Mupirocin-mediated downregulation of claudin-14 enhances chemosensitivity in human colorectal cancer cells.

Claudin-14 (CLDN14), a tight junction protein, contributes to cell proliferation and chemoresistance in human colorectal cancer (CRC)-derived DLD-1 cells. However, small molecules targeting CLDN14 remain unexplored. Here, we identified mupirocin (MUP), a clinically approved topical antibiotic, as a modulator of CLDN14 protein expression. Quartz crystal microbalance analysis revealed interaction between MUP and recombinant CLDN14 protein with a dissociation constant (Kd) of 2.59 ± 0.54 μM. MUP did not alter CLDN14 mRNA levels but reduced CLDN14 protein stability. Pharmacological inhibition of clathrin-mediated endocytosis and lysosomal degradation significantly reversed the MUP-induced reduction of CLDN14 protein. These results suggest MUP accelerates endocytosis-lysosomal degradation of CLDN14 protein. Other antibiotics failed to decrease CLDN14 expression. Functionally, MUP increased paracellular permeability to mineral ions and enhanced the transepithelial flux of doxorubicin (DXR), an anthracycline anticancer drug, and lucifer yellow, an aqueous small compound. In DLD-1 spheroids, MUP reduced intracellular oxidative stress and nuclear factor erythroid 2-related factor 2 (Nrf2) expression. Consequently, MUP promoted intracellular accumulation of DXR and significantly potentiated its cytotoxic effects in spheroids. Moreover, MUP enhanced the antitumor efficacy of other chemotherapeutic agent oxaliplatin. Sulforaphane, an Nrf2 activator, attenuated the MUP-induced enhancement of anticancer efficacy. These findings suggest that MUP enhances anticancer drug sensitivity in CRC through lysosome-dependent downregulation of CLDN14 protein and suppression of oxidative stress signaling.