Decrease in tumor cell contamination and progenitor cell yield in leukapheresis products after consecutive cycles of chemotherapy for breast cancer treatment.

In this retrospective study, we assessed the impact of each of three consecutive cycles of conventional-dose chemotherapy on CD34+ cells, colony-forming units granulocyte-macrophage (CFU-GM), and contaminating breast cancer cells collected in the leukapheresis products of patients with metastatic breast cancer. The patients subsequently underwent high-dose chemotherapy followed by autologous blood progenitor cell transplantation. We analyzed 172 leukapheresis products from 17 patients and have correlated the long-term clinical outcome with tumor cell contamination. The induction chemotherapy regimen consisted of three cycles of cyclophosphamide 750 mg/m2 i.v., epirubicin 100 mg/m2, and 5-fluorouracil (5-FU) 750 mg/m2 i.v., followed by 5 microg/kg body weight of recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) daily until leukapheresis was completed. An average of 10 leukapheresis products (three to four collections after each cycle of chemotherapy) were obtained from each patient. Numbers of CD34+ cells, CFU-GM, and mononuclear cells (MNCs) in the leukapheresis products were determined at the time of collection. Aliquots from the same products were frozen and breast cancer cells were detected by immunocytochemistry with a cocktail of anti-cytokeratin antibodies (AE-1, AE-3, CAM 5.2, Keratin 8+18+19) using a standardized immunoalkaline phosphatase method. A minimum of 10(6) cells were examined by light microscopy and by at least two blinded observers. Cells were considered positive when immunostaining was detected in the cytoplasm and on the cell membrane, and cellular morphology was consistent with a malignant phenotype. Of the 172 samples analyzed, 13 of 57 (23%) leukapheresis products collected after cycle I were positive for tumor cells; 3 of 60 (5%) after cycle II; and 4 of 55 (7%) after cycle III. The likelihood of contamination by breast cancer cells after cycle I was significantly higher than after subsequent cycles of chemotherapy (p = 0.0052). Simultaneously, there was a significant decrease in quantity of CD34+ cells and CFU-GM (p < 0.0001 for both comparisons). Our study indicated that leukapheresis products collected after the second or third cycles of induction chemotherapy carry a significantly lower likelihood of tumor cell contamination, albeit the quantity of CD34+ cells or CFU-GM collected was also significantly reduced.