Use of image cytometry to classify biliary and ampullary adenocarcinomas

To create an objective classification system to perform TNM classification of ampullary adenocarcinoma and cholangiocarcinoma using image cytometric data derived from Feulgen-stained tumor nuclei.

Surgically resected cases of ampullary adenocarcinoma and cholangiocarcinoma with established TNM classifications were selected on the basis of available formalin-fixed, paraffin-embedded tissue. Fifteen numerical variables related to morphometric, densitometric and textural features of each tumor nucleus were recorded. We employed a methodology based on multivariate statistical tools to characterize the association of morphonuclear variables with TNM classification. The first step consisted of identifying and selecting representative nuclei of each T class. From this "purified" data set an objective classification system was created. The classification system was assessed using internal and external validation.

Employing ANOVA, all 15 variables were significantly associated with T classification, 11 of 15 with N and 4 with M. Multivariate analysis was employed to distinguish between T1, T2 and T3 lesions. Our methodology correctly classified 76% of T1 nuclei, 47% of T2 nuclei and 84% of T3 nuclei. Heterogeneity within an individual tumor was defined in 61% of cases included in the training set. Complete concordance between pathologic classification and the classification system was observed in 71% of an independent validation.