Abstract A03: Effects of survivin and lymphoma cell-derived exosomes on natural killer cell function

Tumors alter their microenvironment to promote survival using methods such as angiogenesis promotion, growth signals, and immune suppression. The immune system, whose regular function is to recognize and eradicate infected or abnormal cells, often seems unresponsive to transformed neoplastic cells. Many of the methods employed by tumors to reduce immune function are unclear, but there is evidence for T cell suppression, increased myeloid-derived suppressor cells (MDSCs), and reduced natural killer (NK) cell activity. NK cells are among the first cells to recognize and destroy abnormal cells due to their inherent killing capabilities. However, many types of cancers inhibit the surveillance and cytotoxic abilities of NK cells by releasing exosomes, vesicles that can modulate tumor microenvironment and intercellular communication for the purpose of enhancing tumor malignancy. Exosomes as a means for immunomodulation of tumor microenvironment have been the focus of recent research. These 50-150 nm sized lipid bound vesicles are secreted by many cell types, including immune cells and tumor cells, and the unique protein, lipid, mRNA and miRNA contents contribute to the complex intercellular communication occurring between malignant and normal cells. Cancer patients often have increased numbers of exosomes circulating through their body, including patients with hematological malignancies, such as lymphoma. Recently, cancer cell exosomes have been found to contain Survivin, an Inhibitor of Apoptosis (IAP) protein that prevents cell death and induces Th2 polarity shift and decreases proliferation and cytotoxicity of CD8+ T lymphocytes. The purpose of this study is to explore the effect of Survivin and cancer cell-derived exosomes, which contain Survivin, on the immune functions of NK cells. The hypothesis is that Survivin and exosomes will decrease NK cell functions by suppressing NK cell production or release of cytokines and cytotoxic granules. NK cells were obtained from peripheral blood using magnetic NK cell isolation kit from Miltenyi Biotec and exosomes were obtained from conditioned media from two lymphoma cell lines (WSU-DLCL2 and WSU-FSCCL) using ExoQuickTM (System Biosciences). Flow cytometry methods were used to evaluate degranulation capacity by measuring levels of CD107a, as well as expression of activating receptor NKG2D, cytotoxic granules (perforin, Granzyme B) and cytokines (TNF-α, IFN-γ). RNA message expression was investigated using both block RT-PCR and qPCR. Results showed little significant difference in ability to degranulate when exposed to stimuli, and NKG2D levels did not change after exposure to Survivin or lymphoma exosomes. Although ImageJ analysis of block PCR showed some instances of decreased band densities in some donors treated with Survivin protein, RNA levels in general did not show a trend between NK donors in response to treatment. However, protein expression of Granzyme B, perforin, TNF-α, and IFN-γ did appear to decrease after Survivin treatment. This work is still in the early stages and results are inconsistent due to donor variability. More samples are required, as well as further studies on NK cell functional cytotoxicity. Citation Format: Heather R. Ferguson Bennit, Amber Gonda, Jenniffer Licero Campbell, Laura Oppegard, David Chi, Nathan R. Wall. Effects of survivin and lymphoma cell-derived exosomes on natural killer cell function. [abstract]. In: Proceedings of the AACR Special Conference: Function of Tumor Microenvironment in Cancer Progression; 2016 Jan 7–10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2016;76(15 Suppl):Abstract nr A03.