DETECTION OF BREAST-CANCER CELLS IN BLOOD USING IMMUNOMAGNETIC BEAD SELECTION AND REVERSE TRANSCRIPTION-POLYMERASE CHAIN-REACTION

Background: A sensitive and specific method for the detection of occul t breast cancer cells may prove clinically useful. The purpose of this study was to investigate cytokeratin-19 (CK-19) and gross cystic dise ase fluid protein (GCDFP) as potential reverse transcription-polymeras e chain reaction (RT-PCR) targets for the detection of breast micromet astases. Through positive selection of breast epithelial cells by immu nomagnetic bead separation, two RT-PCR assays were developed. Methods and Results: Positive selection of breast epithelial cells was perform ed using Ber-EP4 monoclonal antibody bound to magnetic beads. RNA was isolated and RT-PCR performed using CK-19 and GCDFP as targets. Detect ion sensitivity was five T47D cells spiked into 10 mL of normal blood for either target. In all, 100% (39/39) of normal bloods were negative for CK-19, whereas only 87% (34/39) were negative with GCDFP. In 15 p atients with metastatic disease including 3 without treatment and 12 o n either tamoxifen or chemotherapy, CK-19 was positive in 67% (10/15) of patients and GCDFP yielded positives in 27% (4/15) of patients test ed. Conclusions: The detection of CK-19 and GCDFP in bloods from patie nts with metastatic breast cancer may be beneficial in determining the course of therapy, as well as having potential prognostic and diagnos tic applications. Although CK-19 appears to be the more sensitive and specific marker, further investigation with both targets is warranted.