Lactate enhances NMNAT1 lactylation to sustain nuclear NAD+ salvage pathway and promote survival of pancreatic adenocarcinoma cells under glucose-deprived conditions.

The aim of this study was to investigate the underlying molecular mechanism behind the promotion of cell survival under conditions of glucose deprivation by l-lactate. To accomplish this, we performed tissue microarray and immunohistochemistry staining to analyze the correlation between the abundance of pan-Lysine lactylation and prognosis. In vivo evaluations of tumor growth were conducted using the KPC and nude mice xenograft tumor model. For mechanistic studies, multi-omics analysis, RNA interference, and site-directed mutagenesis techniques were utilized. Our findings robustly confirmed that l-lactate promotes cell survival under glucose deprivation conditions, primarily by relying on GLS1-mediated glutaminolysis to support mitochondrial respiration. Mechanistically, we discovered that l-lactate enhances the NMNAT1-mediated NAD+ salvage pathway while concurrently inactivating p-38 MAPK signaling and suppressing DDIT3 transcription. Notably, Pan-Kla abundance was significantly upregulated in patients with Pancreatic adenocarcinoma (PAAD) and associated with poor prognosis. We identified the 128th Lysine residue of NMNAT1 as a critical site for lactylation and revealed EP300 as a key lactyltransferase responsible for catalyzing lactylation. Importantly, we elucidated that lactylation of NMNAT1 enhances its nuclear localization and maintains enzymatic activity, thereby supporting the nuclear NAD+ salvage pathway and facilitating cancer growth. Finally, we demonstrated that the NMNAT1-dependent NAD+ salvage pathway promotes cell survival under glucose deprivation conditions and is reliant on the activity of Sirt1. Collectively, our study has unraveled a novel molecular mechanism by which l-lactate promotes cell survival under glucose deprivation conditions, presenting a promising strategy for targeting lactate and NAD+ metabolism in the treatment of PAAD.

Autophagy mediated FTH1 degradation activates gasdermin E dependent pyroptosis contributing to diquat induced kidney injury.

Acute kidney injury (AKI) induced by diquat (DQ) progresses rapidly, leading to high mortality, and there is no specific antidote for this chemical. Our limited knowledge of the pathogenic toxicological mechanisms of DQ has hindered the development of treatments against DQ poisoning. Pyroptosis is a form of programmed cell death and was recently identified as a novel molecular mechanism of drug-induced AKI. To explore the role of pyroptosis in HK-2 cells exposed to DQ, the plasma membrane damage of the cells was detected by LDH release assay. Western blot was performed to detect the cleavage of GSDME. Proteomics analysis was performed to explore the mechanism of DQ induced nephrotoxicity. FerroOrange probe was used to measure the intracellular Fe2+ levels. Herein, we show that DQ induces pyroptosis in HK-2 cells. Mechanistically, DQ induces the accumulation of mitochondrial ROS and initiates the cleavage of gasdermin E (GSDME) in an intrinsic mitochondrial pathway. Knockout of GSDME attenuated DQ-induced cell death. Further analysis revealed that loss of FTH1 induces Fe2+ accumulation, contributing to DQ-induced pyroptosis. Knockdown LC3B could help restore the expression of FTH1 and improve cell viability. Moreover, we found DFO, an iron chelator, could reduce cellular Fe2+ levels and inhibit pyroptosis. Collectively, these findings suggest an unrecognized mechanism for GSDME-dependent pyroptosis in DQ-induced AKI.

LonP1 Links Mitochondria-ER Interaction to Regulate Heart Function.

Interorganelle contacts and communications are increasingly recognized to play a vital role in cellular function and homeostasis. In particular, the mitochondria-endoplasmic reticulum (ER) membrane contact site (MAM) is known to regulate ion and lipid transfer, as well as signaling and organelle dynamics. However, the regulatory mechanisms of MAM formation and their function are still elusive. Here, we identify mitochondrial Lon protease (LonP1), a highly conserved mitochondrial matrix protease, as a new MAM tethering protein. The removal of LonP1 substantially reduces MAM formation and causes mitochondrial fragmentation. Furthermore, deletion of LonP1 in the cardiomyocytes of mouse heart impairs MAM integrity and mitochondrial fusion and activates the unfolded protein response within the ER (UPRER). Consequently, cardiac-specific LonP1 deficiency causes aberrant metabolic reprogramming and pathological heart remodeling. These findings demonstrate that LonP1 is a novel MAM-localized protein orchestrating MAM integrity, mitochondrial dynamics, and UPRER, offering exciting new insights into the potential therapeutic strategy for heart failure.

Human cytomegalovirus infection activates NLRP3 inflammasome by releasing mtDNA into the cytosol in human THP-1 cells.

Human cytomegalovirus (HCMV) infection of monocytes results in the production of inflammatory cytokine through inflammasome. However, the mechanism of NLR family pyrin domain containing 3 (NLRP3) inflammasome activation in HCMV infection remains unclear. In this study, HCMV infection promoted the increase of mitochondrial fusion and caused mitochondrial dysfunction in THP-1 cells, including excessive reactive oxygen species production and decreased mitochondrial membrane potential (Δψm). Meanwhile, the expression of mitochondrial DNA (mtDNA)-binding protein TFAM (transcription factor A, mitochondrial) was decreased and mtDNA content in the cytoplasm was increased. Knockdown of TFAM caused an increase in mtDNA copy number in the cytoplasm and resulted in elevated NLRP3 expression, active caspase-1, and mature IL-1ß. After a 3 h treatment with MCC950, an NLRP3 inhibitor, the increase of cleaved caspase-1 and mature IL-1ß were suppressed. Besides, overexpression of TFAM inhibited the expression of NLRP3, cleaved caspase-1, and mature IL-1ß. In addition, knockdown of NLRP3 inhibited the IL-1ß process after HCMV infection. mtDNA-deficient cells showed a limited ability to produce NLRP3 and process IL-1ß after HCMV infection. In conclusion, HCMV infection of THP-1 cells resulted in decreased mitochondrial TFAM protein expression and increased mtDNA release into the cytoplasm, which eventually led to the activation of NLRP3 inflammasome.

Construction of a metabolism-related gene prognostic model to predict survival of pancreatic cancer patients.

Pancreatic cancer (PC) is one of the most fatal malignant tumors, and is commonly diagnosed at an advanced stage with no effective therapy. Metabolism-related genes (MRGs) and immune-related genes (IRGs) play considerable roles in the tumor microenvironment. Therefore, an effective prediction model based on MRGs and IRGs could aid in the prognosis of PC. In this study, differential expression analysis was performed to gain 25 intersectional genes from 857 differentially expressed MRGs (DEMRGs), and 1353 differentially expressed IRGs, from The Cancer Genome Atlas database of PC. Cox and Lasso regression were applied and a five-DEMRGs prognostic model constructed. Survival analysis, ROC values, risk curve and validation analysis showed that the model could independently predict PC prognosis. In addition, the correlation analysis suggested that the five-DEMRGs prognostic model could reflect the status of the immune microenvironment, including Tregs, M1 macrophages and Mast cell resting. Therefore, our study provides new underlying predictive biomarkers and associated immunotherapy targets.

Human cytomegalovirus-induced immune regulation is correlated with poor prognosis in patients with colorectal cancer.

Evidence suggests that human cytomegalovirus (HCMV) infection may be implicated in the progression of colorectal cancer (CRC). However, the correlation between HCMV infection and survival outcomes in patients with CRC remains unclear. Here, we constructed a flow algorithm to identify HCMV sequences based on the RNA-seq data of patients with CRC derived from the Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO). The patients' clinical information matrix was used to calculate the Euclidean distance to filter out suitable patients not infected with HCMV, combined with patients' survival outcomes, to reveal how HCMV infection is involved in CRC progression. HCMV infection is widespread in patients with CRC, and the prevalence of HCMV infection ranges from 10 to 36% in four independent CRC datasets, with infection being concentrated in carcinoma tissue rather than in normal tissue. In addition, HCMV-positive patients had a poor survival prognosis, with three HCMV genes, UL82, UL42, and UL117, associated with poor patient survival outcomes. Most importantly, we suppose that the regulation of immune function by HCMV may be key to the poor prognosis of patients with CRC. We found that HCMV infection was associated with poor prognosis in CRC patients and identified three prognosis-associated HCMV genes. The regulation of immune function caused by HCMV infection was the key factor, while HCMV-positive patients with CRC mostly presented with a state of immunosuppression. This may provide new ideas for the personalized treatment of patients with CRC, especially with respect to immunotherapy.

A novel small molecule glycolysis inhibitor WZ35 exerts anti-cancer effect via metabolic reprogramming.

BACKGROUND: Liver cancer is the fifth leading cause of cancer death worldwide, but early diagnosis and treatment of liver cancer remains a clinical challenge. How to screen and diagnose liver cancer early and prolong the survival rate is still the focus of researchers. METHODS: Cell experiments were used to detect the effect of WZ35 on the colony formation ability and proliferation activity of hepatoma cells, nude mouse experiment to observe the in vivo anticancer activity and toxic side effects of WZ35; metabolomics analysis, glucose metabolism experiment and Seahorse analysis of liver cancer cells treated with WZ35; cell experiments combined with bioinformatics analysis to explore the mechanism of WZ35-mediated metabolic reprogramming to exert anticancer activity; tissue microarray and case analysis to evaluate the clinical significance of biomarkers for early diagnosis, treatment and prognosis evaluation of liver cancer. RESULTS: WZ35 inhibited the proliferation activity of various cell lines of liver cancer, and showed good therapeutic effect in nude mice model of hepatocellular carcinoma without obvious toxic and side effects; WZ35 inhibited the absorption of glucose in hepatoma cells, and the drug effect glycolysis, phosphorylation and purine metabolism are relatively seriously damaged; WZ35 mainly inhibits YAP from entering the nucleus as a transcription factor activator by activating oxidative stress in liver cancer cells, reducing the transcription of GLUT1, and finally reducing its GLUT1. Tissue microarray and case analysis showed that GLUT1 and YAP were highly expressed and correlated in liver cancer patients, and were associated with poor patient prognosis. The GLUT1-YAP risk model had a high score in predicting prognosis. CONCLUSION: The study confirms that WZ35 is a small molecule glycolysis inhibitor, and through its properties, it mediates metabolic reprogramming dominated by impaired glycolysis, oxidative phosphorylation and purine metabolism to inhibit the proliferation activity of liver cancer cells. Our findings present novel insights into the pathology of liver cancer and potential targets for new therapeutic strategies. GLUT1-YAP has important reference significance for predicting the stages of disease progression in liver cancer patients and have the potential to serve as novel biomarkers for the diagnosis and treatment of liver cancer.

Oxaloacetate acid ameliorates paraquat-induced acute lung injury by alleviating oxidative stress and mitochondrial dysfunction.

Acute lung injury (ALI) is the primary cause of death among patients with acute paraquat (PQ) poisoning, whereby peroxidative damage is an important mechanism underlying PQ-induced lung injury. There is a lack of effective interventional drugs for patients with PQ poisoning. Oxaloacetic acid (OAA) participates in multiple in vivo metabolic processes, whereby it facilitates the clearance of reactive oxygen species (ROS) and improves mitochondrial function. The study aimed to assess the protective effects of OAA on PQ-induced ALI and elucidate the underlying molecular mechanism. Our data demonstrated that OAA treatment significantly alleviated PQ-induced ALI and improved the survival rate of PQ-poisoned mice, and also alleviated PQ-induced cellular oxidative stress and mitochondrial dysfunction. OAA-mediated alleviation of PQ-induced mitochondrial dysfunction depends on the following mechanisms which may explain the above findings: 1) OAA effectively cleared intracellular ROS, inhibited ROS accumulation, and mitochondrial depolarization; 2) OAA inhibited the downregulation of L-OPA1 and MFN2 caused by PQ and promoted a dynamic balance of mitochondrial fusion and fission, and 3) the expression of PGC-1α, TFAM, COX2, and COX4I1, increased significantly following OAA intervention which improved mitochondrial respiratory functions and promoted its biogenesis and energy metabolism in damaged cells. In conclusion, OAA effectively cleared ROS and improved mitochondrial dysfunction, thereby significantly improving ALI caused by PQ poisoning and the animal survival rate. Therefore, OAA may be a potential drug for the treatment of PQ poisoning.

Effects of natural 24-epibrassinolide on inducing apoptosis and restricting metabolism in hepatocarcinoma cells.

BACKGROUND: 24-epibrassinolide (EBR) is a ubiquitous steroidal phytohormone with anticancer activity. Yet the cytotoxic effects and mechanism of EBR on hepatocarcinoma (HCC) cells remain elusive. METHODS: Cell counting kit-8 (CCK-8) assay was performed to evaluate cell viability. Real-time cell analysis (RTCA) technology and colony formation assays were used to evaluate cell proliferation. The apoptosis ratio was measured by flow cytometry. Seahorse XFe96 was applied to detect the effects of EBR on cellular bioenergetics. RNA-seq analysis was performed to investigate differences in gene expression profiles. Western blot and qRT-PCR were used to detect the changes in target molecules. RESULTS: EBR induced apoptosis and caused energy restriction in HCC, both of which were related to insulin-like growth factor-binding protein 1 (IGFBP1). EBR rapidly and massively induced IGBFP1, part of which was transcribed by activating transcription factor-4 (ATF4). The accumulation of secreted and cellular IGFBP1 had different important roles, in which secreted IGFBP1 affected cell energy metabolism by inhibiting the phosphorylation of Akt, while intracellular IGFBP1 acted as a pro-survival factor to resist apoptosis. Interestingly, the extracellular signal-regulated kinase (ERK) inhibitor SCH772984 and MAP/ERK kinase (MEK) inhibitor PD98059 not only attenuated the EBR-induced IGFBP1 expression but also the basal expression of IGFBP1. Thus, the treatment of cells with these inhibitors further enhances the cytotoxicity of EBR. CONCLUSION: Taken together, these findings suggested that EBR can be considered as a potential therapeutic compound for HCC due to its pro-apoptosis, restriction of energy metabolism, and other anti-cancer properties. Meanwhile, the high expression of IGFBP1 induced by EBR in HCC contributes to our understanding of the role of IGFBP1 in drug resistance.

Synthetic oleanane triterpenoid derivative CDDO-Me disrupts cellular bioenergetics to suppress pancreatic ductal adenocarcinoma via targeting SLC1A5.

To investigate the potential antitumor activity of synthetic triterpenoid, methyl-2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oate (CDDO-Me) in pancreatic ductal adenocarcinoma (PDAC), MTT cytotoxicity assay, and xenograft nude mice assay were performed to evaluate tumor growth in vitro and in vivo. Seahorse XFe96 bioenergetics analyzer was applied to determine aerobic glycolysis and mitochondrial respiration. Western blot and quantitative reverse transcription-polymerase chain reactions are used to detect protein and messenger RNA transcripts of SLC1A5 and metabolic enzymes. We confirmed the strong antitumor activity of CDDO-Me in suppressing PDAC growth. Mechanistically, we demonstrated CDDO-Me induced mitochondrial respiration and aerobic glycolysis dysfunction. We also verified CDDO-Me downregulated glutamine transporter SLC1A5, resulting in excessive reactive oxygen species (ROS) levels that suppressed tumor growth. Moreover, we confirmed that SLC1A5 depletion reduced the ratio of glutathione/oxidized glutathione. We also found CDDO-Me could inhibit N-linked glycosylation of SLC1A5, which promotes protease-mediated degradation. Finally, we confirmed SLC1A5 was significantly overexpressed in PDAC and closely correlated with the poor prognosis of PDAC patients. Our work uncovers CDDO-Me is effective at suppressing PDAC cell growth in vitro and in vivo and illuminates CDDO-Me caused excessive ROS and cellular bioenergetics disruption which contributed to CDDO-Me inhibited PDAC growth. Our data highlights CDDO-Me could be considered a potential compound for PDAC therapy, and SLC1A5 could be a novel biomarker for PDAC patients.

Hepatic SIRT6 deficit promotes liver tumorigenesis in the mice models.

SIRT6 belongs to class III sirtuin family with NAD+-dependent histone deacetylase activities and controls multiple processes including aging, metabolism and inflammation. In recent years, increasing studies showed tumor suppressor role of SIRT6 in HCC development. We established a two-stage DEN followed CCl4 induced liver carcinogenesis in the hepatic-specific SIRT6 HKO mice models and found that hepatic SIRT6 deficit significantly promotes liver injury and liver cancer through inhibition of the ERK1/2 pathway. SIRT6 was compensatory upregulated in mice tumor tissues and human HCC cells and overexpressed SIRT6 inhibits tumor growth both in vitro and in vivo. Taken together, we provide a useful mouse model for delineating the molecular pathways involved in chronic liver diseases and primary liver cancer and suggest that SIRT6 can be a promising target for HCC therapies.

N-glycosylation stabilizes MerTK and promotes hepatocellular carcinoma tumor growth.

Despite the evidences of elevated expression of Mer tyrosine kinase (MerTK) in multiple human cancers, mechanisms underlying the oncogenic roles of MerTK in hepatocellular carcinoma (HCC) remains undefined. We explored the functional effects of MerTK and N-Glycosylated MerTK on HCC cell survival and tumor growth. Here, we show that MerTK ablation increases reactive oxygen species (ROS) production and promotes the switching from glycolytic metabolism to oxidative phosphorylation in HCC cells, thus suppressing HCC cell proliferation and tumor growth. MerTK is N-glycosylated in HCC cells at asparagine 294 and 454 that stabilizes MerTK to promote oncogenic transformation. Moreover, we observed that nuclear located non-glycosylated MerTK is indispensable for survival of HCC cells under stress. Pathologically, tissue microarray (TMA) data indicate that MerTK is a pivotal prognostic factor for HCC. Our data strongly support the roles of MerTK N-glycosylation in HCC tumorigenesis and suggesting N-glycosylation inhibition as a potential HCC therapeutic strategy.

6-methoxydihydroavicine, the alkaloid extracted from Macleaya cordata (Willd.) R. Br. (Papaveraceae), triggers RIPK1/Caspase-dependent cell death in pancreatic cancer cells through the disruption of oxaloacetic acid metabolism and accumulation of reactive oxygen species.

BACKGROUND: Many extracts and purified alkaloids of M. cordata (Papaveraceae family) have been reported to display promising anti-tumor effects by inhibiting cancer cell growth and inducing apoptosis in many cancer types. However, no evidence currently exists for anti-pancreatic cancer activity of alkaloids extracted from M. cordata, including a novel alkaloid named 6­methoxy dihydrosphingosine (6-Methoxydihydroavicine, 6-ME) derived from M. cordata fruits. PURPOSE: The aim of this study was to investigate the anti-tumor effects of 6-ME on PC cells and the underlying mechanism. METHODS: CCK-8, RTCA, and colony-formation assays were used to analyze PC cell growth. Cell death ratios, changes in MMP and ROS levels were measured by flow cytometry within corresponding detection kits. A Seahorse XFe96 was employed to examine the effects of 6-ME on cellular bioenergetics. Western blot and q-RT-PCR were conducted to detect changes in target molecules. RESULTS: 6-ME effectively reduced the growth of PC cells and promoted PCD by activating RIPK1, caspases, and GSDME. Specifically, 6-ME treatment caused a disruption of OAA metabolism and increased ROS production, thereby affecting mitochondrial homeostasis and reducing aerobic glycolysis. These responses resulted in mitophagy and RIPK1-mediated cell death. CONCLUSION: 6-ME exhibited specific anti-tumor effects through interrupting OAA metabolic homeostasis to trigger ROS/RIPK1-dependent cell death and mitochondrial dysfunction, suggesting that 6-ME could be considered as a highly promising compound for PC intervention.

A synthesized olean-28,13ß-lactam targets YTHDF1-GLS1 axis to induce ROS-dependent metabolic crisis and cell death in pancreatic adenocarcinoma.

BACKGROUND: Pancreatic adenocarcinoma (PAAD) is a severe malignant with a 5-year survival rate of approximately 9%. Oleanolic acid is a well-known natural triterpenoid which exhibits pharmacological activities. We previously synthesized a series of oleanolic acid derivatives and evaluated the tumor-suppressive activity of olean-28,13ß-lactam (B28) in prostate cancer. However, the detailed mechanism remains to be understood. METHODS: The anti-tumor activity of B28 in PAAD was confirmed by RTCA, colony formation assay and flow cytometry. GO and KEGG enrichment analyses were performed to analyze the differentially expressed genes (DEGs) obtained by RNA sequencing. The effects of B28 on cell bioenergetics were evaluated by seahorse analyzer. Lenti-virus packaged plasmids were performed to knockdown or overexpress target genes. Alteration of mitochondrial membrane potential, ROS and GSH/GSSG were measured by corresponding detection kits according to the manufacturer's protocol. RESULTS: We evaluated and confirmed the promising anti-tumor activity of B28 in vitro. RNA-seq profile indicated that multiple metabolic pathways were interrupted in B28 treated PAAD cells. Next, we demonstrated that B28 induces cellular bioenergetics crisis to inhibit PAAD cells growth and induce cell death. We further validated that cell cycle arrest, inhibition of cell growth, cell apoptosis and cell bioenergetics disruption were functionally rescued by ROS scavenger NAC. Mechanistically, we found glutamine metabolism was inhibited due to B28 administration. Moreover, we validated that down-regulation of GLS1 contributes to ROS generation and bioenergetics interruption induced by B28. Furthermore, we elucidated that YTHDF1-GLS1 axis is the potential downstream target of B28 to induce PAAD cell metabolic crisis and cell death. Finally, we also confirmed the anti-tumor activity of B28 in vivo. CONCLUSIONS: Current study demonstrates B28 disrupts YTDFH1-GLS1 axis to induce ROS-dependent cell bioenergetics crisis and cell death which finally suppress PAAD cell growth, indicating that this synthesized olean-28,13ß-lactam maybe a potent agent for PAAD intervention.

Single-Cell Transcriptome Reveals the Metabolic and Clinical Features of a Highly Malignant Cell Subpopulation in Pancreatic Ductal Adenocarcinoma.

Background: Pancreatic ductal adenocarcinoma (PDAC) is a malignant tumor with a high mortality rate. PDAC exhibits significant heterogeneity as well as alterations in metabolic pathways that are associated with its malignant progression. In this study, we explored the metabolic and clinical features of a highly malignant subgroup of PDAC based on single-cell transcriptome technology. Methods: A highly malignant cell subpopulation was identified at single-cell resolution based on the expression of malignant genes. The metabolic landscape of different cell types was analyzed based on metabolic pathway gene sets. In vitro experiments to verify the biological functions of the marker genes were performed. PDAC patient subgroups with highly malignant cell subpopulations were distinguished according to five glycolytic marker genes. Five glycolytic highly malignant-related gene signatures were used to construct the glycolytic highly malignant-related genes signature (GHS) scores. Results: This study identified a highly malignant tumor cell subpopulation from the single-cell RNA sequencing (scRNA-seq) data. The analysis of the metabolic pathway revealed that highly malignant cells had an abnormally active metabolism, and enhanced glycolysis was a major metabolic feature. Five glycolytic marker genes that accounted for the highly malignant cell subpopulations were identified, namely, EN O 1, LDHA, PKM, PGK1, and PGM1. An in vitro cell experiment showed that proliferation rates of PANC-1 and CFPAC-1 cell lines decreased after knockdown of these five genes. Patients with metabolic profiles of highly malignant cell subpopulations exhibit clinical features of higher mortality, higher mutational burden, and immune deserts. The GHS score evaluated using the five marker genes was an independent prognostic factor for patients with PDAC. Conclusion: We revealed a subpopulation of highly malignant cells in PDAC with enhanced glycolysis as the main metabolic feature. We obtained five glycolytic marker gene signatures, which could be used to identify PDAC patient subgroups with highly malignant cell subpopulations, and proposed a GHS prognostic score.

Identification and Validation of CYBB, CD86, and C3AR1 as the Key Genes Related to Macrophage Infiltration of Gastric Cancer.

Gastric cancer (GC) is rampant around the world. Most of the GC cases are detected in advanced stages with poor prognosis. The identification of marker genes for early diagnosis is of great significance. Studying the tumor environment is helpful to acknowledge the process of tumorigenesis, development, and metastasis. Twenty-two kinds of immune cells were calculated by CIBERSORT from Gene Expression Omnibus (GEO) database. Subsequently, higher infiltration of macrophages M0 was discovered in GC compared with normal tissues. WGCNA was utilized to construct the network and then identify key modules and genes related to macrophages in TCGA. Finally, 18 hub genes were verified. In the PPI bar chart, the top 3 genes were chosen as hub genes involved in most pathways. On the TIMER and THPA websites, it is verified that the expression levels of CYBB, CD86, and C3AR1 genes in tumor tissues were higher than those in normal tissues. These genes may work as biomarkers or targets for accurate diagnosis and treatment of GC in the future. Our findings may be a new strategy for the treatment of GC.

RUNX2 and LAMC2: promising pancreatic cancer biomarkers identified by an integrative data mining of pancreatic adenocarcinoma tissues.

Pancreatic carcinoma (PC) is a severe disease associated with high mortality. Although strategies for cancer therapy have made great progress, outcomes of pancreatic carcinoma patients remain extremely poor. Therefore, it is urgent to find novel biomarkers and therapeutic targets. To identify biomarkers for early diagnosis and therapy, three mRNA microarray datasets and two miRNA datasets were selected, and combinative analysis was performed by GEO2R. Functional and pathway enrichment analysis were performed using DAVID database. MiRTarBase, miRWalk and Diana Tools were used to get key genes. TCGA, HPA and western blotting were used to verify diagnostic and prognostic value of key genes. By integrating mRNA and miRNA expression profiles, we identified 114 differentially expressed genes and 114 differentially expressed miRNAs, respectively. Then, three overlapping key genes, RUNX2, LAMC2 and FBXO32, were found. Their protein levels in pancreatic tissue from PC patients and normal people were analyzed by immunohistochemical staining and western blotting. RUNX2 showed a potential property to identify PC. Aberrant over-expression of LAMC2 was associated with poor prognosis of PC patients, tumor status and subtypes. In summary, our current study identified that RUNX2 and LAMC2 may be promising targets for early diagnosis and therapy of PC patients.

miRNA-382-5p Suppresses the Expression of Farnesoid X Receptor to Promote Progression of Liver Cancer.

BACKGROUND: The dysregulation of microRNAs (miRNAs) and hepatotoxicity due to the aberrant accumulation of bile acids (BAs) are notorious causes that predispose an individual to the development of hepatocellular carcinoma (HCC). Farnesoid X receptor (FXR), encoded by NR1H4 gene, has been identified as a crucial BA receptor to maintain the homeostasis of BA pool and its expression is decreased in HCC. miR-382-5p plays an important role in the pathogenesis of many human malignancies and was reported to promote the proliferation and differentiation of normal liver cells and liver regeneration. However, there is still some controversy about its role in HCC microenvironment. This study aims to explore the expression pattern of miR-382-5p in HCC and its role in regulating FXR during the development of HCC. METHODS: Tissues collected from 30 HCC patients were subjected to extraction of total RNA and quantitative real-time PCR (qRT-PCR) for the analyses of miR-382-5p expression and NR1H4 mRNA levels, and their expressions were verified by analyzing the online HCC-related GSE datasets. The role of miR-382-5p in regulating cellular proliferation and expression of FXR in different HCC cell lines was analyzed by qRT-PCR, Western Blot, real-time cellular analysis (RTCA) and luciferase reporter assays. The role of miR-382-5p in regulating downstream genes of FXR in HCC cells was also analyzed. RESULTS: miR-382-5p was upregulated in HCC tissues and inversely associated with the downregulation of NR1H4 mRNA levels. The luciferase reporter assay proved that miR-382-5p directly targeted the 3'-untranslated region (3'-UTR) of human NR1H4 mRNA. Overexpression of miR-382-5p led to a malignant proliferation of HCC cells by suppressing the expression of FXR. In contrast, blocking the endogenous miR-382-5p was sufficient to suppress the cellular proliferation rate of HCC through increasing FXR expression. Additionally, miR-382-5p inhibited the expression of some target genes of FXR, including SHP, FGF19 and SLC51A, and this inhibitory effect was FXR-dependent. CONCLUSION: Therefore, miR-382-5p promotes the progression of HCC in vitro by suppressing FXR and could serve as a valuable therapeutic target for HCC treatment.

A Novel Mechanism of Cannabidiol in Suppressing Hepatocellular Carcinoma by Inducing GSDME Dependent Pyroptosis.

Cannabidiol (CBD), a phytochemical derived from Cannabis sativa L., has been demonstrated to exhibit promising anti-tumor properties in multiple cancer types. However, the effects of CBD on hepatocellular carcinoma (HCC) cells remain unknown. We have shown that CBD effectively suppresses HCC cell growth in vivo and in vitro, and induced HCC cell pyroptosis in a caspase-3/GSDME-dependent manner. We further demonstrated that accumulation of integrative stress response (ISR) and mitochondrial stress may contribute to the initiation of pyroptotic signaling by CBD. Simultaneously, CBD can repress aerobic glycolysis through modulation of the ATF4-IGFBP1-Akt axis, due to the depletion of ATP and crucial intermediate metabolites. Collectively, these observations indicate that CBD could be considered as a potential compound for HCC therapy.

CXCL10 is a Tumor Microenvironment and Immune Infiltration Related Prognostic Biomarker in Pancreatic Adenocarcinoma.

Pancreatic adenocarcinoma (PAAD) is the 10th most common cancer worldwide and the outcomes for patients with the disease remain extremely poor. Precision biomarkers are urgently needed to increase the efficiency of early diagnosis and to improve the prognosis of patients. The tumor microenvironment (TME) and tumor immune infiltration are thought to impact the occurrence, progression, and prognosis of PAAD. Novel biomarkers excavated originating from the TME and immune infiltration may be effective in predicting the prognosis of PAAD patients. In the current study, the ESTIMATE and CIBERSORT algorithms were applied to estimate the division of immune and stromal components and the proportion of tumor-infiltrating immune cells in 182 PAAD cases downloaded from The Cancer Genome Atlas database. Intersection analyses of the Protein-Protein Interaction networks and Cox regression analysis identified the chemokine (CXC-motif) ligand 10 (CXCL10) as a predictive biomarker. We verified that CXCL10 in the TME negatively correlates with prognosis in PAAD and positively correlates with tumor cell differentiation. GSE62452 from the GEO database and cumulative survival analysis were performed to validate CXCL10 expression as an independent prognostic indicator. We also found that memory B cells, regulatory T cells, and macrophages M0 and M1 were correlated with the expression of CXCL10 indicating that expression of CXCL10 influenced the immune activity of the TME. Our data suggest that CXCL10 is beneficial as a prognostic indicator in PAAD patients and highlights the potential for immune targeted therapy in the treatment of PAAD.