Knockdown of UGDH facilitates epirubicin resistance in MDA-MB-231 cells by regulation of hyaluronan synthesis.

VITALE, DAIANA L.; SPINELLI, FIORELLA M.; SEVIC, INA; CAON, ILARIA; DEMARCHI, GIANINA; VALLA, SOFÍA; CRISTINA, CAROLINA; VIGETTI, DAVIDE; ALANIZ, LAURA

Ciudad Autónoma de Buenos Aires

Simposio; Glycobiology Symposium "GLYCOAR 2019"; 2019

Epirubicin (EPI) is a drug eliminated via glucuronidation by binding to UDP-glucuronic acid, a UDP-sugar formed by the UDP-glucose dehydrogenase enzyme (UGDH) and precursor of glycosaminoglycans like hyaluronan (HA). Since the role of glucuronidation in breast cancer has not been studied yet, the aim was to evaluate the effect of silencing UGDH on EPI response and HA synthesis in MDA-MB-231 cells. The cells were transfected with UGDH-siRNA by electroporation, and after 24h treated with EPI for another 24h. Apoptosis and EPI accumulation were detected by FACS. Cell migration was studied by scratch assay. UGT2B7, ABC-pumps, VEGF, EGF and HAS mRNAs were detected by RT-qPCR. ECM was analyzed by a particle exclusion assay. Secreted HA and VEGF were studied by ELISA and p-Akt expression by Western Blot. Lactate dehydrogenase (LDH) activity and pH were detected by colorimetric assays. Higher EPI accumulation was observed in cells transfected with UGDH-siRNA, but apoptosis was significantly lower. We observed higher expression of ABC-pumps, VEGF, EGF, UGT2B7 and p-Akt after knocked-down UGDH + EPI treatment. Tumor cells increased their migration and ECM, in line with higher HAS-3 mRNA levels. LDH activity was increased according to a decrease in pH of cell supernatants. Knockdown of UGDH increased EPI availability in tumor cells. However, we found an opposite effect in response to EPI accumulation as consequence of mechanisms that involve drug efflux, migration and survival.