Fraser Duthie

Hypermutation In Pancreatic Cancer

Pancreatic cancer is molecularly diverse, with few effective therapies. Increased mutation burden and defective DNA repair are associated with response to immune checkpoint inhibitors in several other cancer types. We interrogated 385 pancreatic cancer genomes to define hypermutation and its causes. Mutational signatures inferring defects in DNA repair were enriched in those with the highest mutation burdens. Mismatch repair deficiency was identified in 1% of tumors harboring different mechanisms of somatic inactivation of MLH1 and MSH2. Defining mutation load in individual pancreatic cancers and the optimal assay for patient selection may inform clinical trial design for immunotherapy in pancreatic cancer.

Ampullary Cancers Harbor ELF3 Tumor Suppressor Gene Mutations and Exhibit Frequent WNT Dysregulation

The ampulla of Vater is a complex cellular environment from which adenocarcinomas arise to form a group of histopathologically heterogenous tumors. To evaluate the molecular features of these tumors, 98 ampullary adenocarcinomas were evaluated and compared to 44 distal bile duct and 18 duodenal adenocarcinomas. Genomic analyses revealed mutations in the WNT signaling pathway among half of the patients and in all three adenocarcinomas irrespective of their origin and histological morphology. These tumors were characterized by a high frequency of inactivating mutations of ELF3, a high rate of microsatellite instability, and common focal deletions and amplifications, suggesting common attributes in the molecular pathogenesis are at play in these tumors. The high frequency of WNT pathway activating mutation, coupled with small-molecule inhibitors of β-catenin in clinical trials, suggests future treatment decisions for these patients may be guided by genomic analysis.

Expression of KOC, S100P, mesothelin and MUC1 in pancreatico-biliary adenocarcinomas: development and utility of a potential diagnostic immunohistochemistry panel

BackgroundPancreatico-biliary adenocarcinomas (PBA) have a poor prognosis. Diagnosis is usually achieved by imaging and/or endoscopy with confirmatory cytology. Cytological interpretation can be difficult especially in the setting of chronic pancreatitis/cholangitis. Immunohistochemistry (IHC) biomarkers could act as an adjunct to cytology to improve the diagnosis. Thus, we performed a meta-analysis and selected KOC, S100P, mesothelin and MUC1 for further validation in PBA resection specimens.MethodsTissue microarrays containing tumour and normal cores in a ratio of 3:2, from 99 surgically resected PBA patients, were used for IHC. IHC was performed on an automated platform using antibodies against KOC, S100P, mesothelin and MUC1. Tissue cores were scored for staining intensity and proportion of tissue stained using a Histoscore method (range, 0–300). Sensitivity and specificity for individual biomarkers, as well as biomarker panels, were determined with different cut-offs for positivity and compared by summary receiver operating characteristic (ROC) curve.ResultsThe expression of all four biomarkers was high in PBA versus normal ducts, with a mean Histoscore of 150 vs. 0.4 for KOC, 165 vs. 0.3 for S100P, 115 vs. 0.5 for mesothelin and 200 vs. 14 for MUC1 (p < .0001 for all comparisons). Five cut-offs were carefully chosen for sensitivity/specificity analysis. Four of these cut-offs, namely 5%, 10% or 20% positive cells and Histoscore 20 were identified using ROC curve analysis and the fifth cut-off was moderate-strong staining intensity. Using 20% positive cells as a cut-off achieved higher sensitivity/specificity values: KOC 84%/100%; S100P 83%/100%; mesothelin 88%/92%; and MUC1 89%/63%. Analysis of a panel of KOC, S100P and mesothelin achieved 100% sensitivity and 99% specificity if at least 2 biomarkers were positive for 10% cut-off; and 100% sensitivity and specificity for 20% cut-off.ConclusionA biomarker panel of KOC, S100P and mesothelin with at least 2 biomarkers positive was found to be an optimum panel with both 10% and 20% cut-offs in resection specimens from patients with PBA.

The role of induction chemotherapy + chemoradiotherapy in localised pancreatic cancer: initial experience in Scotland

BACKGROUND Despite being relatively rare pancreatic cancer is one of the highest causes of death. Even within the potentially resectable group outcomes are poor. We present our initial experiences utilising a neoadjuvant approach to localised pancreatic cancer, evaluating survival, response rates and tolerability. METHODS This was a retrospective analysis of a prospectively maintained database. Patients from 2012 to 2015 referred to a busy regional Hepato-Pancreatic Biliary (HPB) MDT were included. Patients were classified according to respectability criteria (utilising NCCN guidelines) and a treatment plan agreed. Systemic therapy with either FOLFIRINOX or Gem/Cap was delivered followed by chemoradiotherapy if disease remained localised. Toxicity, response, pathological outcomes and survival were all recorded. RESULTS A total of 85 patients were included in the study: 45 had initially resectable disease; 19 required a response for resection and 21 had locally advanced inoperable disease; 34 patients underwent resection. The median survival for the potentially resectable group was 22.2 months while for those undergoing resection it was 37 months. CONCLUSIONS We have demonstrated that a neoadjuvant approach is deliverable and tolerable. In addition we have demonstrated impressive survival results in patients undergoing resection with no detriment in outcome for those not proceeding to surgery.

A Glasgow Tipple—transjugular intrahepatic portosystemic shunt insertion prior to Whipple resection

Abdominal surgery performed in patients with significant liver disease and portal hypertension is associated with high mortality rates, with even poorer outcomes associated with complex pancreaticobiliary operations. We report on a patient requiring portal decompression via transjugular intrahepatic portosystemic shunt (TIPS) prior to a pancreaticoduodenectomy. The 49-year-old patient presented with pain, jaundice and weight loss. At ERCP an edematous ampulla was biopsied, revealing high-grade dysplasia within a distal bile duct adenoma. Liver biopsy was performed to investigate portal hypertension, confirming congenital hepatic fibrosis (CHF). A TIPS was performed to enable a pancreaticoduodenectomy. Prophylactic TIPS can be performed for preoperative portal decompression for patients requiring pancreatic resection. A potentially curative resection was performed when abdominal surgery was initially thought impossible. Notably, CHF has been associated with the development of cholangiocarcinoma in only four previous instances, with this case being only the second reported distal bile duct cholangiocarcinoma.

Abstract 1142: Systematic review and meta-analysis of immunohistochemical diagnostic markers for pancreatic ductal adenocarcinoma.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Background: Pancreatic ductal adenocarcinoma (PDAC) has a five year survival of only 2%. This poor prognosis is partly due to lack of early diagnosis and effective treatment. Surgery is the only curative option but is appropriate for only 10-15% of patients; the remainder have chemotherapy or symptomatic care. Initial diagnosis involves imaging then endoscopy with cytology: diagnosis is important for guiding patient management. The cytological specimens require PDAC to be distinguished from non-neoplastic pancreas. This can be difficult, especially if there is chronic pancreatitis, and because samples are often small. Many immunohistochemical (IHC) biomarkers are differentially expressed between benign and malignant pancreas but none has yet entered routine clinical practice. This study aims to review the evidence on diagnostic biomarkers for PDAC in tissues and cells using IHC. To our knowledge, this is the first meta-analysis on this subject. Material & Methods: The literature was searched using EMBASE and MEDLINE databases from inception to March 2012. We sought publications which assessed the expression of IHC markers in human PDAC and non-neoplastic pancreas (normal and/or chronic pancreatitis). Specimens included both tissue resected at surgery and cytological samples. We catalogued study characteristics including specimen type, IHC details and biomarkers assessed and their staining results including sensitivity and specificity. In the meta-analysis, for each biomarker, coupled forest plots, bivariate summary estimates and combined summary receiver operating characteristic curves were generated, in turn, then compared and ranked according to pooled sensitivity/specificity. Results: 2066 papers were initially identified. 59 studies reporting 33 biomarkers were selected for systematic review. From these, 45 studies reporting 16 biomarkers progressed to meta-analysis. Meta-analysis of IHC biomarkers assessed in resection specimens showed 11 differentiating PDAC from non-neoplastic pancreas. The highest ranked biomarkers according to pooled sensitivity/specificity values were: S100P (100% sensitivity/100% specificity); maspin (92%/97%); KOC (85%/98%); and MUC4 (82%/93%). Meta-analysis of cytology specimens showed seven biomarkers differentiating PDAC from non-neoplastic pancreas. The highest ranked were: KOC (85%/100%); SMAD4 (80%/100%); S100P (91%/91%); and mesothelin (64%/92%). Conclusion: Thirty-three IHC biomarkers for PDAC have been compiled from the literature and the performance of 16 has been quantified and ranked. The highest ranking IHC markers for PDAC were KOC and S100P, then maspin, mesothelin and MUC4. Their reported diagnostic accuracies approach those of optimal conventional cytology. These markers may be worth considering for further clinical validation and potentially routine use in difficult cases. Citation Format: Asif Ali, Zia Ul-Haq, Mohamed Mohamed, Daniel Francis MacKay, Fraser Duthie, Karin Oien. Systematic review and meta-analysis of immunohistochemical diagnostic markers for pancreatic ductal adenocarcinoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1142. doi:10.1158/1538-7445.AM2013-1142

Investigating Various Thresholds as Immunohistochemistry Cutoffs for Observer Agreement.

Background: Clinical translation of immunohistochemistry (IHC) biomarkers requires reliable and reproducible cutoffs or thresholds for interpretation of immunostaining. Most IHC biomarker research focuses on the clinical relevance (diagnostic, prognostic, or predictive utility) of cutoffs, with less emphasis on observer agreement using these cutoffs. From the literature, we identified 3 commonly used cutoffs of 10% positive epithelial cells, 20% positive epithelial cells, and moderate to strong staining intensity (+2/+3 hereafter) to use for investigating observer agreement. Materials and Methods: A series of 36 images of microarray cores stained for 4 different IHC biomarkers, with variable staining intensity and percentage of positive cells, was used for investigating interobserver and intraobserver agreement. Seven pathologists scored the immunostaining in each image using the 3 cutoffs for positive and negative staining. Kappa (&kgr;) statistic was used to assess the strength of agreement for each cutoff. Results: The interobserver agreement between all 7 pathologists using the 3 cutoffs was reasonably good, with mean &kgr; scores of 0.64, 0.59, and 0.62, respectively, for 10%, 20%, and +2/+3 cutoffs. A good agreement was observed for experienced pathologists using the 10% cutoff, and their agreement was statistically higher than for junior pathologists (P=0.02). In addition, the mean intraobserver agreement for all 7 pathologists using the 3 cutoffs was reasonably good, with mean &kgr; scores of 0.71, 0.60, and 0.73, respectively, for 10%, 20%, and +2/+3 cutoffs. For all 3 cutoffs, a positive correlation was observed with perceived ease of interpretation (P<0.003). Finally, cytoplasmic-only staining achieved higher agreement using all 3 cutoffs than mixed staining patterns. Conclusions: All 3 cutoffs investigated achieve reasonable strength of agreement, modestly decreasing interobserver and intraobserver variability in IHC interpretation. These cutoffs have previously been used in cancer pathology, and this study provides evidence that these cutoffs can be reproducible between practicing pathologists.

714 Molecular Pathological Phenotypes and Outcome in Pancreatic Ductal Adenocarcinoma

Introduction: Individuals with pancreatic ductal adenocarcinoma (PDAC) demonstrate a generally poor outcome following resection. Molecular profiling has previously enhanced the identification of phenotypic subtypes of ampullary adenocarcinoma. Furthermore an intestinal subtype of PDAC has been described however the prognostic impact of this variant has not been described in detail. We sought to better characterize the intestinal subgroup of PDAC and assess the impact on outcome. Methods: We assessed the potential clinical utility of molecular pathological phenotypes defined using a combination of histopathology and protein expression (CDX2 [caudal-type homeodomain transcription factor 2] an intestinal marker and MUC1 a pancreaticobiliary marker) assessed by immunohistochemistry (Figure 1) in 95 patients who underwent operative resection for PDAC by pancreaticoduodenectomy at a single institution over a 12 year time period. A tissue microarray was used with at least 4 cores evaluated for each tumor for protein expression analysis in addition towhole section analysis of tumormorphology. Care was taken to exclude all other periampullary malignancies from the analysis. Results: In addition to prognostic impact of T stage, lymph node status, resection margin status, perineural invasion and vascular invasion, a small proportion of tumors had features of an intestinal histological subtype (13%) and a more favorable prognosis. CDX2 and MUC1 expression were significant prognostic variables. Patients with CDX2 negative tumors had a significantly shorter survival (Hazard ratio [HR] = 2.77, 95%CI: 1.5-5.2, P = 0.002 as did those with MUC1 positive tumors (HR = 2.89, 95%CI: 1.7-4.9, P , 0.0001 no survivors at 24 months). Patients with CDX2 negative/ MUC1 negative tumors had an intermediate outcome (Figure 1). In a multivariate analysis lymph node involvement, vascular invasion, positive MUC1 expression and loss of CDX2 expression were independent predictors of poor outcome. Conclusion: Morphological determination of intestinal subtype of PDAC has clinical relevance. Furthermore maintenance of CDX2 expression identifies a group of PDAC patients with a relatively good outcome while MUC1 expression identified patients with a very poor outcome. When combined histopathological and molecular criteria define clinically relevant phenotypes of PDAC with significant implications for prognostication, current therapeutic strategies and may facilitate future trial design.