Summary. Contamination of transplants with tumour cells may contribute to relapse after peripheral blood stem cell transplantation (PBSCT). We studied the feasibility of CD34 þ cell selection from blood-derived autografts obtained following G-CSF-supported cytotoxic chemotherapy in a group of 25 patients with breast cancer (10 with high-risk stage II/III and 15 with stage IV without bone or bone marrow involvement).Using immunomagnetic beads (Isolex 300 SA, Baxter) CD34 þ cells were enriched and released by chymopapain resulting in a median purity of 95% (range 82-99%) and a median recovery of 80% (range 27-132%). The enrichment procedure did not change the proportion of CD34 þ subsets coexpressing HLA-DR, CD38 and Thy-1, while L-selectin was removed from the cell surface following selection. Using a sensitive immunocytological technique with a cocktail of epithelial-specific antibodies (anti-cytokeratin 8, 18 and 19; HEA125; BM7 and BM8), five leukaphereses products contained epithelial cells, whereas the selected CD34 þ cell fraction was free of tumour cells. A neutrophil count of 0·5 × 10 9 /l and a platelet count of 20 × 10 9 /l was reached after a median time of 14 and 10 d following 40 high-dose chemotherapy (HDC) cycles. Our results indicate that immunomagnetic selection of CD34 þ cells yields highly purified autografts devoid of tumour cells whereas the engraftment ability of the progenitor and stem cells is fully retained.

To assess whether measurement of CD34+ cells in the peripheral blood allows one to estimate the progenitor cell yields of subsequent leukapheresis procedures, 733 corresponding blood and leukapheresis samples were analyzed. Peripheral blood progenitor cells of cancer patients were mobilized with hematopoietic growth factors alone or postchemotherapy, and harvested processing 10 liters of blood for each leukapheresis product. The CD34+ cell count (CD34+ cells/microliter blood) correlated most closely with the progenitor cell yield in the corresponding leukapheresis product (CD34+ cells/kg bodyweight, r = 0.80), while the proportion of circulating CD34+ cells to the white blood and mononuclear cells predicted the yield less reliably (r = 0.74 and r = 0.60). The CD34+ cell yield was independent of the white blood count (r = 0.04), whereas a weak correlation was found between the mononuclear cell count and the number of CD34+ cells/kg collected (r = 0.42). It was unlikely to obtain the threshold quantity of 2.5 x 10(6) CD34+ cells/kg required for rapid engraftment when counts below 10 CD34+ cells/microliter blood were detected. At levels between 10 and 30 CD34+ cells/microliter sufficient autografts could be harvested, whereas 30-100 CD34+ cells/microliter were required to achieve this by a single leukapheresis. A surplus of CD34+ cells was likely above 100 CD34+ cells/microliter which could be useful for progenitor cell enrichment techniques. The correlation between the CD34+ cell count and progenitor cell yield was independent of the mobilizing regimen and whether leukaphereses had been performed previously. In conclusion, the number of CD34+ cells/microliter blood allows a reliable prediction of the CD34+ progenitor cell yield in subsequent leukapheresis procedures. However, rare cases of unexpectedly sufficient progenitor cell yields may be observed even at CD34+ cell levels below detection limit.

It was the aim of our study to determine the collection efficiency and yield of CD34+ cells in 88 cancer patients (pts, 44 males/44 females) who underwent 154 large‐volume leukaphereses (LV‐LPs). The diagnoses were as follows: 18 patients had Non‐Hodgkin's lymphoma, 9 Hodgkin's disease, 24 multiple myeloma, 6 acute leukemia, 27 breast cancer, and 4 patients had solid tumors of different types. During the course of LV‐LPs, 20 liters (l) of blood were processed at a median flow‐rate of 85 ml/min (CS 3000 Baxter) and 130 ml/min (COBE Spectra), respectively. Peripheral blood stem cells (PBSC) were collected following granulocyte colony‐stimulating factor (G‐CSF)‐supported cytotoxic chemotherapy. A 31% and 21% mean decrease in the platelet and white blood count was noted at the end of the LV‐LPs when compared with the pre‐leukapheresis values. The aphereses were well tolerated without adverse effects. The level of circulating CD34+ cells was closely related to the number of CD34+ cells contained in the respective leukapheresis product (R = 0.89, P < 0.001). Compared with 270 patients who underwent 838 regular 101 LPs, the yield of CD34+ cells/kg was almost two‐fold greater (4.84 ± 0.63 × 106 [Mean ± SEM] vs 2.60 ± 0.16 × 106, P < 0.001). The antigenic profile of CD34+ cells was assessed in 54 separate products collected on the occasion of 27 LV‐LPs following the processing of 101 and 201, respectively. The intra‐individual comparison included differentiation as well as lineage‐associated markers (CD38, Thy‐1, c‐kit, CD33, CD45RA). No difference in the subset composition was observed between the first and second product, arguing against a preferential release of particular CD34+ cell subsets during the procedure. As shown by molecular biological or immunocytochemical examination, the likelihood of harvesting malignant cells using large‐volume aphereses was not increased in comparison with regular leukaphereses. Single harvests of ≥2.5 × 106 CD34+ cells/kg could be obtained in 74% of the patients, compared with 52% in case of regular LPs. As the majority of patients were autografted with more than 2.5 × 106 CD34+ cells/kg following high‐dose therapy, hematological recovery in general was rapid and not related to the type of apheresis product used. Considering patient comfort and savings in resource utilization, large‐volume leukaphereses have become the standard procedure for PBSC collection in our center. © 1996 Wiley‐Liss, Inc.

Peripheral blood progenitor cells (PBPC) can be mobilized using cytotoxic chemotherapy and cytokines. There is a substantial variability in the yield of hematopoietic progenitor cells between patients. We were looking for predictive parameters indicating a patient's response to a given mobilization regimen. Multiparameter flow-cytometry analysis and clonogenic assays were used to examine the hematopoietic progenitor cells in bone marrow (BM) and peripheral blood (PB) before filgrastim (R-metHuG-CSF; Amgen, Thousand Oaks, CA)-supported chemotherapy and in PB and leukapheresis products (LPs) in the recovery phase. Fifteen patients (four with high-grade non-Hodgkin's lymphoma [NHL], two with low-grade NHL, two with Hodgkin's disease, two with multiple myeloma, three with breast cancer, one with ovarian cancer, and one with germ cell tumor) were included in this study. The comparison of immunofluorescence plots showed a homogenous population of strongly CD34+ cells in steady-state and mobilized PB whereas in steady-state BM, the CD34+ cells ranged from strongly positive with continuous transition to the CD34- population. Consistent with the similarity in CD34 antigen expression, a correlation analysis showed steady-state PB CD34+ cells (r = .81, P < .001) and colony-forming cells (CFCs; r = .69, P < .01) to be a measure of a patient's mobilizable CD34+ cell pool. Individual estimates of progenitor cell yields could be calculated. With a probability of 95%, eg, 0.4 steady-state PB CD34+ cells x 10(6)/L allowed to collect in six LPs 2.5 x 10(6) CD34+ cells/kg, the reported threshold-dose of progenitor cells required for rapid and sustained engraftment after high-dose therapy. For the total steady-state BM CD34+ cell population, a weak correlation (r = .57, P < .05) with the mobilized CD34+ cells only became apparent when an outlier was removed from the analysis. Neither the CD34+ immunologic subgroups defined by the coexpression of the myeloid lineage-associated antigens CD33 or CD45-RA or the phenotypically primitive CD34+/HLA-DR-subset nor the BM CFC count had a predictive value for the mobilization outcome. This may be caused by the additional presence of maturing progenitor cells in BM, which express lower levels of the CD34 antigen and do not circulate. Our results permit us to recognize patients who are at risk to collect low numbers of progenitor cells and those who are likely to achieve sufficient or high progenitor cell yields even before mobilization chemotherapy is administered.