Accomplishments: 1) Approval and implementation of a pancreatic cancer study to collect clinical specimens from PDAC cases: The study includes 500 primary PDAC cases, including 150 cases with surgical resection from which fresh-frozen tumor and surrounding nontumor specimens are being collected at the University of Maryland at Baltimore under NCI-UMD resource contract of the laboratory. Blood samples are being collected from all cases. The collected samples are being used for molecular profiling study with the following specific aims: 1) Functional role of genes and metabolites that are associated with survival in PDAC. 2) Characterizing molecular subgroups in early stage PDAC and screening the subgroup- specific tumor biomarkers in body fluid. 3) Identifying the critical molecular differences between early stage tumors from patients surviving less than 6 months and more than 2 years. 2) Establishment of Multiple independent validation Cohorts: It is important to ensure that our results are broadly applicable to PDAC outside the study population. To ensure the availability of clinical specimens from independent cohorts of PDAC, I established collaborations at different institutes around the world. I have been successful in receiving clinical samples from collaborators at University of Heidelberg and University of Medicine, Goettingen in Germany and Jikei University in Tokyo, Japan. I am continuing my effort to expand the sample size in each of these cohorts. 3) Global-Gene expression and Metabolite Profiling a) Pilot Gene Expression Profiling Study Identified Dipeptidase-1 (DPEP1) as a biologically relevant gene in the progression of PDAC (Zhang et. al., PLoS One, 2012): DPEP1 expression was independently associated with cancer specific mortality when adjusted for tumor stage and resection margin status in both univariable and multivariable analysis. Mechanistic analyses revealed that DPEP1 inhibited pancreatic cancer cell migration and invasion, and increased their sensitivity to gemcitabine, a first-line chemotherapeutic drug for treating pancreatic cancer. We also found that DPEP1 is regulated by EGF/MEK/MAPK pathway, and MEK1/2 inhibitor enhanced DPEP1 expression in vitro. These results show that the analysis of transcriptome in human pancreatic tumors can yield novel prognostic genes with a potential role in disease aggressiveness. b) Integration of Metabolomics and Transcriptomics Revealed a fatty Acid Network Exerting Growth Inhibitory Effects in Human Pancreatic cancer (Zhang et. al., Clin. Cancer Res., 2013, In Press): To identify metabolic pathways that are perturbed in pancreatic ductal adenocarcinoma (PDAC), we investigated gene-metabolite networks with integration of metabolomics and transcriptomics. We have performed global metabolite profiling analysis on two independent cohorts of resected PDAC cases to identify critical metabolites alteration that may contribute to the progression of pancreatic cancer. We then searched for gene surrogates that were significantly correlated with the key metabolites by integrating metabolite and gene expression profiles. Fifty-five metabolites were consistently altered in tumors as compared with adjacent nontumor tissues in a test cohort (N=33) and an independent validation cohort (N=31). Weighted network analysis revealed a unique set of free fatty acids (FFAs) that were highly co-regulated and decreased in PDAC. Pathway analysis of 157 differentially expressed gene surrogates revealed a significantly altered lipid metabolism network, including key lipolytic enzymes PNLIP, CLPS, PNLIPRP1, and PNLIPRP2. Gene expressions of these lipases were significantly decreased in pancreatic tumors as compared with nontumor tissues, leading to reduced FFAs. More importantly, a lower gene expression of PNLIP in tumors was associated with poorer survival in two independent cohorts. We further demonstrated that two saturated FFAs, palmitate and stearate significantly induced TRAIL expression, triggered apoptosis, and inhibited proliferation in pancreatic cancer cells. Our results suggest that impairment in a lipolytic pathway involving lipases and a unique set of FFAs, may play an important role in the development and progression of pancreatic cancer and provide potential targets for therapeutic intervention. Ongoing Studies: 1) Characterizing molecular distinctions in early stage resected PDAC with good and poor survival: Early-stage PDAC, if detected, are resectable and offer relatively better prognosis in resected patients with a median survival of about 2 years. However, more than 80% of the resected cases show recurrence within two years with fatality, in some cases, reported even within 6 months following surgery. In contrast, a small number of the resected cases may survive up to 5 years (12% of the resected cases) and even 10 years (5% of the resected cases). Survival following resection is associated with many clinical prognostic factors including tumor stage, grade (degree of differentiation) and resection margin status but no one factor is consistently prognostic and we do find variable outcomes among cases with similar stage, grade or resection margin status. We hypothesize that molecular characterization of early stage PDAC with markedly different prognoses may identify critical pathways associated with disease aggressiveness and candidate targets for therapeutic intervention. To test this hypothesis we are conducting a pilot study, as shown in the schema 2, by examining and comparing protein coding genes and non-coding miRNA expression profile of tumors in two groups of resected patients: one with a short survival (7 months, N=11) and the other with a longer survival (2-6 years, N=16) following resection in PDAC cases. In this study, we are asking the following questions: 1) What are the differentially expressed protein coding genes and miRNAs in the tumors from short survival as compared with long survival group? 2) Are inflammatory genes expression profile different in short versus long survival groups? 3) What are the functional roles of the most differentially expressed genes and associated pathways in the aggressiveness of pancreatic cancer? 2) Molecular Subgroups of Pancreatic Ductal Adenocarcinoma: Molecular diversity in tumors contributes to the discrepancies in therapeutic response. Examining a large cohort of PDAC for multi-level, global molecular changes including mRNA, miRNA and metabolites, and their integrative analyses may help characterize molecular subgroups, subgroup-specific biomarkers, and candidate therapeutic targets. One of our areas of interest is the role of inflammation in pancreatic cancer. Several of these inflammatory and immune-related genes are shown to have a functional role in the aggressiveness of pancreatic cancer. We hypothesize that Inflammation-associated gene signature defines aggressive subtypes in resectable PDAC. We are conducting a comprehensive transcriptomic (mRNA and microRNA) and metabolomic analyses in a large cohort of PDAC (N=200) with complete clinical, demographic and epidemiological profile. Our goals are as follows: 1) Characterize molecular subgroups of PDAC through integration of transcriptomic and metabolomic profiles. 2) Search for inflammation-associated gene signature in aggressive subtypes. 3) Examine the mechanistic role of subtype-specific genes and metabolites in tumor progression. 4) Evaluate the potential therapeutic significance of subtype specific genes and pathways in pre-clinical in vitro and in vivo models.
|Zhang, Geng; He, Peijun; Gaedcke, Jochen et al. (2013) FOXL1, a novel candidate tumor suppressor, inhibits tumor aggressiveness and predicts outcome in human pancreatic cancer. Cancer Res 73:5416-25|