Radiotherapy enhances natural killer cell cytotoxicity and localization in pre-clinical canine sarcomas and first-in-dog clinical trial

Abstract

BackgroundWe have previously shown that radiotherapy (RT) augments natural killer (NK) functions in pre-clinical models of human and mouse cancers, including sarcomas. Since dogs are an excellent outbred model for immunotherapy studies, we sought to assess RT plus local autologous NK transfer in canine sarcomas.MethodsDog NK cells (CD5dim, NKp46+) were isolated from PBMCs and expanded with irradiated K562-C9-mIL21 feeder cells and 100 IU/mL recombinant human IL-2. NK homing and cytotoxicity ± RT were evaluated using canine osteosarcoma tumor lines and dog patient-derived xenografts (PDX). In a first-in-dog clinical trial for spontaneous osteosarcoma, we evaluated RT and intra-tumoral autologous NK transfer.ResultsAfter 14 days, mean NK expansion and yield were 19.0-fold (±8.6) and 258.9(±76.1) ×106 cells, respectively. Post-RT, NK cytotoxicity increased in a dose-dependent fashion in vitro reaching ~ 80% at effector:target ratios of ≥10:1 (P < 0.001). In dog PDX models, allogeneic NK cells were cytotoxic in ex vivo killing assays and produced significant PDX tumor growth delay (P < 0.01) in vivo. After focal RT and intravenous NK transfer, we also observed significantly increased NK homing to tumors in vivo. Of 10 dogs with spontaneous osteosarcoma treated with focal RT and autologous NK transfer, 5 remain metastasis-free at the 6-month primary endpoint with resolution of suspicious pulmonary nodules in one patient. We also observed increased activation of circulating NK cells after treatment and persistence of labelled NK cells in vivo. ConclusionsNK cell homing and cytotoxicity are increased following RT in canine models of sarcoma. Results from a first-in-dog clinical trial are promising, including possible abscopal effects.Electronic supplementary materialThe online version of this article (10.1186/s40425-017-0305-7) contains supplementary material, which is available to authorized users.

Cellular therapy: Adoptive immunotherapy with expanded natural killer cells

“…In addition to the propagation of NK cells from human peripheral blood and cord blood, the robustness of this expansion platform has been validated across other systems, generating large numbers of NK cells from embryonic and induced pluripotent stem cells, and from canine and macaque peripheral blood for use in comparative oncology and virology models …”

“…97 In addition to the propagation of NK cells from human peripheral blood and cord blood, the robustness of this expansion platform has been validated across other systems, generating large numbers of NK cells from embryonic and induced pluripotent stem cells, 98 and from canine and macaque peripheral blood for use in comparative oncology and virology models. [99][100][101][102] Through our own work and the extensive network of collaborations made possible by sharing the original cell line, the ability to generate large numbers of uniform NK cell populations from limited donor material enabled accelerated research in NK cell biology and F I G U R E 1 Expansion of natural killer (NK) cells by recursive weekly stimulation with mbIL21-expressing feeder cells. After the initial publication, laboratory members recorded NK cell expansion data from their various projects to generate "real-life" expectations.…”

Emerging therapeutic approaches for canine sarcomas: Pushing the boundaries beyond the conventional

“…There is tremendous interest in veterinary oncology to explore the combination of RT and immunotherapy in the dog . In one recent study, NK cells were harvested and expanded from dogs with osteosarcoma and subsequently injected intratumourally to dogs following completion of coarse‐fraction RT (4 × 9 Gy delivered once weekly) . NK cell homing and cytotoxicity were increased following local RT.…”

“…NK cell homing and cytotoxicity were increased following local RT. Five of 10 dogs treated remained free of metastases at 6 months posttreatment and resolution of pulmonary nodules were noted in one dog . As novel drugs become available, it will be an important means in which comparative oncology can help to address several lingering questions regarding the ability to exploit tissue responses to radiation with checkpoint blockade.…”

“…34,133,169,170 In one recent study, NK cells were harvested and expanded from dogs with osteosarcoma and subsequently injected intratumourally to dogs following completion of coarse-fraction RT (4 × 9 Gy delivered once weekly). 170 NK cell homing and cytotoxicity were increased following local RT. Five of 10 dogs treated remained free of metastases at 6 months posttreatment and resolution of pulmonary nodules were noted in one dog.…”