Microbiome Dysbiosis with Enterococcus Presence in the Upper Gastrointestinal Tract is A Risk Factor for Mortality in Patients Undergoing Surgery for Pancreatic Cancer.

BACKGROUND: Recent retrospective studies suggest a role for distinct microbiota in the perioperative morbidity and mortality of pancreatic head resections. OBJECTIVE: We aimed to prospectively investigate the microbial colonization of critical operative sites of pancreatic head resections to identify microbial stratification factors for surgical and long-term oncologic outcomes. METHODS: Prospective biomarker study applying 16S rRNA sequencing and microbial culturing to samples collected from various sites of the GI tract and surgical sites of patients during pancreatic head resections at a German single high-volume pancreatic center. RESULTS: A total of 101 patients were included (38 non-cancer, 63 cancer patients [50 PDAC patients]) in the study. In a first data analysis series, 16S rRNA sequencing data were utilized from 96 patients to assess associations of microbiome profiles with clinical parameters and outcomes. In general, microbiome composition varied according to sampling site, cancer, age or preoperative ERCP intervention, notably for the bile microbiome. In the PDAC subcohort, compositional variance of the bile or periampullary microbiome was significantly associated with postoperative complications such as ICU admission; on a taxonomic level we observed Enterococcus spp. to be significantly more abundant in patients developing deep or organ-space surgical site infections (SSI). Elevated Enterococcus relative abundances in the upper GI tract, in turn, were associated with 6-months mortality rates. In a second step, we focused on microbiological cultures collected from bile aspirates during surgery and investigated associations with perioperative complications and long-term survival. Notably, Enterococcus spp. were among the most prevalent pathobiont isolates observed in cancer patient bile specimens that were associated with severe SSIs, and thereby elevated mortality rates up to 24 months. Clinically relevant postoperative pancreatic fistulas or severe SSI were found as other major variables determining short-term mortality in this cancer patient cohort. In the context of adverse microbiological factors, a preoperative ERCP was also observed to segregate long-term survival, and it appeared to interact with the presence of Enterococcus spp. as highest mortality rates were observed in PDAC patients with both preoperative ERCP and presence of E. faecalis in bile aspirates. CONCLUSIONS: The presence of Enterococcus spp. in bile ducts of PDAC patients undergoing pancreatic surgery represents a significant risk factor for perioperative infections and, thereby, elevated postoperative and long-term mortality. This finding supports previous data on the use of the antibiotic drug piperacillin-tazobactam as appropriate perioperative antibiotic prophylaxis for preventing adverse outcomes after pancreatoduodenectomy.

PPARG activation promotes the proliferation of colorectal cancer cell lines and enhances the antiproliferative effect of 5-fluorouracil.

BACKGROUND: Peroxisome proliferator-activated receptor gamma (PPARG) is a member of the nuclear receptor family. It is involved in the regulation of adipogenesis, lipid metabolism, insulin sensitivity, vascular homeostasis and inflammation. In addition, PPARG agonists, known as thiazolidinediones, are well established in the treatment of type 2 diabetes mellitus. PPARGs role in cancer is a matter of debate, as pro- and anti-tumour properties have been described in various tumour entities. Currently, the specific role of PPARG in patients with colorectal cancer (CRC) is not fully understood. MATERIAL AND METHODS: The prognostic impact of PPARG expression was investigated by immunohistochemistry in a case-control study using a matched pair selection of CRC tumours (n = 246) with either distant metastases to the liver (n = 82), lung (n = 82) or without distant metastases (n = 82). Its effect on proliferation as well as the sensitivity to the chemotherapeutic drug 5-fluorouracil (5-FU) was examined after activation, inhibition, and transient gene knockdown of PPARG in the CRC cell lines SW403 and HT29. RESULTS: High PPARG expression was significantly associated with pulmonary metastasis (p = 0.019). Patients without distant metastases had a significantly longer overall survival with low PPARG expression in their tumours compared to patients with high PPARG expression (p = 0.045). In the pulmonary metastasis cohort instead, a trend towards longer survival was observed for patients with high PPARG expression in their tumour (p = 0.059). Activation of PPARG by pioglitazone and rosiglitazone resulted in a significant dose-dependent increase in proliferation of CRC cell lines. Inhibition of PPARG by its specific inhibitor GW9662 and siRNA-mediated knockdown of PPARG significantly decreased proliferation. Activating PPARG significantly increased the CRC cell lines sensitivity to 5-FU while its inhibition decreased it. CONCLUSION: The prognostic effect of PPARG expression depends on the metastasis localization in advanced CRC patients. Activation of PPARG increased malignancy associated traits such as proliferation in CRC cell lines but also increases sensitivity towards the chemotherapeutic agent 5-FU. Based on this finding, a combination therapy of PPARG agonists and 5-FU-based chemotherapy constitutes a promising strategy which should be further investigated.

TPX2 expression as a negative predictor of gemcitabine efficacy in pancreatic cancer.

BACKGROUND: Targeting protein for Xenopus kinesin-like protein 2 (TPX2) overexpression in human tumours is associated with increased malignancy. Its effect on gemcitabine resistance in pancreatic ductal adenocarcinoma (PDAC) has not been studied yet. METHODS: The prognostic impact of TPX2 expression was examined in the tumour tissue of 139 patients with advanced PDAC (aPDAC) treated within the AIO-PK0104 trial or translational trials and of 400 resected PDAC (rPDAC) patients. The findings were validated using RNAseq data of 149 resected PDAC patients. RESULTS: In the aPDAC cohorts, 13.7% of all samples showed high TPX2 expression, conferring significantly shorter progression-free survival (PFS, HR 5.25, P < 0.001) and overall survival times (OS, HR 4.36, P < 0.001) restricted to gemcitabine-based treated patients (n = 99). In the rPDAC cohort, 14.5% of all samples showed high TPX2 expression, conferring significantly shorter disease-free survival times (DFS, HR 2.56, P < 0.001) and OS times (HR 1.56, P = 0.04) restricted to patients treated with adjuvant gemcitabine. RNAseq data from the validation cohort confirmed the findings. CONCLUSIONS: High TPX2 expression may serve as a negative predictor of gemcitabine-based palliative and adjuvant chemotherapy in PDAC and could be used to inform clinical therapy decisions. CLINICAL TRIAL REGISTRY: The clinical trial registry identifier is NCT00440167.

The impact of adjuvant therapy on outcome in UICC stage I pancreatic cancer.

Adjuvant chemotherapy has become standard of care for pancreatic ductal adenocarcinoma (PDAC) as it improves patient outcome. However, its clinical meaning in early-stage, UICC I tumors remains uncertain. We examined the effect of adjuvant therapy on disease-free survival (DFS) and overall survival (OS) of UICC stage I PDAC patients treated at an academic tertiary care center between 2000 and 2016. Among 124 patients (69 male, 55 female; median age 68 years, range 41-84 years) with UICC stage I disease, adjuvant therapy improved both DFS (19.8 vs 12.8 months, HR 0.59, 95% CI: 0.37-0.94, P = .03) and OS (40.9 vs 20.3 months, HR 0.54, 95% CI: 0.35-0.84, P = .005). Multivariate analyses and propensity score matching confirmed the prognostic impact of adjuvant therapy independent of localization, differentiation and R-status. Thus, every patient with UICC I PDAC should receive adjuvant chemotherapy as it may improve outcome significantly. Our findings support the concept of PDAC as systemic disease from early stages on.

Bcl-xL as prognostic marker and potential therapeutic target in cholangiocarcinoma.

Intrahepatic, perihilar, and distal cholangiocarcinoma (iCCA, pCCA, dCCA) are highly malignant tumours with increasing mortality rates due to therapy resistances. Among the mechanisms mediating resistance, overexpression of anti-apoptotic Bcl-2 proteins (Bcl-2, Bcl-xL , Mcl-1) is particularly important. In this study, we investigated whether antiapoptotic protein patterns are prognostically relevant and potential therapeutic targets in CCA. Bcl-2 proteins were analysed in a pan-cancer cohort from the NCT/DKFZ/DKTK MASTER registry trial (n = 1140, CCA n = 72) via RNA-sequencing and transcriptome-based protein activity interference revealing high ranks of CCA for Bcl-xL and Mcl-1. Expression of Bcl-xL , Mcl-1, and Bcl-2 was assessed in human CCA tissue and cell lines compared with cholangiocytes by immunohistochemistry, immunoblotting, and quantitative-RT-PCR. Immunohistochemistry confirmed the upregulation of Bcl-xL and Mcl-1 in iCCA tissues. Cell death of CCA cell lines upon treatment with specific small molecule inhibitors of Bcl-xL (Wehi-539), of Mcl-1 (S63845), and Bcl-2 (ABT-199), either alone, in combination with each other or together with chemotherapeutics was assessed by flow cytometry. Targeting Bcl-xL induced cell death and augmented the effect of chemotherapy in CCA cells. Combined inhibition of Bcl-xL and Mcl-1 led to a synergistic increase in cell death in CCA cell lines. Correlation between Bcl-2 protein expression and survival was analysed within three independent patient cohorts from cancer centers in Germany comprising 656 CCA cases indicating a prognostic value of Bcl-xL in CCA depending on the CCA subtype. Collectively, these observations identify Bcl-xL as a key protein in cell death resistance of CCA and may pave the way for clinical application.

Protein Kinase D1, Reduced in Human Pancreatic Tumors, Increases Secretion of Small Extracellular Vesicles From Cancer Cells That Promote Metastasis to Lung in Mice.

BACKGROUND & AIMS: Pancreatic tumor cells release small extracellular vesicles (sEVs, exosomes) that contain lipids and proteins, RNA, and DNA molecules that might promote formation of metastases. It is not clear what cargo these vesicles contain and how they are released. Protein kinase D1 (PRKD1) inhibits cell motility and is believed to be dysregulated in pancreatic ductal adenocarcinomas. We investigated whether it regulates production of sEVs in pancreatic cancer cells and their ability to form premetastatic niches for pancreatic cancer cells in mice. METHODS: We analyzed data from UALCAN and human pancreatic tissue microarrays to compare levels of PRKD1 between tumor and nontumor tissues. We studied mice with pancreas-specific disruption of Prkd1 (PRKD1KO mice), mice that express oncogenic KRAS (KC mice), and KC mice with disruption of Prkd1 (PRKD1KO-KC mice). Subcutaneous xenograft tumors were grown in NSG mice from Panc1 cells; some mice were then given injections of sEVs. Pancreata and lung tissues from mice were analyzed by histology, immunohistochemistry, and/or quantitative polymerase chain reaction; we performed nanoparticle tracking analysis of plasma sEVs. The Prkd1 gene was disrupted in Panc1 cells using CRISPR-Cas9 or knocked down with small hairpin RNAs, or PRKD1 activity was inhibited with the selective inhibitor CRT0066101. Pancreatic cancer cell lines were analyzed by gene-expression microarray, quantitative polymerase chain reaction, immunoblot, and immunofluorescence analyses. sEVs secreted by Panc1 cell lines were analyzed by flow cytometry, transmission electron microscopy, and mass spectrometry. RESULTS: Levels of PRKD1 were reduced in human pancreatic ductal adenocarcinoma tissues compared with nontumor tissues. PRKD1KO-KC mice developed more pancreatic intraepithelial neoplasia, at a faster rate, than KC mice, and had more lung metastases and significantly shorter average survival time. Serum from PRKD1KO-KC mice had increased levels of sEVs compared with KC mice. Pancreatic cancer cells with loss or inhibition of PRKD1 increased secretion of sEVs; loss of PRKD1 reduced phosphorylation of its substrate, cortactin, resulting in increased F-actin levels at the plasma membrane. sEVs from cells with loss or reduced expression of PRKD1 had altered content, and injection of these sEVs into mice increased metastasis of xenograft tumors to lung, compared with sEVs from pancreatic cells that expressed PRKD1. PRKD1-deficient pancreatic cancer cells showed increased loading of integrin α6ß4 into sEVs-a process that required CD82. CONCLUSIONS: Human pancreatic ductal adenocarcinoma has reduced levels of PRKD1 compared with nontumor pancreatic tissues. Loss of PRKD1 results in reduced phosphorylation of cortactin in pancreatic cancer cell lines, resulting in increased in F-actin at the plasma membrane and increased release of sEVs, with altered content. These sEVs promote metastasis of xenograft and pancreatic tumors to lung in mice.

Impact of previous transurethral prostate surgery on health-related quality of life after radical prostatectomy: Does the interval between surgeries matter?

PURPOSE: To assess the impact of previous transurethral surgery for benign prostate enlargement (BPE) and time interval between procedures on functional outcomes and health-related quality of life (HRQOL) after radical prostatectomy (RP). METHODS: A propensity score-matched patient cohort [n = 685, (513 without previous BPE surgery, 172 with BPE surgery)] was created and HRQOL was pre- and postoperatively assessed using validated questionnaires (EORTC QLQ-C30). Urinary continence was measured via ICIQ-SF questionnaire and pad usage. Multivariable analysis included binary logistic and Cox regression models (p < 0.05). RESULTS: Median follow-up was 18 months. There was no significant difference in recurrence-free survival in multivariate analysis (HR 0.66, 95%CI 0.40-1.07, p = 0.093). We observe higher mean ICIQ-SF scores (5.7 vs. 8.2, p < 0.001) and daily pad usage (1.3 vs. 2.5, p < 0.001), and decreased continence recovery (OR 0.46, 95%CI 0.30-0.71, p < 0.001) for patients with BPE surgery. Postoperative general HRQOL scores were significantly lower for patients with previous BPE surgery (70.6 vs. 63.4, p = 0.003). In multivariate analysis, continence recovery (OR 5.19, 95%CI 3.10-8.68, p < 0.001) but not previous BPE surgery (0.94, 0.57-1.54, p = 0.806) could be identified as independent predictors of good general HRQOL. There was no significant correlation between time interval between both surgeries and continence (p = 0.408), and HRQOL (p = 0.386) outcomes. CONCLUSIONS: We observe favourable continence outcomes for patients without previous BPE surgery. Our results indicate that RP can be safely performed after transurethral BPE surgery, regardless of the time interval between both interventions.

NGS-guided precision oncology in metastatic breast and gynecological cancer: first experiences at the CCC Munich LMU.

PURPOSE: Comprehensive genomic profiling identifying actionable molecular alterations aims to enable personalized treatment for cancer patients. The purpose of this analysis was to retrospectively assess the impact of personalized recommendations made by a multidisciplinary tumor board (MTB) on the outcome of patients with breast or gynecological cancers, who had progressed under standard treatment. Here, first experiences of our Comprehensive Cancer Center Molecular Tumor Board are reported. METHODS: All patients were part of a prospective local registry. 95 patients diagnosed with metastatic breast cancer or gynecological malignancies underwent extended molecular profiling. From May 2017 through March 2019, the MTB reviewed all clinical cases considering tumor profile and evaluated molecular alterations regarding further diagnostic and therapeutic recommendations. RESULTS: 95 patients with metastatic breast or gynecological cancers were discussed in the MTB (68% breast cancer, 20% ovarian cancer, 5% cervical cancer, 3% endometrial cancer and 4% others). Genes with highest mutation rate were PIK3CA and ERBB2. Overall, 34 patients (36%) received a biomarker-based targeted therapy recommendation. Therapeutic recommendations were implemented in nine cases; four patients experienced clinical benefit with a partial response or disease stabilization lasting over 4 months. CONCLUSION: In the setting of a multidisciplinary molecular tumor board, a small but clinically meaningful group of breast and gynecological cancer patients benefits from comprehensive genomic profiling. Broad and successful implementation of precision medicine is complicated by patient referral at late stage disease and limited access to targeted agents and early clinical trials. TRIAL REGISTRATION NUMBER: 284-10 (03.05.2018).

Bacterial lipopolysaccharide as negative predictor of gemcitabine efficacy in advanced pancreatic cancer - translational results from the AIO-PK0104 Phase 3 study.

BACKGROUND: Gram-negative bacteria mediated gemcitabine resistance in pre-clinical models. We determined if intratumoural lipopolysaccharide (LPS) detection by immunohistochemistry is associated with outcome in advanced pancreatic ductal adenocarcinoma (PDAC) treated with gemcitabine and non-gemcitabine containing 1st-line chemotherapy. METHODS: We examined LPS on tumour tissue from 130 patients treated within the randomised AIO-PK0104 trial and a validation cohort (n = 113) and analysed the association of LPS detection to patient outcome according to treatment subgroups. RESULTS: In 24% of samples from the AIO-PK0104 study LPS was detected; in LPS-positive patients median OS was 4.4 months, compared to 7.3 months with LPS negative tumours (HR 1.732, p = 0.010). A difference in OS was detected in 1st-line gemcitabine-treated patients (n = 71; HR 2.377, p = 0.002), but not in the non-gemcitabine treatment subgroup (n = 59; HR 1.275, p = 0.478). Within the validation cohort, the LPS positivity rate was 23%, and LPS detection was correlated with impaired OS in the gemcitabine subgroup (n = 94; HR 1.993, p = 0.008) whereas no difference in OS was observed in the non-gemcitabine subgroup (n = 19; HR 2.596, p = 0.219). CONCLUSIONS: The detection of intratumoural LPS as surrogate marker for gram-negative bacterial colonisation may serve as a negative predictor for gemcitabine efficacy in advanced PDAC. CLINICAL TRIAL REGISTRY: The Clinical trial registry identifier is NCT00440167.

Systemic but not MDSC-specific IRF4 deficiency promotes an immunosuppressed tumor microenvironment in a murine pancreatic cancer model.

Pancreatic ductal adenocarcinoma is characterized by a strong immunosuppressive network with a dense infiltration of myeloid cells including myeloid-derived suppressor cells (MDSC). Two distinct populations of MDSC have been defined: polymorphonuclear MDSC (PMN-MDSC) and monocytic MDSC (M-MDSC). Several factors influence the development and function of MDSC including the transcription factor interferon regulatory factor 4 (IRF4). Here, we show that IRF4 deficiency accelerates tumor growth and reduces survival, accompanied with a dense tumor infiltration with PMN-MDSC and reduced numbers of CD8+ T cells. As IRF4 has been described to modulate myeloid cell development and function, particularly of PMN-MDSC, we analyzed its role using MDSC-specific IRF4 knockout mice with the Ly6G or LysM knock-in allele expressing Cre recombinase and Irf4flox. In GM-CSF-driven bone marrow cultures, IRF4 deficiency increased the frequency of MDSC-like cells with a strong T cell suppressive capacity. Myeloid (LysM)-specific depletion of IRF4 led to increased tumor weight and a moderate splenic M-MDSC expansion in tumor-bearing mice. PMN cell (Ly6G)-specific depletion of IRF4, however, did not influence tumor progression or MDSC accumulation in vivo in accordance with our finding that IRF4 is not expressed in PMN-MDSC. This study demonstrates a critical role of IRF4 in the generation of an immunosuppressive tumor microenvironment in pancreatic cancer, which is independent of IRF4 expression in PMN-MDSC.

Immune Cell and Stromal Signature Associated With Progression-Free Survival of Patients With Resected Pancreatic Ductal Adenocarcinoma.

BACKGROUND & AIMS: Changes to the microenvironment of pancreatic ductal adenocarcinomas (PDACs) have been associated with poor outcomes of patients. We studied the associations between composition of the pancreatic stroma (fibrogenic, inert, dormant, or fibrolytic stroma) and infiltration by inflammatory cells and times of progression-free survival (PFS) of patients with PDACs after resection. METHODS: We obtained 1824 tissue microarray specimens from 385 patients included in the European Study Group for Pancreatic Cancer trial 1 and 3 and performed immunohistochemistry to detect alpha smooth muscle actin, type 1 collagen, CD3, CD4, CD8, CD68, CD206, and neutrophils. Tumors that expressed high and low levels of these markers were compared with patient outcomes using Kaplan-Meier curves and multivariable recursive partitioning for discrete-time survival tree analysis. Prognostic index was delineated by a multivariable Cox proportional hazards model of immune cell and stromal markers and PFS. Findings were validated using 279 tissue microarray specimens from 93 patients in a separate cohort. RESULTS: Levels of CD3, CD4, CD8, CD68, and CD206 were independently associated with tumor recurrence. Recursive partitioning for discrete-time survival tree analysis identified a high level of CD3 as the strongest independent predictor for longer PFS. Tumors with levels of CD3 and high levels of CD206 associated with a median PFS time of 16.6 months and a median prognostic index of -0.32 (95% confidence interval [CI] -0.35 to -0.31), whereas tumors with low level of CD3 cell and low level of CD8 and high level of CD68 associated with a median PFS time of 7.9 months and a prognostic index of 0.32 (95% CI 0.050-0.32); we called these patterns histologic signatures. Stroma composition, when unassociated with inflammatory cell markers, did not associate significantly with PFS. In the validation cohort, the histologic signature resulted in an error matrix accuracy of predicted response of 0.75 (95% CI 0.64-0.83; accuracy P < .001). CONCLUSIONS: In an analysis of PDAC tissue microarray specimens, we identified and validated a histologic signature, based on leukocyte and stromal factors, that associates with PFS times of patients with resected PDACs. Immune cells might affect the composition of the pancreatic stroma to affect progression of PDAC. These findings provide new insights into the immune response to PDAC.

Evaluation of a Novel Aspergillus Antigen Enzyme-Linked Immunosorbent Assay.

Invasive aspergillosis (IA) is a life-threatening infection that mainly occurs in immunocompromised patients. Here, we compared the novel Aspergillus-specific galactomannoprotein (GP) enzyme-linked immunosorbent assay (ELISA) (Euroimmun Medizinische Labordiagnostika AG) to the established Platelia Aspergillus galactomannan (GM) ELISA (Bio-Rad Laboratories) for the detection of IA. A total of 267 serum samples from 45 cases of proven and 4 episodes of probable IA (according to European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group [EORTC/MSG] criteria) and 156 sera from patients without evidence of IA were tested. Pearson's correlation statistics, as well as sensitivity and specificity, were calculated using manufacturer-recommended (GM) or optimized (GP) cutoff levels. Aspergillus fumigatus was found in 88% of culture-positive infections. When we analyzed all 423 serum samples, GM and GP tests correlated strongly (r = 0.82, P < 0.0001). Among proven IA cases using samples obtained as closely as possible to the day of proven diagnosis, the sensitivity for both tests was 40%. All cases of probable IA (defined by positive GM testing) were also GP positive. Concordant results of the two ELISAs were obtained in 43 of 49 samples (88%). Extending measurements to all sera available in the time frame of 7 days prior to 7 days after the day of proven diagnosis, 47% and 56% of the cases were detected by the GM and GP tests, respectively. Specificity was 99% for GM and 96% for GP testing. For the diagnosis of IA, sensitivity and specificity of the novel GP ELISA are similar to those of the Platelia GM ELISA. The low sensitivities of both tests underline the need for serial testing in patients at risk for IA.

Extended RAS analysis and correlation with overall survival in advanced pancreatic cancer.

BACKGROUND: Mutations in the KRAS gene can be detected in about 70-90% of pancreatic cancer (PC) cases. Whether these mutations have a prognostic or predictive value remains elusive. Furthermore, the clinical relevance of the extended RAS (KRAS+NRAS) mutational status is unclear in PC. METHODS: We prospectively defined a PC patient population who received erlotinib-free chemotherapy regimens. A statistically significant difference between KRAS wild-type and KRAS mutated tumours in at least 160 patients in this population would support the assumption of a rather prognostic role of KRAS. RESULTS: One hundred and seventy-eight tumour samples were collected from prospective clinical studies and successfully analysed for the extended RAS status: 37 tumours were KRAS wild-type (21%), whereas 141 (79%) carried a KRAS mutation; 132 of these mutations were found in KRAS exon 2 (74%), whereas only 9 mutations (5%) were detected in KRAS exon 3. Within KRAS exon 4 and NRAS exons 2-4, no mutations were apparent. There was no significant difference in overall survival for KRAS wild-type vs mutant patients (9.9 vs 8.3 months, P=0.70). CONCLUSIONS: Together with the results of the AIO-PK-0104-trial, the present analysis supports the notion that KRAS mutation status is rather predictive than prognostic in advanced PC.

Loss of desmoglein 2 promotes tumorigenic behavior in pancreatic cancer cells.

The ability to maintain cell-cell adhesion is crucial for tissue integrity and organization. Accordingly, loss of cohesiveness plays a critical role in cancer invasion and metastasis. Desmosomes are cell junctions providing strong intercellular adhesive strength and dysregulation of desmosomal constituents contributes to cancer progression through altered cell signaling pathways. Here, we focused on the desmosomal adhesion molecules Desmoglein 2 (Dsg2) and Desmocollin 2 (Dsc2), and their contribution to migration and invasion in pancreatic cancer cells. Silencing of Dsg2 but not Dsc2 resulted in loss of cell cohesion and enhanced migration, and invasion of pancreatic adenocarcinoma cells. To identify potential pathways regulated by Dsg2, we performed kinase arrays and detected the activity of ERK and growth factor receptors to be significantly enhanced in Dsg2-deficient cells. Consequently, inhibition of ERK phosphorylation in Dsg2 knockdown cells normalized migration. Loss of Dsg2 resulted in reduced levels of the desmosomal adapter protein and transcriptional regulator Plakoglobin (PG) in an ERK-dependent manner, whereas other desmosomal molecules were not altered. Overexpression of PG rescued enhanced migration induced by silencing of Dsg2. These results identify a novel pro-migratory pathway of pancreatic cancer cells in which loss of Dsg2 reduces the levels of PG via deregulated MAPK signaling.

Switch in KRAS mutational status during an unusual course of disease in a patient with advanced pancreatic adenocarcinoma: implications for translational research.

BACKGROUND: Despite the introduction of novel effective treatment regimens like gemcitabine plus nab-paclitaxel and FOLFIRINOX, pancreatic ductal adenocarcinoma (PDAC) remains one of the most aggressive epithelial tumors. Among the genetic alterations frequently found in PDAC, mutations in the KRAS gene might play a prognostic role regarding overall survival and may also have the potential to predict the efficacy of anti-EGFR treatment. CASE PRESENTATION: We report the clinical case of a 69 year old Caucasian female that was diagnosed with histologically confirmed locally advanced PDAC with lymph node involvement in August 2010. At the time of first diagnosis, tumor tissue obtained from an open regional lymph node biopsy showed a poorly differentiated adenocarcinoma with a wild type sequence within exon 2 (codon 12/13) of the KRAS gene. The patient initially received single-agent gemcitabine and a subsequent 5-FU-based chemoradiotherapy with a sequential maintenance chemotherapy with oral capecitabine resulting in a long term disease control. Local disease progression occurred in May 2014 and the patient underwent pancreaticoduodenectomy in September 2014. A novel KRAS gene mutation (c.35G > T, p.G12 V) in exon 2 (codon 12) was detected within the surgical specimen. As of January 2016 the patient is still alive and without evidence of the underlying disease. CONCLUSIONS: Specifically in the context of clinical trials and translational research in PDAC a re-assessment of molecular biomarkers, i. e. KRAS, at defined time points (e. g. relapse, disease progression, unusual clinical course) may be indicated in order to detect a potential switch in biomarker status during the course of disease.

The Impact of SMAD4 Loss on Outcome in Patients with Advanced Pancreatic Cancer Treated with Systemic Chemotherapy.

The role of the tumor suppressor mothers against decapentaplegic homolog 4 (SMAD4) has not yet been defined in patients (pts) with advanced pancreatic cancer (aPC). This translational research study was designed to evaluate the impact of tumoral SMAD4 loss on clinicopathological parameters and outcome in PC patients receiving palliative chemotherapy. Using immunohistochemistry, we examined SMAD4 expression in tumor tissue of 143 aPC pts treated within completed prospective clinical and biomarker trials. In uni- and multivariate analyses, SMAD4 expression status was correlated to clinicopathological patient characteristics and outcome. At chemotherapy initiation, 128 pts had metastatic PC; most pts (n = 99) received a gemcitabine-based regimen. SMAD4 loss was detected in 92 pts (64%); patient characteristics such as gender, age, tumor grading, disease stage or number of metastatic sites had no significant impact on tumoral SMAD4 status. In univariate analyses, SMAD4 loss had no impact on overall survival (hazard ratio (HR) 1.008, p = 0.656); however, we observed a prolonged progression-free survival (HR 1.565, p = 0.038) in pts with tumoral SMAD4 loss. This finding was confirmed in multivariate analyses (HR 1.790, p = 0.040), but only for gemcitabine-treated pts. In contrast to previous studies in resectable PC, loss of SMAD4 expression was not associated with a negative outcome in patients with advanced PC receiving systemic chemotherapy.

Impact of SPARC expression on outcome in patients with advanced pancreatic cancer not receiving nab-paclitaxel: a pooled analysis from prospective clinical and translational trials.

BACKGROUND: Conflicting results on the role of secreted protein acidic and rich in cysteins (SPARC) expression have been reported in resected pancreatic ductal adenocarcinoma (PDAC), and its prognostic and/or predictive role in advanced PDAC (aPDAC) has not been extensively investigated yet. This study was designed to evaluate SPARC expression as a biomarker in aPDAC patients (pts) not receiving nab-paclitaxel. METHODS: Using immunohistochemistry, we examined the stromal as well as the tumoral (i.e., cytoplasmic) SPARC expression in tumour tissue (primary tumours and metastases) of 134 aPDAC pts participating in completed prospective clinical and biomarker trials. The SPARC expression levels were correlated to the pts' clinicopathological parameters and survival times. RESULTS: Sixty-seven per cent of the analysed tumours showed high stromal SPARC expression, which was not associated with overall survival (OS, median 9.1 vs 7.6 months, P=0.316). A positive cytoplasmic SPARC expression was detected in 55% of the tumours and correlated significantly with inferior progression-free survival (PFS, 6.2 vs 8.6 months, P=0.004) and OS (7.8 vs 8.4 months, P=0.032). This association was strongest for pts, where primary tumour tissue was examined (PFS: 6.7 vs 10.8 months, P=0.004; OS: 7.9 vs 11.9 months, P=0.030), whereas no significant correlation was detected for pts, where only metastatic tissue was available (PFS: 5.8 vs 6.6 months, P=0.502; OS: 7.0 vs 7.8 months, P=0.452). In pts receiving gemcitabine-based chemotherapy cytoplasmic SPARC expression was significantly associated with an inferior PFS and OS (PFS: 6.2 vs 9.2 months, P=0.002; OS 7.3 vs 9.9 months, P=0.012), whereas no such association was detected for stromal SPARC expression or for pts receiving fluoropyrimidine-based chemotherapy. CONCLUSION: We identified cytoplasmic SPARC expression in the primary tumour as a biomarker associated with inferior PFS and OS in aPDAC. Cytoplasmic SPARC expression may furthermore act as a negative predictive biomarker in pts treated with gemcitabine-based chemotherapy.

Isolated pulmonary metastases define a favorable subgroup in metastatic pancreatic cancer.

PURPOSE: Liver metastasis represents the first site of dissemination in >80% of metastatic pancreatic cancer (PC) patients. Pulmonary metastasis as first site of dissemination in PC is a rare event and might define a biologically distinct subgroup in metastatic PC. METHODS: Consecutive PC patients who were diagnosed or treated with isolated pulmonary metastases at our high-volume comprehensive cancer center were included in a prospectively maintained database between 2002 and 2015. Medical records and correlating computed tomography findings (CT) were retrospectively analyzed. RESULTS: A total of 40 PC patients with isolated pulmonary metastases were identified. Pulmonary metastases represented disease recurrence after initial resection of PC in 22 patients and disease progression of locally advanced pancreatic cancer in 5 patients. 14 out of 27 PC patients (56%) had received chemoradiotherapy for localized disease prior to pulmonary metastasis. Data on 1st-line treatment for pulmonary metastases was available for 38 patients: most patients (71%) received a gemcitabine-based chemotherapy regimen, 5 patients (13%) received best supportive care. After a median follow-up of 37.3 months, median survival after diagnosis of pulmonary metastasis was estimated with 25.5 months (95% CI 19.1-31.8); a significantly improved survival after diagnosis of pulmonary metastasis was observed for patients with less than 10 lung metastases (31.3 vs 18.7 months, p = 0.003) and for an unilateral localization of lung involvement (31.3 vs 21.8 months, p = 0.03). CONCLUSIONS: Our results suggest a favorable outcome of PC patients with isolated pulmonary metastases. Further research is warranted to elucidate the specific molecular characteristics of this rare subgroup.