The dichotomous role of the glycolytic metabolism pathway in cancer metastasis: Interplay with the complex tumor microenvironment and novel therapeutic strategies.

Cancer metastasis to distant organs is initiated by tumor cells that disseminate from primary heterogeneous tumors. The subsequent growth and survival of tumor metastases depend on different metabolic changes, which constitute one of the enigmatic properties of tumor cells. Aerobic glycolysis, 'the Warburg effect', contributes to tumor energy supply, by oxidizing glucose in a faster manner compared to oxidative phosphorylation, leading to an increased lactate production by lactate dehydrogenase A (LDH-A), which in turn affects the immune response. Surrounding stromal cells contribute to feedback mechanisms further prompting the acquisition of pro-invasive metabolic features. Hence, therapeutic strategies targeting the glycolytic pathway are intensively investigated, with a special interest on their anti-metastatic properties. Various small molecules, such as LDH-A inhibitors, have shown pre-clinical activity against different cancer types, and blocking LDH-A could also help in designing future complimentary therapies. Modulation of specific targets in cells with an altered glycolytic metabolism should indeed result in a milder and distinct toxicity profile, compared to conventional cytotoxic therapy, while a combination treatment with vitamin C leading to increasing reactive oxygen species levels, should further inhibit cancer cell survival and invasion. In this review we describe the impact of metabolic reprogramming in cancer metastasis, the contribution of lactate in this aberrant process and its effect on oncogenic processes. Furthermore, we discuss experimental compounds that target glycolytic metabolism, such as LDH-A inhibitors, and their potential to improve current and experimental therapeutics against metastatic tumors.

Impact of hypoxia on chemoresistance of mesothelioma mediated by the proton-coupled folate transporter, and preclinical activity of new anti-LDH-A compounds.

BACKGROUND: Expression of proton-coupled folate transporter (PCFT) is associated with survival of mesothelioma patients treated with pemetrexed, and is reduced by hypoxia, prompting studies to elucidate their correlation. METHODS: Modulation of glycolytic gene expression was evaluated by PCR arrays in tumour cells and primary cultures growing under hypoxia, in spheroids and after PCFT silencing. Inhibitors of lactate dehydrogenase (LDH-A) were tested in vitro and in vivo. LDH-A expression was determined in tissue microarrays of radically resected malignant pleural mesothelioma (MPM, N = 33) and diffuse peritoneal mesothelioma (DMPM, N = 56) patients. RESULTS: Overexpression of hypoxia marker CAIX was associated with low PCFT expression and decreased MPM cell growth inhibition by pemetrexed. Through integration of PCR arrays in hypoxic cells and spheroids and following PCFT silencing, we identified the upregulation of LDH-A, which correlated with shorter survival of MPM and DMPM patients. Novel LDH-A inhibitors enhanced spheroid disintegration and displayed synergistic effects with pemetrexed in MPM and gemcitabine in DMPM cells. Studies with bioluminescent hypoxic orthotopic and subcutaneous DMPM athymic-mice models revealed the marked antitumour activity of the LDH-A inhibitor NHI-Glc-2, alone or combined with gemcitabine. CONCLUSIONS: This study provides novel insights into hypoxia/PCFT-dependent chemoresistance, unravelling the potential prognostic value of LDH-A, and demonstrating the preclinical activity of LDH-A inhibitors.

Plasma miR-181a-5p Downregulation Predicts Response and Improved Survival After FOLFIRINOX in Pancreatic Ductal Adenocarcinoma.

OBJECTIVE: The aim of the study was to identify plasma microRNA (miRNA) biomarkers for stratifying and monitoring patients with locally advanced or metastatic pancreatic ductal adenocarcinoma (PDAC) treated with FOLFIRINOX, and to investigate their functional roles. SUMMARY BACKGROUND DATA: FOLFIRINOX has become a standard therapy for patients with advanced PDAC and can be used to potentially downstage disease. However, only a subset of patients respond, and biomarkers to guide decision-making are urgently needed. METHODS: We used microarray-based profiling to discover deregulated miRNAs in pre- and postchemotherapy plasma samples from patients based on their progression-free survival (PFS) after FOLFIRINOX. Nine candidate plasma miRNAs were validated in an independent cohort (n = 43). The most discriminative plasma miRNA was correlated with clinicopathological factors and survival, and also investigated in an additional cohort treated with gemcitabine plus nab-paclitaxel. Expression patterns were further evaluated in matched tumor tissues. In vitro studies explored its function, key downstream gene-targets, and interaction with 5-fluorouracil, irinotecan, and oxaliplatin. RESULTS: Plasma miR-181a-5p was significantly downregulated in non-progressive patients after FOLFIRINOX. In multivariate analysis, this decline correlated with improved PFS and overall survival, especially when combined with CA19-9 decline [hazard ratio (HR) = 0.153, 95% confidence interval (CI), 0.067-0.347 and HR = 0.201, 95% CI, 0.070-0.576, respectively]. This combination did not correlate with survival in patients treated with gemcitabine plus nab-paclitaxel. Tissue expression of miR-181a-5p reflected plasma levels. Inhibition of miR-181a-5p coupled with oxaliplatin exposure in pancreatic cell lines decreased cell viability. CONCLUSIONS: Plasma miR-181a-5p is a specific biomarker for monitoring FOLFIRINOX response. Decline in plasma miR-181a-5p and CA19-9 levels is associated with better prognosis after FOLFIRINOX and may be useful for guiding therapeutic choices and surgical exploration.

The occurrence of prion protein in surgically resected pancreatic adenocarcinoma.

BACKGROUND: Among the several new targets for the comprehension of the biology of pancreatic ductal adenocarcinoma (PDAC), Prion proteins (PrPc) deserve particular mention, since they share a marked neurotropism. Actually, PrPc could have also a role in tumorigenesis, as recently demonstrated. However, only few in vitro studies in cell cultures showed the occurrence of PrPc in PDAC cells. We aim to evaluate the presence of PrPc in vivo in PDAC tissues as a potential new biomarker. METHODS: Samples from tumors of 23 patients undergone pancreatic resections from July 2018 to May 2020 at our institution were collected and analyzed. Immunohistochemistry and western blotting of PDAC tissues were compared with control tissues. Immunohistochemistry was used also to evaluate the localization of PrPc and of CD155, a tumoral stem-cell marker. RESULTS: All cases were moderately differentiated PDAC, with perineural invasion (PNI) in 19/23 cases (83%). According to western-blot analysis, PrPc was markedly expressed in PDAC tissues (273.5 ± 44.63 OD) respect to controls (100 ± 28.35 OD, p = 0.0018). Immunohistochemistry confirmed these findings, with higher linear staining of PrPc in PDAC ducts (127.145 ± 7.56 µm vs 75.21 ± 5.01 µm, p < 0.0001). PrPc and CD155 exactly overlapped in ductal tumoral cells, highlighting the possible relationship of PrPc with cancer stemness. Finally, PrPc expression related with cancer stage and there was a potential correspondence with PNI. CONCLUSIONS: Our work provides evidence for increased levels of PrPc in PDAC. This might contribute to cancer aggressiveness and provides a potentially new biomarker. Work is in progress to decipher clinical implications.

Triticum vulgare Extract Modulates Protein-Kinase B and Matrix Metalloproteinases 9 Protein Expression in BV-2 Cells: Bioactivity on Inflammatory Pathway Associated with Molecular Mechanism Wound Healing.

Matrix metalloproteinases (MMPs) are a large family of ubiquitously expressed zinc-dependent enzymes with proteolitic activities. They are expressed in physiological situations and pathological conditions involving inflammatory processes including epithelial to mesenchymal transition (EMT), neuronal injury, and cancer. There is also evidence that MMPs regulate inflammation in tumor microenvironment, which plays an important role in healing tissue processes. Looking at both inflammatory and neuronal damages, MMP9 is involved in both processes and their modulation seems to be regulated by two proteins: tumor necrosis factor-alpha (TNF-alpha) and interleukin 6 (IL-6). However other important genes are involved in molecular regulation of transcription factors, protein-kinase B (AKT), and p65. In addition, Triticum vulgare extract (TVE) modulated the biological markers associated with inflammatory processes, including p65 protein. While there are no evidence that TVE might be involved in the biological modulation of other inflammatory marker as AKT, we would like to assess whether TVE is able to (1) modulate phosphorylation of AKT (pAKT) as an early marker of inflammatory process in vitro and (2) affect MMP9 protein expression in an in vitro model. The BV-2 cells (microglial of mouse) have been used as an in vitro model to simulate both inflammatory and neuronal injury pathologies. Here, MMP9 seems to be involved in cellular migration through inflammatory marker activation. We simulate an inflammatory preclinical model treating BV-2 cells with lipopolysaccharide (LPS) to induce proinflammatory activation affecting pAKT and p65 proteins. TVE is revealed to restore the native expression of AKT and p65. Additionally, TVE extract modulates also the protein concentration of MMP9. Nevertheless, immunofluorescence confocal analyses revealed that both AKT and MMP9 are regulated together, synchronously. This work seems to demonstrate that two important genes can be used to monitor the beginning of an inflammatory process, AKT and MMP9, in which TVE seems able to modulate their expression of inflammation-associated molecules.

Pancreatic serous cystoadenoma (CSA) showing increased tracer uptake at 68-GaDOTA-peptide Positron Emission Tomography (68Ga-DOTA-peptide PET-CT): a case report.

BACKGROUND: Serous cysto-adenoma (SCA) is a rare benign neoplasm of the pancreas. SCA can mimic other pancreatic lesions, such as neuroendocrine tumours. 68Gallium-DOTA-peptide Positron Emission Tomography (PET) is able to image in vivo the over-expression of the somatostatin receptors, playing an important role for the identification of neuroendocrine neoplasms. CASE PRESENTATION: We reported a case of 63-year-old man, with a solid lesion of 7 cm of diameter of the body-tail of the pancreas. Two fine-needle-aspirations (FNA) were inconclusive. A 68Ga-DOTA-peptide PET-CT revealed a pathological uptake of the pancreatic lesion. The diagnosis of a pancreatic neuroendocrine neoplasm was established and a laparoscopic distal splenopancreatectomy and cholecystectomy was performed. Final histopathological report revealed the presence of a micro-cystic SCA. CONCLUSIONS: The current case firstly reports a pancreatic SCA showing increased radiopharmaceutical uptake at 68Ga-DOTA-peptide PET-CT images. This unexpected finding should be taken into account during the diagnostic algorithm of a pancreatic lesion, in order to minimize the risk of misdiagnosis and overtreatment of SCA.

3-(6-Phenylimidazo [2,1-b][1,3,4]thiadiazol-2-yl)-1H-Indole Derivatives as New Anticancer Agents in the Treatment of Pancreatic Ductal Adenocarcinoma.

A new series of imidazo[2,1-b][1,3,4]thiadiazole derivatives was efficiently synthesized and screened for their in vitro antiproliferative activity on a panel of pancreatic ductal adenocarcinoma (PDAC) cells, including SUIT-2, Capan-1 and Panc-1. Compounds 9c and 9l, showed relevant in vitro antiproliferative activity on all three pre-clinical models with half maximal inhibitory concentration (IC50) ranging from 5.11 to 10.8 µM, while the compounds 9e and 9n were active in at least one cell line. In addition, compound 9c significantly inhibited the migration rate of SUIT-2 and Capan-1 cells in the scratch wound-healing assay. In conclusion, our results will support further studies to increase the library of imidazo [2,1-b][1,3,4] thiadiazole derivatives for deeper understanding of the relationship between biological activity of the compounds and their structures in the development of new antitumor compounds against pancreatic diseases.

Robotic-assisted versus open left pancreatectomy for cystic tumours: A single-centre experience.

BACKGROUND: Cystic pancreatic lesions (CPLs) are being identified increasingly, and some benefit from surgical treatment. With the increasing use of robotic-assisted surgery (RAS) for neoplasms of the pancreas, the aim of the present comparative study is to establish whether the RAS offered any advantages over conventional open surgery (OS) in the management of CPLs. PATIENTS AND METHODS: Twenty-seven out of 37 robot-assisted left-sided pancreatectomy (LSP) performed between January 2010 and April 2017 were carried out for CPLs. The surgical outcome and histopathology were compared retrospectively with a control group of 27 patients who had undergone open LSP for CPLs, selected using a one-to-one case-matched methodology (OS-Group) from the prospectively collected institutional database. RESULTS: The spleen was preserved in a significantly higher percentage of patients in the RAS-group (63% vs. 33.3%,P < 0.05). There was no difference in the post-operative course (pancreatic fistula and morbidity) between the two groups. The median post-operative hospital stay was significantly shorter in the RAS-group: 8 days (range 3-25) versus 12 days (range 7-26) in the OS-group (P < 0.01). No conversion to open approach was reported in the RAS-group. CONCLUSIONS: Robotically assisted LSP is a safe and effective procedure. It is accompanied by a significantly higher spleen preservation rate compared to the open approach. In addition, because of the reduced trauma, RAS incurred a shorter post-operative hospital stay and faster return to full recovery, particularly important in patients undergoing surgery for relative indications. However, these benefits of RAS for LSP require confirmation by prospective randomised controlled studies.

Genetic determinants of telomere length and risk of pancreatic cancer: A PANDoRA study.

Telomere deregulation is a hallmark of cancer. Telomere length measured in lymphocytes (LTL) has been shown to be a risk marker for several cancers. For pancreatic ductal adenocarcinoma (PDAC) consensus is lacking whether risk is associated with long or short telomeres. Mendelian randomization approaches have shown that a score built from SNPs associated with LTL could be used as a robust risk marker. We explored this approach in a large scale study within the PANcreatic Disease ReseArch (PANDoRA) consortium. We analyzed 10 SNPs (ZNF676-rs409627, TERT-rs2736100, CTC1-rs3027234, DHX35-rs6028466, PXK-rs6772228, NAF1-rs7675998, ZNF208-rs8105767, OBFC1-rs9420907, ACYP2-rs11125529 and TERC-rs10936599) alone and combined in a LTL genetic score ("teloscore", which explains 2.2% of the telomere variability) in relation to PDAC risk in 2,374 cases and 4,326 controls. We identified several associations with PDAC risk, among which the strongest were with the TERT-rs2736100 SNP (OR = 1.54; 95%CI 1.35-1.76; p = 1.54 × 10-10 ) and a novel one with the NAF1-rs7675998 SNP (OR = 0.80; 95%CI 0.73-0.88; p = 1.87 × 10-6 , ptrend = 3.27 × 10-7 ). The association of short LTL, measured by the teloscore, with PDAC risk reached genome-wide significance (p = 2.98 × 10-9 for highest vs. lowest quintile; p = 1.82 × 10-10 as a continuous variable). In conclusion, we present a novel genome-wide candidate SNP for PDAC risk (TERT-rs2736100), a completely new signal (NAF1-rs7675998) approaching genome-wide significance and we report a strong association between the teloscore and risk of pancreatic cancer, suggesting that telomeres are a potential risk factor for pancreatic cancer.

A propensity score-matched analysis of robotic versus open pancreatoduodenectomy for pancreatic cancer based on margin status.

BACKGROUND: No study has shown the oncologic non-inferiority of robotic pancreatoduodenectomy (RPD) versus open pancreatoduodenectomy (OPD) for pancreatic cancer (PC). METHODS: This is a single institution propensity score matched study comparing RPD and ODP for resectable PC, based on factors predictive of R1 resection (≤ 1 mm). Only patients operated on after completion of the learning curve in both procedures and for whom circumferential margins were assessed according to the Leeds pathology protocol were included. The primary study endpoint was the rate of R1 resection. Secondary study endpoints were as follows: number of examined lymph nodes (N), rate of perioperative transfusions, percentage of patients receiving adjuvant therapies, occurrence of local recurrence, overall survival, disease-free survival, and sample size calculation for randomized controlled trials (RCT). RESULTS: Factors associated with R1 resection were tumor diameter, number of positive N, N ratio, logarithm odds of positive N, and duodenal infiltration. The matching process identified 20 RPDs and 24 OPDs. All RPDs were completed robotically. R1 resection was identified in 11 RPDs (55.0%) and in 10 OPDs (41.7%) (p = 0.38). There was no difference in the rate of R1 at each margin as well as in the proportion of patients with multiple R1 margins. RPD and OPD were also equivalent with respect to all secondary study endpoints, with a trend towards lower rate of blood transfusions in RPD. Based on the figures presented herein, a non-inferiority RCT comparing RPD and OPD having the rate of R1 resection as the primary study endpoint requires 3355 pairs. CONCLUSIONS: RPD and OPD achieved the same rate of R1 resections in resectable PC. RPD was also non-inferior to OPD with respect to all secondary study endpoints. Because of the high number of patients required to run a RCT, further assessment of RPD for PC would require the implementation of an international registry.

Common genetic variants associated with pancreatic adenocarcinoma may also modify risk of pancreatic neuroendocrine neoplasms.

Pancreatic neuroendocrine neoplasms (pNEN) account for less than 5% of all pancreatic neoplasms and genetic association studies on susceptibility to the disease are limited. We sought to identify possible overlap of genetic susceptibility loci between pancreatic ductal adenocarcinoma (PDAC) and pNEN; therefore, PDAC susceptibility variants (n = 23) from Caucasian genome-wide association studies (GWAS) were genotyped in 369 pNEN cases and 3277 controls from the PANcreatic Disease ReseArch (PANDoRA) consortium to evaluate the odds associated with pNEN risk, disease onset and tumor characteristics. Main effect analyses showed four PDAC susceptibility variants-rs9854771, rs1561927, rs9543325 and rs10919791 to be associated with pNEN risk. Subsequently, only associations with rs9543325, rs10919791 and rs1561927 were noteworthy with false positive report probability (FPRP) tests. Stratified analyses considering age at onset (50-year threshold), showed rs2736098, rs16986825 and rs9854771 to be associated with risk of developing pNEN at a younger age. Stratified analyses also showed some single nucleotide polymorphisms to be associated with different degrees of tumor grade, metastatic potential and functionality. Our results identify known GWAS PDAC susceptibility loci, which may also be involved in sporadic pNEN etiology and suggest that some genetic mechanisms governing pathogenesis of these two entities may be similar, with few of these loci being more influential in younger cases or tumor subtypes.

Myoclonus epilepsy, retinitis pigmentosa, leukoencephalopathy and cerebral calcifications associated with a novel m.5513G>A mutation in the MT-TW gene.

We sequenced the mitochondrial genome from a 40-year-old woman with myoclonus epilepsy, retinitis pigmentosa, leukoencephalopathy and cerebral calcifications. Histological and biochemical features of mitochondrial respiratory chain dysfunction were present. Direct sequencing showed a novel heteroplasmic mutation at nucleotide 5513 in the MT-TW gene that encodes tRNATrp. Restriction Fragment Length Polymorphism analysis confirmed that about 80% of muscle mtDNA harboured the mutation while it was present in minor percentages in mtDNA from other tissues. The mutation is predicted to disrupt a highly conserved base pair within the aminoacyl acceptor stem of the tRNA. This is the 17° mutation in MT-TW gene and expands the known causes of late-onset mitochondrial diseases.

Association of genetic polymorphisms with survival of pancreatic ductal adenocarcinoma patients.

Germline genetic variability might contribute, at least partially, to the survival of pancreatic ductal adenocarcinoma (PDAC) patients. Two recently performed genome-wide association studies (GWAS) on PDAC overall survival (OS) suggested (P < 10(-5)) the association between 30 genomic regions and PDAC OS. With the aim to highlight the true associations within these regions, we analyzed 44 single-nucleotide polymorphisms (SNPs) in the 30 candidate regions in 1722 PDAC patients within the PANcreatic Disease ReseArch (PANDoRA) consortium. We observed statistically significant associations for five of the selected regions. One association in the CTNNA2 gene on chromosome 2p12 [rs1567532, hazard ratio (HR) = 1.75, 95% confidence interval (CI) 1.19-2.58, P = 0.005 for homozygotes for the minor allele] and one in the last intron of the RUNX2 gene on chromosome 6p21 (rs12209785, HR = 0.88, 95% CI 0.80-0.98, P = 0.014 for heterozygotes) are of particular relevance. These loci do not coincide with those that showed the strongest associations in the previous GWAS. In silico analysis strongly suggested a possible mechanistic link between these two SNPs and pancreatic cancer survival. Functional studies are warranted to confirm the link between these genes (or other genes mapping in those regions) and PDAC prognosis in order to understand whether these variants may have the potential to impact treatment decisions and design of clinical trials.

Integrated molecular analysis to investigate the role of microRNAs in pancreatic tumour growth and progression.

BACKGROUND: MicroRNAs (miRNAs) are small non-coding RNAs involved in the post-transcriptional regulation of mRNAs and are aberrantly expressed in cancer with important roles in tumorigenesis. A broad analysis of the combined effects of altered activities of miRNAs in pancreatic ductal adenocarcinoma (PDAC) has not been done, and how miRNAs might affect tumour progression or patient outcomes is unclear. METHODS: We combined data from miRNA and mRNA expression profiles from PDAC and normal pancreas samples (each n=9) and used bioinformatic analyses to identify a miRNA-mRNA regulatory network in PDAC. We validated our findings in PDAC cell-lines (PANC-1, MIA PaCa-2, LPc006, and LPc167), subcutaneous PDAC xenografts in mice, and laser capture microdissected PDACs from patients (n=91). We used this information to identify miRNAs that contributed most to tumorigenesis. FINDINGS: We identified three miRNAs (miR-21, miR-23a, and miR-27a) that acted as cooperative repressors of a network of tumour suppressor genes that included PDCD4, BTG2, and NEDD4L. Inhibition of miR-21, miR-23a, and miR-27a had synergistic effects in reducing proliferation of PDAC cells in culture and the growth of xenograft tumours. The level of inhibition was greater than that of silencing oncomiR-21 alone. In PDACs from patients, high levels of the combination of miR-21, miR-23a, and miR-27a was a strong independent predictor of short overall survival after surgical resection (hazard ratio 3·21, 95% CI 1·78-5·78). High expression of this combination was also associated with a more aggressive tumour phenotype: more microscopic tumour infiltration at resection margin and increased perineural invasion. INTERPRETATION: In an integrated data analysis, we identified functional miRNA-mRNA interactions that contribute to PDAC growth. These findings indicate that miRNAs act together to promote tumour progression and that future therapeutic strategies might require inhibition of several miRNAs. Furthermore, high tumour expression of the miR-21, miR-23a, and miR-27a combination could have potential use in the future as a prognostic signature for patients with PDAC. FUNDING: Peel Medical Research Trust, Alliance Family Foundation, Action Against Cancer, National Institute for Health Research, Association for International Cancer Research, Jason Boas Fellowship, Imperial Biomedical Research Centre, Rosetrees Trust, Joseph Ettedgui Charitable Foundation.

Robotic pancreatoduodenectomy with vascular resection.

PURPOSE: This study aims to define the current status of robotic pancreatoduodenectomy (RPD) with resection and reconstruction of the superior mesenteric/portal vein (RPD-SMV/PV). METHODS: Our experience on RPD, including RPD-SMV/PV, is presented along with a description of the surgical technique and a systematic review of the literature on RPD-SMV/PV. RESULTS: We have performed 116 RPD and 14 RPD-SMV/PV. Seven additional cases of RPD-SMV/PV were identified in the literature. In our experience, RPD and RPD-SMV/PV were similar in all baseline variables, but lower mean body mass and higher prevalence of pancreatic cancer in RPD-SMV/PV. Regarding the type of vein resection, there were one type 2 (7.1 %), five type 3 (35.7 %) and eight type 4 (57.2 %) resections. As compared to RPD, RPD-SMV/PV required longer operative time, had higher median estimated blood loss, and blood transfusions were required more frequently. Incidence and severity of post-operative complications were not increased in RPD-SMV/PV, but post-pancreatectomy hemorrhage occurred more frequently after this procedure. In pancreatic cancer, RPD-SMV/PV was associated with a higher mean number of examined lymph nodes (60.0 ± 13.9 vs 44.6 ± 11.0; p = 0.02) and with the same rate of microscopic margin positivity (25.0 % vs 26.1 %). Mean length or resected vein was 23.1 ± 8.08 mm. Actual tumour infiltration was discovered in ten patients (71.4 %), reaching the adventitia in four patients (40.0 %), the media in two patients (20.0 %), and the intima in four patients (40.0 %). Literature review identified seven additional cases, all reported to have successful outcome. CONCLUSIONS: RPD-SMV/PV is feasible in carefully selected patients. The generalization of these results remains to be demonstrated.

TERT gene harbors multiple variants associated with pancreatic cancer susceptibility.

A small number of common susceptibility loci have been identified for pancreatic cancer, one of which is marked by rs401681 in the TERT-CLPTM1L gene region on chromosome 5p15.33. Because this region is characterized by low linkage disequilibrium, we sought to identify whether additional single nucleotide polymorphisms (SNPs) could be related to pancreatic cancer risk, independently of rs401681. We performed an in-depth analysis of genetic variability of the telomerase reverse transcriptase (TERT) and the telomerase RNA component (TERC) genes, in 5,550 subjects with pancreatic cancer and 7,585 controls from the PANcreatic Disease ReseArch (PANDoRA) and the PanScan consortia. We identified a significant association between a variant in TERT and pancreatic cancer risk (rs2853677, odds ratio = 0.85; 95% confidence interval = 0.80-0.90, p = 8.3 × 10(-8)). Additional analysis adjusting rs2853677 for rs401681 indicated that the two SNPs are independently associated with pancreatic cancer risk, as suggested by the low linkage disequilibrium between them (r(2) = 0.07, D' = 0.28). Three additional SNPs in TERT reached statistical significance after correction for multiple testing: rs2736100 (p = 3.0 × 10(-5) ), rs4583925 (p = 4.0 × 10(-5) ) and rs2735948 (p = 5.0 × 10(-5) ). In conclusion, we confirmed that the TERT locus is associated with pancreatic cancer risk, possibly through several independent variants.

MicroRNAs cooperatively inhibit a network of tumor suppressor genes to promote pancreatic tumor growth and progression.

BACKGROUND & AIMS: There has not been a broad analysis of the combined effects of altered activities of microRNAs (miRNAs) in pancreatic ductal adenocarcinoma (PDAC) cells, and it is unclear how these might affect tumor progression or patient outcomes. METHODS: We combined data from miRNA and messenger RNA (mRNA) expression profiles and bioinformatic analyses to identify an miRNA-mRNA regulatory network in PDAC cell lines (PANC-1 and MIA PaCa-2) and in PDAC samples from patients. We used this information to identify miRNAs that contribute most to tumorigenesis. RESULTS: We identified 3 miRNAs (MIR21, MIR23A, and MIR27A) that acted as cooperative repressors of a network of tumor suppressor genes that included PDCD4, BTG2, and NEDD4L. Inhibition of MIR21, MIR23A, and MIR27A had synergistic effects in reducing proliferation of PDAC cells in culture and growth of xenograft tumors in mice. The level of inhibition was greater than that of inhibition of MIR21 alone. In 91 PDAC samples from patients, high levels of a combination of MIR21, MIR23A, and MIR27A were associated with shorter survival times after surgical resection. CONCLUSIONS: In an integrated data analysis, we identified functional miRNA-mRNA interactions that contribute to growth of PDACs. These findings indicate that miRNAs act together to promote tumor progression; therapeutic strategies might require inhibition of several miRNAs.

Contrast enhancement pattern on multidetector CT predicts malignancy in pancreatic endocrine tumours.

OBJECTIVES: Preoperative suspicion of malignancy in pancreatic neuroendocrine tumours (pNETs) is mostly based on tumour size. We retrospectively reviewed the contrast enhancement pattern (CEP) of a series of pNETs on multiphasic multidetector computed tomography (MDCT), to identify further imaging features predictive of lesion aggressiveness. METHODS: Sixty pNETs, diagnosed in 52 patients, were classified based on CEP as: type A showing early contrast enhancement and rapid wash-out; type B presenting even (B1) or only (B2) late enhancement. All tumours were resected allowing pathologic correlations. RESULTS: Nineteen pNETs showed type A CEP (5-20 mm), 29 type B1 CEP (5-80 mm) and 12 type B2 (15-100 mm). All tumours were classified as well differentiated tumours, 19 were benign (WDt-b), 15 with uncertain behaviour (WDt-u) and 26 carcinomas (WDC). None of A lesions were malignant (12 WDt-b; 7 WDt-u), all B2 lesions were WDC, 7 B1 lesions were WDt-b, 8 WDt-u and 14 WDC; 4/34 (12 %) lesions ≤2cm were WDC. CEP showed correlation with all histological prognostic indicators. CONCLUSIONS: Correlating with the lesion grading and other histological prognostic predictors, CEP may preoperatively suggest the behaviour of pNETs, assisting decisions about treatment. Moreover CEP allows recognition of malignant small tumours, incorrectly classified on the basis of their dimension.

Expression of SP7, RUNX1, DLX5, and CTNNB1 in human mesenchymal stem cells cultured on xenogeneic bone substitute as compared with machined titanium.

PURPOSE: The aim of this research was to investigate the gene expression profile of 4 transcription factors in human mesenchymal stem cells (hMSC) cultured with a xenogeneic bone substitute and a support of machined titanium. MATERIAL AND METHODS: In vitro studies were performed on hMSC cells, which grew in contact with cortical porcine bone and machined titanium disks for 10 days. RNA quantification for genes DLX5, CTNNB1, RUNX1, and SP7 was assessed by quantitative real-time polymerase chain reaction. For cells supported by titanium, immunocytochemistry of osteocalcin (OC) was also performed. RESULTS: In the osteoblast-induced cells (OIC), DLX5, CTNNB1, and RUNX1 were significantly upregulated (+2.38-, +3.51-, and +7.08-fold, respectively), whereas SP7 was downregulated (-26.32-fold). None of the genes seemed to be upregulated or downregulated by the corticocancellous porcine bone. In cells grown on titanium support, DLX5 and RUNX1 were respectively upregulated (+3.12-fold) and downregulated (-2.14-fold). For titanium support, the presence of both catenin beta-1 and OC was verified. CONCLUSION: The 2 genes RUNX1 and SP7 resulted differently expressed in cells cultured on metallic supports if compared with the expression recorded for OIC. An induction of the osteogenic phenotype was observed when cells were cultured on machined titanium, but not on xenogeneic material.