The purpose of this study was to assess inter- and intra-observer reproducibility of three different analytic methods to evaluate quantitative dynamic contrast-enhanced computed tomography (DCE-CT) measures from gastroesophageal junctional cancer. Twenty-five DCE-CT studies with gastroesophageal junction cancer were selected from a previous longitudinal study. Three radiologists independently reviewed all scans, and one repeated the analysis eight months later for intraobserver analysis. Review of the scans consisted of three analysis methods: (I) Four, fixed small sized regions of interest (2-dimensional (2D) fixed ROIs) placed in the tumor periphery, (II) 2-dimensional regions of interest (2D-ROI) along the tumor border in the tumor center, and (III) 3-dimensional volumes of interest (3D-VOI) containing the entire tumor volume. Arterial flow, blood volume and permeability (k(trans)) were recorded for each observation. Inter- and intra-observer variability were assessed by Intraclass Correlation Coefficient (ICC) and Bland-Altman statistics. Interobserver ICC was excellent for arterial flow (0.88), for blood volume (0.89) and for permeability (0.91) with 3D-VOI analysis. The 95% limits of agreement were narrower for 3D analysis compared to 2D analysis. Three-dimensional volume DCE-CT analysis of gastroesophageal junction cancer provides higher inter- and intra-observer reproducibility with narrower limits of agreement between readers compared to 2D analysis.