Lorella Torretta

Collection of circulating progenitor cells after epirubicin, paclitaxel and filgrastim in patients with metastatic breast cancer.

The efficacy of high-dose chemotherapy (HDC) and circulating progenitor cell (CPC) transplantation in metastatic breast cancer (MBC) relies mainly on giving this treatment after a response to conventional induction chemotherapy has been achieved. For this reason an optimal mobilization regimen should be therapeutically effective while minimizing the number of leucaphereses required to support the myeloablative therapy. The combination of an anthracycline and paclitaxel in chemotherapy-untreated MBC has produced impressive response rates. We evaluated the CPC-mobilizing capacity of the combination epirubicin (90 mg m(-2)) and paclitaxel (135 mg m(-2)) followed by filgrastim (5 microg kg(-1) day(-1)) starting 48 h after chemotherapy administration in ten patients with MBC who were eligible for an HDC and CPC transplantation programme. Leucaphereses were performed by processing at least two blood volumes per procedure at recovery from neutrophil nadir when CD34+ cells in the peripheral blood exceeded 20 microl(-1). In most patients (six out of 10) more than 2.5 x 10(6) CD34+ cells kg(-1), a threshold considered to be sufficient for haematopoietic reconstitution, were collected with a single apheresis. In the remaining four patients an additional procedure, performed the following day, was enough to reach the required number of progenitors. These data suggest that the epirubicin-paclitaxel combination, besides being a very active regimen in MBC, is effective in releasing large amounts of progenitor cells into circulation.

Adverse effects associated with extracorporeal photochemotherapy.

Adverse effects associated with extracorporeal photochemotherapy The interesting report in TRANSFUSION by McLeod et al.1 described the frequency of immediate, adverse effects associated with therapeutic apheresis, including no adverse reactions in 79 extracorporeal photopheresis (ECP) procedures that were performed with three different instruments. We would like to report the frequency of adverse effects associated with 720 ECP procedures that were performed at our hospital between January 1997 and May 1999. The patients treated had immune or autoimmune diseases, including acute and chronic graft-versus-host disease (23 patients), pemphigus vulgaris (5 patients), interferon-resistant hepatitis C virus infection (1 patient), multiple sclerosis (1 patient), and dermatomyositis (1 patient). There were 18 children and 13 adults. Our ECP procedure consisted of three steps. First, peripheral blood mononuclear cells were collected by the cytapheresis of 2 total blood volumes (Spectra, COBE, Lakewood, CO). The mean number of mononuclear cells collected per ECP was 6.6 × 109. Second, 8-methoxypsoralen (200 ng/mL; 8-MOP, Gerot Pharmazeutika, Vienna, Austria) was added to the collection bag, and this was followed by ultraviolet A radiation (2 J/cm2 of white cell-surface area; UV-MATIC, Vilbert-Lourmat, Marne-la-Vallée, France), as described by Andreu et al.2 Third, the 8-methoxypsoralenphotoactivated cells were returned to the patient by intravenous transfusion within 30 minutes of collection, by the method of Perotti et al.3 Using criteria of the Canadian Apheresis Study Group,4 we graded any adverse effects occurring during the 24 hours after transfusion as mild, moderate, or severe. Nine adverse effects were observed; all were mild, as follows: chills (4 episodes), headache (3), and posttransfusion fever <38°C (2). We believe that the headaches and fevers, all of which occurred after transfusion, are most probably related to the release of cytokines from photoaltered cells. In our experience, ECP was safe, even for critically ill pediatric patients with grade IV graft-versus-host disease. The low incidence and mild nature of adverse effects were important factors in securing patients’ acceptance of long-term treatment. Laura Salvaneschi, MD Cesare Perotti, MD c.perotti@smatteo.pv.it Lorella Torretta, MD Servizio di Immunoematologia e Trasfusione IRCCS Policlinico San Matteo Pavia, Italy

Both cycling and noncycling primitive progenitors continue to be mobilized into the circulation during the leukapheresis of patients pretreated with chemotherapy and G‐CSF

Colony‐forming cells (CFC) and long‐term culture‐initiating cells (LTC‐IC) include a spectrum of progenitor types whose potential contributions to the haemopoietic recovery seen in patients transplanted with mobilized peripheral blood progenitor cells (PBPC) remains unclear. We evaluated both the number and cycling status of the circulating LTC‐IC and CFC harvested from 12 patients treated with chemotherapy and G‐CSF using a modified 6‐week LTC‐IC assay. The frequency of the LTC‐IC and CFC in the mobilized PB samples were increased 45‐ and 750‐fold, respectively. Interestingly, comparison of these values for PB samples, taken just prior to the start of the leukapheresis, with the progenitor content of the 3 h harvest, showed that, on average, the leukapheresis product contained 19 times more LTC‐IC (P < 0.01) than had been detectable in the entire blood volume of the patients at the start of the collection, whereas the number of CFC collected was approximately the same as the number in the initial circulating pool of PBPC. Cycling studies showed many of the LTC‐IC in the apheresis collections to be proliferating although not more so than in the steady‐state marrow LTC‐IC compartment (i.e. per cent kill of mobilized LTC‐IC after 16 h in 3H‐Tdr = 70 ± 8%, n = 9). On the other hand, the majority of the CFC in the apheresis collections were initially quiescent (per cent kill after 16 h in 3H‐Tdr = 37 ± 6%, n = 12). These findings demonstrate the rapidity with which a primitive subset of LTC‐IC may enter the circulation during the early phase of rebound haemopoiesis induced by chemotherapy plus G‐CSF and provide evidence of differences in the mechanisms regulating LTC‐IC and CFC mobilization.

Hematopoietic progenitor cell collection and neoplastic cell contamination in breast cancer patients receiving chemotherapy plus granulocyte-colony stimulating factor (G-CSF) or G-CSF alone for mobilization

BACKGROUND We compared hematopoietic progenitor cell (HPC) collection and neoplastic cell contamination in breast cancer patients given cyclophosphamide (CTX) plus granulocyte-colony stimulating factor (G-CSF) or G-CSF alone for mobilization. PATIENTS AND METHODS In 57 stage II-III breast cancer patients, CD34+ cells, colony-forming units-granulocyte macrophage (CFU-GM), early HPC and breast cancer cells were counted in HPC collections obtained after CTX plus G-CSF (n = 27) or G-CSF-alone mobilization (n = 30). RESULTS The CD34+ cell collection was about two-fold greater after CTX plus G-CSF mobilization (11.0 +/- 7.9 vs. 5.8 +/- 3.5 x 10(6)/kg, P < 0.001). Similarly, the total number of CFU-GM, CD34+CD38- cells and of week-5 cobblestone area forming cells (CAFC) collected was significantly higher in patients mobilized with CTX plus G-CSF. Breast cancer cells were found in the apheresis products of 22% of patients mobilized with CTX plus G-CSF and in 10% of patients mobilized with G-CSF alone (P = 0.36). Of seven patients who failed G-CSF-alone mobilization and eventually underwent chemotherapy plus G-CSF mobilization, none had cytokeratin-positive cells after G-CSF mobilization, whereas four out of seven had cytokeratin-positive cells after chemotherapy plus G-CSF (P = 0.07 by chi 2 test). CONCLUSION The CTX plus G-CSF mobilization protocol was associated with a significantly higher HPC collection. However, this benefit was not accompanied by a reduction in the incidence of tumor-contaminated HPC graft.

Autologous Platelet Collection and Storage to Support Thrombocytopenia in Patients Undergoing High-Dose Chemotherapy and Circulating Progenitor Cell Transplantation for High-Risk Breast Cancer

Objectives: The use of circulating progenitor cell support following high-dose chemotherapy for malignancies decreases but does not entirely abolish platelet transfusion requirement. We investigated the feasibility of supporting the posttransplant thrombocytopenic phase exclusively with autologous platelets collected by apheresis and cryopreserved. Methods: 25 patients underwent plateletpheresis during the platelet rebound occurring after high-dose cyclophosphamide. Autologous platelets were cryopreserved in 5% dimethylsulfoxide, thawed and transfused during the aplastic phase after the myeloablative regimen whenever clinically required. Results: A single plateletpheresis was carried out in all patients, allowing the harvest of a platelet concentrate with a mean value of 7.7 × 1011 platelets. No significant procedure- or transfusion-related side effects were recorded. Mean platelet recovery after freezing and thawing was 63% and the mean number of platelet reinfused was 4.8 × 1011; 23 of 25 patients were fully supported with autologous platelets. Conclusion: Plateletpheresis performed in our selected group of patients was found to be a safe and effective procedure to collect large amounts of autologous platelets; the numbers obtained proved to be sufficient for the transfusion demand of almost all patients.

Peripheral blood stem cell collection from healthy donors for allogeneic transplantation

There is great interest in the use of peripheral blood stem cells (PBSC) for allogeneic transplantation, based on the good results seen with autologous PBSC infusion. Reasonable caution exists regarding the use of allogeneic PBSC for transplantation because of donor toxicities due to rhG-CSF administration and the risk of graft-versus-host-disease (GVHD) in the recipient because of the large number of T-cell infused. We present preliminary data on allogeneic PBSC collections and transplantation in ten patients affected by advanced leukemia (eight patients), severe aplastic anemia (one patient) and sickle cell anemia (one patient). Seven donors were HLA-identical siblings, while the other three were mismatched for three, two and one locus, respectively. All donors received rhG-CSF at a dose of 12 micrograms/kg for a mean of 5 days. Leukaphereses were performed with the aim of collecting a minimum of 5 x 10(6)/kg (recipients weight) CD 34+ cells. Collection timing was determined by monitoring CD 34+ cells in the donors peripheral blood from the second day of rhG-CSF therapy. The PBSC collections yielded a mean of 10.05 x 10(8) MNCs/kg and of 10.48 x 10(6) CD 34+ cells/kg (recipients weight). PBSC were immediately infused after collection in patients given myeloablative therapy. Engraftment was observed in each patient at a mean of 13.2 days for an absolute neutrophil count (ANC) more than 0.5 x 10(9)/L and of 26.5 days for a platelet count of more than 20 x 10(9)/L. Eight patients experienced no or moderate acute GVHD, whereas two patients died of grade 4 GVHD, notwithstanding GVHD prophylaxis with cyclosporine and prednisone. Two other patients died of viral and fungal infections, respectively, despite prophylaxis. The remaining six patients are alive between 58 and 430 days after transplant. Our results document that allogeneic PBSC are capable of engraftment after a myeloablative regimen. Controlled trials are necessary to compare the potential benefits of this approach with the results obtained in allogeneic bone marrow transplantation.