Immunomagnetic enrichment of disseminated tumor cells in bone marrow and blood of breast cancer patients by the Thomsen-Friedenreich-Antigen.
OBJECTIVE
The presence of disseminated tumor cells in the bone marrow (DTC-BM) of breast cancer patients has shown independent prognostic impact. Immunomagnetic enrichment of such cells is an approach to increase the number of detected cells with limited sample volume, especially for circulating tumor cells (CTCs) in blood. The Thomsen-Friedenreich (TF) antigen (CD 176) is a specific oncofetal carbohydrate epitope (Galbeta1-3GalNAcalpha-O) expressed on the surface of various carcinomas. Own studies demonstrated a nearly complete TF expression on DTC-BM, indicating its suitability as marker for immunomagnetic enrichment.


METHODS
BM samples of 65 and peripheral blood samples of 11 breast cancer patients were examined immunocytochemically by staining with the anti-Cytokeratin antibody A45-B/B3 before and after immunomagnetic enrichment. Enrichment was done by incubation with the primary antibody TF 2 (IgM), followed by secondary magnetically labelled rat-anti mouse IgM. Cytospin slides were screened manually by bright-field microscopy.


RESULTS
15/65 pts (23%) showed DTC-BM in primary screening with a median of 2/2 mio cells (range 1-10). By enrichment, a median of 23.3 mio cells (0.8-218) could be analysed, increasing positivity to 72% (47/65 pts) with a med. of 4 DTCs (1-105, P < .0001). Blood from 1/11 pts before and 5/11 (45%) after enrichment showed CTCs (med. 2, 1-20), at a med. of 12.4 mio (2.6-38.5) cells analysed. Comparing BM and blood of the same patients after enrichment, 5 were positive in both compartments, 4 showed DTC-BM without presence of CTCs.


CONCLUSIONS
The positive immunomagnetic enrichment technique with TF-antibodies enables to analyse larger sample volumes and increase tumor cell detection rate. This could allow monitoring and characterisation of CTCs as targets for therapies.
Schindlbeck
Christian
C
First Department of Obstetrics & Gynecology, Ludwig Maximilians University of Munich, Munich, Germany. Christian.Schindlbeck@med.uni-muenchen.de
Stellwagen
Julia
J
Jeschke
Udo
U
Karsten
Uwe
U
Rack
Brigitte
B
Janni
Wolfgang
W
Jückstock
Julia
J
Tulusan
Augustinos
A
Sommer
Harald
H
Friese
Klaus
K
D000328:Adult; D000368:Aged; D015295:Antigens, Tumor-Associated, Carbohydrate / Q000032:analysis*; D014408:Biomarkers, Tumor / Q000032:analysis*; D001853:Bone Marrow / Q000737:chemistry / Q000473:pathology*; D001856:Bone Marrow Examination; D019046:Bone Marrow Neoplasms / Q000175:diagnosis* / Q000378:metabolism / Q000556:secondary; D001943:Breast Neoplasms / Q000737:chemistry / Q000473:pathology*; D005260:Female; D006801:Humans; D007124:Immunoenzyme Techniques; D018189:Immunomagnetic Separation; D008875:Middle Aged; D009360:Neoplastic Cells, Circulating / Q000473:pathology*; D011379:Prognosis
D015295:Antigens, Tumor-Associated, Carbohydrate; D014408:Biomarkers, Tumor; C016467:Thomsen-Friedenreich antigen