Gene expression profiling of ampullary carcinomas classifies ampullary carcinomas into biliary-like and intestinal-like subtypes that are prognostic of outcome

Abstract

Background: Adenocarcinomas of the ampulla of Vater are classified as biliary cancers, though the exact epithelium of origin for these cancers is not known. We sought to molecularly classify ampullary adenocarcinomas in comparison to known adenocarcinomas of the pancreas, bile duct, and duodenum by gene expression analysis.

Methods: We analyzed 32 fresh-frozen resected, untreated periampullary adenocarcinomas (8 pancreatic, 2 extrahepatic biliary, 8 duodenal, and 14 ampullary) using the Affymetrix U133 Plus 2.0 genome array. Unsupervised and supervised hierarchical clustering identified two subtypes of ampullary carcinomas that were molecularly and histologically characterized.

Results: Hierarchical clustering of periampullary carcinomas segregated ampullary carcinomas into two subgroups, which were distinctly different from pancreatic carcinomas. Non-pancreatic periampullary adenocarcinomas were segregated into two subgroups with differing prognoses: 5 year RFS (77% vs. 0%, p = 0.007) and 5 year OS (100% vs. 35%, p = 0.005). Unsupervised clustering analysis of the 14 ampullary samples also identified two subgroups: a good prognosis intestinal-like subgroup and a poor prognosis biliary-like subgroup with 5 year OS of 70% vs. 28%, P = 0.09. Expression of CK7+/CK20- but not CDX-2 correlated with these two subgroups. Activation of the AKT and MAPK pathways were both increased in the poor prognostic biliary-like subgroup. In an independent 80 patient ampullary validation dataset only histological subtype (intestinal vs. pancreaticobiliary) was significantly associated with OS in both univariate (p = 0.006) and multivariate analysis (P = 0.04).

Conclusions: Gene expression analysis discriminated pancreatic adenocarcinomas from other periampullary carcinomas and identified two prognostically relevant subgroups of ampullary adenocarcinomas. Histological subtype was an independent prognostic factor in ampullary adenocarcinomas.

Figure 1

Unsupervised hierarchical clustering of all 32 periampullary adenocarcinoma samples (A).Supervised hierarchical clustering based upon the 133 differentially expressed genes between pancreatic and duodenal adenocarcinomas (B). Clinical characteristics are listed below the figure: age >65 y/o (black), male gender (black), poor differentiation (black), mucinous histology (black), T4 (black), N1 (black), and the presence of a precursor lesion such as an adenoma, dysplasia or pancreatic intraepithelial neoplasia (black). Overall survival by (C) tumor type and (D) gene expression grouping (group 1, n = 9; group 2, n = 13; group 3, n = 10).

Figure 2

Unsupervised hierarchical clustering of ampullary adenocarcinoma samples, n = 14 (A).Clinical and molecular characteristics are listed below the figure: group 3 gene expression grouping (black), poor differentiation (black), mucinous histology (black), T4/T3 (black), N1 (black), presence of an adenoma (black), activation mutations in KRAS, BRAF, PI3K (black), MSI-high status (black), CK7+/CK20− (black), CDX-2+ (black), and histological subtype. Overall survival by gene expression derived biliary-like and intestinal-like ampullary subgroups (B).

Figure 3. Unsupervised hierarchical clustering of the differentially expressed proteins (P<0.05) between gene expression derived biliary-like and intestinal-like ampullary subgroups.

Figure 4. Overall survival and relapse-free survival for the 80 patient ampullary dataset stratified by (A,B) CDX-2 expression status, (C,D) CK7+/CK20− expression status, and (E,F) histological subtype, respectively.