Analysis of cuproptosis-related lncRNA signature for predicting prognosis and tumor immune microenvironment in pancreatic cancer.

Pancreatic cancer (PC) is a highly malignant digestive tract tumor, with a dismal 5-year survival rate. Recently, cuproptosis was found to be copper-dependent cell death. This work aims to establish a cuproptosis-related lncRNA signature which could predict the prognosis of PC patients and help clinical decision-making. Firstly, cuproptosis-related lncRNAs were identified in the TCGA-PAAD database. Next, a cuproptosis-related lncRNA signature based on five lncRNAs was established. Besides, the ICGC cohort and our samples from 30 PC patients served as external validation groups to verify the predictive power of the risk signature. Then, the expression of CASC8 was verified in PC samples, scRNA-seq dataset CRA001160, and PC cell lines. The correlation between CASC8 and cuproptosis-related genes was validated by Real-Time PCR. Additionally, the roles of CASC8 in PC progression and immune microenvironment characterization were explored by loss-of-function assay. As showed in the results, the prognosis of patients with higher risk scores was prominently worse than that with lower risk scores. Real-Time PCR and single cell analysis suggested that CASC8 was highly expressed in pancreatic cancer and related to cuproptosis. Additionally, gene inhibition of CASC8 impacted the proliferation, apoptosis and migration of PC cells. Furthermore, CASC8 was demonstrated to impact the expression of CD274 and several chemokines, and serve as a key indicator in tumor immune microenvironment characterization. In conclusion, the cuproptosis-related lncRNA signature could provide valuable indications for the prognosis of PC patients, and CASC8 was a candidate biomarker for not only predicting the progression of PC patients but also their antitumor immune responses.

ADAMTS12 promotes migration and epithelial-mesenchymal transition and predicts poor prognosis for pancreatic cancer.

BACKGROUND: ADAMTS (a disintegrin and metalloproteinase with thrombospondin-like motifs) family, a group of extracellular multifunctional enzymes, has been proven to play a pivotal role in the tumor. In pancreatic cancer, the role and mechanism of this family remain unclear. The present study aimed to figure out the hub gene of ADAMTSs and explore the exact roles in the prognosis and biological functions in pancreatic ductal adenocarcinoma (PDAC). METHODS: We used several databases to analyze the ADAMTS family and then screen out the hub genes. The expression of ADAMTS12 in 106 pairs of PDAC tumors and adjacent normal tissues was examined by immunohistochemistry, and its correlations with clinical parameters were further analyzed. The impacts of ADAMTS12 on the migration of PDAC cells were predicted by gene set enrichment analysis and confirmed by transwell assays. The potential impacts of ADAMTS12 on the epithelial-mesenchymal transition (EMT) were identified by database analysis and experimental proof of real-time quantitative polymerase chain reaction (qPCR) and Western blotting. RESULTS: Our study found that ADAMTS12 was a crucial gene in PDAC, and it was highly expressed in tumor tissues when compared to that in the adjacent tissues. ADATMS12 had predictive value of a poor prognosis for PDAC. The elevation of ADAMTS12 was parallel to the progression of PDAC. Inhibition of ADAMTS12 suppressed the migration of PDAC cells and interfered with the process of EMT. CONCLUSIONS: ADAMTS12 is a crucial member of ADAMTSs in PDAC and a predictor of poor prognosis. Additionally, based on its impacts on migration and metastasis in PDAC and the relationship with EMT, ADAMTS12 plays a role of an oncogene in PDAC and may be a promising target for treatment.

Increased Serotonin Signaling Contributes to the Warburg Effect in Pancreatic Tumor Cells Under Metabolic Stress and Promotes Growth of Pancreatic Tumors in Mice.

BACKGROUND & AIMS: Desmoplasia and poor vascularity cause severe metabolic stress in pancreatic ductal adenocarcinomas (PDACs). Serotonin (5-HT) is a neuromodulator with neurotransmitter and neuroendocrine functions that contributes to tumorigenesis. We investigated the role of 5-HT signaling in the growth of pancreatic tumors. METHODS: We measured the levels of proteins that regulate 5-HT synthesis, packaging, and degradation in pancreata from KrasG12D/+/Trp53R172H/+/Pdx1-Cre (KPC) mice, which develop pancreatic tumors, as well as in PDAC cell lines and a tissue microarray containing 81 human PDAC samples. We also analyzed expression levels of proteins involved in 5-HT synthesis and degradation by immunohistochemical analysis of a tissue microarray containing 311 PDAC specimens, and associated expression levels with patient survival times. 5-HT level in 14 matched PDAC tumor and non-tumor tissues were analyzed by ELISA. PDAC cell lines were incubated with 5-HT and cell survival and apoptosis were measured. We analyzed expression of the 5-HT receptor HTR2B in PDAC cells and effects of receptor agonists and antagonists, as well as HTR2B knockdown with small hairpin RNAs. We determined the effects of 5-HT stimulation on gene expression profiles of BxPC-3 cells. Regulation of glycolysis by 5-HT signaling via HTR2B was assessed by immunofluorescence and immunoprecipitation analyses, as well as by determination of the extracellular acid ratio, glucose consumption, and lactate production. Primary PDACs, with or without exposure to SB204741 (a selective antagonist of HTR2B), were grown as xenograft tumors in mice, and SB204741 was administered to tumor-bearing KPC mice; tumor growth and metabolism were measured by imaging analyses. RESULTS: In immunohistochemical analysis of a tissue microarray of PDAC specimens, increased levels of TPH1 and decreased level of MAOA, which regulate 5-HT synthesis and degradation, correlated with stage and size of PDACs and shorter patient survival time. We found levels of 5-HT to be increased in human PDAC tissues compared with non-tumor pancreatic tissues, and PDAC cell lines compared with non-transformed pancreatic cells. Incubation of PDAC cell lines with 5-HT increased proliferation and prevented apoptosis. Agonists of HTR2B, but not other 5-HT receptors, promoted proliferation and prevented apoptosis of PDAC cells. Knockdown of HTR2B in PDAC cells, or incubation of cells with HTR2B inhibitors, reduced their growth as xenograft tumors in mice. We observed a correlation between 5-HT and glycolytic flux in PDAC cells; levels of metabolic enzymes involved in glycolysis, the phosphate pentose pathway, and hexosamine biosynthesis pathway increased significantly in PDAC cells following 5-HT stimulation. 5-HT stimulation led to formation of the HTR2B-LYN-p85 complex, which increased PI3K-Akt-mTOR signaling and the Warburg effect by increasing protein levels of MYC and HIF1A. Administration of SB204741 to KPC mice slowed growth and metabolism of established pancreatic tumors and prolonged survival of the mice. CONCLUSIONS: Human PDACs have increased levels of 5-HT, and PDAC cells increase expression of its receptor, HTR2B. These increases allow for tumor glycolysis under metabolic stress and promote growth of pancreatic tumors and PDAC xenograft tumors in mice.

Signet-ring cell carcinoma coexisting with adenocarcinoma arising in a choledochal cyst: report of a case.

Signet-ring cell carcinoma (SRCC) is rare in the biliary system. We report a case of SRCC coexisting with adenocarcinoma, arising in a choledochal cyst of the extrahepatic bile duct. The patient was a 52-year-old man, hospitalized for the investigation of jaundice and pruritus. Abdominal computed tomography and magnetic resonance cholangiopancreatography showed a huge choledochal cyst and distal common bile duct cancer. The patient underwent a pancreaticoduodenectomy with extended lymph node dissection. Histologic examination confirmed an SRCC coexisting with adenocarcinoma arising in a choledochal cyst. Postoperative chemotherapy had to be discontinued after only two cycles because the patient suffered serious side effects. Recurrence was detected in the bilioenteric anastomosis 4 months after surgery, and he died 6 months after surgery. To our knowledge, this represents the first case of SRCC arising in a choledochal cyst of the extrahepatic bile duct ever to be reported.

Effect of CD74 on the prognosis of patients with resectable pancreatic cancer.

BACKGROUND: CD74 is known as a type II transmembrane glycoprotein that is associated with the major histocompatibility complex class II alpha and beta chains. Recent studies have demonstrated that the expression of CD74 is also linked to some forms of tumors. The present study was to assess the effect of CD74 expression on the prognosis of resectable pancreatic ductal adenocarcinoma (PDAC). METHODS: Forty-six patients who had received a curative resection of primary PDAC and postoperative chemotherapy were included in this study. Immunohistochemical staining was conducted of CD74 on paraffin-embedded tumor sample slices. The patients were grouped according to CD74 staining: CD74 (-): CD74 positive tumor cells<25%; and CD74 (+): CD74 positive tumor cells ≥25%. The correlation of CD74 expression level with clinicopathological features and cumulative survival rate was calculated. RESULTS: The numbers of CD74 (+) and (-) patients were 32 and 14, respectively. CD74 (+) patients showed a high rate of perineural invasion (P=0.007). The 3- and 5-year cumulative survival rates of CD74 (-) patients were significantly higher than those of CD74 (+) patients (62% and 41% vs 9% and 0%, P=0.000). Multivariate analysis showed that CD74 expression and lymphatic permeation were the independent prognostic indicators. CONCLUSIONS: The overexpression of CD74 is a key factor associated with perineural invasion. Lower-stage (I and II) PDAC patients with CD74 overexpression have a poor prognosis even if they receive a curative resection. CD74 can be used as a prognostic indicator for resectable PDAC.

Prostacyclin decreases splanchnic vascular contractility in cirrhotic rats.

BACKGROUND: Prostacyclin has been shown to increase portal hypertension, but the mechanism is unclear. This study aimed to investigate whether the overproduction of prostacyclin (PGI2) in cirrhosis participates in the splanchnic vascular hyporesponsiveness to vasoconstrictors in cirrhotic rats. METHODS: Cirrhotic model was created by subcutaneous injection of 60% carbon tetrachloride (CCl4) corn oil solution combined with intermittent drinking of 5% alcohol, and age-matched rats served as controls. The isolated third-generation mesenteric arterioles were used to examine the contractile response to norepinephrine. The changes in vascular diameter were observed under a microscope imaging device. The plasma concentration of 6-ketone-prostaglandin F1alpha (6-keto-PGF1alpha, a stable metabolite of PGI2) was tested via enzyme immunoassays and the expression of cyclooxygenase (COX) in mesenteric arteries was detected by Western blotting. RESULTS: In parallel with the increase of plasma 6-keto-PGF1alpha, the contractile response of arterioles from cirrhotic rats to norepinephrine was significantly impaired compared with that from controls. Inhibition of PGI2 or protein kinase A with indomethacin or Rp-adenosine 3', 5'-cyclic monophosphothioate (Rp-cAMPS) partially reversed the vascular hypo-contractile response to norepinephrine in arterioles from cirrhotic rats. Indomethacin significantly decreased the plasma 6-keto-PGF1alpha. Furthermore, indomethacin significantly attenuated the effect of Rp-cAMPS on arterioles from cirrhotic rats. COX-1 expression was up-regulated in mesenteric arteries from cirrhotic rats, whereas COX-2 was not detectable in the mesenteric arteries from both cirrhotic and control rats. CONCLUSION: Enhanced COX-1 expression in cirrhotic rats resulted in elevated PGI2 production which partially contributed to the splanchnic vascular hyporesponsiveness to a vasoconstrictor via the protein kinase A pathway.

A Nutrient-Deficient Microenvironment Facilitates Ferroptosis Resistance via the FAM60A-PPAR Axis in Pancreatic Ductal Adenocarcinoma.

Ferroptosis, a nonapoptotic form of cell death, is an emerging potential therapeutic target for various diseases, including cancer. However, the role of ferroptosis in pancreatic cancer remains poorly understood. Pancreatic ductal adenocarcinoma (PDAC) is characterized by a poor prognosis and chemotherapy resistance, attributed to its high Kirsten rats arcomaviral oncogene homolog mutation rate and severe nutritional deficits resulting from a dense stroma. Several studies have linked rat sarcoma (RAS) mutations to ferroptosis, suggesting that inducing ferroptosis may be an effective strategy against oncogenic RAS-bearing tumors. We investigated the role of Family With Sequence Similarity 60 Member A (FAM60A) in this study, a protein closely associated with a poor prognosis and highly expressed in PDAC and tumor tissue from KrasG12D/+;Trp53R172H/+; Pdx1-Cre mice, in regulating ferroptosis, tumor growth, and gemcitabine sensitivity in vitro and in vivo. Our results demonstrate that FAM60A regulates 3 essential metabolic enzymes, ACSL1/4 and GPX4, to protect PDAC cells from ferroptosis. Furthermore, we found that YY1 transcriptionally regulates FAM60A expression by promoting its transcription, and the Hippo-YY1 pathway is restricted in the low-amino-acid milieu in the context of nutrient deprivation, leading to downstream suppression of peroxisome proliferator-activated receptor and ACSL1/4 and activation of GPX4 pathways. Importantly, FAM60A knockdown sensitized PDAC cells to gemcitabine treatment. A new understanding of FAM60A transcriptional regulation pattern in PDAC and its dual function in ferroptosis reliever and chemotherapy resistance is provided by our study. Targeting FAM60A may therefore offer a promising therapeutic approach for PDAC by simultaneously addressing 2 major features of the disease (high RAS mutation rate and tumor microenvironment nutrient deficiency) and preventing tumor cell metabolic adaptation.

Prediction of immune infiltration and prognosis for patients with cholangiocarcinoma based on a cuproptosis-related lncRNA signature.

Objective: Cholangiocarcinoma (CHOL) is a malignant disease that affects the digestive tract, and it is characterized by a poor prognosis. This research sought to explore the involvement of cuproptosis-related lncRNAs (CRLs) in the prognostic prediction and immune infiltration of cholangiocarcinoma. Methods: The expression profiles and clinical data of CHOL patients were acquired from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases, and CRLs were defined via co-expression analysis. Two molecular clusters distinguished by cuproptosis-related genes (CRGs) were produced. Then a risk signature consisted by four CRLs was formed, and all samples were separated into low- and high-risk groups using a risk score. Kaplan-Meier survival analysis, principal component analysis, differentially expressed analysis, immune cell infiltration analysis, and sensitivities analysis of chemotherapy drugs were conducted between the two groups. Simultaneously, the expression values of four lncRNAs confirmed by real-time PCR in our own 20 CHOL samples were brought into the risk model. Results: The CHOL samples could be differentiated into two molecular clusters, which displayed contrasting survival times. Additionally, patients with higher risk scores had significantly worse prognosis compared to those in the low-risk group. Furthermore, both immune infiltration and enrichment analysis revealed significant discrepancies in the tumor immune microenvironment (TIME) between different risk groups. Moreover, the predictive power and the correlation with CA19-9 and CEA of risk signature were validated in our own samples. Conclusion: We developed a risk signature which could serve as an independent prognostic factor and offer a promising prediction for not only prognosis but also TIME in CHOL patients.

OCIAD2 promotes pancreatic cancer progression through the AKT signaling pathway.

BACKGROUND: OCIAD2（Ovarian carcinoma immunoreactive antigen-like protein 2） is a protein reported in various cancers. However, the role of OCIAD2 has not been explored in pan-cancer datasets. The purpose of this research lies in analyzing the expression level and prognostic-related value of OCIAD2 in different human cancers, as well as revealing the underlying mechanism in specific cancer type (pancreatic adenocarcinoma, PAAD). METHODS: The correlation between OCIAD2 expression level and clinical relevance in different human cancers was investigated from bioinformatical perspective (GTEx and TCGA). The OCIAD2 expression level and clinical significance in PAAD were explored in GEO datasets and tissue microarray. Functional experiments were used to determine the OCIAD2 cell functions in vitro and in vivo. GSEA, western blot and immunohistochemistry were used to uncover the potential mechanism. RESULTS: OCIAD2 expression level was closely correlated with clinical relevance in many cancer types through pan-cancer analysis, and we found OCIAD2 was highly expressed in PAAD and associated with poorer prognosis. OCIAD2 acted as the promotor of Warburg effect and influenced PAAD cells proliferation, migration and apoptosis. Mechanistically, OCIAD2 upregulation may boost glycolysis in PAAD via activating the AKT signaling pathway in PAAD. CONCLUSIONS: In PAAD, OCIAD2 promotes Warburg effect via AKT signaling pathway and targeting cancer cells metabolic reprogramming could be a potential treatment.

Desensitization of G-protein-coupled receptors induces vascular hypocontractility in response to norepinephrine in the mesenteric arteries of cirrhotic patients and rats.

BACKGROUND: The increased beta-arrestin-2 and its combination with G-protein-coupled receptors (GPCRs) lead to GPCRs desensitization. The latter may be responsible for decreased contractile reactivity in the mesenteric arteries of cirrhotic patients and rats. The present study is to investigate the machinery changes of alpha-adrenergic receptors and G proteins and their roles in the contractility of mesenteric arteries of cirrhotic patients and animal models. METHODS: Patients with cirrhosis due to hepatitis B and cirrhotic rats induced by CCl4 were studied. Mesenteric artery contractility in response to norepinephrine was determined by a vessel perfusion system. The contractile effect of G protein-coupled receptor kinase-2 (GRK-2) inhibitor on the mesenteric artery was evaluated. The protein expression of the alpha1 adrenergic receptor, G proteins, beta-arrestin-2, GRK-2 as well as the activity of Rho associated coiled-coil forming protein kinase-1 (ROCK-1) were measured by Western blot. In addition, the interaction of alpha1 adrenergic receptor with beta-arrestin-2 was assessed by co-immunoprecipitation. RESULTS: The portal vein pressure of cirrhotic patients and rats was significantly higher than that of controls. The dose-response curve to norepinephrine in mesenteric arteriole was shifted to the right, and EC50 was significantly increased in cirrhotic patients and rats. There were no significant differences in the expressions of the alpha1 adrenergic receptor and G proteins in the cirrhotic group compared with the controls. However, the protein expressions of GRK-2 and beta-arrestin-2 were significantly elevated in cirrhotic patients and rats compared with those of the controls. The interaction of the alpha1 adrenergic receptor and beta-arrestin-2 was significantly aggravated. This interaction was significantly reversed by GRK-2 inhibitor. Both the protein expression and activity of ROCK-1 were significantly decreased in the mesenteric artery in patients with cirrhosis compared with those of the controls, and this phenomenon was not shown in the cirrhotic rats. Norepinephrine significantly increased the activity of ROCK-1 in normal rats but not in cirrhotic ones. Norepinephrine significantly increased ROCK-1 activity in cirrhotic rats when GRK-2 inhibitor was used. CONCLUSIONS: beta-arrestin-2 expression and its interaction with GPCRs are significantly upregulated in the mesenteric arteries in patients and rats with cirrhosis. These upregulations result in GPCR desensitization, G-protein dysfunction and ROCK inhibition. These may explain the decreased contractility of the mesenteric artery in response to vasoconstrictors.

Immunosuppression, immune escape, and immunotherapy in pancreatic cancer: focused on the tumor microenvironment.

Pancreatic ductal adenocarcinoma (PDAC), the most common type of pancreatic cancer, is characterized by poor treatment response and low survival time. The current clinical treatment for advanced PDAC is still not effective. In recent years, the research and application of immunotherapy have developed rapidly and achieved substantial results in many malignant tumors. However, the translational application in PDAC is still far from satisfactory and needs to be developed urgently. To carry out the study of immunotherapy, it is necessary to fully decipher the immune characteristics of PDAC. This review summarizes the recent progress of the tumor microenvironment (TME) of PDAC and highlights its link with immunotherapy. We describe the molecular cues and corresponding intervention methods, collate several promising targets and progress worthy of further study, and put forward the importance of integrated immunotherapy to provide ideas for future research of TME and immunotherapy of PDAC.

IRAK2-NF-κB signaling promotes glycolysis-dependent tumor growth in pancreatic cancer.

BACKGROUND: Metabolic reprogramming has emerged as a core hallmark of cancer, and cancer metabolism has long been equated with aerobic glycolysis. Moreover, hypoxia and the hypovascular tumor microenvironment (TME) are major hallmarks of pancreatic ductal adenocarcinoma (PDAC), in which glycolysis is imperative for tumor cell survival and proliferation. Here, we explored the impact of interleukin 1 receptor-associated kinase 2 (IRAK2) on the biological behavior of PDAC and investigated the underlying mechanism. METHODS: The expression pattern and clinical relevance of IRAK2 was determined in GEO, TCGA and Ren Ji datasets. Loss-of-function and gain-of-function studies were employed to investigate the cellular functions of IRAK2 in vitro and in vivo. Gene set enrichment analysis, Seahorse metabolic analysis, immunohistochemistry and Western blot were applied to reveal the underlying molecular mechanisms. RESULTS: We found that IRAK2 is highly expressed in PDAC patient samples and is related to a poor prognosis. IRAK2 knockdown led to a significant impairment of PDAC cell proliferation via an aberrant Warburg effect. Opposite results were obtained after exogenous IRAK2 overexpression. Mechanistically, we found that IRAK2 is critical for sustaining the activation of transcription factors such as those of the nuclear factor-κB (NF-κB) family, which have increasingly been recognized as crucial players in many steps of cancer initiation and progression. Treatment with maslinic acid (MA), a NF-κB inhibitor, markedly attenuated the aberrant oncological behavior of PDAC cells caused by IRAK2 overexpression. CONCLUSIONS: Our data reveal a role of IRAK2 in PDAC metabolic reprogramming. In addition, we obtained novel insights into how immune-related pathways affect PDAC progression and suggest that targeting IRAK2 may serve as a novel therapeutic approach for PDAC.

CD74 promotes perineural invasion of cancer cells and mediates neuroplasticity via the AKT/EGR-1/GDNF axis in pancreatic ductal adenocarcinoma.

Perineural invasion (PNI) is a common feature of pancreatic ductal adenocarcinoma (PDAC) and is one of the important causes of local recurrence in resected pancreatic cancer, but the molecular mechanism remains largely unexplored. Here, we used immunohistochemistry staining to determine the expression of CD74. Then the in vivo PNI model, in vitro neuroplasticity assay, cell proliferation assay, wound healing and Transwell-based invasion assay were performed to examine the function of CD74 in pancreatic cancer cell lines. ChIP assay and Luciferase reporter assay were used to illustrate the mechanism underlying CD74 induced GDNF expression. We confirmed that the expression level of CD74 was an independent predictor of PNI and poor prognosis for PDAC. Moreover, we found that upregulation of CD74 on PDAC enhanced its migration and invasive capabilities and potentiated the secretion of neurotrophic factor GDNF to promote the neuroplasticity. Mechanistically, CD74 promoted GDNF production via the AKT/EGR-1/GDNF axis in PDAC. Taken together, our findings suggest a supportive role of CD74 in the PNI of PDAC, and deepen our understanding of how cancer cells promote neuroplasticity in the microenvironment of PDAC.

SF3B1 mutation in pancreatic cancer contributes to aerobic glycolysis and tumor growth through a PP2A-c-Myc axis.

Hot spot gene mutations in splicing factor 3b subunit 1 (SF3B1) are observed in many types of cancer and create abundant aberrant mRNA splicing, which is profoundly implicated in tumorigenesis. Here, we identified that the SF3B1 K700E (SF3B1K700E ) mutation is strongly associated with tumor growth in pancreatic ductal adenocarcinoma (PDAC). Knockdown of SF3B1 significantly retarded cell proliferation and tumor growth in a cell line (Panc05.04) with the SF3B1K700E mutation. However, SF3B1 knockdown had no notable effect on cell proliferation in two cell lines (BxPC3 and AsPC1) carrying wild-type SF3B1. Ectopic expression of SF3B1K700E but not SF3B1WT in SF3B1-knockout Panc05.04 cells largely restored the inhibitory role induced by SF3B1 knockdown. Introduction of the SF3B1K700E mutation in BxPC3 and AsPC1 cells also boosted cell proliferation. Gene set enrichment analysis demonstrated a close correlation between SF3B1 mutation and aerobic glycolysis. Functional analyses showed that the SF3B1K700E mutation promoted tumor glycolysis, as evidenced by glucose consumption, lactate release, and extracellular acidification rate. Mechanistically, the SF3B1 mutation promoted the aberrant splicing of PPP2R5A and led to the activation of the glycolytic regulator c-Myc via post-translational regulation. Pharmacological activation of PP2A with FTY-720 markedly compromised the growth advantage induced by the SF3B1K700E mutation in vitro and in vivo. Taken together, our data suggest a novel function for SF3B1 mutation in the Warburg effect, and this finding may offer a potential therapeutic strategy against PDAC with the SF3B1K700E mutation.

Single-cell RNA sequencing reveals that targeting HSP90 suppresses PDAC progression by restraining mitochondrial bioenergetics.

Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers, which lacks effective treatment strategies. There is an urgent need for the development of new strategies for PDAC therapy. The genetic and phenotypic heterogeneity of PDAC cancer cell populations poses further challenges in the clinical management of PDAC. In this study, we performed single-cell RNA sequencing to characterize PDAC tumors from KPC mice. Functional studies and clinical analysis showed that PDAC cluster 2 cells with highly Hsp90 expression is much more aggressive than the other clusters. Genetic and pharmacologic inhibition of Hsp90 impaired tumor cell growth both in vitro and in vivo. Further mechanistic study revealed that HSP90 inhibition disrupted the interaction between HSP90 and OPA1, leading to a reduction in mitochondrial cristae amount and mitochondrial energy production. Collectively, our study reveals that HSP90 might be a potential therapeutic target for PDAC.

Evaluation of surgical procedure selection based on intraoperative free portal pressure measurement in patients with portal hypertension.

BACKGROUND: Various surgical procedures can be used to treat liver cirrhosis and portal hypertension. How to select the most appropriate procedure for patients with portal hypertension has become a difficult problem. This study aimed to analyze the relationship between the value of intraoperative free portal pressure (FPP) and postoperative complications, and to explore the significance of intraoperative FPP measurement with respect to surgical procedure selection. METHODS: The clinical data of 187 patients with portal hypertension who received pericardial devascularization and proximal splenorenal shunt combined with devascularization (combined operation) at the Department of General Surgery in our hospital from January 2001 to September 2008 were retrospectively analyzed. Among the patients who received pericardial devascularization, those with a postoperative FPP >or=22 mmHg were included in a high-pressure group (n=68), and those with FPP <22 mmHg were in a low-pressure group (n=49). Seventy patients who received the combined operation comprised a combined group. The intraoperative FPP measurement changes at different times, and the incidence of postoperative complications in the three groups of patients were compared. RESULTS: The postoperative FPP value in the high-pressure group was 27.5+/-2.3 mmHg, which was significantly higher than that of the low-pressure (20.9+/-1.8 mmHg) or combined groups (21.7+/-2.5 mmHg). The rebleeding rate in the high-pressure group was significantly higher than that in the low-pressure and combined groups. The incidence rates of postoperative hepatic encephalopathy and liver failure were not statistically different among the three groups. The mortality due to rebleeding in the low-pressure and combined groups (0.84%) was significantly lower than that of the high-pressure group. CONCLUSIONS: The study demonstrates that FPP is a critical measurement for surgical procedure selection in patients with portal hypertension. A FPP value >or=22 mmHg after splenectomy and devascularization alone is an important indicator that an additional proximal splenorenal shunt needs to be performed.

Perineural Invasion Reprograms the Immune Microenvironment through Cholinergic Signaling in Pancreatic Ductal Adenocarcinoma.

Perineural invasion is a common feature of pancreatic ductal adenocarcinoma (PDAC). Here, we investigated the effect of perineural invasion on the microenvironment and how this affects PDAC progression. Transcriptome expression profiles of PDAC tissues with different perineural invasion status were compared, and the intratumoral T-cell density and levels of neurotransmitters in these tissues were assessed. Perineural invasion was associated with impaired immune responses characterized by decreased CD8+ T and Th1 cells, and increased Th2 cells. Acetylcholine levels were elevated in severe perineural invasion. Acetylcholine impaired the ability of PDAC cells to recruit CD8+ T cells via HDAC1-mediated suppression of CCL5. Moreover, acetylcholine directly inhibited IFNγ production by CD8+ T cells in a dose-dependent manner and favored Th2 over Th1 differentiation. Furthermore, hyperactivation of cholinergic signaling enhanced tumor growth by suppressing the intratumoral T-cell response in an orthotopic PDAC model. Conversely, blocking perineural invasion with bilateral subdiaphragmatic vagotomy in tumor-bearing mice was associated with an increase in CD8+ T cells, an elevated Th1/Th2 ratio, and improved survival. In conclusion, perineural invasion-triggered cholinergic signaling favors tumor growth by promoting an immune-suppressive microenvironment characterized by impaired CD8+ T-cell infiltration and a reduced Th1/Th2 ratio. SIGNIFICANCE: These findings provide a promising therapeutic strategy to modulate the immunosuppressive microenvironment of pancreatic ductal adenocarcinoma with severe perineural invasion.

SFRP4 is a prognostic marker and correlated with Treg cell infiltration in pancreatic ductal adenocarcinoma.

Secreted Frizzled-Related Protein 4 (SFRP4), a member of secreted frizzled-related protein family, has been found as a vital modulator in cell proliferation, cell self-renew and apoptosis through Wnt signaling transduction pathway. In the present study, we re-analyzed the expression pattern of SFRPs in Gene Expression Omnibus (GEO) datasets and evaluated the expression of SFRP4 at protein level in both KrasG12D/+; Trp53R172H/+; Pdx1-Cre; (KPC) mice and human pancreatic ductal adenocarcinoma (PDAC) tissue. We found that the expression of SFRP4 increased gradually in PanINs and PDAC lesions in KPC mice and high expression of SFRP4 was much more common in tumor lesions compared to the adjacent non-tumor tissues. Then we performed Kaplan-Meier survival and Cox regression analysis and found that high expression of SFRP4 in the serum and tumor lesions predicted poor prognosis for pancreatic cancer patients. Furthermore, we demonstrated that SFRP4 positively correlated with FOXP3+ Treg cells infiltration while the down-regulation of SFRP4 in tumor cells impaired the production of cytokines and the recruitments of T cells. This study suggested that SFRP4 can be a novel prognostic biomarker and potential therapeutic target for pancreatic cancer.

Targeting Purinergic Receptor P2Y2 Prevents the Growth of Pancreatic Ductal Adenocarcinoma by Inhibiting Cancer Cell Glycolysis.

PURPOSE: Extensive research has reported that the tumor microenvironment components play crucial roles in tumor progression. Thus, blocking the supports of tumor microenvironment is a promising approach to prevent cancer progression. We aimed to determine whether blocking extracellular ATP-P2RY2 axis could be a potential therapeutic approach for PDAC treatment. EXPERIMENTAL DESIGN: Expression of P2RY2 was determined in 264 human PDAC samples and correlated to patient survival. P2RY2 was inhibited in human PDAC cell lines by antagonist and shRNA, respectively, and cell viability, clonogenicity, and glycolysis were determined. RNA sequencing of PDAC cell line was applied to reveal underlying molecular mechanisms. Multiple PDAC mouse models were used to assess the effects of the P2RY2 inhibition on PDAC progression. RESULTS: P2RY2 was upregulated and associated with poor prognosis in PDAC. Activated P2RY2 by increased extracellular ATP in tumor microenvironment promoted PDAC growth and glycolysis. Further studies showed that the agonist-activated P2RY2 triggered PI3K/AKT-mTOR signaling by crosstalk with PDGFR mediated by Yes1, resulting in elevated expression of c-Myc and HIF1α, which subsequently enhanced cancer cell glycolysis. Genetic and pharmacologic inhibition of P2RY2 impaired tumor cell growth in subcutaneous and orthotopic xenograft model, as well as delayed tumor progression in inflammation-driven PDAC model. In addition, synergy was observed when AR-C118925XX, the selective antagonist of P2RY2 receptor, and gemcitabine were combined, resulting in prolonged survival of xenografted PDAC mice. CONCLUSIONS: These findings reveal the roles of the P2RY2 in PDAC metabolic reprogramming, suggesting that P2RY2 might be a potential metabolic therapeutic target for PDAC.

Clinical significance of programmed death 1/programmed death ligand 1 pathway in gastric neuroendocrine carcinomas.

BACKGROUND: Recently, more and more studies have demonstrated the pivotal role of programmed death 1/programmed death ligand 1 (PD-1/PD-L1) pathway in the immune evasion of tumors from the host immune system. However, the role of PD-1/PD-L1 pathway in gastric neuroendocrine carcinomas (G-NECs) remains unknown. AIM: To investigate the expression of PD-1/PD-L1 and role of PD-1/PD-L1 pathway in G-NECs, which occur rarely but are highly malignant and clinically defiant. METHODS: We investigated the expression of PD-L1 on tumor cells and PD-1+, CD8+, and FOXP3+ T cell infiltration by immunohistochemistry in 43 resected G-NEC tissue specimens. The copy number alterations of PD-L1 were assessed by qRT-PCR. RESULTS: Most of the G-NECs tumor cells exhibited a near-uniform expression pattern of PD-L1, while some showed a tumor-stromal interface enhanced pattern. Of the 43 G-NECs, 21 (48.8%) were classified as a high PD-L1 expression group, and the high expression of PD-L1 was associated with poor overall survival (OS). The high expression of PD-L1 was correlated with abundant PD-1+ tumor infiltrating lymphocytes (TILs) instead of CD8+ TILs and FOXP3+ regulatory T cells (Tregs). Our analysis also suggested that the infiltration of CD8+ TILs tended to be a favorable factor for OS, although the difference did not reach the statistical significance (P = 0.065). Meanwhile, PD-L1 was significantly overexpressed in cases with copy number gain as compared with those without. CONCLUSION: Our data demonstrated for the first time that high expression of PD-L1 in G-NECs is associated with a poor prognosis, while the high expression may be due to the copy number variation of PD-L1 gene or stimulation of TILs. These results provide a basis for the immunotherapy targeting PD-1/PD-L1 pathway in G-NECs.