Enzymatic Digestion of Cell-surface Heparan Sulfate Alters the Radiation Response in Triple-negative Breast Cancer...

Radiation resistance represents a major challenge in the treatment of breast cancer. As heparan sulfate (HS) chains are known to contribute to tumorigenesis, we aimed to investigate the interplay between HS degradation and radiation response in triple-negative breast cancer (TNBC) cells.

HS chains were degraded in vitro as TNBC cells MDA-MB-231 and HCC1806 were treated with heparinase I and III. Subsequently, radioresistance was determined via colony formation assay after doses of 2, 4 and 6 Gy. Cell cycle profile, stem cell characteristics, expression of HS, activation of beta integrins, and apoptosis were determined by flow cytometry. Additionally, cell motility was analyzed via wound-healing assays, and expression and activation of FAK, CDK-6, Src, and Erk1/2 were quantified by western blot pre- and post-irradiation. Finally, the expression of cytokines was analyzed using a cytokine array.

Radiation promoted cell cycle changes, while heparinase treatment induced apoptosis in both cell lines. Colony formation assays showed significantly increased radio-resistance for both cell lines after degradation of HS. Cell migration was similarly upregulated after degradation of HS compared to controls. This effect was even more prominent after irradiation. Interestingly, FAK, a marker of radioresistance, was significantly activated in the heparinase-treated group. Additionally, we found Src to be dysregulated in MDA-MB-231 cells. Finally, we observed differential secretion of GRO, CXCL1, IGFBP1, IL8, Angiogenin, and Osteoprotegerin after HS degradation and radiotherapy.

Our results suggest an influence of HS chains on the development of radioresistance in TNBC.