcia-Martinez E, et al. Radiotherapy cooperates with IL15 to induce antitumor immune responses. Cancer Immunol Res 2020;8(8):1054–1063. doi:10.1158/2326-6066.CIR-19-0338

2. Böttcher JP, Bonavita E, Chakravarty P, et al. NK Cells stimulate recruitment of cDC1 into the tumor microenvironment promoting cancer immune control. Cell 2018;172(5):1022–1037.e14. doi:10.1016/j.cell.2018.01.004

3. Carlyle JR, Jamieson AM, Gasser S, Clingan CS, Arase H, Raulet DH. Missing self-recognition of Ocil/Clr-b by inhibitory NKR-P1 natural killer cell receptors. Proc Natl Acad Sci U S A 2004;101(10):3527–3532. doi:10.1073/pnas.0308304101

4. Williams KJ, Wilson E, Davidson CL, et al. Poxvirus infection-associated downregulation of C-type lectin-related-b prevents NK cell inhibition by NK receptor protein-1B. J Immunol 2012;188(10):4980–4991. doi:10.4049/jimmunol.1103425