Abstract: PD57-02
Sources of Funding: This work is supported by NCI grant nos. U54CA143803, CA163124, CA093900, CA143055 to K.J.P as well as the Prostate Cancer Foundation, the Patrick C. Walsh Fund, and a gift from the Stutt family. E.E.vdT is supported by the Cure for Cancer Foundation. H.J.C. is supported by the Urology Care Foundation's Resident Research Award. T.J.B. received funding from the Greenberg bladder cancer institute.

Introduction

The majority of work performed to date investigating circulating tumor cells (CTCs) as biomarkers of urothelial carcinoma (UC) has utilized the CellSearch test (Jansen Diagnostics, Raritan, NJ). One factor limiting the sensitivity of this assay is its reliance on positive selection of CTCs expressing the cell surface protein EpCAM. In this study, we used a novel selection-free method to enumerate and characterize CTCs in patients with UC across a range of stages.

Methods

Blood samples from 38 patients (9 controls, 8 with non-muscle invasive bladder cancer [NMIBC], 12 with muscle-invasive bladder cancer [MIBC] and 9 with metastatic UC) were processed with the AccuCyte-CyteFinder system (RareCyte, Inc., Seattle, WA). Slides were stained for the white blood cell (WBC) markers CD45 and CD66b, and the epithelial markers EpCAM and pan-cytokeratin (CK). CTCs were defined as nucleated cells positive for CK but negative for the WBC markers. Separately, the more restrictive CellSearch definition was also applied, with the additional requirement of EpCAM positivity. The Kruskal-Wallis ANOVA test was used to compare CTC counts between cancer stage groups.

Results

CTCs were detected in 2/8 (25%) patients with NMIBC, 7/12 (58%) with MIBC, and 6/9 (67%) with metastatic disease. No CTCs were found in any control. Comparing CTC counts between groups, the only statistically significant comparison was between controls and patients with metastatic UC (p=0.009, Fig 1A). Using EpCAM positivity as a requirement for defining a CTC, no CTCs were detected in any patient with NMIBC, and only 2 (17%) patients with MIBC (Fig 1B). CTCs tended to be larger in patients with metastatic UC (Fig 2).

Conclusions

CTCs were detected at all UC stages and exhibited phenotypic diversity for cell size and EpCAM expression. EpCAM negative CTCs that would be missed with the CellSearch test were detected in patients with NMIBC and MIBC.

Funding

This work is supported by NCI grant nos. U54CA143803, CA163124, CA093900, CA143055 to K.J.P as well as the Prostate Cancer Foundation, the Patrick C. Walsh Fund, and a gift from the Stutt family. E.E.vdT is supported by the Cure for Cancer Foundation. H.J.C. is supported by the Urology Care Foundation's Resident Research Award. T.J.B. received funding from the Greenberg bladder cancer institute.