Targeting SOX13 inhibits assembly of respiratory chain supercomplexes to overcome ferroptosis resistance in gastric cancer.

Therapeutic resistance represents a bottleneck to treatment in advanced gastric cancer (GC). Ferroptosis is an iron-dependent form of non-apoptotic cell death and is associated with anti-cancer therapeutic efficacy. Further investigations are required to clarify the underlying mechanisms. Ferroptosis-resistant GC cell lines are constructed. Dysregulated mRNAs between ferroptosis-resistant and parental cell lines are identified. The expression of SOX13/SCAF1 is manipulated in GC cell lines where relevant biological and molecular analyses are performed. Molecular docking and computational screening are performed to screen potential inhibitors of SOX13. We show that SOX13 boosts protein remodeling of electron transport chain (ETC) complexes by directly transactivating SCAF1. This leads to increased supercomplexes (SCs) assembly, mitochondrial respiration, mitochondrial energetics and chemo- and immune-resistance. Zanamivir, reverts the ferroptosis-resistant phenotype via directly targeting SOX13 and promoting TRIM25-mediated ubiquitination and degradation of SOX13. Here we show, SOX13/SCAF1 are important in ferroptosis-resistance, and targeting SOX13 with zanamivir has therapeutic potential.

Preoperative pancreatic CT value is related to pancreatic fistula after pancreaticoduodenectomy: a retrospective study.

Background: Pancreatic fistula (PF) is the main complication in patients undergoing pancreaticoduodenectomy. Computed tomography (CT) value can reflect pancreatic tissue characteristics which is related to PF. This study was designed to study the relationship between the preoperative CT value and pancreatic fistula. Methods: We retrospectively reviewed the clinical and medical data of patients undergoing pancreaticoduodenectomy from 2017 to 2021. The pancreatic CT value and the CT value ratios of the pancreas and abdominal aorta (PCT/ACT) were measured and compared between the PF group and non-PF group. The values in different PF severity groups were compared using variance analysis. A cut-off value was selected by receiver operating characteristic (ROC) curve. Single-factor and multiple-factor analysis were performed to evaluate Correlation between PF and CT. Results: One hundred and twenty-seven cases were included in this study. The PCT/ACT in the PF group was significantly lower than that in the non-PF group (P<0.001), and the PCT/ACT value was correlatively lower in the severe PF group than in the mild PF group (P=0.008). A cutoff value of 0.99 was selected by ROC curves analysis. Further multifactor analysis identified PCT/ACT <0.99 to be an independent preoperative predictor [odds ratio (OR): 11.3, P<0.01]. Conclusions: The preoperative pancreatic CT value can indirectly reflect the histological condition of the pancreas and thus may related to postoperative PF after pancreaticoduodenectomy and provide useful information for surgeons in deciding upon the pancreaticojejunostomy method.

Potential factors affecting success rate and long term outcome in single balloon enteroscopy-assisted therapeutic endoscopic retrograde cholangiopancreatography in patients with pancreaticojejunal anastomotic stenosis: a retrospective study.

Background: Pancreaticojejunal anastomotic stenosis (PJS) after pancreaticoduodenectomy (PD) is difficult to treat. Single-balloon enteroscope-assisted endoscopic retrograde pancreatography (SBE-assisted ERP) is a safe way to treat PJS with the strength of minimally invasion and repeatability, but since its technical difficulty and few patient number, data on long-term outcomes remain limited. The optimal treatment is still unknown. We aim to study the safety, effectiveness, and long-term outcome of single balloon enteroscopy-assisted (SBE-assisted) therapeutic ERP in patients with PJS in this study. Methods: The clinical information of patients undergoing SBE-assisted therapeutic ERP from March 2016 to March 2021 were retrospectively analyzed. All patients were diagnosed as PJS and without any contraindication for therapeutic endoscopy. Treatment details, postoperative complications, factors influencing technical success rate were evaluated. Long-term outcomes results were obtained by clinical or telephone follow-up. Results: Sixteen patients with median age of 51 years were included in this study, surgical reconstruction methods including PD with Whipple reconstruction, PD with Child reconstruction, pylorus-preserving pancreaticoduodenectomy (PpPD) with Whipple reconstruction. Eight patients were successfully treated. No serious complications happened. Risk factors for the failure of pancreaticojejunal anastomotic site identification include the digestive tract reconstruction sequence, pancreaticojejunostomy method, pancreatic duct tube implantation, pancreatic duct width before surgery, and pancreatic fistula during perioperative period. The median follow-up time was 77.2 months, the mean indwelling time of the stent was 62.3 months [interquartile range (IQR), 6.8-153.7 months]. Two of eight patients developed recurrent PJS. The variation in body mass index (BMI) was +2.46 in the non-recurrence group compared to -1.09 in the recurrence group and -2.12 in the endoscopic retrograde cholangiopancreatography (ERCP) treatment failure group. Conclusions: ERP intervention should be carried out early once PJS occurs in order to increase success rate. BMI is a crucial indicator which can reflex PJS rehabilitation degree during follow-up. In order to reduce PJS recurrence rate, a wider pancreatic stent and a longer stent indwelling time are recommended.

Knockdown of IGF2BP3 inhibits the tumorigenesis of gallbladder cancer and modifies tumor microenvironment.

Gallbladder cancer (GBC) ranks seventh among the gastrointestinal cancers. Messenger RNAs (mRNAs) could regulate the progression of GBC. For the purpose of exploring the targets for GBC treatment, RNA sequencing was used to identify the differential expressed mRNAs between GBC and adjacent tissues. Next, CCK8 assay was used to assess the cell viability, and cell proliferation was investigated by colony formation assay. Flow cytometry was performed to evaluate the cell apoptosis. Protein and mRNA expression were analyzed by western blot and RT-qPCR, respectively. Transwell was performed to evaluate the cell metastasis. GBC-derived exosomes were isolated with ultracentrifugation. To evaluate the function of exosomes in GBC, in vivo model of GBC was constructed. The data revealed IGF2BP3 was identified to be upregulated in GBC, and IGF2BP3 silencing was able to decrease GBC cell proliferation by promoting the apoptosis. The migration and invasion of GBC cells were reduced by IGF2BP3 knockdown. Silencing of IGF2BP3 obviously suppressed the level of p-STAT3 in GBC cells. Meanwhile, GBC cell-derived exosomes notably promoted macrophage M2 polarization via carrying IGF2BP3, and then the polarized macrophages promoted the malignant behavior of GBC cells. Furthermore, exosomes markedly promoted the tumor growth of GBC via promoting macrophage M2 polarization. In summary, knockdown of IGF2BP3 suppressed the malignant behavior of GBC cells. Additionally, knockdown of IGF2BP3 modified tumor microenvironment during the progression of GBC. Thus, these findings might provide a new theoretical basis for exploring a strategies against GBC.

Hypoxia inducible lncRNA-CBSLR modulates ferroptosis through m6A-YTHDF2-dependent modulation of CBS in gastric cancer.

Introduction: Tumors are usually refractory to anti-cancer therapeutics under hypoxic conditions. However, the underlying molecular mechanism remains to be elucidated. Objectives: Our study intended to identify hypoxia inducible lncRNAs and their biological function in gastric cancer (GC). Methods: Differentially expressed lncRNAs were determined by microarray analysis between GC cells exposed to hypoxia (1% O2) and normoxia (21% O2) for 24 h. The expression level of CBSLR was manipulated in several GC cell lines to perform molecular and biological analyses both in vitro and in vivo. Results: We identified a hypoxia-induced lncRNA-CBSLR that protected GC cells from ferroptosis, leading to chem-resistance. Mechanically, CBSLR interacted with YTHDF2 to form a CBSLR/YTHDF2/CBS signaling axis that decreased the stability of CBS mRNA by enhancing the binding of YTHDF2 with the m6A-modified coding sequence (CDS) of CBS mRNA. Furthermore, under decreased CBS levels, the methylation of the ACSL4 protein was reduced, leading to protein polyubiquitination and degradation of ACSL4. This, in turn, decreased the pro-ferroptosis phosphatidylethanolamine (PE) (18:0/20:4) and PE (18:0/22:4) content and contributed to ferroptosis resistance. Notably, CBSLR is upregulated, whereas CBS is downregulated in GC tissues compared to matched normal tissues; and GC patients with high CBSLR/low CBS levels have a worse clinical outcome and a poorer response to chemotherapy. Conclusion: Our study reveals a novel mechanism in how HIF1α/CBSLR modulates ferroptosis/chemoresistance in GC, illuminating potential therapeutic targets for refractory hypoxic tumors.

Trends of gallbladder cancer incidence, mortality, and diagnostic approach in urban Shanghai between 1973 and 2009.

PURPOSE: To describe and interpret secular time trends in gallbladder cancer (GBC) incidence, mortality, and diagnostic approach using 37 years of cancer registry data in urban Shanghai. METHODS: Data on registration of GBC in urban Shanghai during 1973 and 2009 were collected by the Shanghai Cancer Registry. To describe time trends and to identify specific time points when significant changes occurred, we used joinpoint regression analysis. RESULTS: The age-standardized rates (ASRs) of incidence increased from 1.1/100,000 (1973-1975) to 2.9/100,000 (2006-2009) in men and from 1.7/100,000 (1973-1975) to 3.9/100,000 (2006-2009) in women. ASRs of incidence increased significantly with estimated annual percent changes (EAPCs) of 2.8% in men and 2.5% in women. The mortality trends increased significantly, with EAPCs of 2.8% in men and 2.5% in women. The increasing incidence and mortality rates were primarily observed in men ⩾60 years of age and in women ⩾70 years of age. Notable downward trends in incidence and mortality were identified among women age 60-69 years over the last decade. The percentage of GBC diagnosed by pathology increased steadily over the years while the percentage of GBC diagnosed by imaging, surgery, and biochemistry sharply increased from 1987 onwards. CONCLUSIONS: Thirty-seven years of cancer registry data document a tremendous increase in incidence/mortality and a slight decline in incidence/mortality over the last decades for GBC, especially among women, in Shanghai. The development of diagnostic approaches and aging population may play important roles.

Circular RNA FOXP1 promotes tumor progression and Warburg effect in gallbladder cancer by regulating PKLR expression.

BACKGROUND: Circular RNAs (circRNAs) have recently been identified as potential functional modulators of the cellular physiology processes. The study aims to uncover the potential clinical value and driving molecular mechanisms of circRNAs in gallbladder cancer (GBC). PATIENTS AND METHODS: We performed RNA sequencing from four GBC and paired adjacent normal tissues to analyze the circRNA candidates. Quantitative real-time polymerase chain reaction (QRT-PCR) was used to measure the circFOXP1 expression from 40 patient tissue samples. Short hairpin RNA mediated knockdown or exogenous expression of circFOXP1 combined with in vitro and in vivo assays were performed to prove the functional significance of circFOXP1. Double luciferase reporter, RNA immunoprecipitation (RIP) and RNA pull-down assays were also performed. RESULTS: By performing RNA sequencing from GBC and paired adjacent normal tissues to analyze the circRNA candidates, we identified that circFOXP1 (hsa_circ_0008234) expression was significantly upregulated in GBC tissues and positively associated with lymph node metastasis, advanced TNM stage and poor prognosis in patients. Short hairpin RNA mediated knockdown or exogenous expression of circFOXP1 combined with in vitro assays demonstrated that circFOXP1 has pleiotropic effects, including promotion of cell proliferation, migration, invasion, and inhibition of cell apoptosis in GBC. In vivo, circFOXP1 promoted tumor growth. Mechanistically, double luciferase reporter, RNA immunoprecipitation (RIP) and biotin-labeled RNA pull-down assays clarified that circFOXP1 interacted with PTBP1 that could bind to the 3'UTR region and coding region (CDS) of enzyme pyruvate kinase, liver and RBC (PKLR) mRNA (UCUU binding bites) to protect PKLR mRNA from decay. Additionally, circFOXP1 acted as the sponge of miR-370 to regulate PKLR, resulting in promoting Warburg effect in GBC progression. CONCLUSIONS: These results demonstrated that circFOXP1 serve as a prognostic biomarker and critical regulator in GBC progression and Warburg effect, suggesting a potential target for GBC treatment.

Long non-coding RNA GBCDRlnc1 induces chemoresistance of gallbladder cancer cells by activating autophagy.

BACKGROUND: Gallbladder cancer is the most common biliary tract malignancy and not sensitive to chemotherapy. Autophagy is an important factor prolonging the survival of cancer cells under chemotherapeutic stress. We aimed to investigate the role of long non-coding RNAs (lncRNAs) in autophagy and chemoresistance of gallbladder cancer cells. METHODS: We established doxorubicin (Dox)-resistant gallbladder cancer cells and used microarray analysis to compare the expression profiles of lncRNAs in Dox-resistant gallbladder cancer cells and their parental cells. Knockdown or exogenous expression of lncRNA combined with in vitro and in vivo assays were performed to prove the functional significance of lncRNA. The effects of lncRNA on autophagy were assessed by stubRFP-sensGFP-LC3 and western blot. We used RNA pull-down and mass spectrometry analysis to identify the target proteins of lncRNA. RESULTS: The drug-resistant property of gallbladder cancer cells is related to their enhanced autophagic activity. And we found a lncRNA ENST00000425894 termed gallbladder cancer drug resistance-associated lncRNA1 (GBCDRlnc1) that serves as a critical regulator in gallbladder cancer chemoresistance. Furthermore, we discovered that GBCDRlnc1 is upregulated in gallbladder cancer tissues. Knockdown of GBCDRlnc1, via inhibiting autophagy at initial stage, enhanced the sensitivity of Dox-resistant gallbladder cancer cells to Dox in vitro and in vivo. Mechanically, we identified that GBCDRlnc1 interacts with phosphoglycerate kinase 1 and inhibits its ubiquitination in Dox-resistant gallbladder cancer cells, which leads to the down-regulation of autophagy initiator ATG5-ATG12 conjugate. CONCLUSIONS: Our findings established that the chemoresistant driver GBCDRlnc1 might be a candidate therapeutic target for the treatment of advanced gallbladder cancer.

Systemic inflammation score is a prognostic marker after curative resection in gastric cancer.

BACKGROUND: The systemic inflammation score (SIS), as calculated from preoperative serum albumin level and lymphocyte-to-monocyte ratio, has been demonstrated to be a prognostic marker in cancer. The present study intended to investigate the prognostic role of SIS in gastric cancer patients after curative gastrectomy in comparison with other prognostic markers. METHODS: Preoperative SIS was retrospectively calculated in patients who underwent curative gastrectomy between 2007 and 2011 in Fudan University Shanghai Cancer Center. The prognostic accuracy of each score was compared utilizing time-dependent receiver operating characteristics analysis. RESULTS: The higher SIS score was associated with older age, larger tumour size, a more advanced tumour-nodes-metastasis stage and lymph node status, deeper tumour invasion, the presence of lymphovascular invasion and a poorer overall survival and disease-free survival. In time-dependent receiver operating characteristics analysis, the SIS had a higher area under the curve for the prediction of 5-year overall survival than the neutrophil lymphocyte ratio. The SIS maintained the predictive accuracy superiority throughout the observation period. CONCLUSION: The SIS is a useful prognostic marker in gastric cancer patients after curative gastrectomy.

Prognostic Value of Inflammatory Biomarkers in Gastric Cancer Patients and the Construction of a Predictive Model.

BACKGROUND: It is known that inflammation promotes cancer development. However, a few studies have evaluated the prognostic significance of inflammatory biomarkers in gastric cancer (GC). METHODS: In this study, 2,334 patients who underwent gastrectomy for GC at Fudan University Shanghai Cancer Center between 2003 and 2007 were retrospectively analyzed, and 1,227 patients were found to be eligible. The preoperative serum neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio, lymphocyte-to-monocyte ratio (LMR), and albumin/globulin ratio (AGR) levels were analyzed. A nomogram was constructed with the Cox proportional hazards regression model in the training set (n = 818) to predict the probability of overall survival (OS) and disease-free survival (DFS). The predictive accuracy and discriminative ability were determined using the concordance index (C-index) and calibration curve. RESULTS: We found that lower AGR and LMR values were correlated with decreased OS, lower LMR values, and higher NLR values with a decreased DFS. Other significant factors were included to construct the nomogram. The discriminative ability of the nomogram was higher than that of the eighth American Joint Committee on Cancer tumor-node-metastasis (TNM) staging system (0.746 for TNM v.s. 0.654 for the nomogram, p < 0.001). CONCLUSIONS: The nomogram yielded a more accurate prognostic prediction in GC patients after gastrectomy, suggesting great clinical value.

lncRNA GCAWKR Promotes Gastric Cancer Development by Scaffolding the Chromatin Modification Factors WDR5 and KAT2A.

Long noncoding RNAs (lncRNAs) have been demonstrated to play a role in carcinogenesis, but their mechanisms of function remain elusive. We explored the mechanisms of the oncogenic role of GCAWKR in gastric cancer (GC) using human tissues and cell lines. The in situ hybridization analysis was utilized to determine GCAWKR levels in samples from 42 GC patients and real-time qPCR in tissues from 123 patients. The GCAWKR levels were modulated in GC cell lines, and relevant biological and molecular analyses were performed. Levels of the GCAWKR were upregulated in GC tissues compared with normal tissues and associated with tumor size, lymph node metastasis, TNM stage, and patient outcomes. GCAWKR affected cell proliferation and cell invasion in multiple GC models. Mechanistically, GCAWKR bound WDR5 and KAT2A and acted as a molecular scaffold of WDR5/KAT2A complexes, modulating the affinity for WDR5/KAT2A complexes in the target gene's promoter region. Thus, our data defined a mechanism of lncRNA-mediated carcinogenesis in GC, suggesting new therapeutic targets in GC.

Arctigenin induced gallbladder cancer senescence through modulating epidermal growth factor receptor pathway.

Gallbladder cancer has poor prognosis and limited therapeutic options. Arctigenin, a representative dibenzylbutyrolactone lignan, occurs in a variety of plants. However, the molecular mechanisms involved in the antitumor effect of arctigenin on gallbladder cancer have not been fully elucidated. The expression levels of epidermal growth factor receptor were examined in 100 matched pairs of gallbladder cancer tissues. A positive correlation between high epidermal growth factor receptor expression levels and poor prognosis was observed in gallbladder cancer tissues. Pharmacological inhibition or inhibition via RNA interference of epidermal growth factor receptor induced cellular senescence in gallbladder cancer cells. The antitumor effect of arctigenin on gallbladder cancer cells was primarily achieved by inducing cellular senescence. In gallbladder cancer cells treated with arctigenin, the expression level of epidermal growth factor receptor significantly decreased. The analysis of the activity of the kinases downstream of epidermal growth factor receptor revealed that the RAF-MEK-ERK signaling pathway was significantly inhibited. Furthermore, the cellular senescence induced by arctigenin could be reverted by pcDNA-epidermal growth factor receptor. Arctigenin also potently inhibited the growth of tumor xenografts, which was accompanied by the downregulation of epidermal growth factor receptor and induction of senescence. This study demonstrates arctigenin could induce cellular senescence in gallbladder cancer through the modulation of epidermal growth factor receptor pathway. These data identify epidermal growth factor receptor as a key regulator in arctigenin-induced gallbladder cancer senescence.

SNORA74B gene silencing inhibits gallbladder cancer cells by inducing PHLPP and suppressing Akt/mTOR signaling.

Small nucleolar RNAs (snoRNAs) have been implicated in the development of many cancers. We therefore examined the differential expression of snoRNAs between gallbladder cancer (GBC) tissues and matched adjacent non-tumor tissues using expression microarray analysis with confirmation by quantitative real-time PCR (qRT-PCR). Western blot analysis showed that SNORA74B levels were higher in GBC than non-tumor tissues. SNORA74B expression was positively associated with local invasion, advanced TNM stage, CA19-9 level, and Ki67 expression in patients with GBC, while it was negatively associated with expression of PHLPP, an endogenous Akt inhibitor. Moreover, SNORA74B expression was prognostic for overall survival (OS) and disease-free survival (DFS). Functional studies revealed that silencing SNORA74B in GBC cells using sh-SNORA74B suppressed cell proliferation, induced G1 arrest, and promoted apoptosis. Preliminary molecular investigation revealed that SNORA74B silencing inhibited activation of the AKT/mTOR signaling pathway, while increasing PHLPP expression. PHLPP depletion using shRNA abrogated sh-SNORA74B suppression of GBC cell proliferation, indicating that the antitumor effects of SNORA74B silencing were mediated by PHLPP. These findings define the important role of SNORA74B in cell proliferation, cell cycle, and apoptosis of GBC, and suggest that it may serve as a novel target for GBC treatment.

Long non-coding RNA LINC00152 promotes gallbladder cancer metastasis and epithelial-mesenchymal transition by regulating HIF-1α via miR-138.

Long non-coding RNA LINC00152 had been reported as an oncogene in gastric and hepatocellular cancer. In this study, we show that LINC00152 is overexpressed in gallbladder cancer (GBC) tissue samples and cell lines. The high LINC00152 levels correlated negatively with the overall survival time in GBC patients. Functionally, LINC00152 dramatically promoted cell migration, invasion and epithelial-mesenchymal transition (EMT) progression in vitro. In vivo, LINC00152 overexpression significantly promoted tumour peritoneal spreading and metastasis. Mechanistic analyses indicated that LINC00152 functions as a molecular sponge for miR-138, which directly suppresses the expression of hypoxia inducible factor-1α (HIF-1α). We revealed that miR-138 is a suppressor of GBC cell metastasis and EMT progression, and a similar phenomenon was observed in HIF-1α knockdown NOZ cells. Through binding to miR-138, LINC00152 has an oncogenic effect on GBC. Overall, our study suggested that the LINC00152/miR-138/HIF-1α pathway potentiates the progression of GBC, and LINC00152 may be a novel therapeutic target.

MiR-31 regulates the cisplatin resistance by targeting Src in gallbladder cancer.

BACKGROUND: Gallbladder cancer (GBC) is a malignant tumor highly resistant to chemotherapy. MicroRNAs (miRNAs) are found extensively involved in modulation of carcinogenesis and chemoresistance. This study aimed to investigate cisplatin (DDP)-susceptibility regulated by expression of the miRNAs and underlying pathways in GBC. RESULTS: The microRNA-31 (miR-31) was selected by microarray due to the biggest fold change between DDP-resistant and parental cells. Ectopic overexpression of miR-31 decreased cell proliferation, viability and invasion capacity, but promoted apoptosis in DDP-resistant cells and in xenograft tumor models. Cell apoptosis and DDP-chemosensitivity was remarkably increased by knockdown of Src proto-oncogene (Src) expression, which was subsequently reversed by rescue of Src expression in miR-31-expressing cells. METHODS: The microarray was used to select the candidate miRNA in two DDP-resistant GBC cell lines. The effect of regulated expression of the miRNA on cell migration, invasion, proliferation and apoptosis was examined by wound healing, transwell assays, CCK-8 assays, colony formation and flow cytometry assays, respectively. Xenograft tumor models were used to validate the function of the downstream target. CONCLUSION: Our results demonstrated that miR-31reduced significantly in GBC cells rendering resistance to cisplatin, and upregulated expression of miR-31 augmented chemosensitivity, presenting a therapeutic potential to overcome drug resistance in GBC.