||HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) exhibits anti-tumor effects on various types of human cancers and is now approved by U.S FDA for clinical cancer treatment. SAHA also suppresses tumor angiogenesis. However, the effect of SAHA on tumor lymphangiogeneis is unclear. My study focuses on anti-lymphangiogenic action of SAHA on breast cancers and contains three parts. In part one, I test whether SAHA affects the production of pro-lymphangiogenesis factor such as VEGF-C to reduce proliferation and migration of lymphatic endothelial cells toward cancer cells. I found that SAHA does-dependently inhibited the expression of VEGF-C in various breast cancer cell lines and the secretion of VEGF-C into conditioned medium was also suppressed. Furthermore, I cloned human VEGF-C gene promoter and demonstrated that SAHA directly suppressed VEGF-C transcription via the -185/+38 promoter region in a Sp1-mediated manner.|
In part two, I aim to study the effect of SAHA on lymphatics endothelial cells (LECs). I established a lymphatic-like endothelial cell line (named as FP01) by overexpressing the master LEC transcription factor PROX1 in EA.hy926 endothelial cells. This cell lines showed similar gene expression pattern and phenotype of primarily cultured LECs. I found that SAHA can suppress proliferation, sprouting and tube formation of LECs. Moreover, SAHA could attenuate the angiopoietin/Tie signaling pathway which is important in the regulation of LEC function. The promoter activity assay revealed that SAHA down-regulated the expression of Tie2 through transcriptional repression. Interestingly, I also found that SAHA could quickly induce the expression of c-Cbl, the E3 ligase for Tie2 ubiquitination leading to Tie2 protein degradation. Knockdown of c-Cbl effectively reversed SAHA-induced Tie2 protein degradation.
In part three, I used breast cancer xenograft model to demonstrate whether SAHA could repress lymphangiogenesis and lymphatic metastasis in vivo. SAHA indeed inhibited tumor formation, lymphangiogenesis and metastasis in MDA-MB-231 luciferase-tagged xenograft model.
Taken together, SAHA not only suppresses proliferation, sprouting and tube formation of LECs and attenuates the Ang/Tie signaling in LECs by down-regulating Tie2 via transcriptional and post-transcriptional mechanism but also inhibits VEGF-C expression in breast cancer cells via transcriptional repression. Breast cancer xenograft model demonstrates that SAHA inhibits tumor formation, lymphangiogenesis and metastasis in vivo. Collectively, this drug exerts anti-lymphangiogenic activity in cancer treatment.