Tumor-Associated Macrophages Enhance Tumor Invasiveness And Promote Cd4+T-Cell Recruitment In Gastric Cancer

Macrophages are one of the major populations of tumor-infiltrating immune cells. To date, several studies have shown that high tumor-associated-macrophages (TAMs) count (CD68high) or high CD68/CD3 ratio in gastric cancer patients is associated to poor surgical outcome or shorter overall survival. Other than macrophages, high CD4+ T cells (i.e. T helper 2 cells and regulatory T cells) are also associated to poor prognosis. Nevertheless, it is still unclear how these immune cells are recruited to the gastric tumor microenvironment and contributing to cancer progression. We hypothesize that gastric tumor secretes soluble factors to recruit monocytes to the lesion and drives their differentiation into tumor macrophages. These TAMs in turn assist pro-tumor immune-cell infiltration and tumor progression. Gastric cancer (GC) cell lines representing intestinal (OCUM-1, NCI-N87, MKN28, MKN45, SNU-5) and diffuse (AZ521, TMK-1, IST-1, CLS145 and YCC18) subtypes were used. CD14+ monocytes, CD4+CD45RA+ and CD4+CD45RO+ T cells were isolated from healthy blood donors and used to examine the gastric tumor ability to recruit these cells. Normal macrophages (MΦ) and GC-macrophages (GC-MΦ) were derived from CD14+ monocytes respectively with macrophage colony-stimulating factor and various gastric tumor-derived soluble factors. Phenotypes of the derived macrophages were examined by cytokine profiling and cell surface marker expression using flow cytometry. Various GC-MΦ secreted factors were also utilized to study the recruitment of CD4+CD45RA+ and CD4+CD45RO+ T cells. To examine the effect of the GC-MΦ on tumor proliferation and invasiveness, gastric tumor cells were conditioned with various GC-MΦ secreted factors prior to MTS assay and transwell assay. Indeed, we observed a significant 2-fold increase in the migration of CD14+ monocytes towards the various gastric tumor-derived soluble factors. We also observed that gastric tumor-derived soluble factors conditioned monocytes towards M2 phenotype macrophages (CD68+/CD206+/CD163+/IL10high/IL12low). Importantly, soluble factors derived from NCI-N87 and TMK-1-conditioned MΦ, but not normal MΦ, significantly increased the proliferation of NCI-N87 and TMK-1 gastric tumor cells by 26.7±10.4% and 14.5±4.0% respectively. In addition, transmigration of these gastric tumor cells were increased following exposure to their respective GC-MΦ derived soluble factors. Furthermore, we observed that the soluble factors derived from GC-MΦ, but not MΦ, were capable of enhancing the recruitment of both CD4+CD45RO+ (memory) and CD4+CD45RA+ (naive) T cells. Notably, the memory T cells were more responsive towards the GC-MΦ-derived soluble factors compared to naive T cells. Taken together, our findings suggest that gastric tumor secretes factors that attract CD14+ monocytes into the tumor lesion and subsequently changing them to M2-like macrophages with pro-tumor properties. These TAMs in turn enhance CD4+ T cell recruitment as well as gastric tumor cell invasion and proliferation. This, therefore, indicates a gastric tumor-dependent influence on immune cells to assist gastric tumor progression. Citation Format: Pin Yan Low, Yaw Chyn Lim, Wei Peng Yong, Bok Yan, Jimmy So. Tumor-associated macrophages enhance tumor invasiveness and promote CD4+ T-cell recruitment in Gastric Cancer. [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 C36.