Despite the development of multimodal therapy consisting of surgery, radiotherapy, and chemotherapy, the prognosis of patients with metastatic or relapsed synovial sarcoma (SS) remains poor. A novel treatment option including immunotherapy is urgently required. Natural killer (NK) cells are innate immune cells that can recognize and kill tumor cells. The cytolytic activity of NK cells is determined by the balance of signals from both inhibitory and activating receptors expressed on cell surface. Activating receptors include NKp30, NKp44, NKp46, NKG2D, and DNAM-1. Furthermore, tumor cells with low HLA class I expression are potentially even more sensitive to NK-mediated cytotoxic killing due to a lack of inhibitory signals. We previously reported that HLA class I expression was down regulated in one third of patients with synovial sarcoma, suggesting that NK cell therapy may be effective for patients with SS. However, the efficacy of NK cells treatment in SS was not well established. In this study, we evaluated surface ligands for NK cell activation on SS cell lines and demonstrated the effectiveness of activated allogenic NK cells for SS cell lines in vitro.The surface expression of ligands for NK cell-activating receptors including NKp30, NKp44, NKp46, NKG2D, and DNAM-1 in three SS cell lines (SYO-I, HS-SY-II, and Yamato-SS) was evaluated using flow cytometry. Peripheral blood from two healthy donors were ficoll-separated to collect peripheral blood mononuclear cells (PBMCs). Then, PBMCs were stimulated with irradiated K562 cells expressing the NK-stimulatory molecules 4-1BB ligand and membrane-bound Interleukin 15 in the presence of human Interleukin-2 in RPMI1640 with 10% FBS. The cells were expanded for 14 days, and the expanded NK cells are CD3-CD56+ NK cells. These expanded cells are reproducible in their function, and we have utilized these cells for in vitro cytotoxicity assays. 2x10E4 of SS cell lines (SYO-I, HS-SY-II, and Yamato-SS) were seeded in a 96 well plates and cultured overnight. Then, allogenic NK cells or activated T cells as a control at various tumor effector ratio. The cells were co-cultured with activated T cells or NK cells in the presence of low dose interleukin2. Forty-eight hours later, cell viabilities were measured using WST-8 assay.Ligands for NKp44, NKp30, NK2GD, were expressed in all three SS cell lines. It suggested that SS cells upregulate activating receptor ligands for NK cells. In addition, cytotoxicity of allogenic NK cells for SS cell lines was significantly higher than that of activated T cells. To determine whether cytotoxic activity was effect of allogenic NK cells, CD107a mobilization assay using flowcytometry was performed. CD107a mobilization assay demonstrated degranulation of NK cells, leading us that NK cells had an antitumoral effect on SS cell lines in vitro. Based on these initial findings, we were able to demonstrate that allogenic NK cells may be a promising immune therapy for SS.

Citation Format: Naoki Oike, Tomohiro Miyazaki, Yudai Murayama, Takashi Ariizumi, Akira Ogose, Minori Baba, Hiroyuki Kawashima, Chihaya Imai. Adoptive natural killer therapy is effective for synovial sarcoma cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5236.