Abstract P2-06-21: RNAi Silencing of CUB-Domaining Containing Protein 1 (CDCP1) in Breast Cancer Cell Lines Results in Decreased Migration and Invasion Capacities In Vitro.
Authors: Seidel J , Kunc K , Possinger K , Lueftner DI. 
Published: April 2011

Abstract
Background: The orphan surface receptor CDCP1 is over-expressed in various cancers and signals via Src family kinases (SFKs). It has been shown to be involved in anchorage independency of distinct tumors [Uekita et al. 2007; Moasser 2009]. Since CDCP1 is also speculated to be involved in tumor cell migration and invasion, the following study investigated the in vitro migration and invasion of breast cancer cell lines. In order to prove if lapatinib treatment might benefit CDCP1-positive but HER2/neu-negative breast cancer patients, we investigated the level of apoptosis after lapatinib application to different cell lines.
Material and Methods: Four breast cancer cell lines were selected according to their CDCP1/HER-2/neu receptor expression (MDA-MB-231high/high; T47Dhigh/low; SKBR3low/high; MCF7neg./high). CDCP1 knock-down was achieved via nucleofection of Stealth™ RNAi (Invitrogen) against CDCP1. In vitro migration and invasion were analyzed for 48hs using trans-well assays (Merck). Migrated cells were labeled with Calcein AM and relative fluorescence units (RFU) were measured at 485/520nm. To investigate the effect of lapatinib, cells were cultured for 48hs in the presence of lapatinib (0,75 µM) as attached cells or as suspension culture on ultra-low adhesion plates, respectively. The enrichment of free nucleosomes was measured and compared between CDCP1-RNAi transfected and parental cells.
Results: When transfected with CDCP1-RNAi, migration of SKBR-3 cells was not affected (96.2% migrating cells compared to control). However, CDCP1-RNAi-transfected MDA-MB-231 cells showed a strong reduction in migration (70.2% (± 21.5%)) compared to mock-transfected cells and the observed effect was even more impressive in T47D cells (58.3± 16.1% migration compared to control; P=0,011)). Invasion of CDCP1- silenced MDA-MB-231 was reduced to 71.4% (± 5.9%) of the mock-transfected control cells. Again, T47D cells also were severely impaired by CDCP1-knock-down (0.32 ± 0.02% of invasive cells compared to control; P&lt;0.01). Lapatinib had no effect on either MDA-MB-231 cells or T47D cells at any culture conditions. However, the level of apoptotic SKBR3 cells significantly increased when cultured in suspension (1.5x) as well as adherent cells (1.8x) in the presence of lapatinib independent of CDCP1-knock down.
Conclusions: The migratory activity of breast cancer cell lines did not strictly correlate with the degree in CDCP1 expression. However, CDCP1- knock down yielded in a clearly reduced migration and invasion of MDA-MB-231 cells and was even more impressive in T47D cell line. However, no effect was observed in SKBR3 cells. Thus, CDCP1 might represent one of a panel of surface receptors that participate in cell movement. However, Lapatinib was shown to have no effect on the viability of CDCP1pos./Her2/neuneg./low cells. Nonetheless, our study reveals that CDCP1 might have a strong impact on migration and invasive capacities of breast cancer cell lines and thus represents a potential target in more individualized breast cancer therapies.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P2-06-21.