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Chronic stress has been reported to be associated with tumor initiation and progression. But the underlying mechanism and the specific role of tumor immunity in this process are still unknown. Herein, we applied the repeated restrain stress model in C57BL/6J mice and found that the tumor growth in stressed mice was accelerated compared with that in control mice. In addition, serotonin, also called 5-hydroxytryptamine (5-HT), in the serum of stressed mice was also elevated. Sertraline, a selective serotonin reuptake inhibitor used in the clinic, can restore the serum 5-HT level in stressed mice and restrain tumor growth. We further explored the distribution of major immune cells, including B lymphocytes cells, T lymphocytes, natural killer cells, dendritic cells, tumor-associated macrophages (TAM) and regulatory T cells (Treg). We found that the infiltration of CD8 + T cells in the tumor microenvironment (TME) decreased significantly in stressed mice. And the extra 5-HT treatment could further decrease the infiltration of CD8 + T cells in the TME. The expression of IFN-γ and Granular enzyme B (GzmB) in CD8 + T cells were also dropped in the stressed mice group, whereas the expression of programmed cell death protein 1 (PD-1) on CD8 + T cells was increased. The T cell deficiency induced by stress can be reversed by sertraline, indicating its promising role in strengthening the efficacy of anti-PDL1/PD-1 immunotherapy. The present study provides new mechanistic insights into the impact of chronic stress on antitumor immunity and implicates a novel combined immunotherapy strategy for cancer patients with chronic stress.

The synergistic effect of trametinib and dipyridamole in CRC cells.

Both the MAPK pathway and mevalonate (MVA) signaling pathway play an increasingly significant role in the carcinogenesis of colorectal carcinoma, whereas the cross-talk between these two pathways and its implication in targeted therapy remains unclear in colorectal carcinoma. Here, we identified that HMGCS1 (3-hydroxy-3-methylglutaryl-CoA synthase 1), the rate-limiting enzyme of the MVA pathway, is overexpressed in colon cancer tissues and positively regulates the cell proliferation, migration, and invasion of colon cancer cells. In addition, HMGCS1 could enhance the activity of pERK independent of the MVA pathway, and the suppression of HMGCS1 could completely reduce the EGF-induced proliferation of colon cancer cells. Furthermore, we found that trametinib, a MEK inhibitor, could only partially abolish the upregulation of HMGCS1 induced by EGF treatment, while combination with HMGCS1 knockdown could completely reverse the upregulation of HMGCS1 induced by EGF treatment and increase the sensitivity of colon cancer cells to trametinib. Finally, we combined trametinib and dipyridamole, a common clinically used drug that could suppress the activity of SREBF2 (sterol regulatory element-binding transcription factor 2), a transcription factor regulating HMGCS1 expression, and identified its synergistic effect in inhibiting the proliferation and survival of colon cancer cells in vitro as well as the in vivo tumorigenic potential of colon cancer cells. Together, the current data indicated that HMGCS1 may be a novel biomarker, and the combination of targeting HMGCS1 and MEK might be a promising therapeutic strategy for patients with colon cancer.

p38δ is a member of p38 mitogen-activated protein kinases (MAPKs) family that displays cell- and tissue-specific expression patterns. Recent studies demonstrate that p38δ is centrally involved in several pathologic events, such as diabetes, neurodegeneration diseases, inflammatory diseases, and cancer, and suggest that it may be a potential target for diagnosis and therapy of specific diseases. A nanobody is a new type of antibody that exhibits high antigen-binding activity, solubility, stability, and easy production. This study utilized phage display to isolate nanobodies specifically against p38δ from a fully synthetic nanobody library. Two of them, nanobodies Nb13-6 and Nb13-1, display high binding activity to p38δ, less cross-reactivity with other p38 MAPKs, and high thermal and pH stabilities. Modeling and docking analysis indicated that Nb13-6 is mostly linked to the activation loop of p38δ. Furthermore, detailed studies revealed that Nb13-6 inhibited the protein kinase activity of p38δ and the growth of cancer cells. Therefore, this study provides p38δ-specific nanobodies that are promisingly exploited for diagnosing and treating p38δ-associated diseases.

Immunotherapy targeting the PD‐L1/PD‐1 pathway is a novel type of clinical cancer treatment, but only small subsets of patients can benefit from it because of multiple factors. PD‐L1/PD‐1 expression is a biomarker for predicting the efficacy of anti‐PD‐L1/PD‐1 therapy, which highlights the importance of understanding the regulatory mechanisms of PD‐L1 expression in cancer cells. Casp8 is an apical caspase protease involved in mediating cell apoptosis, but it also has multiple nonapoptotic functions. Casp8 mutations are associated with increased risks of cancer, and low expression of Casp8 is closely connected with poor prognosis in patients with cancer. In addition, mutations of Casp8 in lymphocytes also lead to human immunodeficiency, thereby causing dysfunction of the innate immune system, but the roles of Casp8 in antitumor immunity remain unclear. Here, we found that knocking down Casp8 in mouse melanoma cells promoted tumor progression in an immune system–dependent manner. Mechanistically, Casp8 induced PD‐L1 degradation by upregulating TNFAIP3 (A20) expression, a ubiquitin‐editing enzyme that results in PD‐L1 ubiquitination. In addition, compared with Casp8fl/fl mice, mice with conditional deletion of Casp8 in natural killer (NK) cells (Ncr1iCre/+Casp8fl/fl mice) showed a decreased frequency of IFN‐γ+ and CD107a+ NK cells but an increased frequency of PD‐1+ and CTLA‐4+ NK cells. Melanoma cells with Casp8 knocked down exhibited sensitivity to anti‐PD‐1 or anti‐CTLA‐4 antibody treatments, particularly in Ncr1iCre/+Casp8fl/fl mice. Together, the results indicate that Casp8 induces PD‐L1 degradation by upregulating A20 expression and that decreased Casp8 expression is a potential biomarker for predicting the sensitivity to anti‐PD‐L1/PD‐1 immunotherapy., Casp8 induces PD‐L1 degradation by upregulating A20 expression, and decreased Casp8 expression is a potential biomarker for predicting the sensitivity to anti‐PD‐L1/PD‐1 immunotherapy.

Sertraline and fluoxetine are the two most commonly used selective serotonin reuptake inhibitors (SSRIs) in the treatment of depression. Accumulating evidence has revealed that SSRIs can reduce the risk of hepatocellular carcinoma (HCC), but their therapeutic effects in HCC have not yet been elucidated. Previous studies have reported that sertraline and fluoxetine can suppress the growth of gastric carcinoma, melanoma and nonsmall cell lung cancers by inhibiting the mammalian target rapamycin (mTOR) activity. In this study, we found that sertraline and fluoxetine blocked the protein kinase B (AKT)/mTOR pathway and suppressed the growth of HCC cells in vitro, in xenografts and in diethylnitrosamine/carbon tetrachloride (DEN/CCL4)-induced primary liver mouse model. Sertraline and fluoxetine can synergize with sorafenib, the first approved standard therapy for advanced HCC, to inhibit the viability of HCC cells in vitro and in vivo. In addition, the combination of sorafenib and SSRIs synergistically inhibited the effects of the AKT/mTOR pathway. These results reveal novel therapeutic effects of a combination of SSRIs and sorafenib in HCC.

Background Tumor metastasis is one of the leading reasons of the dismal prognosis of hepatocellular carcinoma (HCC). Epithelial-mesenchymal transition (EMT) is closely associated with tumor metastasis including HCC. The purpose of this study is to construct and validate an EMT-related gene signature for predicting the prognosis of HCC patients. Methods Gene expression data of HCC patients was downloaded from The Cancer Genome Atlas (TCGA) database. Gene set enrichment analysis (GSEA) was performed to found the EMT-related gene sets which were obviously distinct between normal samples and paired HCC samples. Cox regression analysis was used to develop an EMT-related prognostic signature, and the performance of the signature was evaluated by Kaplan–Meier curves and time-dependent receiver operating characteristic (ROC) curves. A nomogram incorporating the independent predictors was established. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression levels of the hub genes in HCC cell lines, and the role of PDCD6 in the metastasis of HCC was determined by functional experiments. Results An EMT-related 5-gene signature (PDCD6, TCOF1, TRIM28, EZH2 and FAM83D) was constructed using univariate and multivariate Cox regression analysis. Based on the signature, the HCC patients were classified into high- and low-risk groups, and patients in high-risk group had a poor prognosis. Time-dependent ROC and Cox regression analyses suggested that the signature could predict HCC prognosis exactly and independently. The predictive capacity of the signature was also validated in two external cohorts. GSEA results showed that many cancer-related signaling pathways such as PI3K/Akt/mTOR pathway and TGF-β/SMAD pathway were enriched in high-risk group. The result of qRT-PCR revealed that PDCD6, TCOF1 and FAM83D were highly expressed in HCC cancer cells. Among them, PDCD6 were found to promote cell migration and invasion. Conclusion The EMT-related 5-gene signature can serve as a promising prognostic biomarker for HCC patients and may provide a novel mechanism of HCC metastasis.

Background Statins, which are used to lower blood cholesterol levels by inhibiting HMG-CoA reductase, have shown anticancer effects in many cancer cells. However, the role of statins in gastric cancer remains unclear. This study aims to investigate whether the statins could antagonize progression of gastric cancer cells and tried to find the molecule mechanism. Methods Effects of simvastatin on the morphology, proliferation, migration, apoptosis, and invasion of gastric cancer cells were detected and compared. Western blotting, cell viability assay, fluorescence, and transfection were employed to study the molecule mechanism of the effects and the interaction between YAP and β-catenin signaling. Results Simvastatin could inhibit proliferation, migration and invasion, and promote the apoptosis in gastric cancer cells. Mechanistic studies showed that simvastatin treatment could inhibit the expression of β-catenin and the activity of YAP and the downstream targets of YAP and β-catenin in gastric cancer cells. Moreover, we found that YAP and β-catenin could form a positive feedback loop in gastric cancer cells. Further investigation revealed that simvastatin mainly acted through by inhibiting the activity of RhoA to inhibit YAP and β-catenin, and the geranylgeranyl pyrophosphate pathway mediated this regulation. Conclusion Statins represent a promising therapeutic option for gastric cancer by simultaneously targeting YAP and β-catenin signaling.

Both the MAPK pathway and mevalonate (MVA) signaling pathway play an increasingly significant role in the carcinogenesis of colorectal carcinoma, whereas the cross-talk between these two pathways and its implication in targeted therapy remains unclear in colorectal carcinoma. Here, we identified that HMGCS1 (3-hydroxy-3-methylglutaryl-CoA synthase 1), the rate-limiting enzyme of the MVA pathway, is overexpressed in colon cancer tissues and positively regulates the cell proliferation, migration, and invasion of colon cancer cells. In addition, HMGCS1 could enhance the activity of pERK independent of the MVA pathway, and the suppression of HMGCS1 could completely reduce the EGF-induced proliferation of colon cancer cells. Furthermore, we found that trametinib, a MEK inhibitor, could only partially abolish the upregulation of HMGCS1 induced by EGF treatment, while combination with HMGCS1 knockdown could completely reverse the upregulation of HMGCS1 induced by EGF treatment and increase the sensitivity of colon cancer cells to trametinib. Finally, we combined trametinib and dipyridamole, a common clinically used drug that could suppress the activity of SREBF2 (sterol regulatory element-binding transcription factor 2), a transcription factor regulating HMGCS1 expression, and identified its synergistic effect in inhibiting the proliferation and survival of colon cancer cells in vitro as well as the in vivo tumorigenic potential of colon cancer cells. Together, the current data indicated that HMGCS1 may be a novel biomarker, and the combination of targeting HMGCS1 and MEK might be a promising therapeutic strategy for patients with colon cancer.

Protein aggregation and degradation via autophagy (aggrephagy) are major strategies adopted by cells to remove misfolded polypeptides when there is proteasome dysfunction. The functional protein complex consisting of heat shock protein 70 (Hsp70), cochaperone ubiquitin ligase carboxyl-terminal of Hsp70/Hsp90 interacting protein (CHIP), and co-chaperone Bcl-2-associated athanogene 3 (BAG3) has been associated with the activation of protein aggregation. However, data on the mechanisms of action of the complex in the protein degradation remains scant. Here, we report that upon proteasome stress, the M2 isoform of pyruvate kinase (PKM2) promotes the aggregation of ubiquitinated proteins and its knockout or knockdown aggravates the sensitivity of cells to proteasome inhibitors. Besides, following proteasome inhibition, PKM2 promotes the interaction of BAG3 with CHIP and HSP70. Interestingly, re-expression of loss-of-function mutants in PKM2-knockout cells showed that the regulatory function of PKM2 in this progress does not depend on the activity of glycolytic enzymes or protein kinases. Taken together, these findings demonstrate that PKM2 mediates the formation of the CHIP-HSP70-BAG3 protein complex and promotes the aggregation of ubiquitinated misfolded proteins, thus compensating for proteasome stress in cells.

Colorectal cancer (CRC) is a heterogeneous disease at the cellular and molecular levels. Kirsten rat sarcoma (KRAS) is a commonly mutated oncogene in CRC, with mutations in approximately 40% of all CRC cases; its mutations result in constitutive activation of the KRAS protein, which acts as a molecular switch to persistently stimulate downstream signaling pathways, including cell proliferation and survival, thereby leading to tumorigenesis. Patients whose CRC harbors KRAS mutations have a dismal prognosis. Currently, KRAS mutation testing is a routine clinical practice before treating metastatic cases, and the approaches developed to detect KRAS mutations have exhibited favorable sensitivity and accuracy. Due to the presence of KRAS mutations, this group of CRC patients requires more precise therapies. However, KRAS was historically thought to be an undruggable target until the development of KRASG12C allele-specific inhibitors. These promising inhibitors may provide novel strategies to treat KRAS-mutant CRC. Here, we provide an overview of the role of KRAS in the prognosis, diagnosis and treatment of CRC.

Background: The Hippo/YAP pathway plays an important role in the development of cancers. Previous studies have reported that bile acids can activate YAP (Yes Associated Protein) to promote tumorigenesis and tumor progression. Ursodeoxycholic acid (UDCA) is a long-established old drug used for cholestasis treatment. So far, the effect of UDCA on YAP signaling in colorectal cancer (CRC) is not well defined. The study means to explore relationship of UDCA and YAP in CRC.Methods: The effect of UDCA on CRC cells proliferation was examined by MTT and clonogenic assay, western blotting (WB), real-time PCR, seperation of nucleoprotein and cytoplasmic protein and immunofluorescence (IF) assays were used to detect expression of YAP, WB, real-time PCR, GST-pull down assay and rhodamine-labeled phalloidin staining assays were used to detect activity of RhoA. After transfection of YAP5SA plasmids and RhoAV14 plasmids respectively, MTT, clonogenic assay, EdU incorporation assays and WB were performed. WB and ELISA were used to detect activity of cAMP/PKA axis, after blocking cAMP/PKA axis with siRNA and inhibitors, MTT, WB, IF and clonogenic assay were performed to test effect of UDCA on CRC cells. WB, IF and real-time PCR assays were used to detect expression of TGR5 in CRC cells after UDCA treatment, and then TGR5 agonist, TGR5 antagonist and siRNA were used to detect effect of UDCA on CRC cells. AOM/DSS-induced primary model was used to detect effect of UDCA, WB and Immunohistochemistry (IHC) were used to detect expression of related proteins.Results: UDCA suppressed YAP signaling by activating the membrane G‐protein‐coupled bile acid receptor (TGR5). TGR5 mainly regulated cAMP/PKA signaling pathway to inhibit RhoA activity, thereby suppressing YAP signaling. Moreover, the restoration of YAP expression alleviated the inhibitory effect of UDCA on CRC cell proliferation. In AOM/DSS-induced CRC model, UDCA inhibited tumor growth in a concentration-dependent manner and decreased expression of YAP and Ki67.Conclusion: UDCA plays a distinguished role in regulating YAP signaling and CRC growth from the primary bile acids and partial secondary bile acids, demonstrating the importance of maintaining normal intestinal bile acid metabolism in cancer patients. It also presents a potential therapeutic intervention for CRC.

Ferroptosis, a new form of programmed necrosis characterized by iron-dependent lethal accumulation of lipid hydroperoxides, is associated with many human diseases. Targeting amino acid (AA) availability can selectively suppress tumor growth and has been a promising therapeutic strategy for cancer therapy. Compelling studies have indicated that AA metabolism is also involved in ferroptosis, closely regulating its initiation and execution. This manuscript systematically summarizes the latest advances of AA metabolism in regulating ferroptosis and discusses the potential combination of therapeutic strategies that simultaneously target AA metabolism and ferroptosis in cancer to eliminate tumors or limit their invasiveness.

Background Colon cancer is one of the most common cancers worldwide. IFN-stimulated gene 15 (ISG15), a ubiquitin-like molecule, is strongly up-regulated by type I interferon as a crucial response to a variety of microbial and cellular stress stimuli. However, the role of ISG15 in colon cancer remains unclear. Methods The expression of ISG15 in colon cancer tissues and cell lines was detected by using Western blotting and immunohistochemistry. ISG15 expression levels of colon cancer cells treated with trametinib was verified by using the data downloaded from the Gene Expression Omnibus (GEO) databases, quantitative real-time PCR analysis and Western blots assays. ISG15-siRNA was used to silence ISG15 in colon cancer cell line to determine the roles of ISG15 in colon cancer cell proliferation. Results ISG15 was highly expressed in colon cancer tissues and ISG15 upregulation was closely associated with poor prognoses in colon cancer patients. Enhanced ISG15 expression promoted the migration and proliferation of colon cancer cells in vitro, while ISG15 knockdown decreased cell proliferation and metastasis. In addition, we first found that the mRNA and protein expression of ISG15 were up-regulated following trametinib treatment. Further investigation showed that ISG15 knockdown could enhance the anti-cancer effect of trametinib in colon cancer cells. Conclusion We proposed an interesting possibility that ISG15 may be a prognostic bio-marker, and the combined targeting of ISG15 and MEK might be a promising therapeutic strategy for colon cancer.

Background Hepatocellular carcinoma (HCC) is one of the most common malignant tumours worldwide and has a poor prognosis. Sorafenib, the only targeted therapeutic agent for HCC, is a multiple kinase inhibitor with targets including RAF and VEGFR-2/3 that display a very limited ability to extend the survival of patients with advanced metastatic HCC for approximately three months. MEK inhibitors including trametinib and selumetinib have shown promising efficacy in combination with sorafenib in clinical trials. However, the mechanisms about the combined effect of these drugs remain unclear. Methods Two HCC cell lines (Bel7402 and SMMC7721) were used in the experiments. The protein expression of HCC cell lines was quantified via western blotting. Cell viability was examined by cell counting kit-8 and colony formation assays. Drug interactions between sorafenib and trametinib/selumetinib were determined by the combination index (CI) value. Results In this study, we found that short-term sorafenib treatment could inhibit the downstream RAF effectors phosphorylated (p)-MEK and p-ERK in Bel7402 and SMMC7721 cells, while long-term sorafenib treatment could induce a rebound of p-MEK and p-ERK expression in these two human HCC cell lines. We then tested the effect of sorafenib combined with two different FDA-approved MEK inhibitors, trametinib and selumetinib, in the two cell lines. Western blot analysis showed that trametinib/selumetinib could abolish the ERK activation caused by long-term sorafenib treatment. Cell counting kit-8 and colony formation assays indicated that the use of sorafenib or trametinib/selumetinib alone produced a minor effect on the proliferation of these HCC cell lines, while the combination therapies induced strong growth inhibition. CI assays using CompuSyn software indicated that the combined therapies could produce a synergistic effect in these two cell lines. In addition, mechanistic studies revealed that the combination therapies could synergistically reduce the expression of proliferation-related proteins, including cyclin D1 and c-Myc. Conclusions Taken together, our study showed that the rebound of p-ERK induced by long-term sorafenib treatment might limit the benefit of sorafenib monotherapy, and the MEK inhibitors trametinib and selumetinib could enhance the efficacy of sorafenib in HCC cells.

Tumours manage to survive the ablation of mutant KRAS, despite the development of KRAS-targeted drugs. Here we describe that inhibition of mutant KRAS promotes MEK nuclear localization as an alternative mechanism of KRAS-targeted drugs resistance. Tissue microarray analysis in colon tumours shows that aberrant MEK nuclear localization is closely related to YAP levels and tumour malignancy. MEK nuclear localization could sequester β-TrCP from cytoplasmic inactive YAP, then stabilizing YAP. Mutant KRAS restrains MEK within the cytoplasm via IQGAP1, inhibiting MEK nuclear translocation. Trametinib, an allosteric MEK inhibitor, could prevent MEK nuclear localization and subsequently promote YAP degradation. In vitro and in vivo results suggests that inhibition of MEK nuclear localization by trametinib synergizes with KRAS knockdown or deltarasin treatment in suppressing the viability of KRAS mutant colon cancer cells. Our study provides new insights into the mechanisms of resistance to KRAS ablation, and suggests novel strategies for the treatment of KRAS-mutant colon cancers.

Serotonin (5-hydroxytryptamine, 5-HT) metabolism has long been linked to tumorigenesis and tumor progression. Numerous studies have shown the functions of 5-HT and its metabolites in the regulation of tumor biological processes like cell proliferation, invasion, metastasis, tumor angiogenesis and immunomodulatory through multi-step complex mechanisms. Reprogramming of 5-HT metabolism has been revealed in various tumors paving way for development of drugs that target enzymes, metabolites or receptors involved in 5-HT metabolic pathway. However, information on the role of 5-HT metabolism in cancer is scanty. This review briefly describes the main metabolic routes of 5-HT, the role of 5-HT metabolism in cancer and systematically summarizes the most recent advances in 5-HT metabolism-targeted cancer therapy.

Lipid homeostasis plays a fundamental role in the development of hepatocellular carcinoma (HCC). However, the mechanisms that regulate lipid homeostasis to avoid lipotoxicity in HCC remain elusive. Here, we found high-fat diet (HFD) improved the expression of sterol o-acyltransferase1 (SOAT1) and carnitine palmitoyltransferase 1A (CPT1A) in diethylnitrosamine-induced HCC. Bioinformatic analysis showed that SOAT1-mediated fatty acid storage and CPT1A-mediated fatty acids oxidation (FAO) formed a double-negative feedback loop in HCC. We verified that SOAT1 inhibition enhanced CPT1A protein, which shuttled the released fatty acids into the mitochondria for oxidation in vivo and in vitro. Besides, we further confirmed that CPT1A inhibition converted excess fatty acids into lipid drops by SOAT1 in vitro. Simultaneously targeting SOAT1 and CPT1A by the small-molecule inhibitors avasimibe and etomoxir had synergistic anticancer efficacy in HCC in vitro and in vivo. Our study provides new mechanistic insights into the regulation of lipid homeostasis and suggests the combination of avasimibe and etomoxir is a novel strategy for HCC treatment.

Background Serotonin signaling has been associated with tumorigenesis and tumor progression. Targeting the serotonin transporter to block serotonin cellular uptake confers antineoplastic effects in various tumors, including colon cancer. However, the antineoplastic mechanism of serotonin transporter inhibition and serotonin metabolism alterations in the absence of serotonin transporter have not been elucidated, especially in colon cancer, which might limit anti-tumor effects associating with targeting serotonin transporter. Methods The promotion in the uptake and catabolism of extracellular tryptophan and targeting serotonin transporter was detected by using quantitative reverse-transcription polymerase chain reaction, western blotting and liquid chromatography tandem mass spectrometry. Western blotting Immunoprecipitation and immunofluorescence was utilized to research the serotonylation of mTOR by serotonin and serotonin transporter inhibition. The primary mouse model, homograft model and tissue microarry was used to explore the tryptophan pathway in colon cancer. The cell viability assay, western blotting, xenograft and primary colon cancer mouse model were used to identify whether the combination of sertraline and tryptophan restriction had a synergistic effect. Results Targeting serotonin transporter through genetic ablation or pharmacological inhibition in vitro and in vivo induced a compensatory effect by promoting the uptake and catabolism of extracellular tryptophan in colon cancer. Mechanistically, targeting serotonin transporter suppressed mTOR serotonylation, leading to mTOR inactivation and increased tryptophan uptake. In turn, this process promoted serotonin biosynthesis and oncogenic metabolite kynurenine production through enhanced tryptophan catabolism. Tryptophan deprivation, or blocking its uptake by using trametinib, a MEK inhibitor, can sensitize colon cancer to selective serotonin reuptake inhibitors. Conclusions The present study elucidated a novel feedback mechanism involved in the regulation of serotonin homeostasis and suggested innovative strategies for selective serotonin reuptake inhibitors-based treatment of colon cancer.

Additional file 12: Table S1. Primer sequence.

Additional file 1: Table S1. Associations between the clinicopathological characteristics of patients and EMT-related gene signature risk level.

Cholesterol and its metabolites (precursors and derivatives) play an important role in cancer. In recent years, numerous studies have reported the functions of cholesterol metabolism in the regulation of tumor biological processes, especially oncogenic signaling pathways, ferroptosis, and tumor microenvironment. Preclinical studies have over the years indicated the inhibitory effects of blocking cholesterol synthesis and uptake on tumor formation and growth. Besides, some new cholesterol metabolic molecules such as SOAT1, SQLE, and NPC1 have recently emerged as promising drug targets for cancer treatment. Here, we systematically review the roles of cholesterol and its metabolites, and the latest advances in cancer therapy targeting cholesterol metabolism.

Background Primitive electronic waste (e-waste) recycling is ongoing in Guiyu, so toxic heavy metals may continue to threaten the health of children in the area. Objective This study primarily aimed to evaluate the effect of e-waste exposure on haemoglobin (Hb) synthesis in preschool children. Methods Medical examinations were conducted with the permission of children’s guardians and the approval of the Ethics Committee of the Medical College of Shantou University. This study recruited 224 children (aged 3–6 years, exposed group) who lived in Guiyu and 204 children (aged 3–6 years, control group) who lived in a town free of e-waste pollution. Blood levels of lead, Hb, ferritin, folate and vitamin B12 were tested in all children. Furthermore, all children were assessed for thalassemia, and their parents were asked to fill in questionnaires. Results There were no significant differences in the level of ferritin, folate, or vitamin B12 between the exposed and control groups (P > 0.05). No children were identified as having thalassemia in all study participants. Blood lead level (BLL) and the risk of children with BLL ≥ 10 µg/dL in the exposed group were significantly higher than those in the control group (all P P = 0.03), but not in the control group (P = 0.14). Hb levels in group B and group C were also significantly lower in the exposed group than in the control group (Group B: 122.6 ± 9.5 g/L versus 125.8 ± 8.2 g/L, P = 0.01; Group C: 120.3 ± 7.3 g/L versus 123.6 ± 8.3 g/L, P = 0.03). In addition, the prevalence of anaemia associated with BLLs above 10 µg/dL and between 5.0 and 9.9 µg/dL were both significantly higher in the exposed group than in the control group (4.0% vs. 0.5%, 5.4% vs. 1.5%, respectively, both P Conclusion Lead exposure more significantly inhibits Hb synthesis in children who live in e-waste dismantling areas than in those who live in non-e-waste dismantling areas. Other toxins released from e-waste may also contribute to the inhibition of Hb synthesis and may lead to anaemia in local children. Further investigations are needed to provide evidence for the development of relevant protective measures.

In this paper, the working principle and damage mechanism of the high-pressure manifold swivel joints were analyzed. The pressure distribution and velocity variation of the medium in the swivel joint are studied by CFD. The erosion areas and damage patterns of the swivel joints were investigated by practical research and theoretical analysis. Optimization design and improvement of the material, structural and processing technology were suggested. Those results can provide a reference for the design, manufacture and safety assessment of swivel joints.

To evaluate the differential expression profiles of the lncRNAs, miRNAs, mRNAs and ceRNAs, and their implication in the prognosis in clear cell renal cell carcinoma (CCRCC), the large sample genomics analysis technologies were used in this study. The RNA and miRNA sequencing data of CCRCC were obtained from The Cancer Genome Atlas (TCGA) database, and R software was used for gene expression analysis and survival analysis. Cytoscape software was used to construct the ceRNA network. The results showed that a total of 1 570 lncRNAs, 54 miRNAs, and 17 mRNAs were differentially expressed in CCRCC, and most of their expression levels were up-regulated (false discovery rate0.01 and absolute log fold change2). The ceRNA regulatory network showed the interaction between 89 differentially expressed lncRNAs and 9 differentially expressed miRNAs. Further survival analysis revealed that 38 lncRNAs (including COL18A1-AS1, TCL6, LINC00475, UCA1, WT1-AS, HOTTIP, PVT1, etc.) and 2 miRNAs (including miR-21 and miR-155) were correlated with the overall survival time of CCRCC (本文旨在通过大样本基因组学分析技术，评估 lncRNA、miRNA、mRNA 及 ceRNA 在肾透明细胞癌中的差异表达情况及其预后价值。利用 R 软件对 TCGA 数据库中肾透明细胞癌 RNA 和 miRNA 数据进行基因差异表达分析和生存分析，并通过 cytoscape 软件得到差异表达的 lncRNA-miRNA-mRNA 之间的 ceRNA 调控关系网。结果发现共有 1 570 个 lncRNA、54 个 miRNA 和 17 个 mRNA 在肾透明细胞癌组织中存在异常表达，且其表达水平以上调为主（错误发现率0.01；对数差异表达倍数变化绝对值2）。ceRNA 调控网显示共 89 个差异表达的 lncRNA 和 9 个差异表达的 miRNA 之间存在相互作用关系,生存分析共鉴定出 COL18A1-AS1、TCL6、LINC00475、UCA1、WT1-AS、HOTTIP、PVT1 等 38 个有预后价值的 lncRNA 和 2 个有预后价值的 miRNA（miR-21 和 miR-155）（

Background: Both the mitogen-activated protein kinase pathway and mevalonate signaling pathway play a increasingly significant role in the carcinogenesis of colorectal carcinoma (CRC). whereas, the crosstalk between these two pathways and its implication in targeted therapy remains unclear in CRC. Methods: The expression of HMGCS1 in colon cancer tissues and cell lines was detected by using western blotting and immunohistochemistry. A cell viability assay, clonogenic assay, western blot assay were employed to study the interaction between si-RNA of HMGCS1 and EGF treatment. Xenograft studies were used to identify whether the combination of trametinib and dipyridamole had a synergistic effect. Findings: We identified that HMGCS1 is overexpressed in colon cancer tissues and positively regulates the cell proliferation, migration and invasion of colon cancer cells. In addition, HMGCS1 could enhance the activity of pERK independent of the MVA pathway, and the suppression of HMGCS1 could completely reduce the EGF-induced proliferation of colon cancer cells. Furthermore, we found that trametinib could only partially abolish the upregulation of HMGCS1 induced by EGF treatment, while combination with HMGCS1 knockdown could completely reverse the upregulation of HMGCS1 induced by EGF treatment and increase the sensitivity of colon cancer cells to trametinib. Finally, we found that dipyridamole could enhance the cytotoxicities of the trametinib against colon cancer cells in vitro and in vivo. Interpretation: The present data indicated that HMGCS1 may be a novel biomarker, and the combination of targeting HMGCS1 and MEK might be a promising therapeutic strategy for colon cancer patients. Funding Statement: This study was supported by the National Natural Science Foundation of China (81572731). Declaration of Interests: The authors declare that they have no conflicts of interest. Ethics Approval Statement: The animals were handled according to the guidelines of the China Animal Welfare Legislation, as provided by the Committee on Ethics in the Care and Use of Laboratory Animals of Sichuan University West China Hospital. The human colon cancer tissues as provided by the Ethics Committee of Taizhou Hospital of Zhejiang Province.

// Hongwei Xia 1,* , Qingjuan Chen 2,3,* , Ying Chen 2 , Xiaojun Ge 1 , Weibing Leng 2 , Qiulin Tang 1 , Ming Ren 2 , Liang Chen 1 , Dandan Yuan 2 , Yucheng Zhang 1 , Ming Liu 2 , Qiyong Gong 4 and Feng Bi 1,2 1 Laboratory of Signal Transduction & Molecular Targeted Therapy, State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University, and Collaborative Innovation Center for Biotherapy, Chengdu, Sichuan Province, China 2 Department of Medical Oncology, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China 3 Department of Medical Oncology, Xian Yang Central Hospital, Xian Yang City, Shanxi Province, China 4 Department of Radiology, West China Hospital of Sichuan University, Chengdu, Sichuan Province, China * These authors have contributed equally to this work Correspondence to: Feng Bi, email: // Keywords : long non-coding RNA, MALAT1, gastric cancer, metastasis, miR-122 Received : April 29, 2016 Accepted : May 29, 2016 Published : July 29, 2016 Abstract The metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is frequently over-expressed and serves as a prognostic marker in human cancers. However, little is known about the role of MALAT1 in gastric cancer. Here, we reported that the tissue and plasma MALAT1 levels were significantly higher in gastric cancer patients with distant metastasis (P

Aberrant activation of the Src kinase is implicated in the development of a variety of human malignancies. However, it is almost impossible to monitor Src activity in an in vivo setting with current biochemical techniques. To facilitate the noninvasive investigation of the activity of Src kinase both in vitro and in vivo, we developed a genetically engineered, activatable bioluminescent reporter using split-luciferase complementation. The bioluminescence of this reporter can be used as a surrogate for Src activity in real time. This hybrid luciferase reporter was constructed by sandwiching a Src-dependent conformationally responsive unit (SH2 domain-Srcpep) between the split luciferase fragments. The complementation bioluminescence of this reporter was dependent on the Src activity status. In our study, Src kinase activity in cultured cells and tumor xenografts was monitored quantitatively and dynamically in response to clinical small-molecular kinase inhibitors, dasatinib and saracatinib. This system was also applied for high-throughput screening of Src inhibitors against a kinase inhibitor library in living cells. These results provide unique insights into drug development and pharmacokinetics/phoarmocodynamics of therapeutic drugs targeting Src signaling pathway enabling the optimization of drug administration schedules for maximum benefit. Using both Firefly and Renilla luciferase imaging, we have successfully monitored Src tyrosine kinase activity and Akt serine/threonine kinase activity concurrently in one tumor xenograft. This dual luciferase reporter imaging system will be helpful in exploring the complex signaling networks in vivo. The strategies reported here can also be extended to study and image other important kinases and the cross-talks among them.

e13099 Background: YAP and β-catenin play critical roles in the carcinogenesis of several cancers, but their functions in chemoresistance remain unclear in triple-negative breast cancer (TNBC). Herein, we examined the expression and function of the two proteins in the chemoresistance of TNBC. Methods: We examined the expression levels of YAP and β-catenin in TNBC samples by immunohistochemistry (IHC) and their roles in prognosis. Dynamic changes in the two proteins in TNBC tissues were determined by RT-PCR. CCK-8 assays, colony formation assays, flow cytometry, migration assays and nude mouse models were used to investigate the functions of YAP and β-catenin in drug-resistant TNBC cells. The involved signalling pathways were detected by western blot assays. Results: TNBC patients with high YAP and/or β-catenin expression had a higher risk of relapse or mortality than patients with low YAP and/or β-catenin expression, and changes in the expression of the two proteins might be a promising predictive biomarker for neoadjuvant chemotherapy sensitivity. Inhibition of YAP or β-catenin enhanced the cytotoxicity of the anti-microtubule agents docetaxel and vinorelbine against TNBC cells in vitro and in vivo. Moreover, YAP and β-catenin were upregulated in docetaxel- or vinorelbine-resistant TNBC cells, while inhibition of YAP or β-catenin enhanced the sensitivity of drug-resistant cells. Interestingly, aspirin, a kind of acetylsalicylic acid, reversed the drug resistance of cells and in combination with aspirin and docetaxel or vinorelbine significantly inhibited the growth of drug-resistant TNBC cells. The involved mechanism is that aspirin impaired YAP or β-catenin expression by upregulating the E3 ubiquitin ligase β-TrCP. Conclusions: Both YAP and β-catenin act as key biomarkers of prognosis and anti-microtubule drug resistance in TNBC, and the combined use of aspirin and anti-microtubule drugs inhibits YAP and β-catenin, presenting several promising therapeutic approaches for TNBC treatment.

Aberrant activation of the Src kinase is implicated in the development of a variety of human malignancies. However, it is almost impossible to monitor Src activity in an in vivo setting with current biochemical techniques. To facilitate the noninvasive investigation of the activity of Src kinase both in vitro and in vivo, we developed a genetically engineered, activatable bioluminescent reporter using split-luciferase complementation. The bioluminescence of this reporter can be used as a surrogate for Src activity in real time. This hybrid luciferase reporter was constructed by sandwiching a Src-dependent conformationally responsive unit (SH2 domain-Srcpep) between the split luciferase fragments. The complementation bioluminescence of this reporter was dependent on the Src activity status. In our study, Src kinase activity in cultured cells and tumor xenografts was monitored quantitatively and dynamically in response to clinical small-molecular kinase inhibitors, dasatinib and saracatinib. This system was also applied for high-throughput screening of Src inhibitors against a kinase inhibitor library in living cells. These results provide unique insights into drug development and pharmacokinetics/phoarmocodynamics of therapeutic drugs targeting Src signaling pathway enabling the optimization of drug administration schedules for maximum benefit. Using both Firefly and Renilla luciferase imaging, we have successfully monitored Src tyrosine kinase activity and Akt serine/threonine kinase activity concurrently in one tumor xenograft. This dual luciferase reporter imaging system will be helpful in exploring the complex signaling networks in vivo. The strategies reported here can also be extended to study and image other important kinases and the cross-talks among them.