Intratumoral IL-12 and TNF-α-loaded microspheres lead to regression of breast cancer and systemic antitumor immunity

Background: Local, sustained delivery of cytokines at a tumor can enhance induction of antitumor immunity and may be a feasible neoadjuvant immunotherapy for breast cancer. We evaluated the ability of intratumoral poly-lactic-acid-encapsulated microspheres (PLAM) containing interleukin 12 (IL-12), tumor necrosis factor alpha (TNF-alpha), and granulocyte-macrophage colony stimulating factor (GM-CSF) in a murine model of breast cancer to generate a specific antitumor response. Methods: BALB/c mice with established MT-901 tumors underwent resection or treatment with a single intratumoral injection of PLAM containing IL-12, TNF-alpha, or GM-CSF, alone or in combination. Two weeks later, lymph nodes and spleens were harvested, activated with anti-CD3 monoclonal antibodies (mAb) and rhIL-2, and assessed for antitumor reactivity by an interferon gamma (IFNgamma) release assay. Tumor-infiltrating lymphocyte (TIL) analysis was performed on days 2 and 5 after treatment by mechanically processing the tumors to create a single cell suspension, followed by three-color fluorescence-activated cell sorter (FAGS) analysis. Results: Intratumoral injection of cytokine-loaded PLAM significantly suppressed tumor growth, with the combination of IL-12 and TNF-alpha leading to increased infiltration by polymorphonuclear cells and CD8(+) T-cells in comparison with controls. The induction of tumor-specific reactive T-cells in the nodes and spleens, as measured by IFN-gamma production, was highest with IL-12 and TNF-alpha. This treatment resulted in resistance to tumor rechallenge. Conclusions: A single intratumoral injection of IL-12 and TNF-alpha-loaded PLAM into a breast tumor leads to infiltration by polymorphonuclear cells and CD8+ T-cells with subsequent tumor regression. In addition, this local therapy induces specific antitumor T-cells in the lymph nodes and spleens, resulting in memory immune response.