Abstract 2808: Investigating the role of macrophage-mediated suppression in the efficacy of NK cell immunotherapy for metastatic outgrowth of breast cancer cells

Breast cancer is the leading cause of cancer-related death in females worldwide. Although 5-year survival of breast cancer patients in early stages is 89-100%, that of patients with metastatic tumours is reduced to just 21%, suggesting the requirement of more effective therapies for metastatic breast cancer (MBC). As MBC is refractory to current therapies, NK cell infusion has been suggested as an alternative therapy. However, its efficacy is modest in MBC possibly due to the suppressive nature of the tumour microenvironment, especially through the tumour-associated macrophages (TAMs) within it. Here we investigated the effects of TAMs in metastatic tumours on NK cell cytotoxicity, as well as the mechanism behind TAM-mediated NK cell suppression. To evaluate NK cell cytotoxicity, we established a method whereby mouse breast cancer cells were cultured with splenic NK cells, and the resultant tumour cell apoptosis was determined by quantitative fluorescence microscopy. To investigate the effects of TAMs on this NK cytotoxicity, we purified them from lungs with metastatic tumours and added them to this co-culture. We also investigated the suppressive effects of bone marrow-derived macrophages cultured with M-CSF (M-BMMs). To further investigate the mechanism, we depleted TAMs in mice with metastatic tumours and detected inhibitory receptor expression on NK cells as well as NK cell inhibitory ligands on TAMs in the metastatic lung and on M-BMMs by flow cytometry. TAMs from metastatic tumours significantly reduced NK cell cytotoxicity toward tumour cells. M-BMMs (resembling TAMs) also reduced NK cell cytotoxicity. We also found that TAMs and M-BMMs expressed high levels of NK cell inhibitory ligands. There was no difference in the expression of inhibitory receptors on NK cells between tumour bearing lungs with or without TAMs. To conclude, we have shown that TAMs in metastatic tumours, as well as their mimetic M-BMMs, can suppress NK cell cytotoxicity towards breast cancer cells in vitro. Our data suggest that TAMs express higher levels of inhibitory ligands and thereby transmit suppressive signals possibly through direct contact with NK cells. Further investigation of mechanisms behind TAM-mediated NK suppression might lead to the improvement of NK cell-based immunotherapy for MBC. Citation Format: Demi Brownlie, Dahlia Doughty-Shenton, Jeffrey W. Pollard, Takanori Kitamura. Investigating the role of macrophage-mediated suppression in the efficacy of NK cell i