Wn oncogenic signaling pathways. Our RPPA Title Loaded From File analysis was able to recapitulate the molecular heterogeneity identified by whole-genomic transcriptional Title Loaded From File Profiling and concordantly identified two prognostically relevant subgroups of ampullary adenocarcinomas. RPPA analysis demonstrated marked activation of both the PI3K-AKT and RAS-RAF-MAPK pathways in our poor prognosis pancreaticobiliary-like ampullary subgroup. Of note, both ampullary adenocarcinoma samples with mutations in PIK3CA in this cohort of patients occurred in the pancreaticobiliary-like subgroup. These findings support the rationale for functional testing of inhibitors against these pathways in biliary-like ampullary carcinomas. In the intestinal-like subgroup a number of intestinal-specific genes were upregulated, which suggests that this subgroup of ampullary carcinomas may arise from the overlying ampullo-duodenal epithelium of the ampulla of Vater. Since prior studies have inconsistently reported the prognostic relevance of CDX-2 expression, CK7+/CK202 expression, and histological subtypes, we examined these markers. [4,5,9?3] We found that CDX-2 expression status represented an imperfect marker in comparison to our gene expression groupings. CK7+/ CK202 expression pattern and histological subtype were the two factors most closely correlated with our ampullary gene expression groupings. In our validation cohort, only histological subtype demonstrated a consistent prognostic impact and was an independent prognostic factor for OS in patients with ampullary adenocarcinoma. These results are consistent with the recently reported ESPAC-3 periampullary randomized study in which the intestinal as opposed to the pancreaticobiliary subtype of ampullary adenocarcinomas demonstrated an improved DFS (45.7 vs. 20.6 m, p = 0.01) but not OS (56 vs. 43.1 m, p = 0.28). [8] At present reporting the histological subtype of ampullary adenocarcinomas is not standard, as was demonstrated by the ESPAC-3 study in which only 45 of all ampullary adenocarcinomas had histological subtype reported. Our data does differ from a prior report that found statistical significance with the use of CDX-2 and CDX-1 staining in 53 resected ampullary carcinoma patients. [13] However, our data is consistent with two subsequent cohorts of 53 and 71 patients that did not identify CDX-2 expression as a prognostic marker [5,12].Gene Profiling of Periampullary CarcinomasRecently, our clinical approach for both metastatic duodenal and biliary adenocarcinomas has improved with the publication of a phase II study evaluating capecitabine and oxaliplatin in small bowel adenocarcinoma and a phase III study evaluating gemcitabine and cisplatin in biliary adenocarcinoma. [31,32] Interestingly, as a testament to the uncertainty of how to approach ampullary adenocarcinomas, both studies included adenocarcinomas of the ampulla of Vater. The recently completed ESPAC-3 study evaluated the role of adjuvant therapy for ampullary adenocarcinomas and found no difference with regard to the use of either gemcitabine or 5-FU in the adjuvant setting. [8] Though our findings do not provide a direct link between the expression profiling or histological subtype of ampullary adenocarcinoma and chemotherapy benefit, we do feel that histological subtype deserves further study as a potential marker to better select patients for adjuvant therapy and possibly as a means to optimally select chemotherapy, 5-FU-based as opposed to gemcitabine-based. Th.Wn oncogenic signaling pathways. Our RPPA analysis was able to recapitulate the molecular heterogeneity identified by whole-genomic transcriptional profiling and concordantly identified two prognostically relevant subgroups of ampullary adenocarcinomas. RPPA analysis demonstrated marked activation of both the PI3K-AKT and RAS-RAF-MAPK pathways in our poor prognosis pancreaticobiliary-like ampullary subgroup. Of note, both ampullary adenocarcinoma samples with mutations in PIK3CA in this cohort of patients occurred in the pancreaticobiliary-like subgroup. These findings support the rationale for functional testing of inhibitors against these pathways in biliary-like ampullary carcinomas. In the intestinal-like subgroup a number of intestinal-specific genes were upregulated, which suggests that this subgroup of ampullary carcinomas may arise from the overlying ampullo-duodenal epithelium of the ampulla of Vater. Since prior studies have inconsistently reported the prognostic relevance of CDX-2 expression, CK7+/CK202 expression, and histological subtypes, we examined these markers. [4,5,9?3] We found that CDX-2 expression status represented an imperfect marker in comparison to our gene expression groupings. CK7+/ CK202 expression pattern and histological subtype were the two factors most closely correlated with our ampullary gene expression groupings. In our validation cohort, only histological subtype demonstrated a consistent prognostic impact and was an independent prognostic factor for OS in patients with ampullary adenocarcinoma. These results are consistent with the recently reported ESPAC-3 periampullary randomized study in which the intestinal as opposed to the pancreaticobiliary subtype of ampullary adenocarcinomas demonstrated an improved DFS (45.7 vs. 20.6 m, p = 0.01) but not OS (56 vs. 43.1 m, p = 0.28). [8] At present reporting the histological subtype of ampullary adenocarcinomas is not standard, as was demonstrated by the ESPAC-3 study in which only 45 of all ampullary adenocarcinomas had histological subtype reported. Our data does differ from a prior report that found statistical significance with the use of CDX-2 and CDX-1 staining in 53 resected ampullary carcinoma patients. [13] However, our data is consistent with two subsequent cohorts of 53 and 71 patients that did not identify CDX-2 expression as a prognostic marker [5,12].Gene Profiling of Periampullary CarcinomasRecently, our clinical approach for both metastatic duodenal and biliary adenocarcinomas has improved with the publication of a phase II study evaluating capecitabine and oxaliplatin in small bowel adenocarcinoma and a phase III study evaluating gemcitabine and cisplatin in biliary adenocarcinoma. [31,32] Interestingly, as a testament to the uncertainty of how to approach ampullary adenocarcinomas, both studies included adenocarcinomas of the ampulla of Vater. The recently completed ESPAC-3 study evaluated the role of adjuvant therapy for ampullary adenocarcinomas and found no difference with regard to the use of either gemcitabine or 5-FU in the adjuvant setting. [8] Though our findings do not provide a direct link between the expression profiling or histological subtype of ampullary adenocarcinoma and chemotherapy benefit, we do feel that histological subtype deserves further study as a potential marker to better select patients for adjuvant therapy and possibly as a means to optimally select chemotherapy, 5-FU-based as opposed to gemcitabine-based. Th.