Attempts to cure breast cancer by means of adoptive cellular therapy (ACT) have not been successful. This is primarily due to the presence of tumor-induced immune suppressive mechanisms as well as the failure of tumor-reactive T cells to provide long-term memory responses in vivo. In order to address these clinically important challenges we developed an ex vivo protocol for the expansion of tumor-reactive immune cells obtained from tumor-bearing animals prior to or after local radiation therapy. We used an antigen-free protocol which included bryostatin 1/ionomycin (B/I) and sequential common gamma-chain cytokines (IL-7/IL-15 + IL-2). The proposed protocol expanded tumor-reactive T cells as well as activated non-T cells, including NK T cells, NK cells and IFN-γ producing killer dendritic cells (IKDC). Anti-tumor efficacy of T cells depended on the presence of non-T cells. The effector non-T cells also rendered T cells resistant to myeloid-derived suppressor cells (MDSC). Radiation therapy altered phenotypic distribution and differentiation of T cells, as well as their ability to generate central memory T cells (TCM). ACT by means of the expanded cells protected animals from tumor challenge and generated long-term memory responses against the tumor, provided that leukocytes were derived from tumor-bearing animals prior to radiation therapy. The ex vivo protocol was also able to expand HER-2/neu-specific T cells derived from the PBMC of a single patient with breast carcinoma. These data suggest that the proposed ACT protocol should be studied further in breast cancer patients.

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Adoptive T cell immunotherapy (AIT) utilizing ex vivo expanded HER-2/neu-specific T cells is a promising modality in the treatment of patients with breast carcinoma. Previously, our group has described the ability of tumor-sensitized T cells activated ex vivo with bryostatin 1 and ionomycin (B/I) and subsequently expanded with common γ-chain receptor family cytokines IL-2 and IL-7/IL-15 cultured in an alternating manner to induce primary breast tumor regression in FVBN202 transgenic mice. These results, however, required the combination of AIT with in vivo depletion of myeloid-derived suppressor cells (MDSCs) as the presence of these cells abrogated the anti-tumor efficacy of neu-specific lymphocytes. Presently, we report the implementation of a novel ex vivo expansion protocol, in which IL-2 and IL-7/IL-15 are cultured in a sequential manner post B/I with tumor-specific activated cells and support expansion of tumor reactive CD44+CD62L− effector T cells (TE) and CD44+CD62L+ central memory T cells (TCM). We show that tumor-specific cells activated in this manner induce rejection of primary breast tumor and provide long-term protection against the tumor, even in the presence of MDSC. Of particular interest, tumor-reactive CD4+ and CD8+ lymphocytes expanded with this method conferred resistance to the suppressive effects of MDSCs in vitro. Additionally, we demonstrate that innate immune cells are essential in eliciting tumor rejection. Flow cytometric analysis indicates Natural Killer (NK) and Natural Killer T-cell (NKT) as well as the Interferon-γ Producing Killer Dendritic Cell (IKDC) populations significantly increase following ex vivo expansion with sequential common γ-chain cytokines. Depletion of innate immune cells in adoptively transferred cells resulted in the failure of AIT to reject tumor. Together, our data suggest that sequential common γ-chain cytokines can support differentiation of tumor-reactive T cells which are resistant to MDSC, as well as innate immune cells which may be essential in supporting the anti-tumor efficacy of T cells. These results potentially provide a new paradigm in therapeutic AIT of breast carcinoma. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 775. doi:10.1158/1538-7445.AM2011-775