Anti-angiogenic action of leukotriene-C4 induced 15-hydroxyprostaglandin dehydrogenase in colon cancer cells is a TNF-α dependent phenomenon
Authors: S. Satapathy, A. Sjolander
Journal: Annals of Oncology (2018)

Introduction: Colorectal cancer (CRC) is one of the leading causes of cancer-related deaths worldwide. Cyclooxygenase-2 (COX-2), which plays a key role in the biosynthesis of prostaglandin E2 (PGE2), is often up-regulated in CRC and in other types of cancer. PGE2 induces angiogenesis and tumor cell survival, proliferation and migration. The tumor suppressor 15-hydroxyprostaglandin dehydrogenase (15-PGDH) is a key enzyme in PGE2 catabolism, converting it into its inactive metabolite 15-keto-PGE2 and is often down-regulated in cancer. Interesting enough, CRC patients expressing high levels of Cysteinyl leukotriene receptor 2 (CysLTR2) have a good prognosis and therefore, we investigated a potential link between CysLTR2-signaling and the tumor suppressor 15-PGDH in colon cancer cells. In the other hand, TNF-α is considered as the main regulator of COX-2 and mPGES-1 that contribute to the increased synthesis of PGE2, which is inhibited by overexpressed 15-PGDH. Level of the pro-tumorigenic PGE2 is increased in CRC, previously attributed to increased production via TNF-α mediated COX-2 up-regulation but more recently attributed to decreased catabolism due to down-regulation of 15-PGDH. Methods: HT-29 and Caco-2 cells were employed for this study. Unstimulated and LTC4 stimulated cells were compared for various markers through qRT-PCR as well as immunoblotting analysis. Vibrant-DiI labeled HT-29 cells were used for the zebrafish metastasis model. Visualization of the metastatic spread was observed using the light sheet microscopy (SPIM) and tissue sections were analyzed by immunohistochemistry for specific markers. Conditioned media from the unstimulated and LTC4 stimulated HT-29 cells were used to analyze the endothelial tube formation in HUVEC. Results: Elevation of 15-PGDH expression by leukotriene C4 (LTC4), a CysLTR2 ligand, exhibited anti-tumor activity in colon cancer cells with significant phosphorylation of β-catenin and down-regulation of anti-apoptotic marker Bcl-2 with concurrent activation of CASPASE-3 expression. We also observed a dramatic down-regulation of TNF-α on mRNA level and NF-κβ on both mRNA as well as protein level with LTC4 induced 15-PGDH in a TNF-α dependent manner. Moreover, TNF-α regulated anti-angiogenic action of 15-PGDH in HT-29 and Caco-2 colon cancer cells by depleting the mRNA level of MMP-2 and MMP-9 and protein level of VEGFR-1. Furthermore, we also observed disrupted tube formation in HUVEC with LTC4 induced 15-PGDH which is also governed by TNF-α. The angiogenesis study in transgenic zebrafish, Tg(fli1a: EGFP) embryo model suggested significant decrease in early and late metastasis with distinct disruption in the intersegmental vessel in the LTC4 induced 15-PGDH treated group. IHC analysis of Ki-67 exhibited decrease with LTC4 and very interestingly CASPASE-3 showed increase in the stimulated group which supported our in vitro observation. Conclusion: Hence, restoration of 15-PGDH expression through CysLTR2-signaling promotes the anti-angiogenic action against colon cancer cells, indicating an anti-tumor as well as the anti-metastatic efficacy of CysLTR2-signaling.