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• Repeated administration of rcIL-15 was well tolerated and did not cause any serious side effects. • Infusion of rcIL-15 promoted the selective proliferation and activation of CD8+ T lymphocytes, and NK lymphocytes. • Repeated administration of rcIL-15 did not stimulate T reg lymphocytes. • Injection of rcIL-15 stimulated the expression of molecules and transcription factors associated with the activation and effector functions of NK cells. Interleukin-15 (IL-15) is a pleiotropic cytokine that plays pivotal roles in innate and adaptive immunity. It is also a promising cytokine for treating cancer. Despite growing interest in its use as an immunotherapeutic, its safety and immunological effects in dogs have not been reported. In this study, healthy dogs were given recombinant canine IL-15 (rcIL-15) intravenously at a daily dose of 20 μg/kg for 8 days and monitored for 32 days to determine the safety and immunological effects of rcIL-15. The repeated administration of rcIL-15 was well tolerated, did not cause any serious side effects, and promoted the selective proliferation and activation of canine anti-cancer effector cells, including CD3+CD8+ cytotoxic T lymphocytes, CD3+CD5dimCD21–, and non-B/non-T NK cell populations, without stimulating T reg lymphocytes. The rcIL-15 injections also stimulated the expression of molecules and transcription factors associated with the activation and effector functions of NK cells, including CD16, NKG2D, NKp30, NKp44, NKp46, perforin, granzyme B, Ly49, T-bet, and Eomes. These results suggest that rcIL-15 might be a valuable therapeutic adjuvant to improve immunity against cancer in dogs. [ABSTRACT FROM AUTHOR]