Yung Wei Lin

Melatonin inhibits MMP-9 transactivation and renal cell carcinoma metastasis by suppressing Akt-MAPKs pathway and NF-κB DNA-binding activity

Renal cell carcinoma (RCC) is the most lethal of all urological malignancies because of its potent metastasis potential. Melatonin exerts multiple tumor‐suppressing activities through antiproliferative, proapoptotic, and anti‐angiogenic actions and has been tested in clinical trials. However, the antimetastastic effect of melatonin and its underlying mechanism in RCC are unclear. In this study, we demonstrated that melatonin at the pharmacologic concentration (0.5–2 mm) considerably reduced the migration and invasion of RCC cells (Caki‐1 and Achn). Furthermore, we found that melatonin suppressed metastasis of Caki‐1 cells in spontaneous and experimental metastasis animal models. Mechanistic investigations revealed that melatonin transcriptionally inhibited MMP‐9 by reducing p65‐ and p52‐DNA‐binding activities. Moreover, the Akt‐mediated JNK1/2 and ERK1/2 signaling pathways were involved in melatonin‐regulated MMP‐9 transactivation and cell motility. Clinical samples revealed an inverse correlation between melatonin receptor 1A (MTNR1A) and MMP‐9 expression in normal kidney and RCC tissues. In addition, a higher survival rate was found in MTNR1Ahigh/MMP‐9low patients than in MTNR1Alow/MMP‐9high patients. Overall, our results provide new insights into the role of melatonin‐induced molecular regulation in suppressing RCC metastasis and suggest that melatonin has potential therapeutic applications for metastastic RCC.

Quercetin induces mitochondrial-derived apoptosis via reactive oxygen species-mediated ERK activation in HL-60 leukemia cells and xenograft

Abstract Quercetin is a plant-derived bioflavonoid that was recently shown to have multiple anticancer activities in various solid tumors. Here, novel molecular mechanisms through which quercetin exerts its anticancer effects in acute myeloid leukemia (AML) cells were investigated. Results from Western blot and flow cytometric assays revealed that quercetin significantly induced caspase-8, caspase-9, and caspase-3 activation, poly ADP-ribose polymerase (PARP) cleavage, and mitochondrial membrane depolarization in HL-60 AML cells. The induction of PARP cleavage by quercetin was also observed in other AML cell lines: THP-1, MV4-11, and U937. Moreover, treatment of HL-60 cells with quercetin induced sustained activation of extracellular signal-regulated kinase (ERK), and inhibition of ERK by an ERK inhibitor significantly abolished quercetin-induced cell apoptosis. MitoSOX red and 2′,7′-dichlorofluorescin fluorescence, respectively, showed that mitochondrial superoxide and intracellular peroxide levels were higher in quercetin-treated HL-60 cells compared with the control group. Moreover, both N-acetylcysteine and the superoxide dismutase mimetic, MnTBAP, reversed quercetin-induced intracellular reactive oxygen species production, ERK activation, and subsequent cell death. The in vivo xenograft mice experiments revealed that quercetin significantly reduced tumor growth through inducing intratumoral oxidative stress while activating the ERK pathway and subsequent cell apoptosis in mice with HL-60 tumor xenografts. In conclusions, our results indicated that quercetin induced cell death of HL-60 cells in vitro and in vivo through induction of intracellular oxidative stress following activation of an ERK-mediated apoptosis pathway.

Nonautophagic cytoplasmic vacuolation death induction in human PC-3M prostate cancer by curcumin through reactive oxygen species -mediated endoplasmic reticulum stress

The antiapoptotic and antiautophagic abilities of cancer cells constitute a major challenge for anticancer drug treatment. Strategies for triggering nonapoptotic or nonautophagic cell death may improve therapeutic efficacy against cancer. Curcumin has been reported to exhibit cancer chemopreventive properties. Herein, we report that curcumin induced apoptosis in LNCaP, DU145, and PC-3 cells but triggered extensive cytoplasmic vacuolation in PC-3M cells. Electron microscopic images showed that the vacuoles lacked intracellular organelles and were derived from the endoplasmic reticulum (ER). Moreover, curcumin-induced vacuolation was not reversed by an apoptosis- or autophagy-related inhibitor, suggesting that vacuolation-mediated cell death differs from classical apoptotic and autophagic cell death. Mechanistic investigations revealed that curcumin treatment upregulated the ER stress markers CHOP and Bip/GRP78 and the autophagic marker LC3-II. In addition, curcumin induced ER stress by triggering ROS generation, which was supported by the finding that treating cells with the antioxidant NAC alleviated curcumin-mediated ER stress and vacuolation-mediated death. An in vivo PC-3M orthotopic prostate cancer model revealed that curcumin reduced tumor growth by inducing ROS production followed by vacuolation-mediated cell death. Overall, our results indicated that curcumin acts as an inducer of ROS production, which leads to nonapoptotic and nonautophagic cell death via increased ER stress.

Mefloquine exerts anticancer activity in prostate cancer cells via ROS-mediated modulation of Akt, ERK, JNK and AMPK signaling

Mefloquine (MQ) is a prophylactic anti-malarial drug. Previous studies have shown that MQ induces oxidative stress in vitro. Evidence indicates that reactive oxygen species (ROS) may be used as a therapeutic modality to kill cancer cells. This study investigated whether MQ also inhibits prostate cancer (PCa) cell growth. We used sulforhodamine B (SRB) staining to determine cell viability. MQ has a highly selective cytotoxicity that inhibits PCa cell growth. The antitumor effect was most significant when examined using a colony formation assay. MQ also induces hyperpolarization of the mitochondrial membrane potential (MMP), as well as ROS generation. The blockade of MQ-induced anticancer effects by N-acetyl cysteine (NAC) pre-treatment confirmed the role of ROS. This indicates that the MQ-induced anticancer effects are caused primarily by increased ROS generation. Moreover, we observed that MQ-mediated ROS simultaneously downregulated Akt phosphorylation and activated extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and adenosine monophosphate-activated protein kinase (AMPK) signaling in PC3 cells. These findings provide insights for further anticancer therapeutic options.

Interactions between environmental factors and melatonin receptor type 1A polymorphism in relation to oral cancer susceptibility and clinicopathologic development

Background The purpose of this study was to explore the combined effect of melatonin receptor type 1A (MTNR1A) gene polymorphisms and exposure to environmental carcinogens on the susceptibility and clinicopathological characteristics of oral cancer. Methodology and Principal Findings Three polymorphisms of the MTNR1A gene from 618 patients with oral cancer and 560 non-cancer controls were analyzed by real-time polymerase chain reaction (PCR). The CTA haplotype of the studied MTNR1A polymorphisms (rs2119882, rs13140012, rs6553010) was related to a higher risk of oral cancer. Moreover, MTNR1A gene polymorphisms exhibited synergistic effects of environmental factors (betel quid and tobacco use) on the susceptibility of oral cancer. Finally, oral-cancer patients with betel quid-chewing habit who had T/T allele of MTNR1A rs13140012 were at higher risk for developing an advanced clinical stage and lymph node metastasis. Conclusion These results support gene-environment interactions of MTNR1A polymorphisms with smoking and betel quid-chewing habits possibly altering oral-cancer susceptibility and metastasis.

Mefloquine induces cell death in prostate cancer cells and provides a potential novel treatment strategy in vivo

Mefloquine (MQ) is currently in clinical use as a prophylactic treatment for malaria. Previous studies have shown that MQ induces oxidative stress in vitro. The present study investigated the anticancer effects of MQ treatment in PC3 cells. The cell viability was evaluated using sulphorhodamine-B (SRB) staining, while annexin V and propidium iodide (PI) were used as an assay for cell death. Reactive oxygen species (ROS) formation was detected with 2′,7′-dichlorofluorescein-diacetate (DCFH-DA), a sensitive intracellular probe, and the alteration of cellular status was defined by trypan blue staining. The results of the present study indicated that MQ has a high cytotoxicity that causes cell death in PC3 cells. MQ markedly inhibited the PC3 cells through non-apoptotic cell death. MQ also induced significant ROS production. The MQ treatment mediated G1 cell cycle arrest and cyclin D1 accumulation through p21 upregulation in the PC3 cells. Moreover, the use of MQ improved the survival of the treatment group compared with the control group in the experimental mice. The present study indicates that MQ possesses potential therapeutic efficacy for the treatment of prostate cancer (PCa) in vivo. These findings provide insights that may aid the further optimization and application of new and existing therapeutic options.

Quercetin simultaneously induces G0/G1‐phase arrest and caspase‐mediated crosstalk between apoptosis and autophagy in human leukemia HL‐60 cells

Quercetin is a plant‐derived bioflavonoid with high anticancer activity in various tumors. Herein, the molecular mechanisms by which quercetin exerts its anticancer effects against HL‐60 acute myeloid leukemia (AML) cells were investigated. Results showed that quercetin suppressed cell proliferation in the HL‐60 cell line in vitro and in vivo. Quercetin‐induced G0/G1‐phase arrest occurred when expressions of cyclin‐dependent kinase (CDK)2/4 were inhibited and the CDK inhibitors, p16 and p21, were induced. Moreover, quercetin treatment not only activated proapoptotic signaling like poly (ADP ribose) polymerase (PARP)−1 cleavage and caspase activation but also triggered autophagy events as shown by the increased expression of light chain 3 (LC3)‐II, decreased expression of p62, and formation of acidic vesicular organelles. Interestingly, it was found that use of the autophagy inhibitor, 3‐methyladenine, significantly enhanced quercetin‐mediated apoptotic cell death as analyzed by MTS and DNA fragmentation assays. Moreover, pretreatment of HL‐60 cells with the pan‐caspase inhibitor, Z‐VAD‐fmk, dramatically reversed quercetin‐mediated apoptotic and autophagic cell death. Although apoptosis and autophagy are two independent cell death pathways, our findings indicated that quercetin can activate caspases to trigger these two pathways, and both pathways played contrary roles in quercetin‐mediated HL‐60 cell death. In conclusion, besides promoting apoptosis, quercetin also induced cytoprotective autophagy in HL‐60 cells, and inhibition of autophagy may be a novel strategy to enhance the anticancer activity of quercetin in AML.

Efficacy and safety of orally disintegrating tamsulosin tablets in Taiwanese patients with benign prostatic hyperplasia.

Objectives: Tamsulosin is an alpha-1 adrenoceptor antagonist applied in treating lower urinary tract symptoms (LUTS) suggestive of benign prostatic hyperplasia (BPH). This study aimed to evaluate safety, efficacy and acceptance of newly formulated orally disintegrating tamsulosin tablets in Taiwanese patients with LUTS/BPH. Methods: This single center, non-comparative, observational study enrolled 45 male patients over age 50 years. All patients received 0.2 mg tamsulosin orally disintegrating tablets daily and were evaluated at weeks 0, 4, 8, 12 of the 12-week treatment period. Tamsulosin efficacy was evaluated by International Prostate Symptom Score (I-PSS) with 7 questions on urinary symptoms and one disease-specific quality-of-life question, with scores ranging from 0 (no symptoms) to 35 (highly symptomatic). Maximum flow rate (ml/s), voided volume (ml), flow time (s), and mean flow rate (ml/s) were measured. Danish prostatic symptom sexual function scale rated severity and associated concerns of erection quality, ejaculatory function and pain/discomfort were also assessed. Results: Patients’ mean ± SD age was 62.47 ± 7.77 (range: 50–89) and mean ± SD I-PSS was 13.98 ± 5.50. Statistically significant changes from baseline were found in post-test I-PSS and quality of life (both P < 0.001). Mean ± SD I-PSS decreased from 14.30 ± 9.34 to 6.73 ± 0.88 at patients’ final visit. Statistically significant increases in mean maximum flow rate and mean flow rate were found over 12-week study period (P < 0.05). No adverse events were reported. No significant differences were found in pulse, SBP/DBP or sexual function. Conclusion: Orally disintegrating tamsulosin tablets demonstrate acceptable safety and efficacy for acceptance and well tolerance by Taiwanese LUTS/BPH patients.

Tricetin Induces Apoptosis of Human Leukemic HL-60 Cells through a Reactive Oxygen Species-Mediated c-Jun N-Terminal Kinase Activation Pathway

Tricetin is a dietary flavonoid with cytostatic properties and antimetastatic activities in various solid tumors. The anticancer effect of tricetin in nonsolid tumors remains unclear. Herein, the molecular mechanisms by which tricetin exerts its anticancer effects on acute myeloid leukemia (AML) cells were investigated. Results showed that tricetin inhibited cell viability in various types of AML cell lines. Tricetin induced morphological features of apoptosis such as chromatin condensation and phosphatidylserine (PS) externalization, and significantly activated proapoptotic signaling including caspase-8, -9, and -3 activation and poly(ADP-ribose) polymerase (PARP) cleavage in HL-60 AML cells. Of note, tricetin-induced cell growth inhibition was dramatically reversed by a pan caspase and caspase-8- and -9-specific inhibitors, suggesting that this compound mainly acts through a caspase-dependent pathway. Moreover, treatment of HL-60 cells with tricetin induced sustained activation of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK), and inhibition of ERK and JNK by their specific inhibitors respectively promoted and abolished tricetin-induced cell apoptosis. Dichlorofluorescein (DCF) staining showed that intracellular reactive oxygen species (ROS) levels were higher in tricetin-treated HL-60 cells compared to the control group. Moreover, an ROS scavenger, N-acetylcysteine (NAC), reversed tricetin-induced JNK activation and subsequent cell apoptosis. In conclusion, our results indicated that tricetin induced cell death of leukemic HL-60 cells through induction of intracellular oxidative stress following activation of a JNK-mediated apoptosis pathway. A combination of tricetin and an ERK inhibitor may be a better strategy to enhance the anticancer activities of tricetin in AML.

Trichodermin induces c-Jun N-terminal kinase-dependent apoptosis caused by mitotic arrest and DNA damage in human p53-mutated pancreatic cancer cells and xenografts

Pancreatic cancer is an aggressive malignancy, which generally responds poorly to chemotherapy. In this study, trichodermin, an endophytic fungal metabolite from Nalanthamala psidii, was identified as a potent and selective antitumor agent in human pancreatic cancer. Trichodermin exhibited antiproliferative effects against pancreatic cancer cells, especially p53-mutated cells (MIA PaCa-2 and BxPC-3) rather than normal pancreatic epithelial cells. We found that trichodermin induced caspase-dependent and mitochondrial intrinsic apoptosis. Trichodermin also increased apoptosis through mitotic arrest by activating Cdc2/cyclin B1 complex activity. Moreover, trichodermin promoted the activation of c-Jun N-terminal kinase (JNK), and inhibition of JNK by its inhibitor, shRNA, or siRNA significantly reversed trichodermin-mediated caspase-dependent apoptosis. Trichodermin triggered DNA damage stress to activate p53 function for executing apoptosis in p53-mutated cells. Importantly, we demonstrated that trichodermin with efficacy similar to gemcitabine, profoundly suppressed tumor growth through inducing intratumoral DNA damage and JNK activation in orthotopic pancreatic cancer model. Based on these findings, trichodermin is a potential therapeutic agent worthy of further development into a clinical trial candidate for treating cancer, especially the mutant p53 pancreatic cancer.