Genipin-Induced Inhibition of Uncoupling Protein-2 Sensitizes Drug-Resistant Cancer Cells to Cytotoxic Agents

Abstract

Uncoupling protein-2 (UCP2) is known to suppress mitochondrial reactive oxygen species (ROS) production and is employed by drug-resistant cancer cells to mitigate oxidative stress. Using the drug-sensitive HL-60 cells and the drug-resistant MX2 subline as model systems, we show that genipin, a UCP2 inhibitor, sensitizes drug-resistant cells to cytotoxic agents. Increased MX2 cell death was observed upon co-treatment with genipin and different doses of menadione, doxorubicin, and epirubicin. DCFH-DA fluorimetry revealed that the increase in MX2 cell death was accompanied by enhanced cellular ROS levels. The drug-induced increase in ROS was linked to genipin-mediated inhibition of mitochondrial proton leak. State 4 and resting cellular respiratory rates were higher in the MX2 cells in comparison to the HL-60 cells, and the increased respiration was readily suppressed by genipin in the MX2 cells. UCP2 accounted for a remarkable 37% of the resting cellular oxygen consumption indicating that the MX2 cells are functionally reliant on this protein. Higher amounts of UCP2 protein were detected in the MX2 versus the HL-60 mitochondria. The observed effects of genipin were absent in the HL-60 cells pointing to the selectivity of this natural product for drug-resistant cells. The specificity of genipin for UCP2 was confirmed using CHO cells stably expressing UCP2 in which genipin induced an ∼22% decrease in state 4 respiration. These effects were absent in empty vector CHO cells expressing no UCP2. Thus, the chemical inhibition of UCP2 with genipin sensitizes multidrug-resistant cancer cells to cytotoxic agents.

Glutathionylation Acts as a Control Switch for Uncoupling Proteins UCP2 and UCP3

“…S3). Furthermore, we have previously used this range of menadione to induce increases in cellular ROS without inducing cell death (17,32). The regulation of UCP3 by glutathionylation and ROS was confirmed in WT primary myotubes using BioGEE and H 2 O 2 .…”

“…Immediately following H 2 O 2 exposure, WT cells were incubated for 30 min at 37°C in differentiating media containing 20 M dichlorofluorescein diacetate. Fluorescence was determined as described (17).…”

“…Indeed, UCP2 null mice on various genetic backgrounds exhibit oxidative stress in many tissue types and a decrease in the circulating glutathione (GSH)/glutathione disulfide ratio (2,12). Increased expression of UCP2 in cancer cells is associated with the acquisition of drug-resistant phenotypes, a phenomenon related to the ROS-quenching function of UCP2 (17,18). The absence of UCP3 in skeletal muscle increases oxidative damage and perturbs skeletal muscle metabolism (19).…”

UCP2 expression may represent a predictive marker of neoadjuvant chemotherapy effectiveness for locally advanced uterine cervical cancer

“…Loss of UCP2 function may increase ROS production (25), whereas UCP2 overexpression may promote cytoprotection by mitigating oxidative stress (26,27). Furthermore, UCP2 contributes to both carcinogenesis and chemoresistance (28,29). UCP2 is implicated in human colon carcinogenesis (24,30), while mitochondrial uncoupling by UCP2 induces pancreatic cancer cell resistance to gemcitabine (28).…”

“…UCP2 is implicated in human colon carcinogenesis (24,30), while mitochondrial uncoupling by UCP2 induces pancreatic cancer cell resistance to gemcitabine (28). Furthermore, inhibition of UCP2 by genipin sensitizes cancer cells to chemotherapeutic agents (28,29). These findings suggest that UCP2 represents a target for cancer treatment with chemotherapeutic agents that promote oxidative stress.…”