IMMUNOMAGNETIC PURIFICATION OF HUMAN BREAST-CARCINOMA CELLS ALLOWS TUMOR-SPECIFIC DETECTION OF MULTIDRUG-RESISTANCE GENE 1-MESSENGER-RNA BYREVERSE-TRANSCRIPTASE POLYMERASE CHAIN-REACTION IN FINE-NEEDLE ASPIRATES

BACKGROUND: The heterogenous composition of tumors is a major obstacle for the measurement of mRNA levels in cancer cells. We report here a combination of immunomagnetic purification of cancer cells and reverse transcriptase polymerase chain reaction (RT-PCR) that enables highly sensitive detection of multidrug resistance gene 1 (MDR1)-mRNA levels in human breast carcinoma cells obtained from fine needle aspirates (F NA). EXPERIMENTAL DESIGN: Murine mAb 115D8 directed against episialin (MUC1/MAM6, epithelial membrane Ag) was used in combination with goat anti-mouse-coated magnetic microbeads to purify human T47D breast carc inoma cells (115D8(+), MDR1(-)) from different mixtures with COLO320 h uman colon carcinoma cells (115D8(-), MDR1(+)) and to purify carcinoma cells from FNA taken from axillary lymph node metastases in breast ca ncer patients. The efficacy of the purification was determined by FACS -analysis and by measurement of MDR1-mRNA levels by semiquantitative R T-PCR. RESULTS: FACS-analysis demonstrated that T47D cells could be pu rified up to 99.8% from mixtures with COLO320 cells ranging from 3:1 t o 1:3. The MDR1-mRNA level in these enriched mixtures, as detected by RT-PCR, was reduced 250-fold. It was demonstrated that MDR1 expression present in an FNA from a lymph node metastasis of breast carcinoma co uld be attributed completely to the leukocytes present in this FNA, be cause MDR1 expression was no longer detectable after purification of t he tumor cells. CONCLUSION: The combination of immunomagnetic purifica tion of breast carcinoma cells and RT-PCR enables the measurement of c ancer-specific MDR1 mRNA levels in small cell samples obtained by FNA.