Inhibition of Glutamine Uptake Resensitizes Paclitaxel Resistance in SKOV3-TR Ovarian Cancer Cell via mTORC1/S6K Signaling Pathway.

Ovarian cancer is a carcinoma that affects women and that has a high mortality rate. Overcoming paclitaxel resistance is important for clinical application. However, the effect of amino acid metabolism regulation on paclitaxel-resistant ovarian cancer is still unknown. In this study, the effect of an amino acid-deprived condition on paclitaxel resistance in paclitaxel-resistant SKOV3-TR cells was analyzed. We analyzed the cell viability of SKOV3-TR in culture conditions in which each of the 20 amino acids were deprived. As a result, the cell viability of the SKOV3-TR was significantly reduced in cultures deprived of arginine, glutamine, and lysine. Furthermore, we showed that the glutamine-deprived condition inhibited mTORC1/S6K signaling. The decreased cell viability and mTORC1/S6K signaling under glutamine-deprived conditions could be restored by glutamine and α-KG supplementation. Treatment with PF-4708671, a selective S6K inhibitor, and the selective glutamine transporter ASCT2 inhibitor V-9302 downregulated mTOR/S6K signaling and resensitized SKOV3-TR to paclitaxel. Immunoblotting showed the upregulation of Bcl-2 phosphorylation and a decrease in Mcl-1 expression in SKOV3-TR via the cotreatment of paclitaxel with PF-4708671 and V-9302. Collectively, this study demonstrates that the inhibition of glutamine uptake can resensitize SKOV3-TR to paclitaxel and represents a promising therapeutic target for overcoming paclitaxel resistance in ovarian cancer.

Lysine is required for growth factor-induced mTORC1 activation.

Mechanistic target of rapamycincomplex 1 (mTORC1) integrates various environmental signals to regulate cell growth and metabolism. mTORC1 activity is sensitive to changes in amino acid levels. Here, we investigated the effect of lysine on mTORC1 activity in non-small cell lung cancer (NSCLC) cells. Lysine deprivation suppressed mTORC1 activity and lysine replenishment restored the decreased mTORC1 activity in lysine-deprived cells. Supplementing growth factors, such as insulin growth factor-1 or insulin restored the decreased mTORC1 activity in serum-deprived cells. However, in serum/lysine-deprived cells, supplementing growth factors was not sufficient to restore mTORC1 activity, suggesting thatgrowth factors could not activate mTORC1 efficiently in the absence of lysine. General control nonderepressible 2 and AMP-activated protein kinase were involved in lysine deprivation-mediated inhibition of mTORC1. Taken together, these results suggest that lysine might play role in the regulation of mTORC1 activation in NSCLC cells.

Oridonin protects HaCaT keratinocytes against hydrogen peroxide-induced oxidative stress by altering microRNA expression.

microRNAs (miRNAs) have been shown to function as primary regulators of a variety of biological processes, including proliferation, differentiation and apoptosis in human keratinocytes. However, the biological significance of miRNAs in the defense against oxidative stress in keratinocytes remains to be elucidated. In this study, we demonstrate that oridonin, a diterpenoid compound isolated from Rabdosia rubescens with established antioxidant properties, protects HaCaT human keratinocytes from oxidative stress induced by exposure to hydrogen peroxide (H2O2). Our data demonstrate that low doses of oridonin (1-5 µM) protect keratinocytes against H2O2-induced apoptosis in a concentration- and time-dependent manner. Moreover, as shown by our results, oridonin markedly decreased H2O2-induced reactive oxygen species production in HaCaT cells. Oridonin mediated these effects by altering miRNA expression. Bioinformatics analysis identified several putative target genes of the differentially expressed miRNAs. Assessment of their gene ontology annotation revealed that these target genes are likely involved in cell growth and inhibition of apoptosis. Thus, the data from this study establish a role for miRNAs in mediating oridonin-induced protective effects against oxidative stress in human keratinocytes.

Centella asiatica protects against UVB-induced HaCaT keratinocyte damage through microRNA expression changes.

This study aimed to evaluate the protective effects of Centella asiatica (C. asiatica) against ultraviolet B (UVB) damage in human keratinocytes using microRNA (miRNA) expression profiling analysis. Titrated extract of C. asiatica (TECA) demonstrated low cytotoxicity in normal human HaCaT keratinocytes only at low doses (<5 µg/ml). UVB (50 mJ/cm2) irradiation significantly decreased cell viability, and TECA treatment decreased the UVB toxicity. By using miRNA microarrays, we determined that 72 miRNAs had an altered expression following TECA treatment in UVB-irradiated keratinocytes (46 upregulated and 26 downregulated). Using an miRNA target gene prediction tool and Gene Ontology (GO) analysis, we determined that miRNAs with altered expression were functionally related with the inhibition of apoptosis and cell proliferation. Overall, these results provide meaningful information to facilitate the understanding of TECA-mediated UVB protection in human keratinocytes.

Tetraarsenic oxide, a novel orally administrable angiogenesis inhibitor.

Arsenic compounds have been used to treat angiogenic diseases such as cancer, psoriasis, and rheumatoid arthritis in traditional oriental medicine. In recent years, arsenic trioxide (As2O3, diarsenic oxide) has been successfully used to treat acute promyelocytic leukemia. We investigated the antiangiogenic properties of tetraarsenic oxide (As4O6), another trivalent arsenic compound. In in vitro studies, tetraarsenic oxide inhibited the proliferation (IC50 = 99.7 nM), migration into the denuded area (IC50 = 27.4 nM), and invasion through a layer of Matrigel (IC50 = 73.5 nM) of basic fibroblast growth factor (bFGF)-stimulated bovine capillary endothelial (BCE) cells in a dose-dependent manner. Tetraarsenic oxide also inhibited the tube formation of human umbilical vein endothelial cells. Tetraarsenic oxide induced cell cycle arrest of bFGF-stimulated BCE cells in the G2/M phase and inhibited the secretion of matrix metalloproteinase-2 from BCE cells. Orally administered tetraarsenic oxide (50 mg/kg/day) inhibited bFGF-induced new-vessel formation in a rat corneal micropocket assay, and reduced by about 54% the number of experimental pulmonary metastatic nodules in mice implanted with B16F10 melanoma cells. Thus, we provide evidence that tetraarsenic oxide has effective antiangiogenic activities.

Enhancement of radiation response in human cervical cancer cells in vitro and in vivo by arsenic trioxide (As2O3).

Arsenic trioxide (As2O3) inhibits cell growth and induces apoptosis in certain types of cancer cells including acute promyelocytic leukemia, prostate and ovarian carcinomas, but its effect on response of tumor cells to ionizing radiation has never been explored before. Here we demonstrate that As2O3 can sensitize human cervical cancer cells to ionizing radiation both in vitro and in vivo. As2O3 in combination with ionizing radiation have a synergistic effect in decreasing clonogenic survival and in the regression of established human cervical tumor xenografts. Pretreatment of the cells with As2O3 also synergistically enhanced radiation-induced apoptosis. Apoptosis of the cells by combined treatment of As2O3 and radiation was associated with reactive oxygen species generation and loss of mitochondrial membrane potential, resulting in the activation of caspase-9 and caspase-3. The combined treatment also resulted in an increased G2/M cell cycle distribution at the concentration of As2O3 which did not alter cell cycle when applied alone. These results indicate that As2O3 can synergistically enhance radiosensitivity of human cervix carcinoma cells in vitro and in vivo, suggesting a potential clinical applicability of combination treatment of As2O3 and ionizing radiation in cancer therapies.