p62 Suppressed VK3-induced Oxidative Damage Through Keap1/Nrf2 Pathway In Human Ovarian Cancer Cells.

Imbalance of redox homeostasis may be responsible for the resistance of cancer to chemotherapy. Currently, increasing studies demonstrated that vitamin K3 (VK3), which promoted the production of ROS, had potential to be developed as an anti-tumor agent. We found SKOV3/DDP cells with high levels of p62 were insensitive to VK3 compared with SKOV3 cells. Furthermore, Nrf2 downstream antioxidant genes such as HO-1(heme oxygenase 1) and NQO1 (NAD (P) H: quinone oxidoreductase 1) were upregulated in SKOV3/DDP cells with VK3 treatment, which indicated VK3 activated Nrf2 signaling in SKOV3/DDP cells. Moreover, co-localization of p62 and Keap1 was also observed. Suppression of p62 expression increased the apoptosis induced by VK3, and the expression of Nrf2, HO-1 and NQO1 were all downregulated in SKOV3/DDP cells. Our results suggested that overexpressed p62 may protect cells from oxidative damage caused by VK3 through activating Keap1/Nrf2 signaling in ovarian cancer.

Glutamate in cancers: from metabolism to signaling.

Glutamine and glutamate are major bioenergy substrates for normal and cancer cell growth. Cancer cells need more biofuel than normal tissues for energy supply, anti-oxidation activity and biomass production. Genes related to metabolic chains in many cancers are somehow mutated, which makes cancer cells more glutamate dependent. Meanwhile, glutamate is an excitatory neurotransmitter for conducting signals through binding with different types of receptors in central neuron system. Interestingly, increasing evidences have shown involvement of glutamate signaling, guided through their receptors, in human malignancy. Dysregulation of glutamate transporters, such as excitatory amino acid transporter and cystine/glutamate antiporter system, also generates excessive extracellular glutamate, which in turn, activates glutamate receptors on cancer cells and results in malignant growth. These features make glutamate an attractive target for anti-cancer drug development with some glutamate targeted but blood brain barrier impermeable anti-psychosis drugs under consideration. We discussed the relevant progressions and drawbacks in this field herein.

R-Spondin1/LGR5 Activates TGFß Signaling and Suppresses Colon Cancer Metastasis.

Leucine-rich repeat containing G-protein-coupled receptor 5 (LGR5), an intestinal stem cell marker, is known to exhibit tumor suppressor activity in colon cancer, the mechanism of which is not understood. Here we show that R-spondin 1 (RSPO1)/LGR5 directly activates TGFß signaling cooperatively with TGFß type II receptor in colon cancer cells, enhancing TGFß-mediated growth inhibition and stress-induced apoptosis. Knockdown of LGR5 attenuated downstream TGFß signaling and increased cell proliferation, survival, and metastasis in an orthotopic model of colon cancer in vivo Upon RSPO1 stimulation, LGR5 formed complexes with TGFß receptors. Studies of patient specimens indicate that LGR5 expression was reduced in advanced stages and positively correlated with markers of TGFß activation in colon cancer. Our study uncovers a novel cross-talk between LGR5 and TGFß signaling in colon cancer and identifies LGR5 as a new modulator of TGFß signaling able to suppress colon cancer metastasis. Cancer Res; 77(23); 6589-602. ©2017 AACR.

Transforming Growth Factor ß Mediates Drug Resistance by Regulating the Expression of Pyruvate Dehydrogenase Kinase 4 in Colorectal Cancer.

Drug resistance is one of the main causes of colon cancer recurrence. However, our understanding of the underlying mechanisms and availability of therapeutic options remains limited. Here we show that expression of pyruvate dehydrogenase kinase 4 (PDK4) is positively correlated with drug resistance of colon cancer cells and induced by 5-fluorouracil (5-FU) treatment in drug-resistant but not drug-sensitive cells. Knockdown of PDK4 expression sensitizes colon cancer cells to 5-FU or oxaliplatin-induced apoptosis in vitro and increases the effectiveness of 5-FU in the inhibition of tumor growth in a mouse xenograft model in vivo In addition, we demonstrate for the first time that TGFß mediates drug resistance by regulating PDK4 expression and that 5-FU induces PDK4 expression in a TGFß signaling-dependent manner. Mechanistically, knockdown or inhibition of PDK4 significantly increases the inhibitory effect of 5-FU on expression of the anti-apoptotic factors Bcl-2 and survivin. Importantly, studies of patient samples indicate that expression of PDK4 and phosphorylation of Smad2, an indicator of TGFß pathway activation, show a strong correlation and that both positively associate with chemoresistance in colorectal cancer. These findings indicate that the TGFß/PDK4 signaling axis plays an important role in the response of colorectal cancer to chemotherapy. A major implication of our studies is that inhibition of PDK4 may have considerable therapeutic potential to overcome drug resistance in colorectal cancer patients, which warrants the development of PDK4-specific inhibitors.

The BH3 mimetic S1 induces endoplasmic reticulum stress-associated apoptosis in cisplatin-resistant human ovarian cancer cells although it activates autophagy.

SKOV3/DDP human ovarian cancer cells have been shown to be resistant to cisplatin. Although the BH3 mimetic S1 induces cell death in several types of tumor cells, it is unclear whether it induces death in drug-resistant cells. Herein, we found that S1 induced endoplasmic reticulum (ER) stress-associated apoptosis in both SKOV3 and SKOV3/DDP cells. S1 activated autophagy at early time points in SKOV3/DDP cells, and inhibition of autophagy increased ER stress-associated apoptosis. Collectively, our data indicate that autophagy plays a protective role, but it cannot protect against S1-induced cell death in cisplatin-resistant SKOV3/DDP cells.

Regulation of endoplasmic reticulum stress in rat cortex by p62/ZIP through the Keap1-Nrf2-ARE signalling pathway after transient focal cerebral ischaemia.

PRIMARY OBJECTIVE: p62/ZIP as the autophagy receptor can transport the misfolded proteins to a macroautophagy-lysosome system for degradation and also create a positive feedback loop between p62/ZIP and Nrf2. However, the role of p62/ZIP on cerebral ischaemia is unclear. The aim of this study was to evaluate the role of p62/ZIP in the regulation of endoplasmic reticulum(ER) stress induced by cerebral ischaemia/reperfusion. RESEARCH DESIGN: Different ischemic periods were designed by transient middle cerebral artery occlusion (tMCAO) using the suture method. METHODS AND PROCEDURES: At 24 hours after reperfusion, the ischaemic brain tissue was studied histologically and biochemically for autophagic, ER stress and Keap1-Nrf2-ARE signalling pathway markers. MAIN OUTCOMES AND RESULTS: Prolongation of ischaemia significantly increased the cortical injury observed in rats and was associated with a gradual increase in the protein expression of ubiquitin-aggregates, Grp78, GADD153/CHOP and p62/ZIP. Autophagy marker Atg12-Atg5 and LC3-PE increased and then decreased. Moreover, p62/ZIP mRNA expression increased and then decreased and was consistent with Nrf2 activation. CONCLUSIONS: p62/ZIP not only plays a key role in scavenging protein aggregates during autophagy, but it may also be involved in preventing oxidative injury and alleviating ER stress through the Keap1-Nrf2-ARE signalling pathway during cerebral ischaemia/reperfusion injury.

Inhibition of CLIC4 enhances autophagy and triggers mitochondrial and ER stress-induced apoptosis in human glioma U251 cells under starvation.

CLIC4/mtCLIC, a chloride intracellular channel protein, localizes to mitochondria, endoplasmic reticulum (ER), nucleus and cytoplasm, and participates in the apoptotic response to stress. Apoptosis and autophagy, the main types of the programmed cell death, seem interconnected under certain stress conditions. However, the role of CLIC4 in autophagy regulation has yet to be determined. In this study, we demonstrate upregulation and nuclear translocation of the CLIC4 protein following starvation in U251 cells. CLIC4 siRNA transfection enhanced autophagy with increased LC3-II protein and puncta accumulation in U251 cells under starvation conditions. In that condition, the interaction of the 14-3-3 epsilon isoform with CLIC4 was abolished and resulted in Beclin 1 overactivation, which further activated autophagy. Moreover, inhibiting the expression of CLIC4 triggered both mitochondrial apoptosis involved in Bax/Bcl-2 and cytochrome c release under starvation and endoplasmic reticulum stress-induced apoptosis with CHOP and caspase-4 upregulation. These results demonstrate that CLIC4 nuclear translocation is an integral part of the cellular response to starvation. Inhibiting the expression of CLIC4 enhances autophagy and contributes to mitochondrial and ER stress-induced apoptosis under starvation.

The BH3 mimetic S1 induces autophagy through ER stress and disruption of Bcl-2/Beclin 1 interaction in human glioma U251 cells.

Previous results showed that a novel BH3 mimetic S1 could induce cell death in a wide range of cancer types in vitro through Bax/Bak-dependent apoptosis. We demonstrated that in addition to mitochondrial pathway apoptosis, endoplasmic reticulum (ER) stress-associated apoptosis was also induced by S1. Moreover, S1 can induce autophagy in U251 cells, which may occur through ER stress and disruption of the association of Bcl-2 and Beclin 1. Inhibition of autophagy by the autophagic inhibitors 3-methyladenine (3-MA) or chloroquine (CQ) increased S1-induced apoptosis. In conclusion, autophagy plays an important role in S1-induced U251 cell death.

p62/SQSTM1 involved in cisplatin resistance in human ovarian cancer cells by clearing ubiquitinated proteins.

Mechanisms of cisplatin resistance in cancer cells are not fully understood. Here, we show a critical role for the ubiquitin-binding protein p62/SQSTM1 in cisplatin resistance in human ovarian cancer cells (HOCCs). Specifically, we found that cisplatin-resistant SKOV3/DDP cells express much higher levels of p62 than do cisplatin-sensitive SKOV3 cells. The protein p62 binds ubiquitinated proteins for transport to autophagic degradation, reducing apoptosis induced by endoplasmic reticulum (ER) stress in SKOV3/DDP cells. Knockdown of p62 or inhibition of autophagy using 3-methyladenine resensitises SKOV3/DDP cells to cisplatin. Collectively, our data indicate that p62 acts as a receptor or adaptor for autophagic degradation of ubiquitinated proteins, and plays an important role in preventing ER stress-induced apoptosis, leading to cisplatin resistance in HOCCs.