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The mammalian target of rapamycin (mTOR) pathway is abnormally activated in lung cancer. However, the anti-lung cancer effect of mTOR inhibitors as monotherapy is modest. Here, we identified that ginsenoside Rh2, an active component of Panax ginseng C. A. Mey., enhanced the anti-cancer effect of the mTOR inhibitor everolimus both in vitro and in vivo. Moreover, ginsenoside Rh2 alleviated the hepatic fat accumulation caused by everolimus in xenograft nude mice models. The combination of everolimus and ginsenoside Rh2 (labeled Eve-Rh2) induced caspase-independent cell death and cytoplasmic vacuolation in lung cancer cells, indicating that Eve-Rh2 prevented tumor progression by triggering paraptosis. Eve-Rh2 up-regulated the expression of c-MYC in cancer cells as well as tumor tissues. The increased c-MYC mediated the accumulation of tribbles homolog 3 (TRIB3)/P62+ aggresomes and consequently triggered paraptosis, bypassing the classical c-MYC/MAX pathway. Our study offers a potential effective and safe strategy for the treatment of lung cancer. Moreover, we have identified a new mechanism of TRIB3/P62+ aggresomes-triggered paraptosis and revealed a unique function of c-MYC.

Colorectal cancer (CRC) is the third most common cancer and the second leading cause of death due to cancer in the world. Therefore, the identification of novel druggable targets is urgently needed. Tribbles proteins belong to a pseudokinase family, previously recognized in CRC as oncogenes and potential therapeutic targets. Here, we analyzed the expression of TRIB1, TRIB2, and TRIB3 simultaneously in 33 data sets from CRC based on available GEO profiles. We show that all three Tribbles genes are overrepresented in CRC cell lines and primary tumors, though depending on specific features of the CRC samples. Higher expression of TRIB2 in the tumor microenvironment and TRIB3 overexpression in an early stage of CRC development, unveil a potential and unexplored role for these proteins in the context of CRC. Differential Tribbles expression was also explored in diverse cellular experimental conditions where either genetic or pharmacological approaches were used, providing novel hints for future research. This comprehensive bioinformatic analysis provides new insights into Tribbles gene expression and transcript regulation in CRC. info:eu-repo/semantics/publishedVersion

Pulmonary fibrosis (PF) is a chronic, progressive, fatal interstitial lung disease with limited available therapeutic strategies. We recently reported that the protein kinase glycogen synthase kinase-3β (GSK-3β) interacts with and inactivates the ubiquitin-editing enzyme A20 to suppress the degradation of the transcription factor CCAAT/enhancer-binding protein beta (C/EBPβ) in alveolar macrophages (AMs), resulting in a profibrotic phenotype of AMs and promoting the development of PF. Here, we showed that chronic lung injury upregulated the stress response protein tribbles homolog 3 (TRIB3), which interacted with GSK-3β and stabilized GSK-3β from ubiquitination and degradation. Elevated GSK-3β expression phosphorylated A20 to inhibit its ubiquitin-editing activity, causing the accumulation of C/EBPβ and the production of several profibrotic factors in AMs and promoting PF development. Activated C/EBPβ, in turn, increased the transcription of TRIB3 and GSK-3β, thereby establishing a positive feedback loop in AMs. The knockdown of TRIB3 expression or the pharmacologic disruption of the TRIB3‒GSK-3β interaction was an effective PF treatment. Our study reveals an intact profibrotic axis of TRIB3‒GSK-3β‒A20‒C/EBPβ in AMs, which represents a target that may provide a promising treatment strategy for PF.

Tumor‐associated macrophages (TAMs), one of the most common cell components in the tumor microenvironment, have been reported as key contributors to cancer‐related inflammation and enhanced metastatic progression of tumors. To explore the underlying mechanism of TAM‐induced tumor progression, TAMs were isolated from colorectal cancer patients, and the functional interaction with colorectal cancer cells was analyzed. Our study found that coculture of TAMs contributed to a glycolytic state in colorectal cancer, which promoted the stem‐like phenotypes and invasion of tumor cells. TAMs produced the cytokine transforming growth factor‐β to support hypoxia‐inducible factor 1α (HIF1α) expression, thereby upregulating Tribbles pseudokinase 3 (TRIB3) in tumor cells. Elevated expression of TRIB3 resulted in activation of the β‐catenin/Wnt signaling pathway, which eventually enhanced the stem‐like phenotypes and cell invasion in colorectal cancer. Our findings provided evidence that TAMs promoted colorectal cancer progression in a HIF1α/TRIB3‐dependent manner, and blockade of HIF1α signals efficiently improved the outcome of chemotherapy, describing an innovative approach for colorectal cancer treatment., Our findings provided evidence that tumor‐associated macrophages promoted colorectal progression in a HIF1α/TRIB3‐dependent manner, and blockade of HIF1α signals efficiently improved the outcome of chemotherapy, describing an innovative approach for colorectal cancer treatment.

Long non-coding RNA (lncRNA) X-inactive specific transcript (Xist) is involved in apoptosis and inflammatory injury. This study aimed to assess the role of lncRNA Xist in sevoflurane-induced social and emotional impairment and neuronal apoptosis in neonatal mice and hippocampal neuronal cells. The performance in social and emotional tests and the expression levels of lncRNA Xist and microRNA (miR)-98-5p after sevoflurane exposure were measured. Moreover, the effects of suppression of lncRNA Xist on neuronal apoptosis and endoplasmic reticulum (ER) stress were determined. Subsequently, the association among lncRNA Xist, miR-98-5p, and ER degradation-enhancing α-mannosidase-like 1 protein (EDEM1) was explored. Our results showed that lncRNA Xist increased, miR-98-5p decreased, and social and emotional impairment appeared after sevoflurane exposure. Furthermore, suppression of lncRNA Xist improved sevoflurane-induced social and emotional impairment and reduced sevoflurane-induced neuronal apoptosis and ER stress in vivo and in vitro. Moreover, lncRNA Xist negatively regulated miR-98-5p expression, and it contributed to sevoflurane-induced neuronal apoptosis and ER stress by sponging miR-98-5p. Additionally, EDEM1 was identified as a target of miR-98-5p. Our findings revealed that the knockdown of lncRNA Xist ameliorates sevoflurane-induced social and emotional impairment through inhibiting neuronal apoptosis and ER stress by targeting the miR-98-5p/EDEM1 axis., Graphical abstract, Chen and colleagues proved that lncRNA XIST knockdown can ameliorate sevoflurane-induced social and emotional impairment through inhibiting neuron apoptosis and endoplasmic reticulum stress by targeting the miR-98-5p/EDEM1 axis. The understanding of cell death mechanisms in anesthesia-caused social and emotional impairment is vital to improve interventions in patients and reduce neural dysfunctionality.

The current understanding of the molecular pathogenesis of diabetic retinopathy does not provide a mechanistic link between early molecular changes and the subsequent progression of the disease. In this study, we found that human diabetic retinas overexpressed TRIB3 and investigated the role of TRIB3 in diabetic retinal pathobiology in mice. We discovered that TRIB3 controlled major molecular events in early diabetic retinas via HIF1α-mediated regulation of retinal glucose flux, reprogramming cellular metabolism, and governing of inflammatory gene expression. These early molecular events further defined the development of neurovascular deficit observed in mice with diabetic retinopathy. TRIB3 ablation in the streptozotocin-induced mouse model led to significant retinal ganglion cell survival and functional restoration accompanied by a dramatic reduction in pericyte loss and acellular capillary formation. Under hypoxic conditions, TRIB3 contributed to advanced proliferative stages by significant upregulation of GFAP and VEGF expression, thus controlling gliosis and aberrant vascularization in oxygen-induced retinopathy mouse retinas. Overall, our data reveal that TRIB3 is a master regulator of diabetic retinal pathophysiology that may accelerate the onset and progression of diabetic retinopathy to proliferative stages in humans and present TRIB3 as a potentially novel therapeutic target for diabetic retinopathy.

The transcription factor MYC is deregulated in almost all human cancers, especially in aggressive lymphomas, through chromosomal translocation, amplification, and transcription hyperactivation. Here, we report that high expression of tribbles homologue 3 (TRIB3) positively correlates with elevated MYC expression in lymphoma specimens; TRIB3 deletion attenuates the initiation and progression of MYC-driven lymphoma by reducing MYC expression. Mechanistically, TRIB3 interacts with MYC to suppress E3 ubiquitin ligase UBE3B-mediated MYC ubiquitination and degradation, which enhances MYC transcriptional activity, causing high proliferation and self-renewal of lymphoma cells. Use of a peptide to disturb the TRIB3-MYC interaction together with doxorubicin reduces the tumor burden in MycEμ mice and patient-derived xenografts. The pathophysiological relevance of UBE3B, TRIB3 and MYC is further demonstrated in human lymphoma. Our study highlights a key mechanism for controlling MYC expression and a potential therapeutic option for treating lymphomas with high TRIB3-MYC expression., c-MYC is often deregulated in human cancers including lymphomas. Here, the authors show that a member of the pseudokinase family, tribbles homologue 3 (TRIB3), interacts with c-MYC to suppress c-MYC ubiquitination and degradation, leading to increased proliferation and self-renewal of lymphoma cells.

The pseudokinase Tribble 3 (TRIB3) is known as a regulator in cellular responses to a variety of stresses, such as glucose insufficiency and endoplasmic reticulum (ER) stress. TRIB3 is upregulated in various cancer tissues and is closely connected to the poor prognosis of patients. However, the underlying regulation and function of TRIB3 in glioblastoma (GBM) is still largely unknown. In this study, the upregulation of TRIB3 was confirmed both in primary specimens from GBM patients and in vitro with GBM cell lines. Overexpression of specific TRIB3 transcripts promoted cell growth and migration in vitro, while knockdown of TRIB3 expression exerted a repressive effect on these cellular processes. The growth-promoting effect of TRIB3 was also demonstrated in a xenograft mouse model. Mechanistic studies further revealed that TRIB3 was able to suppress autophagic flux and that this suppression was responsible for TRIB3 silencing-induced proliferation and migration of GBM cells. These findings indicate that the suppression of autophagic flux by TRIB3 drives the invasion and proliferation of GBM cells, thus suggesting that TRIB3 is a potential novel therapeutic target for the treatment of glioma.

Background Tribbles pseudokinase 3 (TRIB3) protein is a pseudokinase which plays an important role in cellular stress, metabolism, and tumor progression. However, the expression and function of TRIB3 in ovarian cancer is unknown. Methods TRIB3 expression was detected by immunohistochemistry in the ovarian tissue samples. Following down-regulation of TRIB3 by siRNA, multiple aspects of ovarian cancer cells were detected by the MTT assay, flow cytometry, scratch test and Transwell. Additionally, changes in related molecules and the MEK/ERK pathway were detected by western blotting. Finally, many bioinformatic methods, websites and databases, such as gene set enrichment analysis (GSEA), DVAID, Genemania, TISIDB and cBioPortal were used to study the TRIB3. Results The expression level of TRIB3 was higher in ovarian epithelial malignant tumors as compared to other groups. Patients with a high expression level of TRIB3 had significantly shorter survival times,which was consistent with the results of analysis of the KM-plot database. Down-regulation of TRIB3 expression significantly inhibited the proliferation, invasion, and migration capabilities of ovarian cancer cells, and induced apoptosis and cell cycle arrest. Following TRIB3 siRNA transfection, expression levels of relative proteins were found to be decreased. Additionally, analysis in DAVID website and GSEA revealed that TRIB3 expression was associated with multiple biological processes. Protein phosphorylation levels of MEK and ERK also decreased following TRIB3-siRNA transfection. The Genemania website was used to analyze the proteins that interact with TRIB3. Analysis of TRIB3 in the TISIDB database and cBioPortal website showed that ovarian cancer patients with high levels of mutation in TRIB3 had poor prognosis, and that the expression of TRIB3 was related to immunomodulation. Conclusions The TRIB3 was highly expressed and promoting the malignant behavior of ovarian cancer cells by activating the MEK-ERK signaling pathway. It was also found to be associated with genetic variations and immune modulators.

Objective To explore the biological function and molecular mechanism of Sp2 in hepatocellular carcinoma (HCC). Methods Tissue microarray immunohistochemistry and western blot were used to study the expression of Sp2 in hepatocellular tissue and adjacent non‐neoplastic tissues (ANT). In HCC cell lines, the role of Sp2 was determined by in vitro experiments such as CCK8, clone formation test, Transwell assay, wound‐healing assay, and flow cytometry apoptotic analysis, and its possible mechanism was analyzed. Results Compared with ANT, Sp2 expression in HCC tissues was significantly up‐regulated, which was strongly associated with stage of tumor and poor prognosis of patients. TCGA database were further confirmed these results. Besides, functional studies had shown that Sp2 knockdown not only leads to a decrease in cell proliferation and an increase in cell apoptosis but also inhibits the cells' abilities of migration and invasion. Sp2 silencing could inhibit the expression of TRIB3 protein and down‐regulate the endoplasmic reticulum stress (ERS) level of HCC. Conclusion Sp2 may play a part in promoting cancer by regulating TRIB3 protein, which may be a factor of prognostic and a potential new therapeutic target for HCC., We found that Sp2 expression was significantly up‐regulated in HCC tissues, which was closely related to tumor staging and poor prognosis of patients. Functional studies had shown that Sp2 knockdown not only leads to a decrease in cell proliferation and an increase in cell apoptosis, but also inhibits migration and invasion of hepatoma cells. Sp2 may play a role in promoting cancer by regulating TRIB3 protein, which may be a prognostic factor and a potential new therapeutic target for HCC.

Under the differentiation induction therapy with all-trans retinoic acid and arsenic trioxide, nearly 95% of typical acute promyelocyte leukemia (APL), which is characterized by the presence of PML-RARA, patients can be cured. Though its good prognosis, if left untreated, the natural survival duration of typical APL patients is only 1 month, but some exceptional cases also exist. Occasionally, we have observed the entire natural clinical course of one extremely indolent APL patient, who developed from pre-APL stage (

© 2021 by the authors., [Background]: Tribbles pseudokinase 3 (TRIB3) has been proposed to both promote and restrict cancer generation and progression. However, the precise mechanisms that determine this dual role of TRIB3 in cancer remain to be understood. In this study we aimed to investigate the role of TRIB3 in luminal breast cancer, the most frequent subtype of this malignancy. [Methods]: We genetically manipulated TRIB3 expression in a panel of luminal breast cancer cell lines and analyzed its impact on cell proliferation, and the phosphorylation, levels, or subcellular localization of TRIB3 and other protein regulators of key signaling pathways in luminal breast cancer. We also analyzed TRIB3 protein expression in samples from luminal breast cancer patients and performed bioinfor-matic analyses in public datasets. Results: TRIB3 enhanced the proliferation and AKT phosphoryla-tion in luminal A (HER2-) but decreased them in luminal B (HER2+) breast cancer cell lines. TRIB3 negatively regulated the stability of HER2 in luminal B breast cancer cell lines. TRIB3 expression was associated with increased disease-free survival and a better response to therapy in luminal breast cancer patients. [Conclusions]: Our findings support the exploration of TRIB3 as a potential biomarker and therapeutic target in luminal breast cancer., This work was funded by the Instituto de Salud Carlos III (ISCIII) and confounded by the European Regional Development Fund (ERDF), “A way to make Europe”, grant number PI18/00442 integrated into the State Plan for R & D + I 2017-2020; by the European Commission through the Horizon 2020 European Training Networks program, grant number H2020-MSCA-ITN308 2016 721532; and by the Breast Cancer Now (formerly Breast Cancer Campaign) foundation, grant number 2012NovSP033. Alba Orea-Soufi was supported by a predoctoral FPU contract from Spanish Government. Work in SC-L group is supported by a MINECO/FEDER research grant RTI2018-094130-B-100.

Doublecortin-like kinase 1 (DCLK1) promotes tumour proliferation in human colorectal cancer (CRC). To elucidate the mechanism and clinical relevance of this association, we performed expression analysis using commercially available colon carcinoma cell lines (SW480, HCT116, CaCO2, SW48 and SKCO1) and immunohistochemical analysis of 200 resected CRC samples for correlation with clinical features. DCLK1 showed a high level of expression, especially in SW480 and HCT116 cells. Silencing DCLK1 expression using short hairpin DCLK1 (shDCLK1) RNA inhibited the growth and invasion capacities of these cell lines, which showed signs of entering into the mesenchymal–epithelial transition (MET). We found evidence of a strong correlation of DCLK1 expression with that of Tribbles homolog 3 (TRIB3), and silencing TRIB3 also led to the MET phenotype in these cells. In the clinical samples, compared with samples showing low expression of DCLK1, high expression was associated with poor prognosis in terms of overall and recurrence-free survival (P < 0.0001). The results of univariate and multivariate analysis suggested that high expression of DCLK1 in clinical colon cancer samples was tied to poor prognosis, cancer invasion depth and lymph node metastasis. DCLK1 expression correlates with malignant grade of colon cancer and offers a potential treatment target.

Although dyslipidemia commonly occurs in patients with acute promyelocytic leukemia (APL) in response to anti-APL therapy, the underlying mechanism and the lipid statuses of patients with newly diagnosed APL remain to be addressed. Methods: We conducted a retrospective study to investigate the lipid profiles of APL patients. PML-RARα transgenic mice and APL cells-transplanted mice were used to assess the effects of APL cells on the blood/liver lipid levels. Subsequently, gene set enrichment analysis, western blot and dual luciferase reporter assay were performed to examine the role and mechanism of PML-RARα and TRIB3 in lipid metabolism regulation in APL patients at pretreatment and after induction therapy. Results: APL patients exhibited a higher prevalence of dyslipidemia before anti-APL therapy based on a retrospective study. Furthermore, APL cells caused secretion of triglycerides, cholesterol, and PCSK9 from hepatocytes and degradation of low-density lipoprotein receptors in hepatocytes, which elevated the lipid levels in APL cell-transplanted mice and Pml-Rarα transgenic mice. Mechanistically, pseudokinase TRIB3 interacted with PML-RARα to inhibit PPARγ activity by interfering with the interaction of PPARγ and RXR and promoting PPARγ degradation. Thus, reduced PPARγ activity in APL cells decreased leptin but increased resistin expression, causing lipid metabolism disorder in hepatocytes and subsequent dyslipidemia in mice. Although arsenic/ATRA therapy degraded PML-RARα and restored PPARγ expression, it exacerbated dyslipidemia in APL patients. The elevated TRIB3 expression in response to arsenic/ATRA therapy suppressed PPARγ activity by disrupting the PPARγ/RXR dimer, which resulted in dyslipidemia in APL patients undergoing therapy. Indeed, the PPAR activator not only enhanced the anti-APL effects of arsenic/ATRA by suppressing TRIB3 expression but also reduced therapy-induced dyslipidemia in APL patients. Conclusion: Our work reveals the critical role of the PML-RARα/PPARγ/TRIB3 axis in the development of dyslipidemia in APL patients, potentially conferring a rationale for combining ATRA/arsenic with the PPAR activator for APL treatment.

Background Integrin αvβ3 has been proposed as crucial determinant for tumor sustained progression and a molecular marker for the estimation of tumor angiogenesis. Our study suggested that integrin αvβ3 could efficiently promote lung cancer cell proliferation and stem-like phenotypes in a tribbles homolog 3 (TRIB3) dependent manner. Result Integrin αvβ3 could mediate the activation of FAK/AKT pro-survival signaling pathway. Meanwhile, activated TRIB3 interacted with AKT to upregulated FOXO1 and SOX2 expression, resulting in sustained tumor progression in lung cancer. Our further analysis revealed that TRIB3 was significantly upregulated in lung tumor tissues and correlated with the poor outcome in clinical patients, indicating the potential role of TRIB3 in diagnostic and prognostic estimation for patients with lung cancer. Conclusion Our study showed here for the first time that integrin αvβ3 promote lung cancer development by activating the FAK/AKT/SOX2 axis in a TRIB3 dependent signaling pathway, and interrupting TRIB3/AKT interaction significantly improved the outcome of chemotherapy in tumor-bearing mice, representing a promising therapeutic strategy in lung cancer.

Eccentric exercise training accompanied by a low-fat diet can prevent insulin resistance (IR) and is currently an effective method for the treatment of IR induced by high-fat diet (HFD)-associated obesity. However, the molecular mechanisms underlying this improvement of IR in adipose tissue are still not completely clear. In this study, 5–6-week-old male mice were randomly divided into a standard control diet (SCD) group (SC, n = 12) and a HFD group (HF, n = 72). After 12 weeks, 12 mice in each group were randomly sacrificed. The remaining mice in the HF group were randomly submitted to one of the following experimental protocols for 8 weeks: obesity-HFD-sedentary (OHF-Sed, n = 14), obesity-HFD-exercise (OHF-Ex, n = 16), obesity-SCD-sedentary (OSC-Sed, n = 14), and obesity-SCD-exercise (OSC-Ex, n = 16). All obese mice in the exercise group were subjected to downhill running. Half of the mice in each group received an insulin injection (0.75 U/kg) before sample collection. Epididymal fat was removed and weighed. Adipocyte size and inflammatory cell infiltration were observed by H&E staining. Both basal and insulin-stimulated GLUT4 fluorescence and protein contents were detected by immunofluorescence and Western blot. Levels of IL-1β and IL-10 were detected by ELISA. Protein contents of iNOS, Arg-1, TRIB3, p-AKT, and AKT were determined by Western blot. CD86 and CD206 fluorescence were determined by immunofluorescence. The results showed that a HFD for 12 weeks induced IR accompanied by adipose tissue macrophages M1 polarization (increased iNOS protein content and CD86 fluorescence) and TRIB3-AKT activation. Downhill running accompanied by a low-fat diet attenuated IR (p < 0.01), reduced inflammation levels (increased IL-10 protein content and decreased IL-1β protein content), inhibited adipose tissue macrophages M1 polarization (decreased iNOS protein content and CD86 fluorescence) and promoted M2 polarization (increased Arg-1 protein content and CD206 fluorescence), and suppressed TRIB3-AKT signaling. We concluded that downhill running accompanied by dietary fat regulation attenuates HFD-related IR in mice, which may be associated with reduced TRIB3-AKT signaling and activated M2 macrophages in adipose tissue.

Interleukin (IL-6) is a pleotropic cytokine with both tumor-promoting and -inhibitory effects on breast cancer growth. However, the mechanisms governing the outcome of IL-6 on cancer progression remain to be clarified. Our study unraveled a novel long noncoding RNA (lncRNA) AU021063 downstream of IL-6 signaling. We found that IL-6 induced the expression of AU021063 predominantly in breast cancer compared to other cancer types. Mechanistically, IL-6 induced AT-rich interactive domain 5a (Arid5a) expression, which promotes the transcription of AU021063. In turn, AU021063 promotes breast cancer metastasis through stabilizing tribbles homolog 3 (Trib3) and activating Mek/Erk signaling pathway. Genetic ablation of either Arid5a, AU021063 or Trib3 abolished breast cancer metastasis in vitro and in vivo. Overall, our study highlights the importance of IL-6-Arid5a-AU021063 axis in regulating breast cancer invasiveness and metastasis, which may provide potential novel therapeutics for breast cancer.

© 2021 by the authors., The Tribbles pseudokinases family consists of TRIB1, TRIB2, TRIB3 and STK40 and, although evolutionarily conserved, they have distinctive characteristics. Tribbles members are expressed in a context and cell compartment-dependent manner. For example, TRIB1 and TRIB2 have potent oncogenic activities in vertebrate cells. Since the identification of Tribbles proteins as modulators of multiple signalling pathways, recent studies have linked their expression with several pathologies, including cancer. Tribbles proteins act as protein adaptors involved in the ubiquitin-proteasome degradation system, as they bridge the gap between substrates and E3 ligases. Between TRIB family members, TRIB2 is the most ancestral member of the family. TRIB2 is involved in protein homeostasis regulation of C/EBPα, β-catenin and TCF4. On the other hand, TRIB2 interacts with MAPKK, AKT and NFkB proteins, involved in cell survival, proliferation and immune response. Here, we review the characteristic features of TRIB2 structure and signalling and its role in many cancer subtypes with an emphasis on TRIB2 function in therapy resistance in melanoma, leukemia and glioblastoma. The strong evidence between TRIB2 expression and chemoresistance provides an attractive opportunity for targeting TRIB2., This work was supported by the Spanish Ministry of Science, Innovation and Universities through Grant RTI2018-094629-B-I00 to W.L.

BACKGROUND: Tribbles pseudokinase 3 (TRIB3) is a member of the tribbles-related family, which has been determined in various cancers, including renal cell carcinoma, acute promyelocytic leukemia, colorectal cancer, endometrial cancer, and glioma. However, its role in retinoblastoma (RB) has not yet been explored. METHODS: The expression level of TRIB3 was detected in RB tissues and cell lines using qRT-PCR. The effects of TRIB3 on cell proliferation and invasion capacities were analyzed with MTT, crystal violet, and transwell assays. Western blot and rescue assays were conducted to explore the underlying mechanism. RESULTS: This study found that TRIB3 was upregulated in human RB tissues compared to adjacent normal tissues both at the mRNA and protein levels. Overexpression of TRIB3 significantly promoted cell proliferation and invasion of RB cells, while TRIB3 knockdown inhibited these processes. Moreover, the mechanism deciphering experiments showed that TRIB3 overexpression can increase AKT and mTOR phosphorylation. Conversely, TRIB3 knockdown decreased the phosphorylation of AKT and mTOR. Additionally, MK2206, a potent AKT inhibitor, blocked the promotive effects of TRIB3 in RB cells. CONCLUSION: This study demonstrated that TRIB3 acts as an oncogene and plays a crucial role in the proliferation and invasion of RB cells via regulating the AKT/mTOR signaling pathway. Therefore, TRIB3 may serve as a potential target in the diagnosis and/or treatment of RB.

BackgroundBladder cancer is a common malignant tumor characterized by high mortality and high management costs; however, it lacks useful molecular prognostic markers. Tribbles pseudokinase 3 (TRIB3) is a pseudokinase that participates in cell tumor progression and metabolism and whose function in bladder cancer is not precisely known.Main MethodsWe downloaded transcriptome data and clinical data of bladder cancer from associated databases and extracted the expression matrix of TRIB3 for multiple bioinformatics analysis. RT-PCR detected the expression of TRIB3 in bladder cancer cells. After knockdown of TRIB3 with siRNA, we investigated TRIB3 function using CCK8, Cell Cycle and Transwell assays.Key FindingsKaplan–Meier analysis of TRIB3 in the four cohorts showed that high expression of TRIB3 correlated with poor outcome. Expression of TRIB3 positively correlated with stage and grade and down-regulation of TRIB3 expression significantly inhibited proliferation, migration and cell cycle of bladder cancer cells.SignificanceTRIB3 is a potential prognostic marker and therapeutic target. It can be used to individualize the treatment of bladder cancer.

Various retinal degenerative disorders manifest in alterations of the AKT/mTOR axis. Despite this, consensus on the therapeutic targeting of mTOR in degenerating retinas has not yet been achieved. Therefore, we investigated the role of AKT/mTOR signaling in rd16 retinas, in which we restored the AKT/mTOR axis by genetic ablation of pseudokinase TRB3, known to inhibit phosphorylation of AKT and mTOR. First, we found that TRB3 ablation resulted in preservation of photoreceptor function in degenerating retinas. Then, we learned that the mTOR downstream cellular pathways involved in the homeostasis of photoreceptors were also reprogrammed in rd16 TRB3−/− retinas. Thus, the level of inactivated translational repressor p-4E-BP1 was significantly increased in these mice along with the restoration of translational rate. Moreover, in rd16 mice manifesting decline in p-mTOR at P15, we found elevated expression of Beclin-1 and ATG5 autophagy genes. Thus, these mice showed impaired autophagy flux measured as an increase in LC3 conversion and p62 accumulation. In addition, the RFP-EGFP-LC3 transgene expression in rd16 retinas resulted in statistically fewer numbers of red puncta in photoreceptors, suggesting impaired late autophagic vacuoles. In contrast, TRIB3 ablation in these mice resulted in improved autophagy flux. The restoration of translation rate and the boost in autophagosome formation occurred concomitantly with an increase in total Ub and rhodopsin protein levels and the elevation of E3 ligase Parkin1. We propose that TRB3 may retard retinal degeneration and be a promising therapeutic target to treat various retinal degenerative disorders.

Glioma is the most common primary malignant brain tumor with poor survival and limited therapeutic options. The non-psychoactive phytocannabinoid cannabidiol (CBD) has been shown to be effective against glioma; however, the molecular target and mechanism of action of CBD in glioma are poorly understood. Here we investigated the molecular mechanisms underlying the antitumor effect of CBD in preclinical models of human glioma. Our results showed that CBD induced autophagic rather than apoptotic cell death in glioma cells. We also showed that CBD induced mitochondrial dysfunction and lethal mitophagy arrest, leading to autophagic cell death. Mechanistically, calcium flux induced by CBD through TRPV4 (transient receptor potential cation channel subfamily V member 4) activation played a key role in mitophagy initiation. We further confirmed TRPV4 levels correlated with both tumor grade and poor survival in glioma patients. Transcriptome analysis and other results demonstrated that ER stress and the ATF4-DDIT3-TRIB3-AKT-MTOR axis downstream of TRPV4 were involved in CBD-induced mitophagy in glioma cells. Lastly, CBD and temozolomide combination therapy in patient-derived neurosphere cultures and mouse orthotopic models showed significant synergistic effect in both controlling tumor size and improving survival. Altogether, these findings showed for the first time that the antitumor effect of CBD in glioma is caused by lethal mitophagy and identified TRPV4 as a molecular target and potential biomarker of CBD in glioma. Given the low toxicity and high tolerability of CBD, we therefore propose CBD should be tested clinically for glioma, both alone and in combination with temozolomide. Abbreviations: 4-PBA: 4-phenylbutyrate; AKT: AKT serine/threonine kinase; ATF4: activating transcription factor 4; Baf-A1: bafilomycin A(1); CANX: calnexin; CASP3: caspase 3; CAT: catalase; CBD: cannabidiol; CQ: chloroquine; DDIT3: DNA damage inducible transcript 3; ER: endoplasmic reticulum; GBM: glioblastoma multiforme; GFP: green fluorescent protein; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MTOR: mechanistic target of rapamycin kinase; PARP1: poly(ADP-ribose) polymerase; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; SLC8A1: solute carrier family 8 member A1; SQSTM1: sequestosome 1; TCGA: The cancer genome atlas; TEM: transmission electron microscopy; TMZ: temozolomide; TRIB3: tribbles pseudokinase 3; TRPC: transient receptor potential cation channel subfamily C; TRPV4: transient receptor potential cation channel subfamily V member 4

SALL4, a transcriptional factor involved in embryonic stem cell self-renewal and pluripotency, is overexpressed in gastric cancer (GC). However, the association of SALL4 with the survival of GC patients is not well-understood, and the role of SALL4 in cancer progression is still unknown. In the present study, a total of 1,815 GC patients who underwent radical resection at Peking Cancer Hospital were included consecutively from 2015 to 2018, confirming the prognostic value of SALL4 and validating by data from TCGA and GEO. The protein and mRNA expression levels of SALL4 were evaluated by immunohistochemistry and qPCR, respectively. Besides, GSEA and WGCNA were applied to explore the SALL4-related cancer-promoting signaling pathways and gene modules. Our results showed that overexpression of SALL4 was observed in 16.7% of GC patients. SALL4 positivity was associated with male, older age, mixed-type histology, late stages, lymphatic metastasis, vascular invasion, non-cardia location, high AFP level, and no EBV infection background. SALL4 could be served as a marker for prognostic prediction in GC, and SALL4-positive GC was significantly associated with shortened survival. Further, the bioinformatic analysis indicated that the Wnt/β-catenin signaling pathway was activated in SALL4-high cases compared with SALL4-low cases. Expression of SALL4 was also positively correlated with the expression of multiple co-expressed genes, such as TRIB3, which plays an important role in activating the Wnt/β-catenin pathway. Our findings indicate that SALL4 is associated with clinicopathological features related to cancer progression in GC and its function in the Wnt/β-catenin pathway.

The mechanism by which only some individuals infected with Mycobacterium tuberculosis develop necrotic granulomas with progressive disease while others form controlled granulomas that contain the infection remains poorly defined. Mice carrying the sst1-suscepible (sst1(S)) genotype develop necrotic inflammatory lung lesions, similar to human tuberculosis (TB) granulomas, which are linked to macrophage dysfunction, while their congenic counterpart (B6) mice do not. In this study we report that (a) sst1(S) macrophages developed aberrant, biphasic responses to TNF characterized by superinduction of stress and type I interferon pathways after prolonged TNF stimulation; (b) the late-stage TNF response was driven via a JNK/IFN-β/protein kinase R (PKR) circuit; and (c) induced the integrated stress response (ISR) via PKR-mediated eIF2α phosphorylation and the subsequent hyperinduction of ATF3 and ISR-target genes Chac1, Trib3, and Ddit4. The administration of ISRIB, a small-molecule inhibitor of the ISR, blocked the development of necrosis in lung granulomas of M. tuberculosis–infected sst1(S) mice and concomitantly reduced the bacterial burden. Hence, induction of the ISR and the locked-in state of escalating stress driven by the type I IFN pathway in sst1(S) macrophages play a causal role in the development of necrosis in TB granulomas. Interruption of the aberrant stress response with inhibitors such as ISRIB may offer novel host-directed therapy strategies.

© 2021 by the authors., The Tribbles family of pseudokinases controls a wide number of processes during cancer on-set and progression. However, the exact contribution of each of the three family members is still to be defined. Their function appears to be context-dependent as they can act as oncogenes or tumor suppressor genes. They act as scaffolds modulating the activity of several signaling pathways involved in different cellular processes. In this review, we discuss the state-of-knowledge for TRIB1, TRIB2 and TRIB3 in the development and progression of colorectal cancer. We take a perspective look at the role of Tribbles proteins as potential biomarkers and therapeutic targets. Specifically, we chronologically systematized all available articles since 2003 until 2020, for which Tribbles were associated with colorectal cancer human samples or cell lines. Herein, we discuss: (1) Tribbles amplification and overexpression; (2) the clinical significance of Tribbles overexpression; (3) upstream Tribbles gene and protein expression regulation; (4) Tribbles pharmacological modulation; (5) genetic modulation of Tribbles; and (6) downstream mechanisms regulated by Tribbles; establishing a comprehensive timeline, essential to better consolidate the current knowledge of Tribbles’ role in colorectal cancer., This work was supported by the Spanish Ministry of Science, Innovation and Universities through Grant RTI2018-094629-B-I00 to W.L. This work was supported by the European Commission project TRIBBLES–748585 and Fundação para a Ciência e a Tecnologia (FCT) (grant PTDC/MEDONC/4167/2020 “ENDURING”) to B.I.F.

Simple Summary Tribbles proteins possess the structure of protein kinases but function by forming protein complexes rather than phosphorylating substrates. Here we review the structure–function relationship of TRIB1 in cancers. Of the Tribbles proteins, TRIB1 is currently the most well characterised structurally. TRIB1 has different states that could potentially be targeted by small-molecule inhibitors and well-established relevance in acute myeloid leukaemia through degradation of transcription factors. Less is understood about the role of TRIB1 in solid tumours. Further research is required to fully realise the potential of TRIB1 as either a direct target of small-molecule drugs or a biomarker of treatment response across diverse cancer types. Abstract The Tribbles family of proteins—comprising TRIB1, TRIB2, TRIB3 and more distantly related STK40—play important, but distinct, roles in differentiation, development and oncogenesis. Of the four Tribbles proteins, TRIB1 has been most well characterised structurally and plays roles in diverse cancer types. The most well-understood role of TRIB1 is in acute myeloid leukaemia, where it can regulate C/EBP transcription factors and kinase pathways. Structure–function studies have uncovered conformational switching of TRIB1 from an inactive to an active state when it binds to C/EBPα. This conformational switching is centred on the active site of TRIB1, which appears to be accessible to small-molecule inhibitors in spite of its inability to bind ATP. Beyond myeloid neoplasms, TRIB1 plays diverse roles in signalling pathways with well-established roles in tumour progression. Thus, TRIB1 can affect both development and chemoresistance in leukaemia; glioma; and breast, lung and prostate cancers. The pervasive roles of TRIB1 and other Tribbles proteins across breast, prostate, lung and other cancer types, combined with small-molecule susceptibility shown by mechanistic studies, suggests an exciting potential for Tribbles as direct targets of small molecules or biomarkers to predict treatment response.

Background Altered lipid metabolism is involved in the development of many tumors. However, the role of dissimilar lipid metabolism in head and neck squamous cell carcinoma (HNSCC) is not fully established. Aims Here, we sought to determine the prognostic value of lipid metabolism‐related genes in HNSCC. Methods RNA‐seq data and clinical features of 545 HNSCC cases were obtained from The Cancer Genome Atlas database. A regulatory network of transcription factors‐lipid metabolism genes and a risk prognostic model of lipid metabolism‐related genes was developed using bioinformatics and Cox regression modeling. We used tumor immune estimation resource to analyze immune cell infiltration in patients with HNSCC based on the prognostic index (PI) of lipid metabolism‐related genes. Results A total of 136 differentially expressed lipid metabolism genes were identified. Of these, 23 are related to prognosis. In addition to predicting HNSCC prognosis, 11 lipid metabolism‐related genes (ARSI, CYP27B1, CYP2D6, DGKG, DHCR7, LPIN1, PHYH, PIP5K1B, PLA2G2D, RDH16, and TRIB3) also affect HNSCC clinical features (stage, gender, and pathological stage). The PI of lipid metabolism‐related genes embodied the state of HNSCC tumor immune microenvironment., Lipid metabolism‐related genes effectively predicts head and neck squamous cell carcinoma (HNSCC) prognosis and reflects the state of immune cell in HNSCC.

This study aimed to investigate the role of vascular insulin resistance (VIR) and Tribbles homolog 3 (TRIB3) in the pathogenesis of hypoxia-induced pulmonary hypertension (HPH). Rats were subjected to low air pressure and low oxygen intermittently for 4 weeks to induce HPH. The mean right ventricular pressure (mRVP), mean pulmonary arterial pressure (mPAP), and right ventricular index (RVI) were significantly increased in HPH rats. Pulmonary arteries from HPH rats showed VIR with reduced vasodilating effect of insulin. The protein levels of peroxisome proliferator-activated receptor gamma (PPARγ), phosphoinositide 3-kinase (PI3K), phosphorylations of Akt, and endothelial nitric oxide (NO) synthase (eNOS) were decreased, and TRIB3 and phosphorylated extracellular signal-regulated protein kinases (ERK1/2) were increased in pulmonary arteries of HPH rats. Early treatment of pioglitazone (PIO) partially reversed the development of HPH, improved insulin-induced vasodilation, and alleviated the imbalance of the insulin signaling. The overexpression of TRIB3 in rat pulmonary arterial endothelial cells (PAECs) reduced the levels of PPARγ, PI3K, phosphorylated Akt (p-Akt), and phosphorylated eNOS (p-eNOS) and increased p-ERK1/2 and the synthesis of endothelin-1 (ET-1), which were further intensified under hypoxic conditions. Moreover, TRIB3 knockdown caused significant improvement in Akt and eNOS phosphorylations and, otherwise, a reduction of ERK1/2 activation in PAECs after hypoxia. In conclusion, impaired insulin-induced pulmonary vasodilation and the imbalance of insulin-induced signaling mediated by TRIB3 upregulation in the endothelium contribute to the development of HPH. Early PIO treatment improves vascular insulin sensitivity that may help to limit the progression of hypoxic pulmonary hypertension.

Advances in the understanding of the Tribbles family of pseudokinases (TRIB1, TRIB2 and TRIB3) reveal these proteins as potentially valuable biomarkers of disease diagnosis, prognosis, prediction and clinical strategy. In their role as signalling mediators and scaffolding proteins, TRIBs lead to changes in protein stability and activity, which impact on diverse cellular processes such as proliferation, differentiation, cell cycle and cell death. We review the role of TRIB proteins as promising therapeutic targets, with an emphasis on their role in cancer, and as biomarkers, with potential application across diverse pathological processes.

Rapalogs have become standard-of-care in patients with metastatic breast, kidney, and neuroendocrine cancers. Nevertheless, tumor escape occurs after several months in most patients, highlighting the need to understand mechanisms of resistance. Using a panel of cancer cell lines, we show that rapalogs downregulate the putative protein kinase TRIB3 (tribbles pseudokinase 3). Blood samples of a small cohort of patients with cancer treated with rapalogs confirmed downregulation of TRIB3. Downregulation of TRIB3 was mediated by LRRFIP1 independently of mTOR and disrupted its interaction with the spliceosome, where it participated in rapalog-induced deregulation of RNA splicing. Conversely, overexpression of TRIB3 in a panel of cancer cell lines abolished the cytotoxic effects of rapalogs. These findings identify TRIB3 as a key component of the spliceosome, whose repression contributes significantly to the mechanism of resistance to rapalog therapy. Significance: Independent of mTOR signaling, rapalogs induce cytoxicity by dysregulating spliceosome function via repression of TRIB3, the loss of which may, in the long term, contribute to therapeutic resistance.

Mounting evidence supports that the malignant phenotypes of cancers are defined not only by the intrinsic activity of tumor cells but also by immune cells that are recruited and activated in tumor-related microenvironment. Here, we developed a diagnostic and prognostic model for colon cancer, based on expression profiles of immune-related genes and immune cell component. As a result, we found that B cell infiltration ratio, CD4+ T cells, as well as immune-related genes of TRIB3, CHGA, CASP7, LGALS4, LEP, NOX4, IL17A, and HSPD1 may be highly relevant with clinical outcome of colon cancer.

Tribbles pseudokinase 3 (TRIB3) is a member of the mammalian pseudokinase tribbles family and is involved in multiple biological processes. However, the role of TRIB3 in renal cell carcinoma (RCC) remains unclear. In this study, we aimed to elucidate the biological functions of TRIB3 in RCC and explore its underlying mechanisms. TRIB3 expression and its correlation with clinicopathological features was evaluated in 123 patients with RCC. A series of cytological experiments were performed to clarify the biological functions of TRIB3, and potential molecular regulatory mechanisms were explored using transcriptome sequencing. TRIB3 expression was significantly elevated in RCC tissues compared to that in paracancerous tissues, and high expression of TRIB3 was correlated with both advanced tumor stage and unfavorable prognosis. TRIB3 knockdown markedly inhibited RCC cell proliferation, migration and invasion. Furthermore, overexpression of TRIB3 promoted RCC cell proliferation, migration, invasion and xenograft tumor growth. Notably, TRIB3 expression was modulated by hypoxia-inducible factor-1α (HIF-1α), which enhanced cell viability and invasiveness via targeting the MAPK signaling pathway. This study reveals the potential oncogenic role of TRIB3 in RCC pathogenesis and illustrates the mechanisms underlying TRIB3-mediated tumor progression, providing new insight into the development of TRIB3 as a tumor biomarker and therapeutic target.

Objectives Hypericin is a polycyclic aromatic naphthodianthrone that occurs naturally. It is also an active ingredient in some species of the genus Hypericum. Emerging evidence suggests that hypericin has attracted great attention as a potential anticancer drug and exhibits remarkable antiproliferative effect upon irradiation on various tumour cells. This paper aims to summarise the anticancer effect and molecular mechanisms modulated by hypericin-medicated photodynamic therapy and its potential role in the cancer treatment. Key findings Hypericin-medicated photodynamic therapy could inhibit the proliferation of various tumour cells including bladder, colon, breast, cervical, glioma, leukaemia, hepatic, melanoma, lymphoma and lung cancers. The effect is primarily mediated by p38 mitogen-activated protein kinase (MAPK), JNK, PI3K, CCAAT-enhancer-binding protein homologous protein (CHOP)/TRIB3/Akt/mTOR, TRAIL/TRAIL-receptor, c-Met and Ephrin-Eph, the mitochondria and extrinsic signalling pathways. Furthermore, hypericin-medicated photodynamic therapy in conjunction with chemotherapeutic agents or targeted therapies is more effective in inhibiting the growth of tumour cells. Summary During the past few decades, the anticancer properties of photoactivated hypericin have been extensively investigated. Hypericin-medicated photodynamic therapy can modulate a variety of proteins and genes and exhibit a great potential to be used as a therapeutic agent for various types of cancer.

Identifying disease-causing pathways and drugs that target them in Parkinson's disease (PD) has remained challenging. We uncovered a PD-relevant pathway in which the stress-regulated heterodimeric transcription complex CHOP/ATF4 induces the neuron prodeath protein Trib3 that in turn depletes the neuronal survival protein Parkin. Here we sought to determine whether the drug adaptaquin, which inhibits ATF4-dependent transcription, could suppress Trib3 induction and neuronal death in cellular and animal models of PD. Neuronal PC12 cells and ventral midbrain dopaminergic neurons were assessed in vitro for survival, transcription factor levels and Trib3 or Parkin expression after exposure to 6-hydroxydopamine or 1-methyl-4-phenylpyridinium with or without adaptaquin co-treatment. 6-hydroxydopamine injection into the medial forebrain bundle was used to examine the effects of systemic adaptaquin on signaling, substantia nigra dopaminergic neuron survival and striatal projections as well as motor behavior. In both culture and animal models, adaptaquin suppressed elevation of ATF4 and/or CHOP and induction of Trib3 in response to 1-methyl-4-phenylpyridinium and/or 6-hydroxydopamine. In culture, adaptaquin preserved Parkin levels, provided neuroprotection and preserved morphology. In the mouse model, adaptaquin treatment enhanced survival of dopaminergic neurons and substantially protected their striatal projections. It also significantly enhanced retention of nigrostriatal function. These findings define a novel pharmacological approach involving the drug adaptaquin, a selective modulator of hypoxic adaptation, for suppressing Parkin loss and neurodegeneration in toxin models of PD. As adaptaquin possesses an oxyquinoline backbone with known safety in humans, these findings provide a firm rationale for advancing it towards clinical evaluation in PD.

Metformin has beneficial effects of preventing and treating cancers on type 2 diabetic patients. However, the role of metformin in non-diabetic cancer patients and the precise molecular mechanisms against cancer have not yet been sufficiently elucidated. We recently reported that the pseudokinase protein TRIB3 acts as a stress sensor linking metabolic stressors to cancer promotion by inhibiting autophagy and ubiquitin-proteasomal degradation systems; genetically abrogating of TRIB3 expression reduces tumourigenesis and cancer progression. Thus, TRIB3 is a potential therapeutic target for diverse cancers. In this study, we found that metformin attenuates melanoma growth and metastasis by reducing TRIB3 expression in non-diabetic C57BL/6 mice and diabetic KK-Ay mice; overexpression of TRIB3 protects metformin from the activation of autophagic flux, the clearance of accumulated tumour-promoting factors and the attenuation of tumour progression. We further elucidated that TRIB3 acts as an adaptor to recruit lysine acetyltransferase 5 (KAT5) to SMAD3 and induce a phosphorylation-dependent K333 acetylation of SMAD3, which sustains transcriptional activity of SMAD3 and subsequently enhances TRIB3 transcription. Metformin suppresses SMAD3 phosphorylation and decreases the KAT5/SMAD3 interaction, to attenuate the KAT5-mediated K333 acetylation of SMAD3, reduce the SMAD3 transcriptional activity and subsequent TRIB3 expression, thereby antagonizes melanoma progression. Together, our study not only defines a molecular mechanism by which metformin protects against melanoma progression by disturbing the KAT5/TRIB3/SMAD3 positive feedback loop in diabetes and non-diabetes mice, but also suggests a candidate diverse utility of metformin in tumour prevention and therapy because of suppressing stress protein TRIB3 expression.

While acute promyelocytic leukemia has a good prognosis with all-trans retinoic acid (ATRA) treatment, ATRA resistance is a major obstacle. It is now demonstrated that TRIBBLES 3 (TRIB3) stabilizes TWIST1, leading to ATRA resistance. Peptides that disrupt this interaction lead to the degradation of TWIST1 and overcome ATRA resistance. See related article by Lin et al., p. 6228

Numerous animal studies have suggested that sevoflurane can induce neuronal cell death and damage social and emotional behaviors and endoplasmic reticulum stress is a crucial underlying mechanism. The goal of this study was to investigate the role and the mechanisms of long noncoding RNA X-inactive specific transcript (XIST) in sevoflurane-induced social and emotional impairment and neuron apoptosis in neonatal mice. The performance in social and emotional tests and the expression levels of lncRNA-XIST after sevoflurane exposure in neonatal mice and hippocampal neuronal cells were measured. Also, the effects of the suppression of lncRNA-XIST on neuron apoptosis and endoplasmic reticulum stress were determined. Subsequently, the association between lncRNA-XIST and miR-98-5p as well as EDEM1 in hippocampal neuronal cells was explored. Our results showed that social and emotional behaviors were impaired in neonatal mice and lncRNA-XIST and EDEM1were increasingly expressed in hippocampal neuronal cells after sevoflurane exposure and suppression of lncRNA-XIST decreased sevoflurane-induced neuron apoptosis and endoplasmic reticulum stress in vitro. Further in vitro studies showed that overexpression of miR-98-5p decreased apoptosis and endoplasmic reticulum stress while lncRNA Xist negative regulated miR-98-5p expression and the effects of lncRNA Xist on apoptosis and endoplasmic reticulum stress through sponging miR-98-5p. Moreover, suppression of lncRNA-XIST improved sevoflurane-induced social and emotional impairment and alleviated sevoflurane-induced apoptosis and endoplasmic reticulum stress accompanied in neonatal mice. Our findings reveal that knockdown of lncRNA-XIST may contribute to ameliorating sevoflurane-induced social and emotional impairment via inhibiting neuron apoptosis and endoplasmic reticulum stress, which may be achieved by regulating miR-98-5p/EDEM1 axis. Funding Statement: This work was supported by the National Natural Science Foundation of China (81400929, 81471240, 81641042 and 81603545), the Bureau of Chinese Medicine, Zhejiang, China (2018ZB065) and the Innovative Talents Project of Zhejiang province (2016). Declaration of Interests: There is no conflict of interest in the authors. Ethics Approval Statement: The study protocol was approved by the Zhejiang University Institutional review board of Animal studies (ZJU2019-16778). The Guidance Recommendations for Experimental Animal Care and Use of the Ministry of Science and Technology of the People’s Republic of China supervised all experimental procedures.

Background Recent evidence indicates that TRIB3 and miR-124 levels have been deregulated in type 2 diabetes (T2D); however, the simultaneous evaluation of these markers in diabetic patients has not been investigated to date. Methods This case-control study included 50 T2D patients and 40 age-gender matched controls. The circulation level of miR-124a was assessed by real-time PCR. TRIB3 plasma level was measured using the enzyme-linked im-munosorbent assay. Results Our findings revealed that the TRIB3 plasma level was significantly increased (p = 0.025), while miR-124a plasma levels were significantly reduced (p = 0.028) in diabetic patients compared to healthy subjects. ROC analysis showed that TRIB3 and miR-124a levels could discriminate control subjects and diabetic patients. Interestingly, a significant negative correlation was found between the TRIB3 and miR-124a plasma levels. Furthermore, there was a significant positive correlation between the TRIB3 plasma level with fasting blood glucose and insulin resistance. Conclusions In this study, we showed deregulation of TRIB3 level in diabetic patients and its association with miR-124a circulating level and clinical parameters. These findings suggest that miR-124a may affect T2D incidence and progression by modulating the expression of TRIB3 protein level.

Here, we aim to explore whether tribbles pseudokinase 3 (TRIB3) enhances glioma cell stemness. TRIB3 was overexpressed in glioma tissues and cell-formed spheres, positively correlated with the size and grade. Additionally, TRIB3 expression displayed a negative correlation with the overall survival rate of glioma patients. Moreover, TRIB3 knockdown reduced the stemness of nonadherent spheres, evident by the decreased sphere-forming ability, stemness master expression, and ALDH1 activity, while TRIB3 overexpression enhanced the stemness of adherent cells, which was rescued by β-catenin knockdown. Mechanistically, TRIB3 activated β-catenin signaling via physically interacting with β-catenin. This study suggests that the TRIB3-β-catenin interaction is responsible for glioma cell stemness.

Laura Martinez Campesino; Jessica M Johnston; Sheila E Francis; Endre Kiss-Toth; Heather L Wilson. Department of Infection, Immunity & Cardiovascular Disease, Medical School, Beech Hill Road, University of Sheffield, UK. Background Metabolic dysfunction and chronic inflammation under obesity are key factors promoting atherosclerosis. Macrophages are the major immune cell type found in atherosclerotic lesions, where plaque macrophages engulf oxidised lipids forming foam cells and also regulate plaque stability. Tribbles (Trib) 3 is a pseudokinase known to be involved in the regulation of adipose homeostasis, lipid and glucose metabolism and macrophage lipid uptake. We propose that Trib3 has a role in regulating metabolic and immune responses to obesity ultimately affecting atherosclerosis development. Methodology. Trib3KO and Trib3WT mice were injected with rAAV8/mPCSK9 and fed with western diet for 12 weeks to promote atherosclerosis development. Atherosclerotic plaques were quantified using Oil-red-O and aortic root lesions were characterised histologically. Primary macrophages from bone marrow (BMDMs) were isolated from these strains and human monocyte derived macrophages (MDMs) were used to further characterise the effects of Trib3 regulating macrophage function. Results En face analysis of the aortae showed no statistical differences in atheroma burden, while histological examination of the plaques indicated an increase in plaque stability in the absence of Trib3. Trib3KO BMDMs showed differential expression of genes involved in macrophage metabolism and regulation of the cytoskeleton. The differential expression of these genes was assessed in human MDMs following control or siTRIB3 treatment. Conclusion These results suggest a role for Trib3 in the regulation of macrophage function promoting plaque stability, and may also have an impact on atherosclerosis development. Conflict of interest No COI

Purpose: To develop a multi-RNA-based model to provide survival risk prediction for colon cancer by constructing a competing endogenous RNAs (ceRNAs) network. Methods: The prognostic information and expression of the lncRNAs, miRNAs, and mRNAs in colon cancer specimens from The Cancer Genome Atlas (TCGA) were assessed. Constructing prognostic models used the differentially expressed RNAs. Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses and Gene Ontology were used to identify the functional role of the ceRNA network in the prognosis of colon cancer. Results: Five lncRNAs (AC007384.1, AC002511.1, AC012640.1, C17orf82, and AP001619.1), 8 miRNAs (hsa-mir-141, hsa-mir-150, hsa-mir-375, hsa-mir-96, hsa-mir-107, hsa-mir-106a, hsa-mir-200a, and hsa-mir-1271), and 5 mRNAs (BDNF, KLF4, SESN2, SMOC1, and TRIB3) were highly correlated with tumor status and tumor stage. Three prognostic models based on the 5 lncRNAs, 8 miRNAs, and 5 mRNAs were constructed. The prognostic ability was 0.850 for the lncRNA-based model, 0.811 for the miRNA-based model, and 0.770 for the mRNA-based model. Patients with high-risk scores revealed worse overall survival. The KEGG pathways were significantly enriched in the "neuroactive ligand-receptor interaction." Conclusion: This study identified several potential prognostic biomarkers to construct a multi-RNA-based prognostic model for colon cancer.

TRIB3 roles in tumor progression have been revealed with similar or opposite results. Here, we found that TRIB3 expression was highly expressed in lung cancer tissues and correlated with tumor grades and metastasis. Functional experiments showed that TRIB3 knockdown (KD) inhibited lung cancer cell migration, invasion, EMT (epithelial-mesenchymal transition) process and stemness. Mechanistic studies demonstrated that TRIB3 physically interacted with β-catenin and increased the recruitment of β-catenin to the promoter region of genes regulated by Wnt. Re-activation of β-catenin attenuated the inhibition of TRIB3 KD on lung cancer progression. These results suggest that TRIB3 interacts with β-catenin and thus activates β-catenin signaling, which is responsible for lung cancer progression, and blocking TRIB3 activity might be developed to treat lung cancer.

INTRODUCTION AND OBJECTIVES:Tribbles pseudokinase 3 (TRIB3) is a member of the mammalian pseudokinase tribbles family and is involved in multiple biological processes. However, the role of TRIB3 in...

The Tribbles (TRIB) family of pseudokinase proteins has been shown to play key roles in cell cycle, metabolic diseases, chronic inflammatory disease, and cancer development. A better understanding of the mechanisms of TRIB pseudokinases could provide new insights for disease development and help promote TRIB proteins as novel therapeutic targets for drug discovery. At the 2nd International Symposium on Tribbles and Diseases held on May 7-9, 2018 in Beijing, China, a group of leading Tribbles scientists reported their findings and ongoing studies about the effects of the different TRIB proteins in the areas of immunity, metabolism, fundamental cell biology and cancer. Here, we summarize important and insightful overviews from 4 keynote lectures, 13 plenary lectures and 8 short talks that took place during this meeting. These findings may offer new insights for the understanding of the roles of TRIB pseudokinases in the development of various diseases. (C) 2019 Chinese Pharmaceutical Association and Institute of Materia Medica, Chinese Academy of Medical Sciences. Production and hosting by Elsevier B.V. Dr. Zhuowei Hu' s work was supported by National Key R&D Program of China (Grant No. 2017YFA0205400, China), the National Natural Science Foundation of China (Grant Nos. 81530093 and 81773781, China), Chinese Academy of Medical Sciences (CAMS) Innovation Fund for Medical Sciences (Grant No. 2016-12M-1-007, China) and CAMS Central Public-interest Scientific Institution Basic Research Fund (Grant No. 2017PT3104, China); Dr. Bing Cui' s work was supported by grants of the National Natural Science Foundation of China (Grant No. 81874316, China), the CAMS Innovation Fund for Medical Sciences (Grant No. 2016-I2M-3-008, China); Dr. Patrick A. Eyers' s work was supported by grants of from the BBSRC and NWCR (Grant Nos. 1088 and 1097, UK); Dr. Leonard L. Dobens' s work was supported by grants of NSF (Grant No. IOS-1456023, USA), NIH (Grant No. NIH R21 CA197317, USA); Dr. Nguan Soon Tan' s work was supported by grants of Ministry of Education, Singapore (Grant Nos. MOE2014-T2-1-012 and 2012-T1-001-036, Singapore); Dr. Peter D. Mace' s work was supported by grants from the Health Research Council of New Zealand, and additionally supported by a Rutherford Discovery Fellowship from the New Zealand government administered by the Royal Society of New Zealand; Dr. Wolfgang Link' s work was supported by Fundacao para a Ciencia e a Tecnologia (FCT) Research Center Grant UID/KM/04773/2013 Centre for Biomedical Research 1334, by a research grant from Liga Portuguesa Contra o Cancro NUcleo Regional do Sul (LPCC/NRS, Portugal), Terry Fox, by a FCT 2014 research grant SFRH/BPD/100434/2014 and a ProRegem grant PD/BD/114258/2016 (Portugal); Dr. Endre Kiss-Toth' s work was supported by European Marie Sklodowska Curie ITN Project TRAIN-TRIBBLES Research and Innovation Network (Grant No. 721532, EU)and Innovation Network and the British Heart Foundation (PG/16/44/32146, UK); Dr. Karen Keeshan' s work was supported by grants from The Howat Foundation Ltd. (UK), Children with Cancer UK, Bloodwise and the Friends of Paul O' Gorman (UK); Dr. Takuro Nakamura' s work was supported by grants of P-CREATE from Japan Agency for Medical Research and Development; Dr. Warren S. Pear' s work was supported by grants from the NIH (NIAID, USA), Alex' s Lemonade Stand Foundation (USA) and the Samuel Waxman Cancer Research Foundation (USA); Yodit Feseha is supported by European Marie Sklodowska Curie ITN Project TRAIN-TRIBBLES Research and Innovation Network (Grant No. 721532, EU). Yodit Feseha' s work is conducted in the context of the "Fondation Centaure" (RTRS), which supports a French transplantation research network, the 1HU-Cesti project, the DHU Oncogreffe and the LabEX IGO thanks to French government financial support managed by the National Research Agency via the "Investment into the Future" program (Grant Nos. ANR-10-IBHU-005 and ANR-11-LABX-0016-01, France). The IHU-Cesti project is also supported by Nantes Metropole and Region Pays de la Loire (France); Dr. Jessica Johnston's work was supported by grants of the British Heart Foundation (PG/16/44/32146, UK); Dr. Arkatiz Carracedo' s work was supported by European Marie Sklodowska Curie ITN Project TRAIN-TRIBBLES Research and Innovation Network (Grant No. 721532, EU); Dr. Marcel Scheideler' s work was supported by European Marie Sklodowska Curie ITN Project TRAIN-TRIBBLES Research and Innovation Network (Grant No. 721532, EU); Dr. Zabran llyas' s work was supported by a joint Ph.D studentship beween the A* Star Institute and the University of Sheffield (UK); Dr. Robert C. Bauer s work was supported by funding from the National Institutes of Health National Heart, Lung, and Blood Institute (R01HL141745, USA); Dominika Grzesik' s work was supported by European Marie Sklodowska Curie ITN Project TRAIN-TRIBBLES Research and Innovation Network (Grant No. 721532, EU); Juan Salamanca-Viloria's work was supported by European Marie Sklodowska Curie ITN Project TRAIN-TRIBBLES Research and Innovation Network (Grant No. 721532, EU); Dr. Xiaoxi Lv' s work was supported by the National Natural Science Foundation of China (Grant No. 81503128, China) and CAMS Innovation Fund for Medical Sciences (Grant No. 201642M-1-008, China); Dr. Yishi Jin' s work was supported by National Institute of Health (NS R01-035546, USA); Dr. Ke Li' s work was supported by the National Natural Science Foundation of China (Grant No. 81400140, China), and CAMS Innovation Fund for Medical Sciences (Grant No. 201642M-1-011, China); Dr. Guillermo Velasco' s work was supported by European Marie Sklodowska Curie ITN Project TRAIN-TRIBBLES Research and Innovation Network (Grant No. 721532, EU); Dr. Jose M. Lizcano's work was supported by Spanish Ministry of Economy and Competitiveness (MINECO) and Fondo Europeo de desarrollo Regional (FEDER) (Grant No. INNPACTO/IPT-2012-0614-010000, Spain); Dr. Xiaowei Zhang' s work was supported by the National Natural Science Foundation of China (Grant Nos. 81400286 and 81530093, China), and the CAMS Innovation Fund for Medical Sciences (Grant No. 201642M-1-010, China); Dr. Fang Hua' s work was supported by the National Natural Science Foundation of China (Grant Nos. 81472717 and 81673474, China), Beijing Natural Science Foundation (Grant No. 7162133, China), and the CAMS Innovation Fund for Medical Sciences (Grant No. 201642M-1-007, China); Dr. Jiaojiao Yu' s work was supported by the National Natural Science Foundation of China (Grant No. 81703564, China); Dr. Jinmei Yu' s work was supported by the National Natural Science Foundation of China (Grant No. 81603129, China). In addition, the Meeting Organizing Committee was greatly thankful to PerkinElmer and Innovent Biologics for their generous financial supports, as well as Dr. Zhuowei Hu' s laboratory members for their assistance in preparing and organizing the conference.

Purpose: The resistance to differentiation therapy and early death caused by fatal bleeding endangers the health of a significant proportion of patients with acute promyelocytic leukemia (APL). This study aims to investigate the molecular mechanisms of all-trans retinoic acid (ATRA) resistance and uncover new potential therapeutic strategies to block the rapid progression of early death. Experimental Design: The important role of TWIST1 in APL leukemogenesis was first determined by gain- and loss-of-function assays. We then performed in vivo and in vitro experiments to explore the interaction of TWIST1 and TRIB3 and develop a potential peptide-initiated therapeutic opportunity to protect against early death and induction therapy resistance in patients with APL. Results: We found that the epithelial–mesenchymal transition (EMT)-inducing transcription factor TWIST1 is highly expressed in APL cells and is critical for leukemic cell survival. TWIST1 and TRIB3 were highly coexpressed in APL cells compared with other subtypes of acute myeloid leukemia cells. We subsequently demonstrated that TRIB3 could bind to the WR domain of TWIST1 and contribute to its stabilization by inhibiting its ubiquitination. TRIB3 depletion promoting TWIST1 degradation reverses resistance to induction therapy and improves sensitivity to ATRA. On the basis of a detailed functional screen of synthetic peptides, we discovered a peptide analogous to the TWIST1 WR domain that specifically represses APL cell survival by disrupting the TRIB3/TWIST1 interaction. Conclusions: Our data not only define the essential role of TWIST1 as an EMT-TF in patients with APL but also suggest that disrupting the TRIB3/TWIST1 interaction reverses induction therapy resistance and blocks rapid progression of APL early death. See related commentary by Peeke and Gritsman, p. 6018

Post-translational modifications directly control protein activity and, thus, they represent an important means to regulate the responses of cells to different stimuli. Protein SUMOylation has recently been recognised as one such modification, and it has been associated with various diseases, including different types of cancer. However, the precise way that changes in SUMOylation influence the tumorigenic properties of cells remains to be fully clarified. Here, we show that blocking the SUMO pathway by depleting SUMO1 and UBC9, or by exposure to ginkgolic acid C15:1 or 2-D08 (two different SUMOylation inhibitors), induces cell death, also inhibiting the invasiveness of tumour cells. Indeed, diminishing the formation of SUMO1 complexes induces autophagy-mediated cancer cell death through increasing the expression of Tribbles pseudokinase 3 (TRIB3). Moreover, we found that blocking the SUMO pathway inhibits tumour cell invasion by decreasing RAC1 SUMOylation. These findings shed new light on the mechanisms by which SUMO1 modifications regulate the survival, and the migratory and invasive capacity of tumour cells, potentially establishing the bases to develop novel anti-cancer treatments based on the inhibition of SUMOylation., Summary: Blockade of the SUMO pathway inhibits tumorigenesis through two different mechanisms: (i) stimulation of autophagy-mediated cancer cell death via TRIB3 upregulation; and (ii) inhibition of invasiveness via abrogation of RAC1 SUMOylation.

Impaired macroautophagy/autophagy is involved in the pathogenesis of hepatic fibrosis. However, how aberrant autophagy promotes fibrosis is far from understood. Here, we aimed to define a previously unrevealed pro-fibrotic mechanism for the stress protein TRIB3 (tribbles pseudokinase 3)-mediated autophagy dysfunction. Human fibrotic liver tissues were obtained from patients with cirrhosis who underwent an open surgical repair process. The functional implications of TRIB3 were evaluated in mouse models of hepatic fibrosis induced by bile duct ligation (BDL) or thioacetamide (TAA) injection. Human fibrotic liver tissues expressed higher levels of TRIB3 and selective autophagic receptor SQSTM1/p62 (sequestosome 1) than nonfibrotic tissues and the elevated expression of TRIB3 and SQSTM1 was positively correlated in the fibrotic tissues. Silencing Trib3 protected against experimentally induced hepatic fibrosis, accompanied by restored autophagy activity in fibrotic liver tissues. Furthermore, TRIB3 interacted with SQSTM1 and hindered its binding to MAP1LC3/LC3, which caused the accumulation of SQSTM1 aggregates and obstructed autophagic flux. The TRIB3-mediated autophagy impairment not only suppressed autophagic degradation of late endosomes but also promoted hepatocellular secretion of INHBA/Activin A-enriched exosomes which caused migration, proliferation and activation of hepatic stellate cells (HSCs), the effector cells of liver fibrosis. Disrupting the TRIB3-SQSTM1 interaction with a specific helical peptide exerted potent protective effects against hepatic fibrosis by restoring autophagic flux in hepatocytes and HSCs. Together, stress-elevated TRIB3 expression promotes hepatic fibrosis by interacting with SQSTM1 and interfering with its functions in liver-parenchymal cells and activating HSCs. Targeting this interaction is a promising strategy for treating fibroproliferative liver diseases. Abbreviations: 3-MA: 3-methyladenine; AAV: adeno-associated virus; ACTA2/α-SMA: actin, alpha 2, smooth muscle, aorta; BDL: bile duct ligation; BECN1/Beclin 1: beclin 1, autophagy related; CHX: cycloheximide; CQ: chloroquine; Edu: 5-ethynyl-2-deoxyuridine; ESCRT: endosomal sorting complexes required for transport; HSC: hepatic stellate cell; ILV: intralumenal vesicle; LAMP1: lysosomal-associated membrane protein 1; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MVB: multivesicular body; PIK3C3: phosphatidylinositol 3-kinase, catalytic subunit type 3; PPI: protein-protein interaction; SQSTM1/p62: sequestosome 1; TAA: thioacetamide; TEM: transmission electron microscopy; TGFB1/TGFβ1: transforming growth factor, beta 1; TLR2: toll-like receptor 2; TRIB3: tribbles pseudokinase 3

Type 2 diabetes mellitus is a complex disease. Our previous study revealed that TRIB3 genetic variations were strongly associated with diabetic vascular complications, although TRIB3 regulation pathways remain poorly understood. We used two extreme treatment groups from a 2 × 2 factorial randomized controlled trial to identify a positive association, which was further validated in patients receiving cross treatment to test the effect of genetic polymorphisms among the different treatment groups. A gene-centric score (GS)-weighted model including the three associated genetic variations TRIB3 rs2295490, ATF6 rs12086247, and SMARCD3 rs58125572 was used. The results of the GS model indicated a 46% reduction in the risk of primary vascular complications in patients bearing more than two risk alleles [hazard ratio (HR) 0.54, 95% confidence interval (CI) 0.38–0.76, p < 0.001], following intensive glucose control treatment when compared with patients who received standard glucose control treatment. Furthermore, these patients benefited from active blood pressure-lowering treatment (HR 0.39, 95% CI 0.24–0.64, p < 0.001). However, no significant difference was observed between the two interventions in patients with fewer than two risk alleles (HR 1.09, 95% CI 0.86–1.39, p = 0.47). These results indicate that genetic variants in these three genes may be useful biomarkers for individualized drug therapy in diabetic patients.