Silencing of Dicer enhances dacarbazine resistance in melanoma cells by inhibiting ADSL expression.

Dacarbazine (DTIC) is the primary first-line treatment for advanced-stage metastatic melanoma; thus, DTIC resistance is poses a major challenge. Therefore, investigating the mechanism underlying DTIC resistance must be investigated. Dicer, a type III cytoplasmic endoribonuclease, plays a pivotal role in the maturation of miRNAs. Aberrant Dicer expression may contribute to tumor progression, clinical aggressiveness, and poor prognosis in various tumors. Dicer inhibition led to a reduction in DTIC sensitivity and an augmentation in stemness in melanoma cells. Clinical analyses indicated a low Dicer expression level as a predictor of poor prognosis factor. Metabolic alterations in tumor cells may interfere with drug response. Adenylosuccinate lyase (ADSL) is a crucial enzyme in the purine metabolism pathway. An imbalance in ADSL may interfere with the therapeutic efficacy of drugs. We discovered that DTIC treatment enhanced ADSL expression and that Dicer silencing significantly reduced ADSL expression in melanoma cells. Furthermore, ADSL overexpression reversed Dicer silencing induced DTIC resistance and cancer stemness. These findings indicate that Dicer-mediated ADSL regulation influences DTIC sensitivity and stemness in melanoma cells.

γ-aminobutyric acid B2 receptor: A potential therapeutic target for cholangiocarcinoma in patients with diabetes mellitus.

BACKGROUND: The association between diabetes mellitus (DM) and the increased risk and progression of cholangiocarcinoma (CCA) has been reported with unclear underlying mechanisms. Previous studies showed that γ-aminobutyric acid (GABA) B2 receptor (GABBR2) was upregulated in CCA cells cultured in high glucose (HG) conditions. Roles of GABA receptors in CCA progression have also been studied, but their association with DM and hyperglycemia in CCA remains unclarified. AIM: To investigate the effects of hyperglycemia on GABBR2 expression and the potential use of GABBR2 as a CCA therapeutic target. METHODS: CCA cells, KKU-055 and KKU-213A, were cultured in Dulbecco Modified Eagle's Medium supplemented with 5.6 mmol/L (normal glucose, NG) or 25 mmol/L (HG) glucose and assigned as NG and HG cells, respectively. GABBR2 expression in NG and HG cells was investigated using real-time quantitative polymerase chain reaction and western blot. Expression and localization of GABBR2 in CCA cells were determined using immunocytofluorescence. GABBR2 expression in tumor tissues from CCA patients with and without DM was studied using immunohistochemistry, and the correlations of GABBR2 with the clinicopathological characteristics of patients were analyzed using univariate analysis. Effects of baclofen, a GABA-B receptor agonist, on CCA cell proliferation and clonogenicity were tested using the MTT and clonogenic assays. Phospho-kinases arrays were used to screen the affected signaling pathways after baclofen treatment, and the candidate signaling molecules were validated using the public transcriptomic data and western blot. RESULTS: GABBR2 expression in CCA cells was induced by HG in a dose- and time-dependent manner. CCA tissues from patients with DM and hyperglycemia also showed a significantly higher GABBR2 expression compared with tumor tissues from those with euglycemia (P < 0.01). High GABBR2 expression was significantly associated with a poorer non-papillary histological subtype but with smaller sizes of CCA tumors (P < 0.05). HG cells of both tested CCA cell lines were more sensitive to baclofen treatment. Baclofen significantly suppressed the proliferation and clonogenicity of CCA cells in both NG and HG conditions (P < 0.05). Phospho-kinase arrays suggested glycogen synthase kinase 3 (GSK3), ß-catenin, and the signal transducer and activator of transcription 3 (STAT3) as candidate signaling molecules under the regulation of GABBR2, which were verified in NG and HG cells of the individual CCA cell lines. Cyclin D1 and c-Myc, the common downstream targets of GSK3/ß-catenin and STAT3 involving cell proliferation, were accordingly downregulated after baclofen treatment. CONCLUSION: GABBR2 is upregulated by HG and holds a promising role as a therapeutic target for CCA regardless of the glucose condition.

Hyperglycemia Alters O-GlcNAcylation Patterns of Hepatocytes in Mice Treated With Hepatoxic Carcinogen.

BACKGROUND/AIM: Diabetes mellitus (DM) is an established risk for hepatocellular carcinoma (HCC), with unclarified mechanisms. This study investigated the effects of hyperglycemia on O-GlcNacylation in hepatocytes and its associations with hepatocarcinogenesis. MATERIALS AND METHODS: Mouse and human HCC cell lines were used in an in vitro model of hyperglycemia. Western blotting was used to determine the effects of high glucose on O-GlcNacylation in HCC cells. Twenty 4-week-old C3H/HeNJcl mice were randomized into four groups: non-DM control, non-DM plus diethylnitrosamine (DEN), DM, and DM plus DEN. DM was induced using intraperitoneal injection of a single high dose of streptozotocin. DEN was used to induce HCC. All mice were euthanized at week 16 after DM induction, and the liver tissues were histologically examined using hematoxylin and eosin, and immunohistochemistry. RESULTS: High glucose increased O-GlcNacylated proteins in mouse and human HCC cell lines compared with those cultured at normal glucose concentration. Mice with hyperglycemia or DEN treatment had increased O-GlcNacylated proteins in hepatocytes. No gross tumors were evident at the end of the experiment but hepatic morbidity was observed. Mice with hyperglycemia and DEN treatment showed greater histological morbidity in their livers, i.e. increased nuclear size, hepatocellular swelling and sinusoidal dilatation, compared with mice in the DM group or treated with DEN alone. CONCLUSION: Hyperglycemia increased O-GlcNAcylation in both in vitro and animal models. Increased O-GlcNAcylated proteins may be associated with hepatic histological morbidities which then promote HCC development in carcinogen-induced tumorigenesis.

A Mitochondrion-Targeting Protein (B2) Primes ROS/Nrf2-Mediated Stress Signals, Triggering Apoptosis and Necroptosis in Lung Cancer.

The betanodavirus B2 protein targets mitochondria and triggers mitochondrion-mediated cell death signaling in lung cancer cells; however, its molecular mechanism remains unknown. In this study, we observed that B2 triggers hydrogen peroxide/Nrf2-involved stress signals in the dynamic regulation of non-small lung cancer cell (NSCLC)-programmed cell death. Here, the B2 protein works as a necrotic inducer that triggers lung cancer death via p53 upregulation and RIP3 expression, suggesting a new perspective on lung cancer therapy. We employed the B2 protein to target A549 lung cancer cells and solid tumors in NOD/SCID mice. Tumors were collected and processed for the hematoxylin and eosin staining of tissue and cell sections, and their sera were used for blood biochemistry analysis. We observed that B2 killed an A549 cell-induced solid tumor in NOD/SCID mice; however, the mutant ΔB2 did not. In NOD/SCID mice, B2 (but not ΔB2) induced both p53/Bax-mediated apoptosis and RIPK3-mediated necroptosis. Finally, immunochemistry analysis showed hydrogen peroxide /p38/Nrf2 stress strongly inhibited the production of tumor markers CD133, Thy1, and napsin, which correlate with migration and invasion in cancer cells. This B2-triggered, ROS/Nrf2-mediated stress signal triggered multiple signals via pathways that killed A549 lung cancer tumor cells in vivo. Our results provide novel insight into lung cancer management and drug therapy.

Aspartame consumption during pregnancy impairs placenta growth in mice through sweet taste receptor-reactive oxygen species-dependent pathway.

The prevalence of obesity has risen dramatically over recent years, and so has the prevalence of adverse obesity-associated pregnancy outcomes. To combat obesity, the calorie contents of many foods and beverages may be reduced by the use of artificial sweeteners, such as aspartame. However, animal studies suggest that aspartame and its metabolites may exhibit toxicity, and the effects of aspartame on pregnancy are largely unknown. In this study, we treated pregnant mice with aspartame by oral gavage and found that the treatment decreased fasting blood glucose level, whereas systolic blood pressure was elevated. Importantly, the aspartame-treated animals also had low placenta and fetus weights, as well as reduced thickness of the placenta decidua layer. Moreover, aspartame decreased the expression of epithelial-mesenchymal transition proteins and manganese superoxide dismutase (MnSOD) in mouse placentae. In order to clarify the mechanisms though which aspartame affects placenta, we performed experiments on 3A-sub-E trophoblasts. In the cells, aspartame treatments induced cell cycle arrest and reduced the proliferation rate, epithelial-mesenchymal transition, migration activity and invasion activity. We also found that aspartame increased reactive oxygen species (ROS) levels to hyper-activate Akt and downregulate MnSOD expression. Pretreatment with antioxidants or sweet taste receptor inhibitors reversed the effects of aspartame on trophoblast function. We also found that the aspartame metabolite phenylalanine similarly induced ROS production and affected proliferation of trophoblasts. Taken together, our data suggest that aspartame consumption during pregnancy may impact the structure, growth and function of the placenta via sweet taste receptor-mediated stimulation of oxidative stress.

Organic cation transporter 2 activation enhances sensitivity to oxaliplatin in human pancreatic ductal adenocarcinoma.

Oxaliplatin, a third-generation platinum derivative, has become one of the main chemotherapeutic treatments for esophagus, gastric and colorectal cancer; however, it is still unclear the potential effectiveness for pancreatic ductal adenocarcinoma (PDAC) with gemcitabine resistance. Here, we observed that PDAC tumors have low level of organic cation transporter 2 (OCT2, also known as SLC22A2) compared with non-tumor tissues and identified that OCT2 expression is positively correlated with oxaliplatin sensitivity in PDAC cells. Treatment of OCT2 inhibitors or knockdown of OCT2 expression significantly decreased the sensitivity to oxaliplatin in PANC-1 cells. In addition, bisulfite sequencing polymerase chain reaction analysis revealed that higher methylation frequency represses OCT2 expression in gemcitabine-resistant PANC-1 (PANC-1/GR) cells. Moreover, we found that treatment of DNA methyltransferase (DNMT) inhibitors, decitabine or 5-azacytidine recover OCT2 expression and oxaliplatin sensitivity in PANC-1/GR cells, and DNMT1 level has inverse correlation with OCT2 expression in PDAC cells and tumors. Our findings jointly suggest that OCT2 expression is a potential and predictive marker for evaluating oxaliplatin sensitivity and developing alternative treatments for PDAC patients with gemcitabine resistance.

Evasion of NK cell immune surveillance via the vimentin-mediated cytoskeleton remodeling.

Cancer immunotherapy uses the immune system to achieve therapeutic effects; however, its effect is still limited. Therefore, in addition to immune checkpoint-based treatment, the development of other strategies that can inhibit cancer cells from resisting immune cytotoxicity is important. There are currently few studies on the mechanism of tumors using cytoskeletal proteins reorganization to participate in immune escape. In this study, we identified cancer cell lines that were sensitive or resistant to natural killer cells in urothelial and lung cancer using the natural killer cell sensitivity assay. We found that immunoresistant cancer cells avoid natural killer cell-mediated cytotoxicity by upregulation of vimentin and remodeling of actin cytoskeleton. Immunofluorescence staining showed that immune cells promoted the formation of actin filaments at the immune synapse, which was not found in immunosensitive cancer cells. Pretreatment of the actin polymerization inhibitors latrunculin B increased the cytotoxicity of natural killer cells, suggesting that cytoskeleton remodeling plays a role in resisting immune cell attack. In addition, silencing of vimentin with shRNA potentiated the cytotoxicity of natural killer cells. Interestingly, the upregulation and extension of vimentin was found in tumor islands of upper tract urothelial carcinoma infiltrated by natural killer cells. Conversely, tumors without natural killer cell invasion showed less vimentin signal. The expression level of vimentin was highly correlated with natural killer cell infiltration. In summary, we found that when immune cells attack cancer cells, the cancer cells resist immune cytotoxicity through upregulated vimentin and actin reorganization. In addition, this immune resistance mechanism was also found in patient tumors, indicating the possibility that they can be applied to evaluate the immune response in clinical diagnosis.

Phosphomimetic Dicer S1016E triggers a switch to glutamine metabolism in gemcitabine-resistant pancreatic cancer.

OBJECTIVE: Dicer is an enzyme that processes microRNAs (miRNAs) precursors into mature miRNAs, which have been implicated in various aspects of cancer progressions, such as clinical aggressiveness, prognosis, and survival outcomes. We previously showed that high expression of Dicer is associated with gemcitabine (GEM) resistance in pancreatic ductal adenocarcinoma (PDAC); thus, in this study, we aimed to focus on how Dicer is involved in GEM resistance in PDAC, including cancer prognosis, cell proliferation, and metabolic regulation. METHODS: We generated stable shRNA knockdown of Dicer in GEM-resistant PANC-1 (PANC-1 GR) cells and explored cell viability by MTT and clonogenicity assays. Metabolomic profiling was employed to investigate metabolic changes between parental cells, PANC-1, and PANC-1 GR cells, and further implied to compare their sensitivity to the glutaminase inhibitor, CB839, and GEM treatments. To identify putative phosphorylation site involves with Dicer and its effects on GEM resistance in PDAC cells, we further generated phosphomimetic or phosphomutant Dicer at S1016 site and examined the changes in drug sensitivity, metabolic alteration, and miRNA regulation. RESULTS: We observed that high Dicer levels in pancreatic ductal adenocarcinoma cells were positively correlated with advanced pancreatic cancer and acquired resistance to GEM. Metabolomic analysis indicated that PANC-1 GR cells rapidly utilised glutamine as their major fuel and increased levels of glutaminase (GLS): glutamine synthetase (GLUL) ratio which is related to high Dicer expression. In addition, we found that phosphomimetic Dicer S1016E but not phosphomutant Dicer S1016A facilitated miRNA maturation, causing an imbalance in GLS and GLUL and resulting in an increased response to GLS inhibitors. CONCLUSION: Our results suggest that phosphorylation of Dicer on site S1016 affects miRNA biogenesis and glutamine metabolism in GEM-resistant pancreatic cancer.

Dicer-mediated miR-200b expression contributes to cell migratory/invasive abilities and cancer stem cells properties of breast cancer cells.

Distant metastasis is the leading cause of death in patients with breast cancer. Despite considerable treatment advances, the clinical outcomes of patients with metastatic breast cancer remain poor. CSCs can self-renew, enhancing cancer progression and metastasis. Dicer, a microRNA (miRNA) processing-related enzyme, is required for miRNA maturation. Imbalanced Dicer expression may be pivotal in cancer progression. However, whether and how Dicer affects the stemness of metastatic breast cancer cells remains unclear. Here, we hypothesized that Dicer regulates the migration, invasion, and stemness of breast cancer cells. We established highly invasive cell lines (MCF-7/I-3 and MDA-MB-231/I-3) and observed that Dicer expression was conspicuously lower in the highly invasive cells than in the parental cells. The silencing of Dicer significantly enhanced the cell migratory/invasive abilities and CSCs properties of the breast cancer cells. Conversely, the overexpression of Dicer in the highly invasive cells reduced their migration, invasion, and CSCs properties. Our bioinformatics analyses demonstrated that low Dicer levels were correlated with increased breast cancer risk. Suppression of Dicer inhibited miR-200b expression, whereas miR-200b suppression recovered Dicer knockdown-induced migration, invasion, and cancer stem cells (CSCs) properties of the breast cancer cells. Thus, our findings reveal that Dicer is a crucial regulator of the migration, invasion, and CSCs properties of breast cancer cells and is significantly associated with poor survival in patients with breast cancer.

Protein disulfide isomerase a4 promotes lung cancer development via the Stat3 pathway in stromal cells.

BACKGROUND: Protein disulfide isomerases a4 (Pdia4) is known to be involved in cancer development. Our previous publication showed that Pdia4 positively promotes cancer development via its inhibition of procaspase-dependent apoptosis in cancer cells. However, nothing is known about its role in the cancer microenvironment. RESULTS: Here, we first found that Pdia4 expression in lung cancer was negatively correlated with patient survival. Next, we investigated the impact of host Pdia4 in stromal cells during cancer development. We showed that Pdia4 was expressed at a low level in stromal cells, and this expression was up-regulated akin to its expression in cancer cells. This up-regulation was stimulated by tumour cell-derived stimuli. Genetics studies in tumour-bearing wild-type and Pdia4-/- mice showed that host Pdia4 promoted lung cancer development in the mice via cancer stroma. This promotion was abolished in Rag1-/- mice which lacked T and B cells. This promotion could be restored once T and B cells were added back to Rag1-/- mice. In addition, host Pdia4 positively regulated the number and immunosuppressive function of stromal cells. Mechanistic studies showed that host Pdia4 positively controlled the Stat3/Vegf pathway in T and B lymphocytes via its stabilization of activated Stat3 in a Thioredoxin-like domain (CGHC)-dependent manner. CONCLUSIONS: These findings identify Pdia4 as a possible target for intervention in cancer stroma, suggesting that targeting Pdia4 in cancer stroma is a promising anti-cancer approach.

Association Between Essential and Non-essential Metals, Body Composition, and Metabolic Syndrome in Adults.

Growing evidence indicates that metal exposure is associated with metabolic syndrome (MetS); however, mixed results have been reported. The aim of this study was to clarify associations of exposure to essential and non-essential metals with body composition and risks of obesity and MetS. Anthropometry and blood biochemistry of metabolic parameters were obtained from 150 middle-aged Taiwanese adults. Plasma metals were assessed using inductively coupled plasma mass spectrometry, and body compositions were measured by a bioelectrical impedance analysis (BIA). The essential metals of copper (Cu), manganese (Mn), and chromium (Cr) were positively correlated with the body fat mass but inversely correlated with the skeletal muscle mass (all p < 0.05). An adjusted logistic regression showed that Mn [odds ratio (OR) = 1.624 (95% confidence interval 1.072, 2.462), p = 0.02] and, to a lesser extent, Cu [OR = 1.501 (0.985, 2.292), p = 0.059] predicted abdominal obesity, while plasma Cu [OR = 2.211 (1.146, 4.266), p = 0.02] and zinc (Zn) [OR = 2.228 (1.048, 4.736) p = 0.04] predicted MetS. Significant correlations between dyslipidemia and lithium [OR = 1.716 (1.080, 2.726)], Cu [OR = 2.210 (1.415, 3.454)], Mn [OR = 2.200 (1.320, 3.666)], molybdenum [OR = 1.853 (1.160, 2.958)], and Zn [OR = 1.993 (1.186, 3.349)], and between boron [OR = 2.583 (1.137, 5.868)] and hyperglycemia were observed (all p < 0.05). Exposure to essential metals may affect the body composition and metabolic profiles, exacerbating the risk of MetS.

A case-control study in Taiwanese cohort and meta-analysis of serum ferritin in pancreatic cancer.

Pancreatic cancer is one of the most lethal diseases which lack an early diagnostic marker. We investigated whether serum ferritin (SF) reflects risk for pancreatic cancer and potential genes that may contribute ferritin and pancreatic cancer risks. We performed a meta-analysis of relevant studies on SF and pancreatic cancer risk by searching articles in PUBMED and EMBASE published up to 1 March 2020. We also collected serum samples from Taipei Medical University Joint Biobank and compared SF levels in 34 healthy controls and 34 pancreatic cancer patients. An Oncomine database was applied as a platform to explore a series of genes that exhibited strong associations between ferritin and pancreatic cancer. Herein, we show that high levels of SF can indicate risk of pancreatic cancer, suggesting SF as the new tumor marker that may be used to help pancreatic cancer diagnosis. We also found that expressions of iron homeostasis genes (MYC, FXN) and ferroptosis genes (ALOX15, CBS, FDFT1, LPCAT3, RPL8, TP53, TTC35) are significantly altered with pancreatic tumor grades, which may contribute to differential expression of ferritin related to pancreatic cancer prognosis.

Aldehyde Dehydrogenase Mutation Exacerbated High-Fat-Diet-Induced Nonalcoholic Fatty Liver Disease with Gut Microbiota Remodeling in Male Mice.

Mitochondrial aldehyde dehydrogenase 2 (ALDH2) is a critical enzyme involved in ethanol clearance in acetaldehyde metabolism and plays a key role in protecting the liver. The ALDH2*2 mutation causes a significant decrease in acetaldehyde scavenging capacity, leading to the accumulation of acetaldehyde after consuming alcohol. The prevalence of the ALDH2*2 variant is in 45% of Taiwanese individuals. ALDH2 reportedly has protective properties on myocardial damage, stroke, and diabetic retina damage. However, the effects of ALDH2 in the modulation of metabolic syndromes remain unclear. This study evaluates the roles of ALDH2 in a high-fat-diet-induced metabolic syndrome in mice. Male (M) and female (F) wild-type (WT) and ALDH2 knock-in C57BL/6J mice (4-5 weeks old) were fed a high-fat diet for 16 weeks. Results showed that the body and white-adipose-tissue weights were significantly increased in ALDH2-M compared to those in the other groups. We observed markedly elevated serum levels of alanine transaminase and glucose. Oral glucose-tolerance test and homeostasis-model assessment of insulin resistance (HOMA-IR) values were significantly higher in ALDH2-M mice than those in WT-M mice, with no observable differences in female mice. Abundant steatosis and inflammatory cells were observed in ALDH2-M, with significantly decreased expression of hepatic genes IRS2, GLUT4, and PGC-1α compared to that in WT-M. ALDH2 gene mutation also affected the ß-diversity of gut microbiota in ALDH2-M resulting in the decreased abundance of Actinobacteria and an increase in Deferribacteres. Our results suggest that potential changes in gut microbiota may be associated with the defective ALDH2 exacerbation of high-fat-diet-induced liver diseases in male mice. However, female mice were not affected, and sex hormones may be an important factor that requires further investigation.

Eicosapentaenoic Acid Inhibits KRAS Mutant Pancreatic Cancer Cell Growth by Suppressing Hepassocin Expression and STAT3 Phosphorylation.

BACKGROUND: The oncogenic Kirsten rat sarcoma viral oncogene homolog (KRAS) mutation was reported to be the signature genetic event in most cases of pancreatic ductal adenocarcinoma (PDAC). Hepassocin (HPS/FGL1) is involved in regulating lipid metabolism and the progression of several cancer types; however, the underlying mechanism of HPS/FGL1 in the KRAS mutant PDAC cells undergoing eicosapentaenoic acid (EPA) treatment remains unclear. METHODS: We measured HPS/FGL1 protein expressions in a human pancreatic ductal epithelial (HPNE) normal pancreas cell line, a KRAS-wild-type PDAC cell line (BxPC-3), and KRAS-mutant PDAC cell lines (PANC-1, MIA PaCa-2, and SUIT-2) by Western blot methods. HEK293T cells were transiently transfected with corresponding KRAS-expressing plasmids to examine the level of HPS expression with KRAS activation. We knocked-down HPS/FGL1 using lentiviral vectors in SUIT-2 cells and measured the cell viability by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and clonogenicity assays. Furthermore, a lipidomic analysis was performed to profile changes in lipid metabolism after HPS/FGL1 knockdown. RESULTS: We found that the HPS/FGL1 level was significantly upregulated in KRAS-mutated PDAC cells and was involved in KRAS/phosphorylated (p)-signal transduction and activator of transcription 3 (STAT3) signaling, and the knockdown of HPS/FGL1 in SUIT-2 cells decreased cell proliferation through increasing G2/M cell cycle arrest and cyclin B1 expression. In addition, the knockdown of HPS/FGL1 in SUIT-2 cells significantly increased omega-3 polyunsaturated fatty acids (PUFAs) and EPA production but not docosahexaenoic acid (DHA). Moreover, EPA treatment in SUIT-2 cells reduced the expression of de novo lipogenic protein, acetyl coenzyme A carboxylase (ACC)-1, and decreased p-STAT3 and HPS/FGL1 expressions, resulting in the suppression of cell viability. CONCLUSIONS: Results of this study indicate that HPS is highly expressed by KRAS-mutated PDAC cells, and HPS/FGL1 plays a crucial role in altering lipid metabolism and increasing cell growth in pancreatic cancer. EPA supplements could potentially inhibit or reduce ACC-1-involved lipogenesis and HPS/FGL1-mediated cell survival in KRAS-mutated pancreatic cancer cells.

Betulinic Acid Affects the Energy-Related Proteomic Profiling in Pancreatic Ductal Adenocarcinoma Cells.

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive disease with a 5-year survival rate of <8%. Therefore, finding new treatment strategies against PDAC cells is an imperative issue. Betulinic acid (BA), a plant-derived natural compound, has shown great potential to combat cancer owing to its versatile physiological functions. In this study, we observed the impacts of BA on the cell viability and migratory ability of PDAC cell lines, and screened differentially expressed proteins (DEPs) by an LC-MS/MS-based proteomics analysis. Our results showed that BA significantly inhibited the viability and migratory ability of PDAC cells under a relatively low dosage without affecting normal pancreatic cells. Moreover, a functional analysis revealed that BA-induced downregulation of protein clusters that participate in mitochondrial complex 1 activity and oxidative phosphorylation, which was related to decreased expressions of RNA polymerase mitochondrial (POLRMT) and translational activator of cytochrome c oxidase (TACO1), suggesting that the influence on mitochondrial function explains the effect of BA on PDAC cell growth and migration. In addition, BA also dramatically increased Apolipoprotein A1 (APOA1) expression and decreased NLR family CARD domain-containing protein 4 (NLRC4) expression, which may be involved in the dampening of PDAC migration. Notably, altered expression patterns of APOA1 and NLRC4 indicated a favorable clinical prognosis of PDAC. Based on these findings, we identified potential proteins and pathways regulated by BA from a proteomics perspective, which provides a therapeutic window for PDAC.

TARBP2 Suppresses Ubiquitin-Proteasomal Degradation of HIF-1α in Breast Cancer.

TAR (HIV-1) RNA binding protein 2 (TARBP2) is an RNA-binding protein participating in cytoplasmic microRNA processing. Emerging evidence has shown the oncogenic role of TARBP2 in promoting cancer progression, making it an unfavorable prognosis marker for breast cancer. Hypoxia is a hallmark of the tumor microenvironment which induces hypoxia-inducible factor-1α (HIF-1α) for transcriptional regulation. HIF-1α is prone to be rapidly destabilized by the ubiquitination-proteasomal degradation system. In this study, we found that TARBP2 expression is significantly correlated with induced hypoxia signatures in human breast cancer tissues. At a cellular level, HIF-1α protein level was maintained by TARBP2 under either normoxia or hypoxia. Mechanistically, TARBP2 enhanced HIF-1α protein stability through preventing its proteasomal degradation. In addition, downregulation of multiple E3 ligases targeting HIF-1α (VHL, FBXW7, TRAF6) and reduced ubiquitination of HIF-1α were also induced by TARBP2. In support of our clinical findings that TARBP2 is correlated with tumor hypoxia, our IHC staining showed the positive correlation between HIF-1α and TARBP2 in human breast cancer tissues. Taken together, this study indicates the regulatory role of TARBP2 in the ubiquitination-proteasomal degradation of HIF-1α protein in breast cancer.

ERK-mediated transcriptional activation of Dicer is involved in gemcitabine resistance of pancreatic cancer.

Gemcitabine has been a commonly used therapeutic agent for treatment of pancreatic cancer. In the clinic, a growing resistance to gemcitabine has been observed in patients with pancreatic cancer, and investigation of the underlying mechanism of gemcitabine resistance is urgently required. The microRNA (miRNA)-producing enzyme, Dicer, is crucial for the maturation of miRNAs, and is involved in clinical aggressiveness, poor prognosis, and survival outcomes in various cancers, however, the role of Dicer in acquired gemcitabine resistance of pancreatic cancer is still not clear. Here, we found that Dicer expression was significantly increased in gemcitabine-resistant PANC-1 (PANC-1/GEM) cells compared with parental PANC-1 cells and observed a high level of Dicer correlated with increased risk of pancreatic cancer. Suppression of Dicer obviously decreased gemcitabine resistance in PANC-1/GEM cells; consistently, overexpression of Dicer in PANC-1 cells increased gemcitabine resistance. Moreover, we identified that transcriptional factor Sp1 targeted the promoter region of Dicer and found ERK/Sp1 signaling regulated Dicer expression in PANC-1/GEM cells, as well as positively correlated with pancreatic cancer progression and suggest that targeting the ERK/Sp1/Dicer pathway has potential therapeutic value for pancreatic cancer with acquired resistance to gemcitabine.

The novel camptothecin derivative, CPT211, induces cell cycle arrest and apoptosis in models of human breast cancer.

OBJECTIVE: Breast cancer is the second leading cause of cancer deaths in women worldwide and represents a highly aggressive nature with limited therapeutic options; thus, investigating novel therapeutic agents for breast cancer is much needed. In this study, we investigated the anticancer effects of a novel camptothecin derivative, CPT211, against human breast cancer. METHODS: We used hormone receptor-positive MCF-7, triple-negative (TNBC) MDA-MB-231, and HER2-positive BT-474 human breast cancer cells to examine cytotoxicity of CPT211. We measured cell viability with dose dependence of CPT211 treatments by an MTT assay and investigated the potential underlying mechanism through flow cytometric and Western blot methods. Furthermore, we evaluated the efficacy of the treatment combination of CPT211 and doxorubicin in a mouse model bearing MDA-MB-231 xenografts. RESULTS: CPT211 treatment led to dose-dependent decreases in cell viability of both MCF-7 and MDA-MB-231 cells, but not BT-474 cells. Analysis of the underlying molecular mechanism revealed that CPT211 activated p53-mediated apoptosis, by triggering intrinsic and extrinsic apoptotic pathways in MCF-7 cells. Additionally, CPT211 induced apoptosis and cell cycle arrest of MDA-MB-231 cells by activating Fas/FADD/caspase-8 signaling, suggesting that CPT211-mediated MDA-MB-231 cell apoptosis may occur through an extrinsic apoptosis pathway. CPT211 treatment with doxorubicin in mice bearing MDA-MB-231 xenografts was shown to enhance caspase-8 and caspase-7 activation, resulting in significant inhibition of tumor growth. CONCLUSIONS: These results indicate that Fas/FADD/caspase-8 activation plays an important role in CPT211-mediated tumor growth suppression in TNBC, and the novel camptothecin derivative, CPT211, can be exploited for specific targeted therapies and potentially improve approaches to combination treatments for human breast cancer.

Changes in post-oral glucose challenge pancreatic polypeptide hormone levels following metabolic surgery: A comparison of gastric bypass and sleeve gastrectomy.

BACKGROUND: Pancreatic polypeptide (PP) hormone is a 36-amino-acid peptide released from the pancreas, the serum levels of which have been shown to rise upon food intake. The underlying mechanism for metabolic surgery in the treatment of patients with type 2 diabetes mellitus (T2DM) remains intriguing. We compared post-oral glucose challenge PP levels between patients undergoing laparoscopic gastric bypass (GB) and sleeve gastrectomy (SG) at 1 year after surgery. METHODS: This hospital-based, prospective study followed up a total of 12 laparoscopic GB and 12 laparoscopic SG patients and evaluated their glucose homeostasis. One year after metabolic surgery, 75-g oral glucose tolerance tests (OGTTs) were performed in the patients in the GB and SG groups and the blood levels of PP were evaluated. RESULTS: The laparoscopic GB group had stable serum PP levels within 120 min after OGTT; however, the levels were significantly higher in the laparoscopic SG group at 30 min after OGTT. The patients with complete T2DM remission did not exhibit significantly different PP levels at fasting and post-OGTT than those in patients without remission after GB. Similarly, after SG, patients with T2DM remission did not show significantly different PP levels at fasting and post-OGTT than those in patients without T2DM remission. CONCLUSIONS: No significant difference was found in plasma PP levels after OGTT in T2DM patients that received either GB or SG, or in T2DM remitters or non-remitters 1 year after metabolic surgery.

Housing condition-associated changes in gut microbiota further affect the host response to diet-induced nonalcoholic fatty liver.

Diet-induced obesity is the most widely used animal model for studying nonalcoholic fatty liver disease (NAFLD). However, the physiological effects of a high-fat diet (HFD) are inconsistent between different studies. To elucidate this mystery, mice raised with conventional (CONV), specific pathogen-free (SPF) and gentamicin (G) treatments and fed with standard diet (STD) or HFD were analyzed in terms of their physiology, gut microbiota composition, hepatic steatosis and inflammation. Serum biochemistry showed increased levels of cholesterol and aspartate aminotransferase in the G-STD and CONV-HFD groups, respectively. The CONV-HFD group exhibited more inflammatory foci compared to the SPF-HFD and G-HFD groups. Furthermore, immunohistochemistry staining revealed the infiltration of Kupffer cells in the liver, consistent with increased mRNA levels of MCP-1, CD36 and TLR4. Principal coordinate analysis and the cladogram of LEfSe showed that the distinguished clusters of gut microbiota were dependent on housing conditions. The Rikenellaceae, F16 and Desulfovibrionaceae were strongly correlated with hepatic inflammation. Otherwise, higher NAFLD activity score correlated with altered relative abundances of Bacteroidetes and Firmicutes. In conclusion, gut microbiota varying with housing condition may be pivotal for the host response to HFD.