Takanori Abe

Factors associated with granulocyte colony-stimulating factor-induced peripheral blood stem cell yield in healthy donors

Background and Objectives  Poor collection results are a clinical problem in granulocyte‐colony stimulating factor (G‐CSF)‐induced peripheral blood stem cell (PBSC) collection in healthy donors. It would be beneficial to be able to predict the PBSC yield from allogeneic donors before mobilization or harvesting.

Toxicities associated with cryopreserved and thawed peripheral blood stem cell autografts in children with active cancer

To evaluate the safety of cryopreserved and thawed peripheral blood stem cell (PBSC) autografts in children with active cancer, a toxicity assessment was made of 54 PBSC transfusions to 52 children (aged 1–16 years; median, 9 years). Patients were conditioned with high‐dose chemotherapy without total body irradiation. The volume of PBSCs transfused varied from 46 to 500 mL (219.6 +/− 118.4 mL, mean +/− SD), with a mean of 0.91 g per kg of dimethyl sulfoxide. Insignificant and transient toxicities included hemoglobinuria in 40 patients (74%), headache in 38 (70%), nausea in 37 (69%), and vomiting in 25 patients (46%). Significant shock developed in 8 patients (15%), but they recovered quickly, whether they had supportive therapy or not. Vomiting and hyperbilirubinemia were the only toxicities that showed a correlation with the amount of PBSCs transfused. The data suggest that transient toxicity associated with PBSC autografts is rather common in children, and close observation of patients for possible serious morbidity is required.

Effectiveness of high-dose MCNU therapy and hematopoietic stem cell autografts treatment of childhood acute leukemia/lymphoma with high-risk features.

Clinical and pharmacokinetic studies were performed regarding the toxicity of methyl 6‐[3‐(2‐chloroethyl)‐3‐nitrosoureido]‐6‐deoxy‐α‐D‐glucopyranoside (MCNU) with other drugs, in conjunction with a peripheral blood stem cell autograft (PBSCT), for treating 26 children with acute leukemia or lymphoma associated with high‐risk features. In the early phase of the study, MCNU (300 to 500 mg/m2) was administered with cytosine arabinoside (Ara‐C) (1.6 to 16 g/m2),etoposide (VP‐16) (0.8 to 1.6 g/m2), cyclophosphamide (CY) (100 to 200 mg/kg), or busulfan (16 mg/kg). No acute toxicity was noticed after this high‐dose therapy. The dose‐limiting factor of the regimens was significant but reversible interstitial pneumonitis (IP). In a subsequent trial with an MCNU/VP‐16/Ara‐C/CY (MCVAC) regimen in which the dose of MCNU was reduced, the risk of IP diminished. This study is still in progress, but the clinical response has so far been encouraging. Fifteen of 26 children are alive and well in unmaintained complete remission (CR) with a median follow‐up period of 11 months (range, 3 to 34 months) after transplantation.

Peripheral blood stem cell mobilization by granulocyte colony-stimulating factor alone and engraftment kinetics following autologous transplantation in children and adolescents with solid tumor

In 56 pediatric and adolescent patients (median age 7 years, range 1–21) with various solid tumors, peripheral blood stem cells (PBSC) were mobilized with granulocyte colony-stimulating factor (G-CSF) alone, and the yields of PBSC and engraftment kinetics following autologous peripheral blood stem cell transplantation (PBSCT) were evaluated retrospectively. Granulocyte colony-stimulating factor (10 μg/kg) was injected subcutaneously for mobilization when patients showed no influence of previous chemotherapy, and administration was continued for 5 days. The peaks of CD34+ cells and colony-forming units-granulocyte/macrophage in the blood were observed on days 4 through 6 of G-CSF administration in all patients. Peripheral blood stem cell harvest was commenced on day 5 of G-CSF treatment. Compared to the results in patients mobilized by chemotherapy plus G-CSF (N=18), the progenitor cell yields were lower in patients mobilized with G-CSF alone. However, there were no significant differences in WBC and ANC engraftment compared to the chemotherapy plus G-CSF mobilization group. Platelet recovery following autologous PBSCT was delayed in patients mobilized with G-CSF alone. The median time taken for ANC and platelet counts to reach 0.5 × 109 and 20 × 109/l was 12 days (range: 9–28) and 15 days (8–55), respectively, in all patients who received PBSC mobilized by G-CSF alone. In summary, mobilization with G-CSF alone can mobilize sufficient CD34+ cells for successful autografting and sustained hematological reconstitution in pediatric and adolescent patients with solid tumors, and even in heavily pre-treated patients.

Allogeneic peripheral blood stem cell transplantation in children with hematologic malignancies from HLA-matched siblings

BACKGROUND Despite the ethical problem of using granulocyte colony-stimulating factor (G-CSF) in normal children, allogeneic peripheral blood stem cell transplantation (PBSCT) might have advantages over allogeneic bone marrow transplantation (BMT). PROCEDURE Eleven HLA-matched sibling donors aged 2-16 years received 10 microg/kg/day G-CSF for 5 days and underwent apheresis to harvest peripheral blood stem cells (PBSC). PBSC were then cryopreserved until infusion. The 11 corresponding patients aged 8 months to 14 years with high-risk hematological malignancies received busulfan (16 mg/kg or 600 mg/m(2)) and melphalan (210 mg/m(2)) as a preparative regimen. Graft-versus-host disease (GVHD) prophylaxis consisted of cyclosporine and methylprednisolone. RESULTS All of the donors tolerated G-CSF administration and apheresis procedures. The patients received a median of 5.8 (range 1. 4-11.5) x 10(6)/kg CD34(+) cells, 17.2 (3.8-36.0) x 10(5)/kg colony forming units-granulocyte/macrophage (CFU-GM), and 3.5 (1.4-7.1) x 10(8)/kg CD3(+) cells. All of the patients showed prompt engraftment, with a median time to reach an absolute neutrophil count (ANC) above 0.5 x 10(9)/liter of 10 (9-13) days. Grade I acute GVHD occurred in seven patients (64%), whereas grade II-IV acute GVHD was not seen. Chronic GVHD occurred in four patients (40%) among 10 patients evaluable for chronic GVHD. Three patients showed extensive chronic GVHD. Currently, eight patients (73%) are alive and disease-free for a median follow-up of 775 (103-1,069) days. CONCLUSIONS Allogeneic PBSCT is feasible in the pediatric population, and PBSC harvest is an alternative to BM harvest in donors who are not eligible for BM harvest. Furthermore, PBSC were successfully collected in pediatric donors with peripheral access. The choice of a stem cell source should be based on the risk/benefit assessment for both patients and donors.

Cryopreservation of mobilized blood stem cells at a higher cell concentration without the use of a programmed freezer

Cryopreservation of peripheral blood stem cells (PBSC) mobilized by chemotherapy combined with or without granulocyte colony-stimulating factor (G-CSF) is an essential part of procedure for anti-cancer strategies. We evaluated whether a higher cell concentration (2×108/ml) without the use of a programmed freezer was acceptable for the storage of mobilized PBSC in an autologous setting. Mobilized PBSC were enriched to mononuclear cells (MNC) by Percoll separation and then frozen at cell concentrations of 2–5×107/ml (group I, n=20) or 2×108/ml (group II, n=44) without the use of a programmed freezer using 5% DMSO, 6% hydroxy ethyl starch, and 4% autologous serum or human albumin. CD34+ cells purified by ISOLEX300 were frozen at 2×107/ml (group III, n=22) using the same method. The median recovery rates of CD34+ cells and CFU-GM were, respectively, n.d. (not determined) and 88% in group I, 103 and 64% in group II, and 98 and 53% in group III. There was a statistical significance between the recovery rate of CFU-GM in group III and that in group I (p=0.02). The median percentage of cell viability after thawing in each group was 89, 87, and 75%, respectively. The median numbers of days after PBSCT to achieve a WBC of >1.0×109/l, an absolute neutrophil count of >0.5×109/l, and a platelet count of >50×109/l were, respectively, 11, 11 and 15 in group I; 12, 12 and 16 in group II; and 12, 12 and 27 in group III. These results suggest that enriched MNC from mobilized PBSC could be frozen at a higher cell concentration (2×108/ml) without the use of a programmed freezer, leading to reduction of the toxicities associated with infusion of thawed cells and of costly space required for cell storage.

Intra-apheresis recruitment of blood progenitor cells in children

BACKGROUND: Determination of the optimal duration of apheresis requires a careful examination of blood progenitor cell (BPC) kinetics during apheresis. Intra‐apheresis recruitment of BPCs should be evaluated.

Syndrome of Inappropriate Antidiuretic Hormone Secretion (Siadh) in Children Undergoing High-Dose Chemotherapy and Autologous Peripheral Blood Stem Cell Transplantation

The incidence of SIADH (the syndrome of inappropriate antidiuretic hormone secretion) was analyzed retrospectively in 43 children who received marrow-ablative chemotherapy before autografts with peripheral blood stem cells for lymphoid malignancies. SIADH was documented in three children (ages 3, 13, and 13 years) who received chemotherapy, which included high-dose methyl 6-[3-(chloroethyl)-3-nitrosoureido]-6-deoxy-alpha-D-glucopyranoside (MCNU) and cyclophosphamide, under a concomitant overhydration protocol. SIADH was manifested as frequent vomiting in two patients and as generalized seizure in one. Hyponatremia (< 125 mEq/L), hypo-osmolality (< 260 mOsm/kgH2O), and continued urinary excretion of sodium (> 30 mEq/L) were used to diagnose SIADH in these three patients. All signs and symptoms subsided within 24 hours either by fluid restriction alone (n = 1) or by supportive care including anticonvulsant and D-mannitol, or hyperhydration with saline plus 5% glucose and diuretic. None of the patients died. Careful monitoring of the serum sodium level, as well as the osmolality of plasma and urine, should be incorporated into the patient management protocol for this type of high-dose chemotherapy.

Granulocyte/Macrophage Origin of Glomerular Mesangial Cells

We previously demonstrated the ability of hematopoietic stem cells (HSCs) to generate glomerular mesangial cells by transplanting clonal populations of cells derived from a single enhanced green fluorescent protein (EGFP)-positive HSC into lethally irradiated mice. To define more precisely the hematopoietic differentiation pathway through which mesangial cells are derived, we studied the relationship between mesangial cell expression and individual hematopoietic lineages by means of a transplantation strategy. In a series of clonal HSC transplantation experiments, we generated 3 mice engrafted predominantly by granulocytes and macrophages (GMs) and 4 mice engrafted with B-cells or with B-cells and T-cells. When the kidneys of these mice were analyzed, the mice exhibiting high GM lineage engraftment revealed much higher levels of EGFP-positive mesangial cells than those with predominantly lymphocyte engraftment. Fluorescence in situ hybridization analysis of the kidneys from a male recipient of an EGFP-positive female donor excluded cell fusion as the cause for the observed differentiation. These results support the notion that glomerular mesangial cells share their origin with GMs.

New consecutive high-dose chemotherapy modality with fractionated blood stem cell support in the treatment of high-risk pediatric solid tumors: a feasibility study.

For the treatment of childhood solid tumors, we performed a pilot feasibility study of consecutive high-dose therapies, in which each course was followed by transplantation with granulocyte colony-stimulating factor-mobilized peripheral blood cells which had been separated into CD34-positive and -negative fractions by an Isolex system (Baxter). Positive selection of CD34+ cells has been associated with inevitable cell loss. To overcome this loss, CD34+ cells that had migrated into the negative fraction were saved and used for the first transplant, which was followed by a second transplant after a 3- to 5-month interval. In this phase I feasibility study, the results in six children were evaluated for safety and engraftment. Multi-drug cytoreductive regimens using ranimustine (MCNU), melphalan, thiotepa, carboplatin, cyclophosphamide or VP-16 were comparable between the two transplant procedures in terms of their intensity. The number of CD34+ cells in the ‘CD34(+) fraction’ was 3.31 × 106/kg (0.63–4.3 × 106/kg), while this number in the ‘CD34(−) fraction’ could not be evaluated correctly due their scarcity (<0.1%). the median numbers of infused mnc and cfu-gm were, respectively, 4.2 × 106/kg and 1.75 × 105/kg in the CD34(+) fraction, and 4.8 × 108/kg and 3.35 × 105/kg in the CD34(−) fraction. The number of days required to achieve an ANC >0.5 × 109/l and a platelet count >20 × 109/l and >50 × 109/l were, respectively, 14.5, 15.0 and 19.5 in the first transplant with CD34− cells, and 13.5, 18.0 and 25.0 in the second transplant with CD34+ cells, with no essential difference between the two treatments. Although the small number of patients, the variation in clinical status and treatment, and the short follow-up invalidate any evaluation of the therapeutic benefit of this strategy, the encouraging results support the feasibility of this strategy, which enables an escalation of dose intensity with an improved cost/benefit ratio.