Triple-negative breast cancer (TNBC) affects about 20% of breast cancer patients and is characterized by a high recurrence rate within three years. Sadly, after recurrence, survival rates for TNBC are lower than other breast cancer types. In addition, hormone therapy and the molecular targeted drug Herceptin therapy, which are commonly used in the treatment of breast cancer, are less effective, making treatment difficult in many cases and causing many patients to suffer from side effects from anticancer drugs.
Fucoidan, on the other hand, is a complex carbohydrate extracted from brown seaweed, and it is distinguished by its sulfated structure. Research has shown that this substance triggers programmed cell death (apoptosis) in multiple breast cancer cell lines without any reported adverse effects.
In this blog, I would like to share the following study: “Fucoidan inhibits epithelial-to-mesenchymal transition via regulation of the HIF-1α pathway in mammary cancer cells under hypoxia” by Weiwei Li et al. The study investigated the effect of fucoidan on epithelial-mesenchymal transition (EMT) of human triple-negative breast cancer (TNBC) cell lines in a hypoxic microenvironment.
The study focused on exploring how fucoidan influences epithelial-mesenchymal transition (EMT) and the underlying molecular mechanisms in triple-negative breast cancer cell lines, specifically MDA-MB-231 cells. This research also aimed to determine if fucoidan directly impacts the expression of hypoxia-inducible factor-1α (HIF-1α) in triple-negative breast cancer (TNBC) cell lines, since hypoxia can enhance epithelial-mesenchymal transition (EMT) in these cells by modifying the expression of EMT regulators.
First, the effect of fucoidan on breast cancer cell proliferation under hypoxic conditions was measured using MTT assay. Figure 1 shows that fucoidan treatment reduced breast cancer cell proliferation (P<0.05). After treatment with fucoidan at a concentration of 25µg/ml for 72 hours, cell proliferation was reduced by 53.2%. This indicated that fucoidan treatment inhibited MDA-MB-231 cell proliferation under hypoxic conditions.
In the next stage of the experiment, wound healing assays and Matrigel-coated transwell invasion assays were conducted to determine if fucoidan intervention exhibited any effect on the epithelial-mesenchymal transition (EMT) of MDA-MB-231 cells. As shown in Figure 2A, fucoidan treatment reduced the migration ability of breast cancer cells (P<0.001). After treatment with fucoidan at a concentration of 12.5 or 25µg/ml for 24 hours, cell invasion was still inhibited (Figure 2B). Fucoidan inhibits breast cancer cell migration and invasion.
Fucoidan treatment reduced the expression levels of mesenchymal markers N-cadherin and vimentin. The epithelial markers ZO-1 and E-cadherin were barely expressed in the membrane of MDA-MB-231 cells, but the expression levels of ZO-1 and E-cadherin were significantly increased after fucoidan treatment, which found that fucoidan affects the expression levels of EMT markers.
The study examined the impact of fucoidan on HIF-1α expression and its transfer to the nucleus using Western blotting. When fucoidan was administered in a low-oxygen environment, the amount of HIF-1α protein in the nucleus was reduced, as depicted in Figure 3A. Activation of HIF-1α was also evaluated. As shown in Figure 3B, the administration of fucoidan reduced HIF-1α activation. This indicates that fucoidan inhibits nuclear protein accumulation and HIF-1α activation.
Hypoxia-inducible factor (HIF) can regulate the expression of numerous genes involved in tumor growth, metastasis, and metabolic reprogramming. HIF-1α can directly or indirectly regulate the EMT regulators TWIST, Snail, CAIX, and GLUT-1. The expression levels of TWIST, Snail, CAIX, and GLUT-1 were detected by Western blotting. As shown in Figure. 4, following, fucoidan treatment suppressed the expression levels of TWIST-1, Snail, CAIX, and GLUT-1 which indicated fucoidan down-regulates the expression of HIF-1α target genes.
To investigate whether the anti-EMT effect of fucoidan depends on HIF-1α, HIF-1α overexpression plasmid (HIF-1α-pcDNA3.0) was transfected into MDA-MB-231 cells. Empty plasmid (pcDNA3.0) was used as a control. The expression of HIF-1α and its target genes (TWIST and Snail) in MDA-MB-231 cells was increased by HIF-1α-pcDNA3.0 transfection. The mRNA expression level of HIF-1α in breast cancer cells was measured by RT-qPCR. After transfection with HIF-1α-pcDNA3.0, the mRNA expression level of HIF-1α was increased. After transfection with HIF-1α-pcDNA3.0, breast cancer cells were incubated with 25 µg/ml fucoidan to detect cell invasion and migration. Wound healing assays and Matrigel-coated transwell invasion assays revealed that overexpression of HIF-1α reversed the fucoidan-induced inhibition of cell migration and invasion.
The research uncovered a previously unknown mechanism by which fucoidan suppresses breast cancer metastasis and EMT through the HIF-1α signaling pathway. Fucoidan prevented HIF-1α from being activated and entering the nucleus, and reduced the levels of EMT regulators downstream of HIF-1α, such as the transcription factors TWIST, Snail, CAIX, and GLUT-1. (Glucose transporters-1) The study found that under low oxygen conditions, the levels of N-cadherin and vimentin, proteins associated with EMT, were reduced in TNBC cells. Simultaneously, the levels of ZO-1 and E-cadherin, markers of cell-cell adhesion, were increased. This suggests that hypoxia inhibits EMT and migration in TNBC cells. Thus, this study provides new insights into the mechanism of fucoidan in breast cancer treatment.
Source: Oncol Lett. 2019 Jul; 18(1): 330–338. doi: 10.3892/ol.2019.10283