Probing the Anti-Cancer Potency of Sulfated Galactans on Cholangiocarcinoma Cells Using Synchrotron FTIR Microspectroscopy, Molecular Docking, and In Vitro Studies

Sulfated galactans (SG) isolated from red alga Gracilaria fisheri have been reported to inhibit the growth of cholangiocarcinoma (CCA) cells, which was similar to the epidermal growth factor receptor (EGFR)-targeted drug, cetuximab. Herein, we studied the anti-cancer potency of SG compared to cetuximab. Biological studies demonstrated SG and cetuximab had similar inhibition mechanisms in CCA cells by down-regulating EGFR/ERK pathway, and the combined treatment induced a greater inhibition effect. The molecular docking study revealed that SG binds to the dimerization domain of EGFR, and this was confirmed by dimerization assay, which showed that SG inhibited ligand-induced EGFR dimer formation. Synchrotron FTIR microspectroscopy was employed to examine alterations in cellular macromolecules after drug treatment. The SR-FTIR-MS elicited similar spectral signatures of SG and cetuximab, pointing towards the bands of RNA/DNA, lipids, and amide I vibrations, which were inconsistent with the changes of signaling proteins in CCA cells after drug treatment. Thus, this study demonstrates the underlined anti-cancer mechanism of SG by interfering with EGFR dimerization. In addition, we reveal that FTIR signature spectra offer a useful tool for screening anti-cancer drugs’ effect.

Antibacterial Activity of Sulfated Galactans from Eucheuma serra and Gracilari verrucosa against Diarrheagenic Escherichia coli via the Disruption of the Cell Membrane Structure

Seaweed sulfated polysaccharides have attracted significant attention due to their antibacterial activity. This work investigated the antibacterial activity and mechanism of depolymerized sulfated galactans from Eucheuma serra (E. serra) and Gracilaria verrucosa (G. verrucosa) against enterotoxigenic Escherichia coli (ETEC) K88. The results show that removing the metal ions improves the anti-ETEC K88 activity of the galactans. The fluorescence labeling study confirmed that the sulfated galactans penetrated the cell walls and eventually reached the interior of the ETEC K88. Nucleic acid staining and intracellular protein leakage were also observed, indicating the destruction of permeability and integrity of the cell membrane. Interestingly, the two polysaccharides exhibited no effect on the proliferation of the selected Gram-positive bacteria and yeast. This indicates that the cell wall structure of the microorganisms could influence the bacteriostatic activity of the sulfated polysaccharides, as well. These results suggest that the sulfated seaweed polysaccharides might have potential application value in antibacterial diarrhea.

Immunostimulatory Effect of Sulfated Galactans from the Green Seaweed Caulerpa cupressoides var. flabellata

Sulfated polysaccharides (SPs) obtained from green seaweeds are structurally heterogeneous molecules with multifunctional bioactivities. In this work, two sulfated and pyruvated galactans were purified from Caulerpa cupressoides var. flabellata (named SP1 and SP2), and their immunostimulatory effect was evaluated using cultured murine macrophage cells. Both SPs equally increased the production of nitric oxide, reactive oxygen species, and the proinflammatory cytokines TNF-α and IL-6. NMR spectroscopy revealed that both galactans were composed primarily of 3)-β-d-Galp-(1→3) units. Pyruvate groups were also found, forming five-membered cyclic ketals as 4,6-O-(1’carboxy)-ethylidene-β-d-Galp residues. Some galactoses are sulfated at C-2. In addition, only SP2 showed some galactose units sulfated at C-4, indicating that sulfation at this position is not essential for the immunomodulatory activity of these galactans. Overall, the data showed that the galactans of C. cupressoides exhibited immunostimulating activity with potential therapeutic applications, which can be used in the development of new biomedical products.

Inhibitory Effect of Depolymerized Sulfated Galactans from Marine Red Algae on the Growth and Adhesion of Diarrheagenic Escherichia coli

Active polysaccharides as safe and natural polymers against bacterial diarrhea have been reconsidered as an alternative to antibiotics. This work investigated the inhibiting effect of depolymerized sulfated galactans from Eucheuma serra and Gracilaria verrucosa on the growth and adhesion of diarrheagenic enterotoxigenic Escherichia coli (ETEC) K88. Results showed that the sulfated polysaccharides with molecular weight distribution ≤20.0 kDa exhibited antibacterial activity against ETEC K88. A structure–activity study revealed that the anti-ETEC K88 activity of sulfated polysaccharides is strictly determined by their molecular weight distribution, sulfate group content, and monosaccharide composition. In addition, the promoted nucleic acid release and the fluorescence quenching of membrane proteins were observed after the treatment with selected polysaccharides. Scanning electron microscopy further confirmed that the depolymerized sulfated galactans can effectively inhibit ETEC K88 adhesion. In conclusion, depolymerized sulfated galactans exhibited an inhibitory effect on the growth and adhesion of ETEC K88.

Anti-Thrombin, Anti-Adhesive, Anti-Migratory, and Anti-Proliferative Activities of Sulfated Galactans from the Tropical Green Seaweed, Udotea flabellum

In this study, sulfated polysaccharide-rich extracts were isolated from 22 tropical seaweeds (4 red, 11 brown, and 7 green) found in northeastern Brazil, and evaluated for the role of anticoagulant agents. Fifteen of the extracts showed anticoagulant activity, including all the extracts from green seaweeds. Udotea flabellum (a green seaweed) extract was the most potent, requiring an amount of only 3 µg to double the plasma coagulation time in the activated partial thromboplastin time test. A similar result was obtained with 1 µg of heparin. Two sulfated homogalactans with anticoagulant activity, F-I (130 kDa) and F-II (75 kDa), were isolated from this extract using several bio-guided purification steps. Their anticoagulant activity, as well as properties related to antitumor activity (anti-proliferative, anti-adhesive, and anti-migratory), were accessed. Their anticoagulant activities were close to that of heparin. We found that F-I and F-II (0.5–10 μg/mL) were not able to directly inhibit thrombin. In the presence of anti-thrombin, F-I (0.5 μg/mL) was more effective than heparin (0.5 μg/mL) in inhibiting thrombin, while F-II showed similar effects as heparin. F-I and F-II also inhibited B16-F10 (murine melanoma cells) adhesion, migration, and proliferation on a fibronectin-coated surface, but not on laminin- or collagen I-coated surfaces. Except for the antiproliferative activity, the other effects of F-I and F-II were eliminated upon their desulfation (~50%), indicating that the degree of sulfation is not as important for F-I and F-II anti-proliferative activity as the sulfation position. Taken together, the results provide strong evidence for the potential utility of sulfated galactans from U. flabellum, making these compounds an interesting option for future investigations that aim to design new anticoagulant/antitumor agents.