Abstract P2-01-19: Sphingosine-1-phosphate affects tumor-associated macrophages in breast cancer patients
Authors: Tsuchida J , Nagahashi M , Moro K , Ikarashi M , Koyama Y , Sakata J , Kobayashi T , Kameyama H , Qi Q , Yan L , Takabe K , Wakai T 
Published: May 2019

Abstract
Background: Tumor-associated macrophages (TAMs) are considered to be one of the key players in the tumor microenvironment, which regulates cancer invasion and metastases. TAMs can be divided into two phenotypes with opposite functions. While M1 macrophages are known to exert anti-tumor activity by promoting pro-inflammatory effects and immune responses such as intereukin-6 (IL-6) and tumor necrosis factor-α (TNF-α), M2 macrophages influence an anti-inflammatory response, wound healing, and pro-tumorigenic properties. A bioactive lipid mediator, sphingosine-1-phosphate (S1P) has emerged as a key regulatory molecule in cancer progression. We previously demonstrated that S1P generated by sphingosine kinase 1 (SPHK1), is a crucial mediator of breast cancer-induced angiogenesis and lymphangiogenesis, and promotes its metastasis. In particular, we found that SPHK1 is highly expressed in HER2 negative breast cancer, and the patients who developed lymph node metastasis demonstrated significantly higher levels of S1P (J Surg Res 2016). Although we have previously reported the role of S1P in recruitment of TAMs in vivo (Cancer Res 2018), its relevance in patients is yet to be uncovered. Here, we test our hypothesis that S1P signaling affects TAMs in human patients with breast cancer.
Materials and Methods: The expression level of each enzyme-encoding gene involved in S1P production was evaluated by retrieving RNA sequencing and gene expression quantification data using the Genomics Data Commons (GDC) data portal of the The Cancer Genome Atlas cohort. Gene expression levels were derived using normalization methods provided in the DESeq2 package. We compared the difference in expression levels of tumor associated macrophage related genes, including CD68, CD163, IL-6, andTNF-α between SPHK1-high breast tissue, and SPHK1-low breast tissue in the group of HER2 negative or positive patients. Unpaired t-tests were performed to compare expression differences between SPHK1-high and SPHK1-low breast tissue. All tests were two-sided and P values &lt; 0.05 were considered statistically significant.
Results: CD68, pan-macrophage marker, is significantly increased in SPHK1-high breast cancer tissues both in HER2 negative and positive breast cancer patients (p=&lt;0.001, &lt;0.01). CD163 which is a scavenger receptor that is regarded as highly specific for M2 macrophages is significantly increased in SPHK1-high breast cancer tissues in HER2 negative breast cancer patients, but not in HER2 positive breast cancer patients (p=&lt;0.001, 0.2). IL-6, which characterize M1 phenotype is significantly increased in SPHK1-high breast cancer tissues both in HER2 negative and positive breast cancer patients (p=&lt;0.001, &lt;0.001). TNF-α, which also characterizes M1 phenotype, is significantly increased in SPHK1-high breast cancer tissues in HER2 negative breast cancer patients, but not in HER2 positive breast cancer patients (p=&lt;0.001, 0.05).
Conclusion: Our results suggest that S1P affects TAMs in breast cancer patients, which implicate the important roles of S1P in the complicated immune system related to tumor progression. Our results also indicate that S1P have a large role in HER2 negative breast cancer patients. Further investigations are needed to understand the underlying mechanisms.
Citation Format: Tsuchida J, Nagahashi M, Moro K, Ikarashi M, Koyama Y, Sakata J, Kobayashi T, Kameyama H, Qi Q, Yan L, Takabe K, Wakai T. Sphingosine-1-phosphate affects tumor-associated macrophages in breast cancer patients [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P2-01-19.