Glycolysis and tumor progression promoted by the m6A writer VIRMA via m6A-dependent upregulation of STRA6 in pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive and lethal malignancies, highlighting the urgent need to elucidate the underlying oncogenic mechanisms. VIRMA is a classic isoform of methyltransferases that participates in epigenetic transcriptomic modification in eukaryotic mRNAs. However, the exact roles of VIRMA in PDAC remain unclear. Here, we identified that VIRMA is highly expressed in PDAC, and histone modifications of the promoter may partly account for this dysregulation. Moreover, VIRMA is closely related to glycolysis and poor prognosis in PDAC. We further determined that STRA6 is a direct downstream target of VIRMA in PDAC by RNA sequencing (RNA-seq) and m6A sequencing (m6A-seq). VIRMA is involved in gene expression regulation via 3' UTR targeting of STRA6 mRNA. Furthermore, the m6A reader IGF2BP2 was shown to critically contribute to the stability of STRA6 mRNA. We describe the role of VIRMA in promoting signaling via the STRA6/STAT3 axis, which results in increased levels of HIF-1α, a key activator of glycolysis. In vivo and in vitro experiments reveal that the VIRMA-STRA6-STAT3-HIF-1α axis plays an instrumental role in glycolysis and tumor progression in PDAC. In conclusion, we demonstrate that VIRMA can increase glycolysis in PDAC by upregulating STRA6, a cell surface membrane protein that stimulates the STAT3 pathway, thereby activating HIF-1α and leading to pancreatic cancer malignancy. Overall, our data strongly suggest that the VIRMA-STRA6-STAT3-HIF-1α axis is a viable therapeutic target in PDAC.

Tumor-associated macrophages drive glycolysis through the IL-8/STAT3/GLUT3 signaling pathway in pancreatic cancer progression.

Glycolytic metabolism is a hallmark of pancreatic ductal adenocarcinoma (PDAC), and tumor-associated stromal cells play important roles in tumor metabolism. We previously reported that tumor-associated macrophages (TAMs) facilitate PDAC progression. However, little is known about whether TAMs are involved in regulating glycolysis in PDAC. Here, we found a positive correlation between CD68+ TAM infiltration and FDG maximal standardized uptake (FDG SUVmax) on PET-CT images of PDAC. We discovered that the glycolytic gene set was prominently enriched in the high TAM infiltration group through Gene Set Enrichment Analysis using The Cancer Genome Atlas database. Mechanistically, TAMs secreted IL-8 to promote GLUT3 expression in PDAC cells, enhancing tumor glycolysis both in vitro and in vivo, whereas this effect could be blocked by the IL-8 receptor inhibitor reparixin. Furthermore, IL-8 promoted the translocation of phosphorylated STAT3 into the nucleus to activate the GLUT3 promoter. Overall, we demonstrated that TAMs boosted PDAC cell glycolysis through the IL-8/STAT3/GLUT3 signaling pathway. Our cumulative findings suggest that the abrogation of TAM-induced tumor glycolysis by reparixin might exhibit an antitumor impact and offer a potential therapeutic target for PDAC.

Cancer-specific survival in patients with cholangiocarcinoma after radical surgery: a Novel, dynamic nomogram based on clinicopathological features and serum markers.

BACKGROUND: This study aims to (1) identify preoperative testing-based characteristics associated with enhanced prognosis and survival for cholangiocarcinoma patients, and (2)create a distinctive nomogram to anticipate each patient's cancer-specific survival (CSS). METHODS: Retrospective analysis was performed on 197 CCA patients who underwent radical surgery at Sun Yat-sen Memorial Hospital; they were divided into a 131-person "training cohort" and a 66-person "internal validation cohort." The prognostic nomogram was created following a preliminary Cox proportional hazard regression search for independent factors influencing the patients' CSS. Its applicable domain was examined via an external validation cohort, which included 235 patients from the Sun Yat-sen University Cancer Center. RESULTS: The median follow-up period for the 131 patients in the training group was 49.3 months (range, 9.3 to 133.9 months). One-, three-, and five-year CSS rates were 68.7%, 24.5%, and 9.2%, respectively, with the median CSS length being 27.4 months (range: 1.4 to 125.2 months). PLT, CEA, AFP, tumor location, differentiation, lymph node metastasis, chemotherapy, and TNM stage were determined to be independent risk factors for CCA patients by univariate and multivariate Cox proportional hazard regression analysis. We were able to accurately predict postoperative CSS after incorporating all of these characteristics into a nomogram. The AJCC's 8th edition staging method's C-indices were statistically substantially (P < 0.001) lower than the nomogram's C-indices (0.84, 0.77, and 0.74 in the training, internal and external validation cohorts respectively). CONCLUSIONS: A realistic and useful model for clinical decision-making and the optimization of therapy is presented as a nomogram that includes serum markers and clinicopathologic features for predicting postoperative survival in cholangiocarcinoma.

Development and validation of a radiomics model of magnetic resonance for predicting liver metastasis in resectable pancreatic ductal adenocarcinoma patients.

BACKGROUND: Nearly one fourth of patients with pancreatic ductal adenocarcinoma (PDAC) occur to liver metastasis after surgery, and liver metastasis is a risk factor for prognosis for those patients with surgery therapy. However, there is no effective way to predict liver metastasis post-operation. METHOD: Clinical data and preoperative magnetic resonance imaging (MRI) of PDAC patients diagnosed between July 2010 and July 2020 were retrospectively collected from three hospital centers in China. The significant MRI radiomics features or clinicopathological characteristics were used to establish a model to predict liver metastasis in the development and validation cohort. RESULTS: A total of 204 PDAC patients from three hospital centers were divided randomly (7:3) into development and validation cohort. Due to poor predictive value of clinical features, MRI radiomics model had similar receiver operating characteristics curve (ROC) value to clinical-radiomics combing model in development cohort (0.878 vs. 0.880, p = 0.897) but better ROC in validation dataset (0.815 vs. 0.732, p = 0.022). Radiomics model got a sensitivity of 0.872/0.750 and a specificity of 0.760/0.822 to predict liver metastasis in development and validation cohort, respectively. Among 54 patients randomly selected with post-operation specimens, fibrosis markers (α-smooth muscle actin) staining was shown to promote radiomics model with ROC value from 0.772 to 0.923 (p = 0.049) to predict liver metastasis. CONCLUSION: This study developed and validated an MRI-based radiomics model and showed a good performance in predicting liver metastasis in resectable PDAC patients.

CCL3 secreted by hepatocytes promotes the metastasis of intrahepatic cholangiocarcinoma by VIRMA-mediated N6-methyladenosine (m6A) modification.

BACKGROUND: Intrahepatic cholangiocarcinoma (ICC) is a malignant disease characterized by onset occult, rapid progression, high relapse rate, and high mortality. However, data on how the tumor microenvironment (TME) regulates ICC metastasis at the transcriptomic level remains unclear. This study aimed to explore the mechanisms and interactions between hepatocytes and ICC cells. METHODS: We analyzed the interplay between ICC and liver microenvironment through cytokine antibody array analysis. Then we investigated the role of N6-methyladenosine (m6A) modification and the downstream target in vitro, in vivo experiments, and in clinical specimens. RESULTS: Our study demonstrated that cytokine CCL3, which is secreted by hepatocytes, promotes tumor metastasis by regulating m6A modification via vir-like m6A methyltransferase associated (VIRMA) in ICC cells. Moreover, immunohistochemical analyses showed that VIRMA correlated with poor outcomes in ICC patients. Finally, we confirmed both in vitro and in vivo that CCL3 could activate VIRMA and its critical downstream target SIRT1, which fuels tumor metastasis in ICC. CONCLUSIONS: In conclusion, our results enhanced our understanding of the interaction between hepatocytes and ICC cells, and revealed the molecular mechanism of the CCL3/VIRMA/SIRT1 pathway via m6A-mediated regulation in ICC metastasis. These studies highlight potential targets for the diagnosis, treatment, and prognosis of ICC.

Glutamate from nerve cells promotes perineural invasion in pancreatic cancer by regulating tumor glycolysis through HK2 mRNA-m6A modification.

BACKGROUND: Perineural invasion (PNI) has a high incidence and poor prognosis in pancreatic ductal adenocarcinoma (PDAC). Our study aimed to identify the underlying molecular mechanism of PNI and propose effective intervention strategies. METHODS: To observe PNI in vitro and in vivo, a Matrigel/ dorsal root ganglia (DRG) model and a murine sciatic nerve invasion model were respectively used. Magnetic resonance (MR) imaging and positron emission tomography/computed tomography (PET-CT) imaging were also used to evaluate tumor growth. Publicly available datasets and PDAC tissues were used to verify how the nerve cells regulate PDAC cells' PNI. RESULTS: Our results showed that glutamate from nerve cells could cause calcium influx in PDAC cells via the N-methyl-d-aspartate receptor (NMDAR), subsequently activating the downstream Ca2+ dependent protein kinase CaMKII/ERK-MAPK pathway and promoting the mRNA transcription of gene METTL3. Next, METTL3 upregulates the expression of hexokinase 2 (HK2) through N6-methyladenosine (m6A) modification in mRNA, enhances the PDAC cells' glycolysis, and promotes PNI. Furthermore, the IONPs-PEG-scFvCD44v6-scAbNMDAR2B nanoparticles dual targeting CD44 variant isoform 6 (CD44v6) and t NMDAR subunit 2B (NMDAR2B) on PDAC cells were synthesized and verified showing a satisfactory blocking effect on PNI. CONCLUSIONS: Here, we firstly provided evidence that glutamate from the nerve cells could upregulate the expression of HK2 through mRNA m6A modification via NMDAR2B and downstream Ca2+ dependent CaMKII/ERK-MAPK pathway, enhance the glycolysis in PDAC cells, and ultimately promote PNI. In addition, the dual targeting nanoparticles we synthesized were verified to block PNI effectively in PDAC.

A LETM2-Regulated PI3K-Akt Signaling Axis Reveals a Prognostic and Therapeutic Target in Pancreatic Cancer.

Pancreatic ductal adenocarcinoma (PDAC) is one of the highest mortalities malignant tumors, which is characterized by difficult diagnosis, rapid progression and high recurrence rate. Nevertheless, PDAC responds poorly to conventional therapies, which highlights the urgency to identify novel prognostic and therapeutic targets. LEMT2 was a newly discovered protein-encoding gene with little cancer research and an unclear mechanism. Thus, this study aimed to illustrate LETM2 as the crucial oncogene for tumor progression in PDAC. In this study, we analyzed the expression level and prognostic value of LETM2 in multiple cancers using pan-cancer analysis. The analyses based on the TCGA-GTEx dataset indicated that the LETM2 expression was obviously elevated in several cancers, and it was the most significantly related to the dismal prognosis of PDAC. Subsequently, we demonstrated the functional role and mechanism of LETM2 by clinical sample evaluation, and in in vitro and in vivo experiments. Immunohistochemical analyses showed that high expression of LETM2 was correlated with poor outcomes of PDAC. Moreover, we demonstrated that LETM2 knockdown significantly inhibited tumor proliferation and metastasis, and promoted cell apoptosis, while LETM2 overexpression exerted the opposite effects. Finally, the impairment caused by LETM2-knockdown could be recovered via excitation of the PI3k-Akt pathway in vitro and in vivo animal models, which suggested that LETM2 could activate the downstream PI3K-Akt pathway to participate in PDAC progression. In conclusion, the study enhanced our understanding of LETM2 as an oncogene hallmark of PDAC. LETM2 may facilitate tumor progression by activating the PI3K-Akt signaling pathway, which provides potential targets for the diagnosis, treatment, and prognosis of pancreatic cancer.

Arsenic trioxide-loaded nanoparticles enhance the chemosensitivity of gemcitabine in pancreatic cancer via the reversal of pancreatic stellate cell desmoplasia by targeting the AP4/galectin-1 pathway.

Pancreatic stellate cells (PSCs) constitute the fibrotic tumor microenvironment composed of the stroma matrix, which blocks the penetration of gemcitabine (GEM) in pancreatic adenocarcinoma (PDAC) and results in chemoresistance. We analyzed the expression of α-SMA, collagen type I, and fibronectin by immunohistochemistry of pancreatic cancer tissues and demonstrated that the abundant interstitial stroma is associated with dismal survival. Two desmoplastic pancreatic tumor models are treated with arsenic trioxide (ATO) and GEM to confirm the sensitizing effect of ATO on GEM. RNA-seq was performed to analyze the potential fibrotic genes regulated by ATO. Western blotting, CCK-8 methods, colony formation, and wound healing and transwell assays were utilized to verify that ATO attenuates the tumor-promoting ability of PSCs by inhibiting its activation and decreasing matrix secretion via the PI3K/AKT/AP4/galectin-1 pathway. Furthermore, we developed targeted ATO-loaded nanoparticles self-assembled by poly (D,L-lactide) and poly(ethylene glycol) (PEG-PDLLA) and modified by the single-chain antibody of FAP-α (scAbFAP-α) (scAb-ATO-NPs) to promote the delivery efficiency of ATO to PSCs and enhance anti-tumor effects with gemcitabine. Herein, we elucidate the mechanism of how ATO inhibits the activation of PSCs and enhances the therapeutic effect of GEM. We propose a novel cocktail therapy including scAb-ATO-NPs and GEM, indicating a new perspective in the treatment of PDAC.

TIMP1 derived from pancreatic cancer cells stimulates Schwann cells and promotes the occurrence of perineural invasion.

Perineural invasion (PNI) occurs in most pancreatic ductal adenocarcinomas (PDACs). The relationship between cancer cells and peripheral nerves, however, is unknown. Therefore, we focused on the cooperation of PDAC cells and peripheral nerve astrocytes, Schwann cells (SCs), in PNI. The mutual tumor-supportive secretory cytokines between SCs (sNF96.2) and PDAC cells (PANC-1, BxPC-3) were screened by human cytokine arrays and verified. The prognostic value of selected cytokines and SC-associated markers was confirmed in PDAC patients. TIMP1 and CCL7 were found to form a paracrine feedback loop between PDAC cells and SCs. PDAC cell-derived TIMP1 promotes SCs proliferation and migration via CD63/PI3K/AKT signaling. CCL7 secreted from SCs enhances PDAC cell migration, invasion and expression of TIMP1 via CCR2/STAT3. PDAC cell-SC cooperation in PNI was blocked when TIMP1 knockdown in vitro and in vivo. Finally, TIMP1, CCL7 and SC-associated markers were correlated with PNI and prognosis in PDAC patients. In conclusion, SCs collaborate with PDAC cells through the TIMP1-CCL7 paracrine feedback loop to promote PNI. TIMP1 knockdown in PDAC cells suppresses PNI. Strategies to disrupt the TIMP1-CCL7 feedback loop might be developed to inhibit PNI in PDAC.

GLUT1 Regulates the Tumor Immune Microenvironment and Promotes Tumor Metastasis in Pancreatic Adenocarcinoma via ncRNA-mediated Network.

Pancreatic adenocarcinoma (PAAD) is a digestive tumor with extremely high malignancy. Previous studies have reported that Glucose transporter 1 (GLUT1) contributes to the aggressive tumor progression in various cancer types and indicates an unfavorable prognosis. However, the function of GLUT1 in PAAD remains largely unclear. Through pan-cancer analysis of GLUT1 expression, GLUT1 expression was significantly higher in several cancer types including PAAD. Survival analysis based on the GLUT1 expression showed that GLUT1 could serve as a predictor of poor prognosis. We further predicted and screened the candidate non-coding RNAs (ncRNAs) upstream of the GLUT1 mRNA through correlation analysis, and found that the CASC19/miR-140-5p axis contributing to the regulation of GLUT1 expression. Our study suggested a link exists between GLUT1 expression and selected immunity-related indicators. Subsequent analysis revealed overexpression of GLUT1 in pancreatic cancer specimens and patients with highly expressed GLUT1 expression had worse prognosis. Based on the significantly different expression of GLUT1, the possibility that GLUT1 participated in tumor progression was identified. Using online public databases, genes co-expressed with GLUT1 were screened and enriched to metastasis-related pathways by enrichment analysis. Additionally, functional assays verified that GLUT1 could function in the metastatic process of PAAD cancer cells. Therefore, we proposed that GLUT1 might serve as a role in tumor immunity and tumor metastasis, and was expected to be a prognostic factor in PAAD.

Cascade and Fusion: A Deep Learning Approach for Camouflaged Object Sensing.

The demand for the sensor-based detection of camouflage objects widely exists in biological research, remote sensing, and military applications. However, the performance of traditional object detection algorithms is limited, as they are incapable of extracting informative parts from low signal-to-noise ratio features. To address this problem, we propose Camouflaged Object Detection with Cascade and Feedback Fusion (CODCEF), a deep learning framework based on an RGB optical sensor that leverages a cascaded structure with Feedback Partial Decoders (FPD) instead of a traditional encoder-decoder structure. Through a selective fusion strategy and feedback loop, FPD reduces the loss of information and the interference of noises in the process of feature interweaving. Furthermore, we introduce Pixel Perception Fusion (PPF) loss, which aims to pay more attention to local pixels that might become the edges of an object. Experimental results on an edge device show that CODCEF achieved competitive results compared with 10 state-of-the-art methods.

Bioinformatics-Based Identification of Tumor Microenvironment-Related Prognostic Genes in Pancreatic Cancer.

OBJECTIVE: Growing evidence has highlighted that the immune and stromal cells that infiltrate in pancreatic cancer microenvironment significantly influence tumor progression. However, reliable microenvironment-related prognostic gene signatures are yet to be established. The present study aimed to elucidate tumor microenvironment-related prognostic genes in pancreatic cancer. METHODS: We applied the ESTIMATE algorithm to categorize patients with pancreatic cancer from TCGA dataset into high and low immune/stromal score groups and determined their differentially expressed genes. Then, univariate and LASSO Cox regression was performed to identify overall survival-related differentially expressed genes (DEGs). And multivariate Cox regression analysis was used to screen independent prognostic genes and construct a risk score model. Finally, the performance of the risk score model was evaluated by Kaplan-Meier curve, time-dependent receiver operating characteristic and Harrell's concordance index. RESULTS: The overall survival analysis demonstrated that high immune/stromal score groups were closely associated with poor prognosis. The multivariate Cox regression analysis indicated that the signatures of four genes, including TRPC7, CXCL10, CUX2, and COL2A1, were independent prognostic factors. Subsequently, the risk prediction model constructed by those genes was superior to AJCC staging as evaluated by time-dependent receiver operating characteristic and Harrell's concordance index, and both KRAS and TP53 mutations were closely associated with high risk scores. In addition, CXCL10 was predominantly expressed by tumor associated macrophages and its receptor CXCR3 was highly expressed in T cells at the single-cell level. CONCLUSIONS: This study comprehensively investigated the tumor microenvironment and verified immune/stromal-related biomarkers for pancreatic cancer.

Peripheral blood monocytes predict clinical prognosis and support tumor invasiveness through NF-κB-dependent upregulation of Snail in pancreatic cancer.

BACKGROUND: The tumor inflammatory microenvironment plays a vital role in the initiation and progression of pancreatic cancer (PC). Both the lymphocyte-to-monocyte ratio (LMR) and preoperative peripheral blood monocytes are related to the prognosis of PC patients. However, the direct effect of monocytes on PC cells is not fully understood. The current study aimed to assess the effect of monocytes on PC and explore its potential mechanism. METHODS: The cutoff value of peripheral blood monocytes was evaluated by the receiver operating characteristic (ROC) curve. Transwell migration and invasion assays were used to detect the mobility of PC cells. The cytokines derived from monocytes were measured by quantitative real-time polymerase chain reaction (qRT-PCR). Western blotting was utilized to assess the expression of epithelial-mesenchymal transition (EMT) related markers. The expression level of Snail in PC tissue was determined by immunohistochemical (IHC) staining. RESULTS: A high monocyte count was inversely correlated with lymph node status and 5-year overall survival in PC. The PC cells underwent a cellular morphology change and increased cell motility after coculture with THP-1 monocytes. The THP-1 monocytes secreted various proinflammatory cytokines, including tumor necrosis factor-α (TNF-α) and interleukin-1α (IL-1α), which activated the nuclear factor-κB (NF-κB) signaling pathway leading to the upregulation of Snail and thereby promoting the EMT of PC cells. The expression level of Snail correlated significantly with the density of peripheral blood monocytes, and their level status was significantly associated with 5-year overall survival. CONCLUSIONS: These findings indicated that elevated monocytes counts were a poor prognostic marker in PC, and monocytes could directly induce the EMT process of PC cells by upregulating Snail expression through the NF-κB signaling pathway.

TIMP1 down-regulation enhances gemcitabine sensitivity and reverses chemoresistance in pancreatic cancer.

The therapeutic effect of gemcitabine (GEM) in pancreatic ductal adenocarcinoma (PDAC) is limited due to low drug sensitivity and high drug resistance. Tissue inhibitor of matrix metalloprotease 1 (TIMP1) is reportedly associated with GEM resistance in PDAC. However, the effect of TIMP1 down-regulation in combination with GEM treatment is unknown. We analyzed the expression of TIMP1 in human PDAC tissue using western blot, quantitative real-time polymerase chain reaction (qRT-PCR), and immunohistochemistry. TIMP1 was highly expressed in PDAC specimens. Kaplan-Meier survival analysis suggested that a higher level of TIMP1 was correlated with poorer overall survival in 103 PDAC patients. The mRNA and protein expression profiles of TIMP1 were explored in the HTERT-HPNE human pancreatic ductal epithelium cell line, five PDAC cell lines (MIA PaCa-2, PANC-1, BxPC-3, Capan2, and SW1990), and two GEM-resistant PDAC cell lines (MIA PaCa-2R and PANC-1R). Compared with HTERT-HPNE, TIMP1 was highly expressed in the PDAC cell lines. In addition, TIMP1 was upregulated in GEM-resistant PDAC cell lines compared with their parental cells. When TIMP1 was knocked-down using short hairpin RNA, GEM-induced cytotoxicity and apoptosis were increased, while colony formation was repressed in MIA PaCa-2, PANC-1, and their GEM-resistant cells. When Bax was activated by BAM7 or Bcl-2 was inhibited by venetoclax, CCK-8 assays demonstrated that GEM sensitivity was restored in GEM-resistant cells. When Bax was down-regulated by siRNA, CCK-8 assays verified that GEM sensitivity was decreased in PDAC cells. The observations that TIMP1 knockdown enhanced GEM sensitivity and reversed chemoresistance by inducing cells apoptosis indicated cooperative antitumor effects of shTIMP1 and GEM therapy on PDAC cells. The combination may be a potential strategy for PDAC therapy.

Suppression MGP inhibits tumor proliferation and reverses oxaliplatin resistance in colorectal cancer.

Matrix Gla protein (MGP), an extracellular matrix protein, has been widely reported to participate in the tumorigenic process and is abnormally expressed in several tumors. However, the role of MGP in colorectal cancer (CRC) remains unknown. Chemotherapy resistance represents a significant limitation in the treatment of CRC. Here, a comprehensive bioinformatics analysis revealed that MGP, which is overexpressed in CRC, might act as one of the critical genes conferring resistance to oxaliplatin (OXA). Furthermore, we found that MGP overexpression in tumor tissue might be correlated with cancer stage and patient prognosis, consistent with the bioinformatics analysis. The upregulation of MGP may act as an independent risk factor for CRC. The knockdown of MGP or inhibition of MGP expression significantly increased the sensitivity of the CRC cell lines to OXA. Suppression of MGP may reverse OXA resistance by upregulating copper transporter 1 (CTR1) and downregulating ATP7A and ATP7B. When used in combination with OXA, the inhibition of MGP reduced cancer cell proliferation, invasion, and migration and increased cell apoptosis in vitro. Suppression of MGP or OXA treatment alone significantly inhibited tumor growth in the CRC mouse model. Additionally, we found that OXA might promote the antitumor immune response in vivo. In summary, our study is the first to provide evidence that MGP expression confers OXA chemotherapy resistance in CRC and provides novel strategies to overcome chemotherapy resistance in CRC.

Bmi-1-induced miR-27a and miR-155 promote tumor metastasis and chemoresistance by targeting RKIP in gastric cancer.

BACKGROUND: We previously reported an inverse relationship between B cell-specific Moloney murine leukemia virus integration site 1 (Bmi-1) and Raf kinase inhibitory protein (RKIP), which is associated with the prognosis of gastric cancer (GC). In this study, we further explored the microRNA (miRNA) regulatory mechanism between Bmi-1 and RKIP. METHODS: Microarray analysis was first carried out to identify miRNA profiles that were differentially expressed in cells overexpressing Bmi-1. Then, miRNAs that could regulate RKIP were identified. Quantitative real-time PCR (qRT-PCR) and Western blotting were performed to measure the expression of Bmi-1, miR-155, miR-27a and RKIP. RKIP was confirmed as a target of miR-27a and miR-155 through luciferase reporter assays, qRT-PCR and Western blotting. The effects of the Bmi-1/miR-27a/RKIP and Bmi-1/miR-155/RKIP axes on tumor growth, proliferation, migration, invasion, colony-formation ability, metastasis and chemoresistance were investigated both in vitro and in vivo. RESULTS: The downregulation of RKIP by Bmi-1 occurred at the protein but not mRNA level. This indicates probable posttranscriptional regulation. miRNA expression profiles of cells with ectopic expression of Bmi-1 were analyzed and compared to those of control cells by microarray analysis. A total of 51 upregulated and 72 downregulated miRNAs were identified. Based on publicly available algorithms, miR-27a and miR-155 were predicted, selected and demonstrated to target RKIP. Bmi-1, miR-27a and miR-155 are elevated in human GC and associated with poor prognosis of GC, while RKIP is expressed at lower levels in GC and correlated with good prognosis. Then, in vitro tests shown that in addition to regulating RKIP expression via miR-27a and miR-155, Bmi-1 was also able to regulate the migration, invasion, proliferation, colony-formation ability and chemosensitivity of GC cells through the same pathway. Finally, the in vivo test showed similar results, whereby the knockdown of the Bmi-1 gene led to the inhibition of tumor growth, metastasis and chemoresistance through miR-27a and miR-155. CONCLUSIONS: Bmi-1 was proven to induce the expression of miR-27a and miR-155 and thus promote tumor metastasis and chemoresistance by targeting RKIP in GC. Overall, miR-27a and miR-155 might be promising targets for the screening, diagnosis, prognosis, treatment and disease monitoring of GC.

Development and validation of a novel nomogram for pretreatment prediction of liver metastasis in pancreatic cancer.

PURPOSE: The diagnostic value of nomogram in pancreatic cancer (PC) with liver metastasis (PCLM) is still largely unknown. We sought to develop and validate a novel nomogram for the prediction of liver metastasis in patients with PC. METHOD: About 604 pathologically confirmed PC patients from the Sun Yat-sen University Cancer Center (SYSUCC) between July, 2001 and December, 2013 were retrospectively studied. The SYSUCC cohort was randomly assigned to as the training set and internal validation set. Using these two sets, we derived and validated a prognostic model by using concordance index and calibration curves. Another two independent cohorts between August, 2002 and December, 2013 from the Sun Yat-sen Memorial Hospital (SYSMH, n = 335) and Guangdong General Hospital (GDGH, n = 503) was used for external validation. RESULT: Computed tomography (CT) reported liver metastasis status, carcinoembryonic antigen (CEA) level and differentiation type were identified as risk factors for PCLM in the training set. The final diagnostic model demonstrated good calibration and discrimination with a concordance index of 0.97 and had a robust internal validation. The score ability to diagnose PCLM was further externally validated in SYSMH and GDGH with a concordance index of 0.93. The model showed better calibration and discrimination than CT, CEA and differentiation in each cohort. CONCLUSION: Based on a large multi-institution database and on the routinely observed CT-reported status, CEA level and tumor differentiation in clinical practice, we developed and validated a novel nomogram to predict PLCM.

Nanoparticles modified by triple single chain antibodies for MRI examination and targeted therapy in pancreatic cancer.

Precise diagnosis and effective treatment are crucial to the prognosis of pancreatic ductal adenocarcinoma (PDAC). Magnetic iron oxide nanoparticles (IONPs) are superior magnetic resonance imaging (MRI) contrast agents, while antibodies are significant immunotherapy reagents. Herein, we firstly generated a novel nanocomposite combining triple single chain antibodies (scAbs) and IONPs for the detection and treatment of PDAC. METHODS: Triple scAbs (scAbMUC4, scAbCEACAM6, scFvCD44v6, MCC triple scAbs) were conjugated to the surface of polyethylene glycol modified IONPs (IONPs-PEG), forming the IONPs-PEG-MCC triple scAbs nanocomposite. Characterization of the nanocomposite was performed, and its cytotoxicity, specificity, and apoptosis induction were evaluated. In vivo MRI study and anti-pancreatic cancer effect assessment were performed in tumor-bearing nude mice. RESULTS: The size of the IONPs-PEG-MCC triple scAbs nanocomposite was about 23.6 nm. The nanocomposite was non-toxic to normal pancreatic ductal epithelial cells, and could specifically bind to and be internalized by MUC4/CEACAM6/CD44v6-expressing PDAC cells. With an r2 relaxivity of 104.2 mM-1 s-1, the IONPs-PEG-MCC triple scAbs nanocomposite could significantly shorten the MRI T2-weighted signal intensity both in vitro and in vivo. The IONPs-PEG-MCC triple scAbs nanocomposite also showed a favorable anti-pancreatic cancer effect. CONCLUSION: In the present study, the IONPs-PEG-MCC triple scAbs nanocomposite was firstly confirmed as a bi-functional nanocomposite in both MRI and treatment, providing its critical clinical transformation potential in PDAC detection and treatment.

Expression of the PTEN/FOXO3a/PLZF signalling pathway in pancreatic cancer and its significance in tumourigenesis and progression.

Pancreatic cancer (PC) is one of the most lethal gastrointestinal malignancies. The PTEN/AKT signalling pathway is closely related to the tumourigenesis and progression of PC. The downstream effectors, FOXO3a, PLZF and VEGF, are reported to be involved in angiogenesis, lymph node metastasis and poor survival in PC. By using tissue microarrays and immunohistochemistry, we found, that PTEN, FOXO3a and PLZF expression was significantly decreased in PC specimens compared with that in chronic pancreatitis (CP) specimens, while VEGF expression was significantly increased. Furthermore, the expression of PTEN was positively correlated with that of FOXO3a and PLZF but negatively correlated with that of VEGF. Our results suggest that the PTEN/FOXO3a/PLZF signalling pathway may negatively regulate VEGF expression in PC. Through clinical analysis of 69 PC patients, PTEN, FOXO3a and PLZF expression was found to be significantly decreased in specimens from PC patients with lymph node metastasis and poor prognosis, while VEGF expression was significantly increased. Taken together, these reaults suggest that the PTEN/FOXO3a/PLZF signalling pathway may be capable of inhibiting growth and metastasis in PC by regulating VEGF-mediated angiogenesis, which requires further in vivo and in vitro studies and can potentially be a therapeutic target for PC.

Prognostic and metastasis-related factors in colorectal neuroendocrine tumors: A cross-sectional study based on the Surveillance, Epidemiology and End Results.

The incidence of colorectal neuroendocrine tumors (NETs) is gradually increasing with the increasing availability of colonoscopy and computed tomography. However, prognostic and metastatic factors for colorectal NETs are unknown. The aim of the present study was to identify clinicopathological prognostic and metastasis-related risk factors for colorectal NETs. Based on the Surveillance, Epidemiology and End Results (SEER) program, the data of 3,093 patients with colorectal NETs were included in the present study. χ2 test, crosstabulation, Kaplan-Meier method, Cox proportional hazard regression model, nomogram and logistic regression analysis were used to evaluate the risk factors. Systemic analysis of data from 3,093 patients revealed that age (P<0.001), marital status (P<0.001), number of malignant tumors (P<0.001), histopathological grade (P<0.001) and clinical stage (P<0.001) were independent prognostic factors of cancer-specific survival (CSS). Among them, married patients exhibited longer CSS compared with unmarried patients (P<0.05). A nomogram based on these five factors with an area under the curve of 0.921 [95% confidence interval (CI), 0.908-0.934; P<0.001] was constructed. Age (P=0.010), primary tumor site (P<0.001), surgery (P<0.001), tumor size (P<0.001) and histopathological grade (P<0.001) were risk factors for distant metastasis of colorectal NETs. In conclusion, age, marital status, number of malignant tumors, histopathological grade and clinical stage may be independent prognostic factors for CSS of colorectal NETs, and the nomogram may have higher predictive efficiency compared with clinical stage. Age, primary tumor site, surgery, tumor size and histopathological grade may be risk factors for the distant metastasis of colorectal NETs.