Intermittent cyclophosphamide and vinorelbine combined with anti-PD-1 induce transferable T-cell-mediated antitumor immunity and long-term memory in TNBC mouse models

Cell-based immunotherapies offer a powerful means to harness treatment-induced antitumor immunity for durable tumor control. We previously developed a Triple-Therapy (TT) regimen combining vinorelbine-mediated activation of antigen-presenting cells (APCs) with cyclophosphamide-driven induction of TCF1⁺ stem-cell-like T cells (scTs), markedly enhancing anti-PD-1 efficacy in resistant tumor models. The TT protocol consists of three intermittent doses of vinorelbine, and cyclophosphamide administered every six days, together with five anti-PD-1 doses delivered every two days.In a Non-Hodgkin Lymphoma (NHL) model, TT not only achieved strong tumor regression but also conferred long-term immunological memory, as evidenced by protection upon tumor rechallenge. Extending these findings to triple-negative breast cancer (TNBC), TT induced comparable antitumor activity. To determine whether TT-elicited immunity could be adoptively transferred, immunocompetent BALB/c mice bearing 4T1 TNBC were treated with TT, and immune tissues-including blood, spleen, and lymph nodes-were collected and transferred into syngeneic BALB/c nude mice inoculated with the same tumor.Strikingly, transfer of peripheral blood from TT-treated donors significantly delayed tumor growth, indicating that circulating immune cells are sufficient to convey protection. In contrast, lymphocyte depletion abolished this effect. Selective ablation of CD4⁺ or CD8⁺ T cells-performed either by flow cytometric sorting or using neutralizing antibodies-also eliminated the therapeutic benefit, confirming their essential role in the transferable antitumor response.Overall, these findings demonstrate that TT induces systemic, memory-forming T-cell responses capable of mediating cell-transferable tumor control. Current work focuses on identifying the tumor antigens recognized by TT-induced T cells, with the ultimate goal of developing targeted therapeutic vaccines to extend these benefits to broader patient populations.