clean-room facilities, pumps, sensors, tubing), making them less appealing to conventional cellular and molecular biology labs.29,42 Here we presented a model that does not require clean room processing and can be operated with no other additional gear rather than micropipettes. were able to detect the presence of the tumor spheroid several hundreds of microns away and penetrate the spheroid faster than the antibodies. Once inside the spheroid, natural killer cells were able AB05831 to eliminate tumor cells at the spheroid periphery and, importantly, also at the innermost layers. Finally, the combination of antibody-cytokine conjugates and natural killer cells led to an enhanced cytotoxicity located mostly at the spheroid periphery. Overall, these results demonstrate the power of the model for informing immunotherapy of solid tumors. KEYWORDS: Natural Killer cell, Antibody-dependent cell cytotoxicity, solid tumor, microfluidics, organotypic model Introduction In recent years, there has been a growing body of literature demonstrating the great potential of the immune system in fighting malignancy.1?4 Some immunotherapies are based on modulating the immune system Mouse monoclonal to Fibulin 5 (i.e. immunomodulation), injecting exogenous molecules like cytokines (e.g. IL-2) or antibodies targeting the tumor-immune cell conversation (e.g. PD-1/PD-L1 or HER-2).5 Other immunotherapies rely on the direct injection of na?ve or engineered immune cells into the bloodstream (cell-based immunotherapy) to engage and destroy the tumor cells. Most AB05831 of the cell-based immunotherapies rely on cytotoxic T cells (also known as CD8+ T cells) and natural killer (NK) cells given their cytotoxic capacity.6,7 Both cell types destroy tumor cells by opening pores in the tumor cell membranes (by secreting perforins) or secreting different apoptosis-inducing proteins (e.g. granzymes, TRAIL).8 Additionally, NK cells express antibody receptors (i.e. CD16), allowing them to recognize antibody-coated tumor cells as a target in a process known as antibody-dependent cell cytotoxicity (ADCC).9,10 Therefore, antibodies targeting molecules that are overexpressed in tumor cells are broadly used in the clinic (e.g. trastuzumab, cetuximab, pembrolizumab), combining both immunomodulation and cell-based immunotherapy for optimal results.9 Although these approaches have worked well for hematological cancers, their success in treating solid tumors has been modest. Multiple studies have highlighted different factors that may dampen the immune response in solid tumors.11 In this context, during the development of a solid tumor, immune cells and antibodies need to extravasate from your blood vessels; diffuse and migrate through the matrix; penetrate through the solid tumor; and finally destroy the tumor cells.12 However, sound tumors are dense structures that hinder drug penetration and cell migration; raising the question about whether NK cells and antibodies can reach the innermost layers of the tumor. Multiple studies have shown the presence of NK cells in solid tumors (e.g. melanoma, colon and breast malignancy).13-16 However, the number of NK cells and their penetration capacity AB05831 in the tumor is very heterogeneous among patients. Furthermore, different NK cell populations have been observed in solid tumors (e.g. CD56 bright vs CD56 dim NK cells), showing a positive or unfavorable correlation with individual end result depending on the specific NK cell populace observed.16 In addition, sound tumors generate a very harsh microenvironment characterized by hypoxia, nutrient starvation, waste product accumulation and pH gradients.17,18 These factors affect NK cell cytotoxic capacity as well as tumor cell sensitivity to NK cells.19 More specifically, hypoxia has been shown to reduce the expression of activating AB05831 receptors on NK cell surface (e.g. NKG2D, NKp46, NKp30 and NKp44).20,21 Likewise, hypoxia also downregulates the expression of NK cell ligands around the tumor cell membrane (e.g. MICA); rendering the tumor cells invisible to NK cells.22 Interestingly, hypoxia does not impact the ADCC response; CD16 expression levels remain constant under hypoxic conditions and NK cells continue destroying tumor cells if they are coated with antibodies.21 However, recent evidence suggests tumor cells have the potential to remove the antibody ligands as a camouflage mechanism.23 Finally, NK cells can be genetically engineered to thrive in the tumor.