Detection Of Circulation Tumor Cells Using Telomerase-Selective Adenoviral Marker (Obp-401 (R)) In Breast Cancer Patients.

Abstract Background: The detection of circulating tumor cells (CTCs) is a potential method to predict survival of metastatic breast cancer patients as well as outcomes of early breast cancer patients. However, no method for CTCs has yet proven to be the golden standard. We developed a new approach for detecting CTCs using telomerase-specific replication-competent adenovirus expressing green fluorescent protein (GFP) (OBP-401®, Oncolys Biopharma™).Methods: OBP-401® contains the replication cassette, in which the human telomerase reverse transcriptase promoter drives expression of E1 genes, and the GFP gene for monitoring viral replication. This system is consisted of the following steps; (1) virus-infection for 7.5 ml whole blood (incubated with 4 X 108 Plaque Forming Unit OBP-401 virus for 24 hours at 37 Celsius), (2) dead cell staining using L23102 (invitrogen™), (3) virus-inactivation and RBC elimination, and (4) detection of GFP expressing cells using fluorescence microscopy. In a preclinical study, the sensitivity of this system was assessed using cell lines. Next, we conducted feasibility studies for CTCs detection in 80 healthy individuals, 50 metastatic, and 27 early breast cancer patients. In metastatic and early breast cancer patients, we compared the sensitivity of this system with that of CellSearch® (Veridex™) and tumor makers (CEA and CA15-3). GFP-positive cells (viable CTCs) and L23102 expressing cells measuring ≥ 20µm in diameter (dead CTCs) were considered as CTCs in this system.Results: The sensitivity of this system, which was determined by a serial dilution of MDA-MB-468 cells against healthy volunteer's blood, was 1 cell per 7.5 ml. No CTCs were detected in any of healthy controls. Of 50 metastatic patients, 12% were primary breast cancers with stage IV disease, 24% were in the 1st line chemotherapy setting, and 42% were heavily pretreated with chemotherapy. The sensitivities of tumor markers, CellSearch®, and OBP-401® were 78%, 54%, and 66%, respectively. Neither CTCs detected with CellSearch® nor OBP-401® were significantly associated with clinicopathologic parameters. However, CTCs-positivity detected with CellSearch® were strongly associated with CA15-3 positivity (p = 0.003). Of 14 patients with normal CA15-3 levels, CellSearch® detected CTCs in three patients (21%) but OBP-401® in nine patients (64%). The sensitivity of the combination of tumor markers and OBP-401® was 92%. In 27 early breast cancers (Stage1 7, StageII 17, StageIII 3), three patients were treated with neoadjuvant chemotherapy (NAC). All blood samples were drawn before surgery or NAC. The sensitivities of tumor markers, CellSearch®, and OBP-401® were 7%, 0%, 67%, respectively. There were no significant correlations between CTCs detected with OBP-401® and clinicopathologic features.Conclusion: OBP-401® showed no false positive in healthy controls, and a high sensitivity for CTCs detection, particular in metastatic breast cancer patients with normal 15-3 levels and early breast cancer patients, compared with CellSearch®. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 3012.