Elevated 2-oxoglutarate antagonizes DNA damage responses in cholangiocarcinoma chemotherapy through regulating aspartate beta-hydroxylase.

Cholangiocarcinoma (CCA) is resistant to systemic chemotherapies that kill malignant cells mainly through DNA damage responses (DDRs). Recent studies suggest that the involvement of 2-oxoglutarate (2-OG) dependent dioxygenases in DDRs may be associated with chemoresistance in malignancy, but how 2-OG impacts DDRs in CCA chemotherapy remains elusive. We examined serum 2-OG levels in CCA patients before receiving chemotherapy. CCA patients are classified as progressive disease (PD), partial response (PR), and stable disease (SD) after receiving chemotherapy. CCA patients classified as PD showed significantly higher serum 2-OG levels than those defined as SD and PR. Treating CCA cells with 2-OG reduced DDRs. Overexpression of full-length aspartate beta-hydroxylase (ASPH) could mimic the effects of 2-OG on DDRs, suggesting the important role of ASPH in chemoresistance. Indeed, the knockdown of ASPH improved chemotherapy in CCA cells. Targeting ASPH with a specific small molecule inhibitor also enhanced the effects of chemotherapy. Mechanistically, ASPH modulates DDRs by affecting ATM and ATR, two of the major regulators finely controlling DDRs. More importantly, targeting ASPH improved the therapeutic potential of chemotherapy in two preclinical CCA models. Our data suggested the impacts of elevated 2-OG and ASPH on chemoresistance through antagonizing DDRs. Targeting ASPH may enhance DDRs, improving chemotherapy in CCA patients.

Molecular profiling reveals potential targets in cholangiocarcinoma.

BACKGROUND: Cholangiocarcinoma (CCA) is a devastating malignancy and has a very poor prognosis if tumors spread outside the liver. Understanding the molecular mechanisms underlying the CCA progression will likely yield therapeutic approaches toward treating this deadly disease. AIM: To determine the molecular pathogenesis in CCA progression. METHODS: In silico analysis, in vitro cell culture, CCA transgenic animals, histological, and molecular assays were adopted to determine the molecular pathogenesis. RESULTS: The transcriptomic data of human CCA samples were retrieved from The Cancer Genome Atlas (TGCA, CHOL), European Bioinformatics Institute (EBI, GAD00001001076), and Gene Expression Omnibus (GEO, GSE107943) databases. Using Gene set enrichment analysis, the cell cycle and Notch related pathways were demonstrated to be significantly activated in CCA in TCGA and GEO datasets. We, through differentially expressed genes, found several cell cycle and notch associated genes were significantly up-regulated in cancer tissues when compared with the non-cancerous control samples. The associated genes, via quantitative real-time PCR and western blotting assays, were further examined in normal human cholangiocytes, CCA cell lines, mouse normal bile ducts, and mouse CCA tumors established by specifically depleting P53 and expressing KrasG12D mutation in the liver. Consistently, we validated that the cell cycle and Notch pathways are up-regulated in CCA cell lines and mouse CCA tumors. Interestingly, targeting cell cycle and notch pathways using small molecules also exhibited significant beneficial effects in controlling tumor malignancy. More importantly, we demonstrated that several cell cycle and Notch associated genes are significantly associated with poor overall survival and disease-free survival using the Log-Rank test. CONCLUSION: In summary, our study comprehensively analyzed the gene expression pattern of CCA samples using publicly available datasets and identified the cell cycle and Notch pathways are potential therapeutic targets in this deadly disease.

Tumor Cell Derived Lnc-FSD2-31:1 Contributes to Cancer-Associated Fibroblasts Activation in Pancreatic Ductal Adenocarcinoma Progression through Extracellular Vesicles Cargo MiR-4736.

Pancreatic ductal adenocarcinoma (PDAC) presents with high mortality and short overall survival. Cancer-associated fibroblasts (CAFs) act as refuge for cancer cells in PDAC. Mechanisms of intracelluar communication between CAFs and cancer cells need to be explored. Long noncoding RNAs (lncRNAs) are involved in the modulation of oncogenesis and tumor progression of PDAC; however, specific lncRNAs and their mechanism of action have not been clarified clearly in tumoral microenvironment. This work aims to identify novel lncRNAs involved in cellular interaction between cancer cells and CAFs in PDAC. To this end, differentially expressed lncRNAs between long-term and short-term survival PDAC patients are screened. Lnc-FSD2-31:1 is found to be significantly increased in long-term survival patients. This work then discovers that tumor-derived lnc-FSD2-31:1 restrains CAFs activation via miR-4736 transported by extracellular vesicles (EVs) in vitro and in vivo. Mechanistically, EVs-derived miR-4736 suppresses autophagy and contributes to CAFs activation by targeting ATG7. Furthermore, blocking miR-4736 suppresses tumor growth in genetically engineered KPC (LSL-KrasG12D/+, LSL-Trp53R172H/+, and Pdx-1-Cre) mouse model of PDAC. This study demonstrates that intratumoral lnc-FSD2-31:1 modulates autophagy in CAFs resulting in their activation through EVs-derived miR-4736. Targeting miR-4736 may be a potential biomarker and therapeutic target for PDAC.

The Prediction of Steel Bar Corrosion Based on BP Neural Networks or Multivariable Gray Models.

The corrosion of steel bars in concrete has a significant impact on the durability of constructed structures. Based on the gray relational analysis (GRA) of the accelerated corrosion data and practical engineering data using MATLAB, a back propagation neural network (BPNN) model, a multivariable gray prediction model (GM (1, N)), and an optimization multivariable gray prediction model (OGM (1, N)) of steel corrosion were established by using a sequence of the key affecting factors. By comparing the prediction results of the three models, it is found that the GM (1, N) model has larger fitting and prediction errors for steel corrosion, while the OGM (1, N) model has smaller prediction errors in the accelerated corrosion data; the BPNN model offers more accurate predictions of the practical engineering data. The results show that the BPNN and OGM (1, N) models are all suitable for the prediction of steel bar corrosion in concrete structures.

Adaptive antitumor immune response stimulated by bio-nanoparticle based vaccine and checkpoint blockade.

BACKGROUND: Interactions between tumor and microenvironment determine individual response to immunotherapy. Triple negative breast cancer (TNBC) and hepatocellular carcinoma (HCC) have exhibited suboptimal responses to immune checkpoint inhibitors (ICIs). Aspartate ß-hydroxylase (ASPH), an oncofetal protein and tumor associated antigen (TAA), is a potential target for immunotherapy. METHODS: Subcutaneous HCC and orthotopic TNBC murine models were established in immunocompetent BALB/c mice with injection of BNL-T3 and 4 T1 cells, respectively. Immunohistochemistry, immunofluorescence, H&E, flow cytometry, ELISA and in vitro cytotoxicity assays were performed. RESULTS: The ASPH-MYC signaling cascade upregulates PD-L1 expression on breast and liver tumor cells. A bio-nanoparticle based λ phage vaccine targeting ASPH was administrated to mice harboring syngeneic HCC or TNBC tumors, either alone or in combination with PD-1 blockade. In control, autocrine chemokine ligand 13 (CXCL13)-C-X-C chemokine receptor type 5 (CXCR5) axis promoted tumor development and progression in HCC and TNBC. Interactions between PD-L1+ cancer cells and PD-1+ T cells resulted in T cell exhaustion and apoptosis, causing immune evasion of cancer cells. In contrast, combination therapy (Vaccine+PD-1 inhibitor) significantly suppressed primary hepatic or mammary tumor growth (with distant pulmonary metastases in TNBC). Adaptive immune responses were attributed to expansion of activated CD4+ T helper type 1 (Th1)/CD8+ cytotoxic T cells (CTLs) that displayed enhanced effector functions, and maturation of plasma cells that secreted high titers of ASPH-specific antibody. Combination therapy significantly reduced tumor infiltration of immunosuppressive CD4+/CD25+/FOXP3+ Tregs. When the PD-1/PD-L1 signal was inhibited, CXCL13 produced by ASPH+ cancer cells recruited CXCR5+/CD8+ T lymphocytes to tertiary lymphoid structures (TLSs), comprising effector and memory CTLs, T follicular helper cells, B cell germinal center, and follicular dendritic cells. TLSs facilitate activation and maturation of DCs and actively recruit immune subsets to tumor microenvironment. These CTLs secreted CXCL13 to recruit more CXCR5+ immune cells and to lyse CXCR5+ cancer cells. Upon combination treatment, formation of TLSs predicts sensitivity to ICI blockade. Combination therapy substantially prolonged overall survival of mice with HCC or TNBC. CONCLUSIONS: Synergistic antitumor efficacy attributable to a λ phage vaccine specifically targeting ASPH, an ideal TAA, combined with ICIs, inhibits tumor growth and progression of TNBC and HCC.

Bio-nanoparticle based therapeutic vaccine induces immunogenic response against triple negative breast cancer.

Triple negative breast cancer (TNBC) is more aggressive and has a poorer prognosis than other sub-types of breast tumors. This study elucidates how aspartate beta-hydroxylase (ASPH) network promotes drug resistance, and immunotherapy targeting ASPH may improve the efficacy of Doxorubicin (DOX) therapy. An orthotopic model of breast cancer generated by 4T1 cells in immunocompetent mice was used to explore efficacy of immunotherapy in combination with DOX chemotherapy. We evaluated mRNA and protein expression in cultured tumor cells and tissue, as well as assessed cell proliferation, apoptosis, soluble factors/cytokine production, immune cell population diversity and function. We observed that ASPH expression enables TNBC cells to exhibit primary resistance to DOX induced single-/double-strand breaks (SSB/DSB) and enhanced proliferation and survival. Specific bio-nanoparticle based therapeutic vaccine (BNP-TV) promoted ASPH uptake by and maturation of DCs. This BNP-TV combined with DOX induces immunogenic cell death (ICD) in orthotopic xenograft tumors and significantly suppressed primary mammary tumor growth and distant multi-organ metastases. Immunogenic cell death induced by BNP-TV targeting ASPH combined with DOX provides opportunities to treat a highly resistant and metastatic form of breast cancer.

Integrating Genetic and Transcriptomic Data to Reveal Pathogenesis and Prognostic Markers of Pancreatic Adenocarcinoma.

Pancreatic adenocarcinoma (PAAD) is one of the deadliest malignancies and mortality for PAAD have remained increasing under the conditions of substantial improvements in mortality for other major cancers. Although multiple of studies exists on PAAD, few studies have dissected the oncogenic mechanisms of PAAD based on genomic variation. In this study, we integrated somatic mutation data and gene expression profiles obtained by high-throughput sequencing to characterize the pathogenesis of PAAD. The mutation profile containing 182 samples with 25,470 somatic mutations was obtained from The Cancer Genome Atlas (TCGA). The mutation landscape was generated and somatic mutations in PAAD were found to have preference for mutation location. The combination of mutation matrix and gene expression profiles identified 31 driver genes that were closely associated with tumor cell invasion and apoptosis. Co-expression networks were constructed based on 461 genes significantly associated with driver genes and the hub gene FAM133A in the network was identified to be associated with tumor metastasis. Further, the cascade relationship of somatic mutation-Long non-coding RNA (lncRNA)-microRNA (miRNA) was constructed to reveal a new mechanism for the involvement of mutations in post-transcriptional regulation. We have also identified prognostic markers that are significantly associated with overall survival (OS) of PAAD patients and constructed a risk score model to identify patients' survival risk. In summary, our study revealed the pathogenic mechanisms and prognostic markers of PAAD providing theoretical support for the development of precision medicine.

Multi-organ metastasis as destination for breast cancer cells guided by biomechanical architecture.

A majority of breast cancer patients die of widespread aggressive multidrug-resistant tumors. Aspartate ß-hydroxylase (ASPH) is an α-ketoglutarate-dependent dioxygenase and oncofetal antigen involved in embryogenesis. To illustrate if ASPH could be targeted for metastatic breast cancer, embedded and on-top three-dimensional (3-D) cultures, 3-D invasion, mammosphere formation, immunofluorescence, immunohistochemistry, Western blot, co-IP and microarray were conducted. In vitro metastasis was developed to imitate how cancer cells invade basement membrane at the primary site, transendothelially migrate, consequently colonize and outgrow at distant sites. Orthotopic and experimental pulmonary metastatic (tail vein injection) murine models were established using stable breast cancer cell lines. Cox proportional hazards regression models and Kaplan-Meier plots were applied to assess clinical outcome of breast cancer patients. In adult non-cancerous breast tissue, ASPH is undetectable. Pathologically, ASPH expression re-emerged at ductal carcinoma in situ (DCIS), and enhanced with disease progression, from early-stage invasive ductal carcinoma (IDC) to late-stage carcinoma. ASPH at moderate to high levels contribute to aggressive molecular subtypes, early relapse or more frequent progression and metastases, whereas substantially shortened overall survival and disease-free survival of breast cancer patients. Through direct physical interactions with A disintegrin and metalloproteinase domain-containing protein (ADAM)-12/ADAM-15, ASPH could activate SRC cascade, thus upregulating downstream components attributed to multifaceted metastasis. ASPH-SRC axis initiated pro-invasive invadopodium formation causing breakdown/disorganization of extracellular matrix (ECM), simultaneously potentiated epithelial-mesenchymal transition (EMT), induced cancer stem cell markers (CD44 and EpCAM), enhanced mammosphere formation and intensified 3-dimentional invasion. Oncogenic SRC upregulated matrix metallopeptidases (MMPs) were assembled by invadopodia, acting as executive effectors for multi-step metastasis. ASPH-SRC signal guided multi-organ metastases (to lungs, liver, bone, spleen, lymph nodes, mesentery or colon) in immunocompromised mice. Malignant phenotypes induced by ASPH-SRC axis were reversed by the third-generation small molecule inhibitor (SMI) specifically against ß-hydroxylase activity of ASPH in pre-clinical models of metastatic breast cancer. Collectively, ASPH could activate ADAMs-SRC-MMPs cascades to promote breast cancer tumor progression and metastasis. ASPH could direct invadopodium construction as a biomechanical sensor and pro-metastatic outlet. ASPH-mediated cancer progression could be specifically/efficiently subverted by SMIs of ß-hydroxylase activity. Therefore, ASPH emerges as a therapeutic target for breast cancer.

Targeting Aspartate Beta-Hydroxylase with the Small Molecule Inhibitor MO-I-1182 Suppresses Cholangiocarcinoma Metastasis.

BACKGROUND: Cholangiocarcinoma is a devastating disease with a 2% 5-year survival if the disease has spread outside the liver. The enzyme aspartate beta-hydroxylase (ASPH) has been demonstrated to be highly expressed in cholangiocarcinoma but not in normal bile ducts and found to stimulate tumor cell migration. In addition, it was found that targeting ASPH inhibits cholangiocarcinoma malignant progression. However, it is not clear whether targeting ASPH with the small molecule inhibitor MO-I-1182 suppresses cholangiocarcinoma metastasis. The current study aims to study the efficacy of MO-I-1182 in suppressing cholangiocarcinoma metastasis. METHODS: The analysis was performed in vitro and in vivo with a preclinical animal model by using molecular and biochemical strategies to regulate ASPH expression and function. RESULTS: Knockdown of ASPH substantially inhibited cell migration and invasion in two human cholangiocarcinoma cell lines. Targeting ASPH with a small molecule inhibitor suppressed cholangiocarcinoma progression. Molecular mechanism studies demonstrated that knockdown of ASPH subsequently suppressed protein levels of the matrix metalloproteinases. The ASPH knockdown experiments suggest that this enzyme may modulate cholangiocarcinoma metastasis by regulating matrix metalloproteinases expression. Furthermore, using an ASPH inhibitor in a rat cholangiocarcinoma intrahepatic model established with BED-Neu-CL#24 cholangiocarcinoma cells, it was found that targeting ASPH inhibited intrahepatic cholangiocarcinoma metastasis and downstream expression of the matrix metalloproteinases. CONCLUSION: ASPH may modulate cholangiocarcinoma metastasis via matrix metalloproteinases expression. Taken together, targeting ASPH function may inhibit intrahepatic cholangiocarcinoma metastasis and improve survival.

Ten-Eleven Translocation 1 Promotes Malignant Progression of Cholangiocarcinoma With Wild-Type Isocitrate Dehydrogenase 1.

BACKGROUND AND AIMS: Cholangiocarcinoma (CCA) is a highly lethal disease without effective therapeutic approaches. The whole-genome sequencing data indicate that about 20% of patients with CCA have isocitrate dehydrogenase 1 (IDH1) mutations, which have been suggested to target 2-oxoglutarate (OG)-dependent dioxygenases in promoting CCA carcinogenesis. However, the clinical study indicates that patients with CCA and mutant IDH1 have better prognosis than those with wild-type IDH1, further complicating the roles of 2-OG-dependent enzymes. APPROACH AND RESULTS: This study aimed to clarify if ten-eleven translocation 1 (TET1), which is one of the 2-OG-dependent enzymes functioning in regulating 5-hydroxymethylcytosine (5hmC) formation, is involved in CCA progression. By analyzing The Cancer Genome Atlas (TCGA) data set, TET1 mRNA was found to be substantially up-regulated in patients with CCA when compared with noncancerous bile ducts. Additionally, TET1 protein expression was significantly elevated in human CCA tumors. CCA cells were challenged with α-ketoglutarate (α-KG) and dimethyl-α-KG (DM-α-KG), which are cosubstrates for TET1 dioxygenase. The treatments with α-KG and DM-α-KG promoted 5hmC formation and malignancy of CCA cells. Molecular and pharmacological approaches were used to inhibit TET1 activity, and these treatments substantially suppressed 5hmC and CCA carcinogenesis. Mechanistically, it was found that knockdown of TET1 may suppress CCA progression by targeting cell growth and apoptosis through epigenetic regulation. Consistently, targeting TET1 significantly inhibited CCA malignant progression in a liver orthotopic xenograft model by targeting cell growth and apoptosis. CONCLUSIONS: Our data suggest that expression of TET1 is highly associated with CCA carcinogenesis. It will be important to evaluate TET1 expression in CCA tumors before application of the IDH1 mutation inhibitor because the inhibitor suppresses 2-hydroxyglutarate expression, which may result in activation of TET, potentially leading to CCA malignancy.

Prometastatic secretome trafficking via exosomes initiates pancreatic cancer pulmonary metastasis.

To demonstrate multifaceted contribution of aspartate ß-hydroxylase (ASPH) to pancreatic ductal adenocarcinoma (PDAC) pathogenesis, in vitro metastasis assay and patient derived xenograft (PDX) murine models were established. ASPH propagates aggressive phenotypes characterized by enhanced epithelial-mesenchymal transition (EMT), 2-D/3-D invasion, extracellular matrix (ECM) degradation/remodeling, angiogenesis, stemness, transendothelial migration and metastatic colonization/outgrowth at distant sites. Mechanistically, ASPH activates Notch cascade through direct physical interactions with Notch1/JAGs and ADAMs. The ASPH-Notch axis enables prometastatic secretome trafficking via exosomes, subsequently initiates MMPs mediated ECM degradation/remodeling as an effector for invasiveness. Consequently, ASPH fosters primary tumor development and pulmonary metastasis in PDX models, which was blocked by a newly developed small molecule inhibitor (SMI) specifically against ASPH's ß-hydroxylase activity. Clinically, ASPH is silenced in normal pancreas, progressively upregulated from pre-malignant lesions to invasive/advanced stage PDAC. Relatively high levels of ASPH-Notch network components independently/jointly predict curtailed overall survival (OS) in PDAC patients (log-rank test, Ps < 0.001; Cox proportional hazards regression, P < 0.001). Therefore, ASPH-Notch axis is essential for propagating multiple-steps of metastasis and predicts prognosis of PDAC patients. A specific SMI targeting ASPH offers a novel therapeutic approach to substantially retard PDAC development/progression.

ASPH-notch Axis guided Exosomal delivery of Prometastatic Secretome renders breast Cancer multi-organ metastasis.

BACKGROUND: Aspartate ß-hydroxylase (ASPH) is silent in normal adult tissues only to re-emerge during oncogenesis where its function is required for generation and maintenance of malignant phenotypes. Exosomes enable prooncogenic secretome delivering and trafficking for long-distance cell-to-cell communication. This study aims to explore molecular mechanisms underlying how ASPH network regulates designated exosomes to program development and progression of breast cancer. METHODS: Stable cell lines overexpressing or knocking-out of ASPH were established using lentivirus transfection or CRISPR-CAS9 systems. Western blot, MTT, immunofluorescence, luciferase reporter, co-immunoprecipitation, 2D/3-D invasion, tube formation, mammosphere formation, immunohistochemistry and newly developed in vitro metastasis were applied. RESULTS: Through physical interactions with Notch receptors, ligands (JAGs) and regulators (ADAM10/17), ASPH activates Notch cascade to provide raw materials (especially MMPs/ADAMs) for synthesis/release of pro-metastatic exosomes. Exosomes orchestrate EMT, 2-D/3-D invasion, stemness, angiogenesis, and premetastatic niche formation. Small molecule inhibitors (SMIs) of ASPH's ß-hydroxylase specifically/efficiently abrogated in vitro metastasis, which mimics basement membrane invasion at primary site, intravasation/extravasation (transendothelial migration), and colonization/outgrowth at distant sites. Multiple organ-metastases in orthotopic and tail vein injection murine models were substantially blocked by a specific SMI. ASPH is silenced in normal adult breast, upregulated from in situ malignancies to highly expressed in invasive/advanced ductal carcinoma. Moderate-high expression of ASPH confers more aggressive molecular subtypes (TNBC or Her2 amplified), early recurrence/progression and devastating outcome (reduced overall/disease-free survival) of breast cancer. Expression profiling of Notch signaling components positively correlates with ASPH expression in breast cancer patients, confirming that ASPH-Notch axis acts functionally in breast tumorigenesis. CONCLUSIONS: ASPH-Notch axis guides particularly selective exosomes to potentiate multifaceted metastasis. ASPH's pro-oncogenic/pro-metastatic properties are essential for breast cancer development/progression, revealing a potential target for therapy.

Anti-tumor activity of antibody drug conjugate targeting aspartate-ß-hydroxylase in pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is an extremely aggressive malignancy with very limited treatment options. Antibody drug conjugates (ADCs) are promising cytotoxic agents capable of highly selective delivery. Aspartate-ß-hydroxylase (ASPH) is a type II transmembrane protein highly expressed in PDACs (97.1%) but not normal pancreas. We investigated anti-tumor effects of an ADC guided by a human monoclonal antibody (SNS-622) against ASPH in human PDAC cell lines and derived subcutaneous (s.c.) xenograft as well as a patient-derived xenograft (PDX) murine model with spontaneous pulmonary metastasis. The cytotoxic effects exhibited by several candidate payloads linked to SNS-622 antibody targeting ASPH+ PDACs were analyzed. After i.v. administration of SNS-622-emtansine (DM1) ADC, the primary PDAC tumor growth and progression (number and size of pulmonary metastases) were determined. The PDAC cell lines, s.c. and PDX tumors treated with ADC were tested for cell proliferation, cytotoxicity and apoptosis by MTS and immunohistochemistry (IHC) assays. SNS-622-DM1 construct has demonstrated optimal anti-tumor effects in vitro. In the PDX model of human PDAC, SNS-622-DM1 ADC exerted substantially inhibitory effects on tumor growth and pulmonary metastasis through attenuating proliferation and promoting apoptosis.

Surveillance and management for serous cystic neoplasms of the pancreas based on total hazards-a multi-center retrospective study from China.

BACKGROUND: Serous cystic neoplasms (SCN) rarely have malignant potential, so accurate diagnosis of SCN is crucial for proper clinical management, especially to avoid unnecessary surgeries. However, the misdiagnosis of other pancreatic cystic neoplasm instead of SCN may highly increase the risk of malignancy in patients who receive no surgery. METHODS: Data from a total of 678 patients with pathologically confirmed to have SCN at sixteen institutions in China from January 1st, 2006 to December 31st, 2016 were retrieved to evaluate the malignancy risk of SCN. RESULTS: Among the 678 patients confirmed to have SCN with postoperative pathologic analysis, 649 patients (95.7%) had only one lesion and the average maximum diameter was 3.8±2.47 cm. Four patients were pathologically verified as having serous cystadenocarcinoma, so the SCN actual malignancy rate was 0.6%, while the mortality due to pancreatic surgery in these high-volume centers was nearly 0.2-2%. However, among the 99 SCN patients based on preoperative radiology, three were confirmed to have intraductal papillary mucinous neoplasms (IPMN), nine as mucinous cystic neoplasms (MCN), and four as solid pseudopapillary tumors (SPT) after postoperative pathological analysis. Thus, the total theoretical malignancy rate resulting from preoperative misdiagnosis was elevated to approximately 2.9%, higher than the risk of perioperative mortality. CONCLUSIONS: When SCN can't be accurately distinguished from cystic tumors of pancreas, the malignant risk of cystic tumors may be higher than perioperative risk. However, if it can be diagnosed as SCN accurately, surgery can be avoided as well.

Aspartate ß-hydroxylase promotes pancreatic ductal adenocarcinoma metastasis through activation of SRC signaling pathway.

BACKGROUND: Signaling pathways critical for embryonic development re-emerge in adult pancreas during tumorigenesis. Aspartate ß-hydroxylase (ASPH) drives embryonic cell motility/invasion in pancreatic development/differentiation. We explored if dysregulated ASPH is critically involved in pancreatic cancer pathogenesis. METHODS: To demonstrate if/how ASPH mediates malignant phenotypes, proliferation, migration, 2-D/3-D invasion, pancreatosphere formation, immunofluorescence, Western blot, co-immunoprecipitation, invadopodia formation/maturation/function, qRT-PCR, immunohistochemistry (IHC), and self-developed in vitro metastasis assays were performed. Patient-derived xenograft (PDX) models of human pancreatic ductal adenocarcinoma (PDAC) were established to illustrate in vivo antitumor effects of the third-generation small molecule inhibitor specifically against ASPH's ß-hydroxylase activity. Prognostic values of ASPH network components were evaluated with Kaplan-Meier plots, log-rank tests, and Cox proportional hazards regression models. RESULTS: ASPH renders pancreatic cancer cells more aggressive phenotypes characterized by epithelial-mesenchymal transition (EMT), 2-D/3-D invasion, invadopodia formation/function as demonstrated by extracellular matrix (ECM) degradation, stemness (cancer stem cell marker upregulation and pancreatosphere formation), transendothelial migration (mimicking intravasation/extravasation), and sphere formation (mimicking metastatic colonization/outgrowth at distant sites). Mechanistically, ASPH activates SRC cascade through direct physical interaction with ADAM12/ADAM15 independent of FAK. The ASPH-SRC axis enables invadopodia construction and initiates MMP-mediated ECM degradation/remodeling as executors for invasiveness. Pharmacologic inhibition of invadopodia attenuates in vitro metastasis. ASPH fosters primary tumor development and pulmonary metastasis in PDX models of PDAC, which is blocked by a leading compound specifically against ASPH enzymatic activity. ASPH is silenced in normal pancreas, progressively upregulated from pre-malignant lesions to invasive/advanced stages of PDAC. Expression profiling of ASPH-SRC network components independently/jointly predicts clinical outcome of PDAC patients. Compared to a negative-low level, a moderate-very high level of ASPH, ADAM12, activated SRC, and MMPs correlated with curtailed overall survival (OS) of pancreatic cancer patients (log-rank test, ps < 0.001). The more unfavorable molecules patients carry, the more deleterious prognosis is destinated. Patients with 0-2 (n = 4), 3-5 (n = 8), 6-8 (n = 24), and 9-12 (n = 73) unfavorable expression scores of the 5 molecules had median survival time of 55.4, 15.9, 9.7, and 5.0 months, respectively (p < 0.001). CONCLUSION: Targeting the ASPH-SRC axis, which is essential for propagating multi-step PDAC metastasis, may specifically/substantially retard development/progression and thus improve prognosis of PDAC.

Recent advances in research on aspartate ß-hydroxylase (ASPH) in pancreatic cancer: A brief update.

Pancreatic cancer (PC) is a highly aggressive tumor, often difficult to diagnose and treat. Aspartate ß-hydroxylase (ASPH) is a type II transmembrane protein and the member of α-ketoglutarate-dependent dioxygenase family, found to be overexpressed in different cancer types, including PC. ASPH appears to be involved in the regulation of proliferation, invasion and metastasis of PC cells through multiple signaling pathways, suggesting its role as a tumor biomarker and therapeutic target. In this review, we briefly summarize the possible mechanisms of action of ASPH in PC and recent progress in the therapeutic approaches targeting ASPH.

RB1CC1-enhanced autophagy facilitates PSCs activation and pancreatic fibrogenesis in chronic pancreatitis.

Chronic pancreatitis (CP) is described as a progressive fibro-inflammatory disorder of the exocrine disease, which eventually leads to damage of the gland. Excessive activation of pancreatic stellate cells (PSCs) is a critical participant in the initiation of CP. Autophagy is involved in multiple degeneration and inflammation in acute pancreatitis and CP. In our study, we report that retinoblastoma coiled coil protein 1 (RB1CC1) expression and the autophagic level are elevated in activated PSCs. RB1CC1 is positively correlated with pancreatic fibrogenesis in tissues and plasma of CP patients. Knockdown of RB1CC1 restrains alpha smooth muscle actin (α-SMA) and collagen expressions, and autophagy in activated PSCs in vitro. Furthermore, we show that RB1CC1 induces PSC activation via binding to ULK1 promoter and the direct interaction with ULK1 protein. These suppress ULK1 expression and its kinase activity. In mice, knockdown of RB1CC1 blocks autophagy and then inhibits the pancreatic duct ligation-induced pancreatic fibrosis. Consequently, our study highlights that RB1CC1-mediated autophagy is a key event for the activation of PSCs. Inhibition of RB1CC1 alleviates autophagy, which plays a critical role in anti-fibrotic activation in PSCs and CP progression. RB1CC1 could be a novel strategy for the treatment of pancreatic fibrosis.

Identification and characterization of a novel neuropeptide (neuropeptide Y-HS) from leech salivary gland of Haemadipsa sylvestris.

The present study was designed to identify immunomodulatory components from the leech salivary gland of Haemadipsa sylvestris. The Sephadex G-50, Resource(TM) S column chromatography and reverse-phase high performance liquid chromatography (RP-HPLC) were used to isolate and purify the salivary gland extracts (SGE). Structural analysis of isolated compounds was based on Edman degradation and matrix assisted laser desorption ionization time-of-flight mass spectrometer (MALDI-TOF-MS). The cDNA encoding the precursor of the compound was cloned from the cDNA library of the salivary gland of H. sylvestris. The levels of inflammatory mediators, including tumor necrosis factor-α (TNF-α), interferon γ (IFN-γ), interleukin-6 (IL-6), and monocyte chemotactic protein-1 (MCP-1) were assayed using an enzyme-linked immunosorbent assay (ELISA). The effects on cell proliferation and cell viability were observed using MTT assay. A novel neuropeptide Y (Neuropeptide Y-HS) from the leech salivary gland of H. sylvestris was purified and characterized. It was composed of 36 amino acid residues and the amino acid sequence was determined to be FLEPPERPAVFTSVEQMKSYIKALNDYYLLLGRPRF-NH2, containing an amidated C-terminus. It showed significant inhibitory effects on the production of inflammatory cytokines including TNF-α, IFN-γ, IL-6, and MCP-1. Neuropeptide Y was identified from leeches for the first time. The presence of neuropeptide Y-HS in leech salivary gland may help get blood meal from hosts and inhibit inflammation.

Long Noncoding RNA MALAT1 Promotes Aggressive Pancreatic Cancer Proliferation and Metastasis via the Stimulation of Autophagy.

Recently, pancreatic ductal adenocarcinoma (PDAC) has emerged as one of the most aggressive malignant tumors with the worst prognosis. Previous studies have demonstrated that long noncoding RNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is increased in pancreatic cancer and is identified as a diagnostic biomarker. Nonetheless, the molecular mechanism of elevated MALAT1 levels and tumor aggressiveness remains unknown. In this study, MALAT1 was found to be highly expressed in PDAC tissues, and elevated expression was associated with poorer prognoses. In addition, MALAT1 was positively linearly correlated with the expression of LC3B mRNA. Furthermore, several molecules involved in cellular autophagic flux were modulated following the downregulation of MALAT1, including LC3, P62, and LAMP-2. Mechanistically, we found that MALAT1 interacted with RNA binding protein HuR, and silencing of MALAT1 greatly enhanced the posttranscriptional regulation of TIA-1 and had further effects on inhibiting autophagy. MALAT1 was speculated to regulate tumorigenesis via HuR-TIA-1-mediated autophagic activation. Hence, we investigated the biological properties of MALAT1 in terms of tumor proliferation and metastasis by promoting autophagy in vitro In brief, these data demonstrate that MALAT1 could facilitate the advanced progression of tumors in vivo Our study highlights the new roles of MALAT1 on protumorigenic functioning and anticancer therapy via activating autophagy in pancreatic cancer. Mol Cancer Ther; 15(9); 2232-43. ©2016 AACR.