D. Seong

Clinical toxicity and laboratory effects of granulocyte-colony-stimulating factor (filgrastim) mobilization and blood stem cell apheresis from normal donors, and analysis of charges for the procedures.

BACKGROUND: Apheresis of granulocyte‐colony‐stimulating factor (filgrastim)‐mobilized blood stem cells from normal donors is now being used in place of a marrow harvest in transplantation. How the adverse effects of and charges for this procedure compare with those of the standard marrow harvest is not known.

Dual Alu polymerase chain reaction primers and conditions for isolation of human chromosome painting probes from hybrid cells

A method for rapid and efficient production of chromosome- and chromosome-region specific probes for fluorescent in situ hybridization (FISH) detectable by simple fluorescent microscopy is described. The procedure is based on simultaneous use of two inter-Alu-polymerase chain reaction (PCR) primers for extraction of highly heterogeneous human DNA from interspecific somatic cell hybrids containing the chromosome regions of interest. Probes so produced do not hybridize to centromeric sequences and simultaneously band the target chromosomes, making them useful for unambiguous identification of chromosomal elements and breakpoints associated with cancer.

Collection of peripheral blood stem cells from normal donors 60 years of age or older

We report 14 normal peripheral blood stem cell (PBSC) donors ≥ 60 years of age who had cytokine mobilization followed by PBSC apheresis for allogeneic transplantation. Mobilization was achieved with filgrastim (6 μg/kg twice daily). Their median age was 63.5 years (range 60–77), and 43% had a positive medical history, mainly hypertension and/or cardiac problems. Their median pre‐apheresis leucocyte count (×109/l) was 38.6 (range 29.6–63.4). The median apheresis yield (×106 CD34+ cells/litre blood processed, first apheresis) was 27.9 (range 1.6–54.8). The target cell dose (≥4& times; 106 CD34+ cells/kg recipient) was reached with one procedure in eight (57%) donors. Filgrastim‐related adverse events were acceptable and apheresis was well tolerated. When compared to younger donors (<60 years of age), a trend to a lower CD34+ apheresis yield and a requirement for more than one apheresis to achieve the collection target (≥4 ×106 CD34+ cells/kg) was evident. Although older (≥60 years) donors seem to mobilize less effectively, these data suggest that PBSC collection from them is feasible and has an acceptable short‐term safety profile.

Duration of filgrastim mobilization and apheresis yield of CD34+ progenitor cells and lymphoid subsets in normal donors for allogeneic transplantation.

Seventy‐seven normal donors underwent leukapheresis for peripheral blood progenitor cell collection beginning on day 4 (n = 45) or day 5 (n = 32) of filgrastim mobilization (12 μg/kg/d). The two groups were comparable for age, weight, blood volumes processed during leukapheresis and target CD34+ cell dose to be collected. The day 5 schedule allowed a more consistent achievement of the target cell dose with one apheresis (P = 0.005) and resulted in the initial collection of a significantly larger number of CD34+ cells (P = 0.009). There was no statistically significant difference in the leukapheresis yield of lymphoid subsets and natural killer cells.

Phase I trial of cyclosporine-induced autologous graft-versus-host disease in patients with multiple myeloma undergoing high-dose chemotherapy with autologous stem-cell rescue.

PURPOSE To determine the feasibility and toxicity of inducing autologous graft-versus-host disease (GVHD) with cyclosporine in patients with multiple myeloma undergoing autologous stem-cell transplantation. PATIENTS AND METHODS Fourteen multiple myeloma patients with a median age of 50 years (range, 41 to 63) were enrolled. The median time from diagnosis to transplant was 651 days (range, 229 to 3,353). Ten patients had primary refractory disease, two were in first remission, and two were responsive to salvage therapy. The preparative regimen consisted of thiotepa, busulfan, and cyclophosphamide. Cyclosporine was administered daily for 28 days after the stem-cell infusion, and the dose was adjusted to maintain whole-blood cyclosporine levels between 50 and 150 ng/dL in the first seven patients (low-level group) and between 150 and 300 ng/dL in the other seven patients (high-level group). RESULTS All patients achieved neutrophil engraftment a median of 11 days after transplant. Four patients developed > or = grade 2 hepatic toxicity, six developed > or = grade 2 nephrotoxicity, and four developed reversible cardiac toxicity. Only one treatment-related death occurred. Cyclosporine was withheld in seven patients for a median of 6 days because of renal and/or liver dysfunction. One patient developed clinical skin GVHD, which responded to corticosteroid therapy. Six patients developed histologic evidence of GVHD without clinical signs of GVHD (subclinical GVHD). The incidence of clinical and subclinical GVHD was similar in both cyclosporine groups. Three of 11 patients assessable for response achieved remissions. Three patients experienced disease progression 80, 160, and 354 days after transplant. Ten patients are alive without progression between 56 and 444 days after transplant. CONCLUSION Induction of autologous GVHD by posttransplant cyclosporine is feasible and well tolerated in patients with multiple myeloma.

Peripheral blood stem cell apheresis in normal donors : feasibility and yield of second collections

We report 13 normal peripheral blood stem cell (PBSC) donors who had a second PBSC collection for allogeneic transplantation performed after the first. The median interval between the first and second collection was 5 months. Mobilization was achieved with filgrastim (12 μg/kg/d). No significant difference was found in the median pre‐apheresis leucocyte count (×109/l) between the two donations (40.2 v 38.5; P = 0.91). The median apheresis yield (×106 CD34+ cells/litre blood processed, first apheresis) was also similar (28 v 27.3; P = 0.91). Filgrastim‐related adverse events were comparable. These data suggest that second PBSC collections are feasible, similarly tolerated and provide comparable apheresis yields.

Transient neutropenia in normal donors after G-CSF mobilization and stem cell apheresis

Thirteen normal adult donors underwent daily leukapheresis for peripheral blood progenitor cell collection for allografting beginning on day 4 or 5 of G‐CSF mobilization (12 μg/kg/d). They had complete blood counts performed 7–10 d after the completion of the procedure. A reduction in the total leucocyte count below baseline levels, accounted for by a decrease in the absolute neutrophil (ANC) and lymphocyte counts, was noted. Neutropenia (ANC < 1.5 × 109/l) occurred in two (15%) of the donors. The lowest ANC observed was 0.6 × 109/l. The neutropenia was asymptomatic and resolved on follow‐up evaluation.

Usefulness of detection of minimal residual disease by ‘hypermetaphase’ fluorescent in situ hybridization after allogeneic BMT for chronic myelogenous leukemia

Hypermetaphase FISH (HMF), a molecular cytogenetic procedure combining the long term mitotic arrest of bone marrow cultures with detection of a specific chromosomal rearrangement by fluorescence in situ hybridization (FISH), has recently been shown to be effective in determining the frequency of Philadelphia chromosome positive (Ph+) cells in the bone marrow of chronic myelogenous leukemia (CML) patients undergoing treatment. By combining the probe for the Ph chromosome with one for the detection of the X chromosome, we used HMF to monitor the presence of malignant cells within the emerging host cell population in the marrow of a CML patient that had undergone sex-mismatched allogeneic bone marrow transplantation. In successive studies, we detected 0.5% and 1.75% Ph+ cells, respectively, confirmed by Western blot analysis for p210 protein. These readings occurred concordantly with a repopulation of host-derived diploid female cells. Standard G-band cytogenetic analyses did not detect any Ph+ cells at these time points. Intervention with donor lymphocyte infusion reinduced complete remission. This experience indicates that HMF is useful to identify low levels of repopulation by Ph+ cells in the marrow post-BMT at a stage when intervention is most efficacious.

Detection of variant Ph-positive chronic myelogenous leukemia involving chromosomes 1, 9, and 22 by fluorescence in situ hybridization

Abstract Probes made by inter- Alu -PCR from an interspecific somatic cell hybrid containing only the distal half of the 9q as its only human genomic content when used for fluorescence in situ hybridization (FISH) on the cells of a chronic myelogenous leukemia (CML) patient detected involvement of chromosome 9 in a complex translocation that was not observed by standard G-band cytogenetics.

New Directions in the Biology and Therapy of Chronic Myeloid Leukemia

Great excitement surrounded the initial discovery of the Philadelphia chromosome translocation and the molecular rearrangement of the abl protooncogene which is generated by this translocation in chronic myelogenous leukemia (CML). In this review, we will discuss how molecular analysis of the biological abnormalities generated in the CML cells by this molecular rearrangement can be applied to new directions in therapy for this disease.