Immature reticulocytes as an early predictor of engraftment in autologous and allogeneic bone marrow transplantation

The purpose of this study was to evaluate reticulocyte parameters by means of flow cytometric reticulocyte counting in a group of patients who had undergone autologous and allogeneic bone marrow transplantation (BMT). The pattern of reticulocyte response and the predictive value of absolute neutrophil count (ANC), platelet count, number of CD34+ cell infused and graft source for reticulocyte response were studied. We compared absolute reticulocyte count (RetAbs), mean fluorescence index (MFI) and mean reticulocyte volume/mean corpuscular volume (MRV/MCV) ratio with conventional criteria (ANC and platelet count) in 22 allogeneic and 20 autologous BMT recipients. An abrupt increase in MRV/MCV ratio or a rise in MFI value were the earliest signs of erythropoietic recovery following allogeneic transplantation (63.6 and 22.8% of cases, respectively). In 13.6% of the cases, both parameters were observed simultaneously. All but three autologous transplant recipients showed changes in reticulocyte parameters earlier than ANC recovery. Granulocyte recovery and peripheral blood progenitor cells (PBPC) graft were predictive variables for RetAbs response in allogeneic transplant recipients. In the autologous group, predictive variables for RetAbs response were a high number of CD34+ infused cells and platelet recovery. An increase in the immature reticulocyte population is the earliest sign of haematopoietic recovery following BMT.     

Evaluation of erythropoiesis after bone marrow transplantation: quantitative reticulocyte counting

Erythroid regeneration is an important and separate element in the engraftment process in allogeneic and autologous bone marrow transplantation (alloBMT, autoBMT). Qualitative visual reticulocyte counting has proved inadequate in the evaluation of erythropoiesis after BMT but automated flow cytometry now allows the reliable quantitation of reticulocytes even to very low levels. Reticulocyte counts and highly fluorescent reticulocyte (HFR) counts (very early reticulocytes) were estimated daily in recipients of 22 autoBMT and 14 alloBMT using a Sysmex R-1000 automated reticulocyte counter. Marrow ablation caused an immediate and rapid fall in both the reticulocyte count and the HFR. Measurable numbers of reticulocytes persisted throughout the hypoplastic period, but HFR fell to zero in the majority of both the autoBMT and alloBMT. HFR rose significantly after a median time of 14 d post-autoBMT, and 12 d post-alloBMT. Attainment of 15 x 10(9)/l reticulocytes and 0.5 x 10(9)/l HFR at day 21 post-transplant was associated with ultimate engraftment in 100% cases. Inadequate engraftment was seen in the majority of patients whose responses fell below these levels. Graft-versus-host disease was associated with a transient slight reduction in reticulocyte count. Neither episodes of infection nor blood transfusions had any significant impact on trends of reticulocytes or HFR. Automated flow cytometric reticulocyte counting has been shown to provide an accessible measure of erythroid activity which may be of predictive value in the management of patients following bone marrow transplantation.     

Assessment of hematologic progenitor engraftment by complete reticulocyte maturation parameters after autologous and allogeneic hematopoietic stem cell transplantation

BACKGROUND AND OBJECTIVES: Hematopoietic restoration after marrow ablation is initiated by the erythroid compartment. However, the absolute microscope counts or corrected percentage of reticulocytes have proven to be poor markers of hematopoietic engraftment. Some reports have highlighted the usefulness of automatic flow cytometry methods to determine highly fluorescent reticulocytes, or mean fluorescence index. In this series of 60 hematopoietic stem cell transplants, we sought the normal kinetics throughout the post-transplant period of the following reticulocyte maturing parameters: highly fluorescent reticulocytes (RETH), immature reticulocyte fraction (IRF), mean fluorescence index (MFI) and also mean reticulocyte volume (MRV). DESIGN AND METHODS: Sixty consecutive patients undergoing allogeneic bone marrow (30 cases) and autologous mobilized stem cell transplantation (30 cases) were studied. Parameters of reticulocyte maturation were measured every other day from the beginning of the conditioning regimen until myeloid engraftment. RESULTS: Nadir values for the analyzed reticulocyte parameters were found between days +4 and +7 and thereafter, increases in these reticulocyte parameters appeared earlier than the rise in neutrophils. We considered erythroid engraftment to have occurred on the day when RETH reached 3%, IRF 10%, MFI 10 and MRV 110 fL. These cut-offs were assigned considering the 25% quartile for each parameter on the day that the myeloid engraftment occurred. The median engraftment days for RETH were +9 and +16, for IRF +9 and +13, for MFI +9 and +13 and for MRV +11 and +13 in autologous and allogeneic procedures, respectively. When compared to standard neutrophil engraftment, IRF and MFI engraftment occurred significantly earlier in all patients. Remarkably, we found a statistical correlation between the day a reticulocyte parameter reached its cut-off and the subsequent day of absolute neutrophil count (ANC) recovery for MFI after allogeneic transplants and for MRV after autologous procedures (p < 0.001 and p= 0.02, respectively). Of all the clinical parameters tested, only the number of infused CD34 cells showed a statistical influence on erythroid engraftment in autologous transplant. INTERPRETATION AND CONCLUSIONS: Early reticulocytes appear sooner than neutrophils after both autologous and allogeneic transplants, and any determined reticulocyte parameter can reliably measure this fraction. Nevertheless, our results show that MRV and MFI cut-offs are useful for determining subsequent myeloid engraftment. These findings could be relevant to decision-making in those patients with primary graft failure heralded by an absence of increasing values of MFI and MRV, indicating very low production of reticulocytes from the graft, who could, therefore, benefit from earlier rescue therapy.     

Recovery from neutropenia can be predicted by the immature reticulocyte fraction several days before neutrophil recovery in autologous stem cell transplant recipients

The duration of neutropenia (absolute neutrophil count (ANC) < or = 100/microl) identifies cancer patients at risk for infection. A test that precedes ANC > or = 100/microl would be of clinical value. The immature reticulocyte fraction (IRF) reflects erythroid engraftment and hence a recovering marrow. We evaluated the IRF as predictor of marrow recovery among 90 myeloma patients undergoing their first and second (75 patients) melphalan-based autologous stem cell transplantation (Mel-ASCT). The time to IRF doubling (IRF-D) preceded ANC > or = 100/microl in 99% of patients after the first Mel-ASCT by (mean+/-s.d.) 4.23+/-1.96 days and in 97% of the patients after the second Mel-ASCT by 4.11+/-1.95 days. We validated these findings in a group of 117 myeloma patients and 99 patients with various disorders undergoing ASCT with different conditioning regimens. We also compared the time to hypophosphatemia and to absolute monocyte count > or = 100/microl to the time to ANC > or = 100/microl. These markers were reached prior to this ANC end point in 55 and 25% of patients but were almost always preceded by IRF-D. We conclude that the IRF-D is a simple, inexpensive and widely available test that can predict marrow recovery several days before ANC> or = 100/microl.     

Frequency of the development of RNA-rich reticulocytes in allogeneic bone marrow transplant recipients]

We measured appearance rates of RNA-rich reticulocytes in 8 patients undergoing allogeneic bone marrow transplantation, using a Sysmex R-1000 reticulocyte counter which utilizes laser flow cytometry. The changes in proportion of RNA-rich reticulocytes (high fluorescence ratio: HFR) and maturation index (MI: HFR + middle fluorescence ratio/low fluorescence ratio) were compared with those of WBC, neutrophil or reticulocyte counts. Engraftment was defined as an HFR of greater than or equal to 5% or a MI of greater than or equal to 15%. Engraftment was confirmed significantly earlier by HFR (13.5 +/- 2.4 days) and MI (13.1 +/- 2.9 days) than by neutrophils (17.1 +/- 3.2 days) or reticulocytes (20.4 +/- 6.2 days). The maturation of reticulocytes would be a useful indicator for engraftment or recovery from marrow aplasia in cases of bone marrow transplantation.     

采用未成熟网织红细胞指数监测恶性肿瘤患者放疗前、后骨髓功能恢复的临床价值

目的　探讨采用未成熟网织红细胞指数 (IRF)监测恶性肿瘤患者放疗前、后骨髓功能恢复的临床价值。方法　采用XE 2 10 0型血液细胞分析仪检测未成熟网织红细胞指数 (IRF)、网织红细胞比率 (RET)和嗜中性粒细胞计数 (NEUT )。结果　患者组放疗后第 10天以上 3项指标均显著低于对照组 (P <0 .0 1) ,放疗后第 15天IRF开始增高 ,并显著高于第 10天的水平 (P <0 .0 1) ,而RET、NEUT未发生显著改变 (P >0 .0 5 ) ,患者组放疗后第 2 0天IRF继续升高 ,并显著高于第 15天的水平 (P <0 .0 1) ,RET和NEUT也开始增高 ,但与第 15天比较 ,无显著性差异 (P >0 .0 5 )。患者组放疗后第 2 5天 ,IRF已经基本恢复至放疗前水平 ,RET和NEUT也开始显著升高 ,但仍低于放疗前水平 (P <0 .0 1)。结论　肿瘤患者IRF升高较RET和NEUT要早。未成熟网织红细胞指数是放、化疗时反映骨髓抑制和恢复较敏感的指标     

Circulating erythropoietin levels after bone marrow transplantation: inappropriate response to anemia in allogeneic transplants.

We studied 24 recipients of autologous bone marrow transplantation (ABMT) or allogeneic BMT (BMT) to determine whether impaired erythropoietin (Epo) response to anemia could delay full erythropoietic recovery. Observed Epo levels were compared with predicted levels based on the relationship between Epo and hematocrit in 125 control subjects. Circulating Epo levels were normal during conditioning and the early posttransplant period. Between days 21 and 180, Epo levels remained normal in ABMT patients but were inappropriately low for the degree of anemia in BMT patients. Median time to full erythropoietic engraftment was longer in BMT than in ABMT recipients. Circulating Epo returned to appropriate levels after day 180, except in patients with active cytomegalovirus infection. We conclude that impaired Epo response to anemia can contribute to delayed erythropoietic recovery after allogenic BMT. Renal toxicity of ciclosporin, interaction between host and donor marrow, and cytomegalovirus infection might play a role. This study could support the use of recombinant human Epo to accelerate erythropoietic engraftment after BMT.     

Optimal timing of granulocyte colony-stimulating factor (G-CSF) administration after bone marrow transplantation

The positive role of G-CSF in hastening the myeloid recovery of patients undergoing allogeneic bone marrow transplantation (ALLO-BMT) or autologous bone marrow transplantation (ABMT) has recently been established. Considerable knowledge about adequate doses and route of administration has been accumulated in the past few years. Nonetheless, the optimal time to start growth-factor administration remains undetermined. We have performed a stratified study according to the source of hematopoietic progenitors (ALLO-BMT or ABMT), underlying disease and its stage, and acute graft-versus-host disease (GVHD) prophylaxis regimen and randomized patients in two arms: group A, which started G-CSF on day 0 (36 patients), and group B, which started on day +7 post-BMT (39 patients). The same dose (5 Μg/kg/day) and route of administration were employed in both groups. We found no significant differences in the time to reach an absolute neutrophil count (ANC) of 0.1, 0.5, and 1×10⁹/l and 50×10⁹ platelets/l (medians: 10 and 11, 14.5 and 14, 17 and 16, 23 and 24 days, respectively, in groups A and B). We did not find differences in the days of fever or days on antibiotic treatment with less than 1×10⁹/l ANC, rate of bacteriemia, or days of hospitalization in both groups. In contrast, a considerable saving of GCSF in B group was found (mean days of infusion in group A, 18, versus 11 in group B) (p<0.0001). This is equivalent to a saving of 1120 $US per patient. Therefore, early use of G-CSF after BMT is useless and more expensive and provides no advantage over delayed administration.     

Bone marrow transplantation with interleukin-2-activated bone marrow followed by interleukin-2 therapy for acute myeloid leukemia in mice

We have investigated approaches to induce graft-versus-leukemia (GVL) effect in autologous bone marrow transplantation (ABMT) without graft-versus-host disease to improve survival and cure in leukemia. The present study shows that bone marrow transplantation (BMT) using syngeneic bone marrow activated with interleukin-2 (ABM) for 24 hours in vitro, followed by interleukin-2 (IL-2) therapy, was superior to BMT with fresh, syngeneic bone marrow (FBM) in terms of survival and cure in mice with acute myeloid leukemia (P less than .001) and led to normal hematopoietic reconstitution. Addition of IL-2 therapy after BMT with FBM did not improve the results over BMT with FBM alone (P = .98). These results suggest that the GVL effect of ABMT can be enhanced by using ABM for BMT followed by IL-2 therapy without compromising engraftment.     

Characteristics of engraftment after repeated autologous bone marrow transplantation

The rate of engraftment after autologous bone marrow transplantation (ABMT) is extremely variable and largely unpredictable. To identify factors influencing engraftment, we studied 35 patients with refractory germ cell tumors undergoing high-dose chemotherapy with carboplatin (900-2000 mg/m2) and etoposide (1200 mg/m2) with bone marrow rescue. Prior to the initiation of chemotherapy, bone marrow sufficient for two marrow infusions was harvested (range 0.86-4.82 x 10(8) nucleated cells per kg). All 35 patients received half of the collected bone marrow 3 days after the last dose of chemotherapy; 23 responders received a second round of the same chemotherapy followed by infusion of the second half of the bone marrow. Eighteen patients could be compared for the two transplant episodes. The "rate of engraftment" was defined as the unweighted mean of four parameters: 1) the number of days until the absolute granulocyte count surpassed 0.2 x 10(9)/liter, 2) the number of days until the absolute granulocyte count surpassed 0.5 x 10(9)/liter, 3) the number of days until the last platelet transfusion, and 4) the number of days until the reticulocyte count surpassed 25 x 10(9)/liter. No significant correlation was found between rate of engraftment and such factors as the number of nucleated cells per kg infused, the dose of chemotherapy, extent of prior chemotherapy, tumor response to the high-dose chemotherapy, age of the patient, or the days of granulocytopenic fever (all p greater than 0.20). In contrast, a close correlation was found for the number of units of platelets (p = 0.005) and red blood cells (p = 0.006) transfused following each of the two transplants. There was no significant difference between rate of engraftment after first and second transplantation. Comparison of these data with the results obtained in reported ABMT with separate harvests suggests that the characteristics of the infused marrow determine the rate of engraftment after ABMT. This model of repeated transplantation could provide an important tool for assessing the therapeutic efficacy of hematopoietic growth factors.     

Bone marrow graft engineering by counterflow centrifugal elutriation: results of a phase I-II clinical trial

In an attempt to reduce the incidence and severity of acute graft-versus-host disease (GVHD), we have decreased the number of bone marrow (BM) lymphocytes in the donor marrow graft before bone marrow transplantation (BMT) using counterflow centrifugal elutriation (CCE). In a phase I-II clinical trial, 23 patients received lymphocyte-depleted BM allografts from their HLA-identical, mixed lymphocyte culture (MLC)-nonreactive sibling donors. The patients entered in the study were deemed to be at high risk for treatment failure on the basis of age (greater than 30 years; median, 39 years) and the result of our skin explant assay predictive of acute GVHD. Patients predicted not to develop acute GVHD by this assay were excluded from this study. All patients received a standard lymphocyte dose of 0.5 x 10(6) morphologic lymphocytes per kilogram ideal body weight (IBW) in the marrow graft and were maintained on cyclosporine A (CsA) immunosuppression for 170 days after BMT. Prompt hematopoietic recovery occurred in 22 of 23 patients with a median time to an absolute neutrophil count (ANC) greater than or equal to 500/microL of 21 days. Donor cell engraftment was subsequently verified by cytogenetic and/or DNA analysis in all of 21 evaluable patients. No patient developed systemic acute GVHD. Only five (22%) developed cutaneous GVHD (clinical stage 1) that required steroid treatment, including one patient who failed to engraft. The median follow-up of the patients enrolled in this study is 14 months (range, 5 to 20 months). Actuarial survival 1 year after BMT is 83%. Thus, in two consecutive clinical trials using CCE to deplete donor BM of alloreactive lymphocytes (1.0 x 10(6) versus 0.5 x 10(6) lymphocytes/kg), we have demonstrated that the procedure does not interfere with BM engraftment and is effective in decreasing the incidence and severity of acute GVHD. Furthermore, comparison of these studies has revealed a differential dose response relationship between the number of graft lymphocytes, protection of engraftment, and induction of acute GVHD. Although there appears to be a modest relationship between lymphocyte dose and time to hematopoietic recovery, the 50% reduction in lymphocyte dose from that used in our previous trial resulted in a marked decrease in acute GVHD without compromising engraftment.     

Recombinant human granulocyte-macrophage colony-stimulating factor after autologous bone marrow transplantation for malignant lymphoma: a British National Lymphoma Investigation double-blind, placebo-controlled trial

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is active in enhancing the production of mature myeloid cells in vitro and several phase I/II clinical trials have suggested that its administration may accelerate neutrophil recovery after autologous bone marrow transplantation (ABMT). We have conducted a multicentre randomized double-blind placebo controlled trial in patients with poor prognosis malignant lymphoma receiving an identical high-dose combination chemotherapy regimen with ABMT. 61 patients were entered and 29 in each arm of the trial were evaluated. Treatment with GM-CSF did not affect the period of severe neutropenia (absolute neutrophil count (ANC) of < 0.1 x 10(9)/l) but accelerated recovery to an ANC of 0.5 x 10(9)/l (median 14 d v 20 d in controls, P = 0.001). There was no significant difference in platelet recovery between the groups (GM-CSF group platelet dependent for 25 d v control 19 d, P = NS). The number of positive blood cultures was similar in both groups (GM-CSF 14 v placebo 13) and there were no differences in days of fever > 37.5 degrees C (median 8 v 6) or days on parenteral antibiotics (11 v 10). Patients receiving GM-CSF had a median period of hospitalization following BMT of 24 d (control 25). No significant major toxicity attributable to GM-CSF administration was detected. We have confirmed in a randomized trial that GM-CSF accelerates neutrophil but not platelet recovery following ABMT. We were unable to demonstrate any accompanying changes in clinical outcome and believe that further trials are necessary to assess the clinical value of GM-CSF in BMT.     

Automated analyzer evaluation of reticulocytes in bone marrow and peripheral blood of hematologic disorders

The R-3000 reticulocyte analyzer uses flow cytometry with an argon laser as its light source. This analyzer stains residual RNA with auramine O to provide a reticulocyte maturation differential. Using the R-3000, we analyzed 119 samples of bone marrow (BM) and peripheral blood (PB) from 111 patients with hematologic disorders. Parameters were reticulocytes, immature reticulocyte fraction (IRF) percentage in BM and PB, BM/PB reticulocyte ratio, and BM/PB IRF ratio. Reticulocytes and IRF percentage in BM were significantly higher than in PB (p < 0.01). There was also a good correlation between reticulocyte percentages in BM and in PB (r = 0.81). Patients were classified into a normal group (without anemia) and an anemia group. Furthermore, the anemia group was classified into three groups: group 1: cases with hematopoietic dysfunction; group 2: cases in bone marrow recovery phase after chemotherapy and hematopoietic stem cell transplantation, and hematologic disorders with bone marrow accelerative phase, and group 3: cases with ineffective hematopoiesis (myelodysplastic syndrome). The mean reticulocyte percentage of the normal group was 2.3 +/- 1.1%, which was close to the normal value in BM. The BM/PB reticulocyte ratio of group 3 was statistically higher than that of groups 1 and 2. This indicates that group 3 had ineffective erythropoiesis and that the BM/PB ratio is a useful indicator for the diagnosis of myelodysplastic syndrome.     

Difference in kinetics of hematopoietic reconstitution between ALL and ANLL after autologous bone marrow transplantation with marrow treated in vitro with mafosfamide (ASTA Z 7557)

The kinetics of hematopoietic recovery after autologous bone marrow transplantation (ABMT) reflect the hematopoietic capacity of the infused marrow. In vitro treatment of marrow with high doses of mafosfamide (ASTA Z 7557) alters the hematopoietic regenerative capacity of the graft. Thirty-two patients with acute leukemia (12 acute lymphoblastic leukemia (ALL) and 20 acute non-lymphoblastic leukemia (ANLL] with 27 in complete remission and five in partial remission were consolidated with cyclophosphamide (60 mg/kg x 2) and total body irradiation (10 Gy), followed by reinfusion of autologous marrow treated in vitro with mafosfamide. The marrow of each patient had been incubated with the highest tolerable dose of mafosfamide, individually predetermined from a preincubation test. We report here that the kinetics of engraftment are strikingly different in ANLL and ALL patients. In the ANLL group recovery to 0.1% reticulocytes took a median of 20.5 days (range 14-32) versus 15 (11-28) in the ALL group; 33.5 days (18-45) versus 19 (15-30) for leukocytes to reach 1.0 x 10(9)/l; 35 (19-60) versus 20.5 (15-30) for neutrophils to reach 0.5 x 10(9)/l; 110+ (45-480+) versus 50 (23-90) for platelets to reach 50 x 10(9)/l (p less than 0.01 and p less than 0.05). Detection of granulocyte-macrophage progenitors (CFU-GM) regeneration in marrow aspirates post-ABMT was delayed in ANLL (p less than 0.05). Neither the nature of the previous induction therapy, nor the status of the blood or bone marrow at the time of collection (CFU-GM and erythroid burst-forming units/ml) nor the stem cell sensitivity to mafosfamide, nor the doses of progenitor cells infused could explain these differences. We interpreted these observations as suggesting that the engraftment potential has been more severely altered in ANLL than in ALL, which may reflect both the intensity of the in vitro treatment and the intrinsic fragility of the stem cell pool in ANLL.     

Influence of recombinant human granulocyte colony-stimulating factor (filgrastim) on hematopoietic recovery and outcome following allogeneic bone marrow transplantation (BMT) from volunteer unrelated donors.

Effects of recombinant human granulocyte colony-stimulating factor (rhG-CSF, filgrastim) on hematopoietic recovery and clinical outcome in patients undergoing allogeneic bone marrow transplantation (BMT) from volunteer unrelated donors (VUD) were analyzed retrospectively. Additionally, the influence of baseline patient and transplant characteristics on hematopoietic recovery was evaluated. From January 1994 to March 1996, 47 consecutive adult patients received VUD-BMT. GVHD prophylaxis was cyclosporin A/short course methotrexate/prednisolone, and in four patients additional ATG. Post-transplantation, cohorts of patients received rhG-CSF (5 microg/kg/day) (n = 22) or no rhG-CSF (n = 25) in a non-randomized manner. The patient groups with and without rhG-CSF were rather comparable with respect to baseline patient and transplant characteristics. Median time to neutrophil counts (ANC) >500/microl was 14 days with rhG-CSF vs 16 days without rhG-CSF (P = 0.048), to ANC >1000/microl was 15 vs 18 days (P = 0.084). Neutrophil recovery was accelerated in patients receiving more than the median MNC dose of 2.54 x 10(8)/kg with a median time to ANC >1000/microl of 13 days vs 19 days (P = 0.017). RhG-CSF did not influence platelet recovery and incidence of infectious complications. Incidence of acute GVHD II-IV was 50% with rhG-CSF and 28% without rhG-CSF (P = 0.144), but death before acute GVHD II-IV occurred in 9% of patients with and 20% of patients without rhG-CSF. The median follow-up time was 38 and 36 months in patients with and without rhG-CSF, respectively. Survival at 2 years post-transplant was 39% (95% confidence interval (18%, 60%)) in patients with rhG-CSF and 24% (95% confidence interval (7%, 41%)) in patients without rhG-CSF. Administration of rhG-CSF after VUD-BMT may lead to more rapid neutrophil recovery, but did not influence the incidence of infectious complications. Patients receiving rhG-CSF showed a slightly higher incidence of acute GVHD II-IV. Higher numbers of MNC in the marrow graft accelerated hematopoietic engraftment.     

Autologous bone marrow transplantation for acute myeloid leukemia using monoclonal antibody-purged bone marrow

We report our experience from a clinical trial of autologous bone marrow transplantation (ABMT) in the treatment of 30 patients with acute myeloid leukemia (AML) using monoclonal antibody (MoAb) and complement-treated bone marrow. All patients were in complete remission (CR) at the time of transplant: 6 patients were in first CR, 18 in second CR, and 6 in third CR. The median age of all patients was 42 years (range 11 to 57 years). For marrow ablation, 28 patients were treated with cyclophosphamide and total body irradiation. One patient was treated with busulfan and cyclophosphamide and one was treated with busulfan and VP-16. Each patient was then transfused with autologous bone marrow that had been harvested previously and treated with two MoAbs, PM-81 and AML-2-23, and rabbit complement. Median time to recovery of neutrophils (500/microL) was 30 days, and platelets (20,000/microL) was 45 days. Median time for initial erythrocyte engraftment, assessed by a flow cytometric reticulocyte assay, was 13 days. Median overall and relapse-free survival of first CR patients was at least 17.4 months post-ABMT and the 2- and 3-year actuarial overall and relapse-free survival was 67% (+/- 19%). Median survival for the 24 patients in second or third CR was 6.8 months post-ABMT and 9.3 months since CR; however, six patients survived disease-free from 16 to 61 months post-ABMT. For the second and third CR group it was observed that six patients (5 of the 6 survivors) showed "inversions," when their post-ABMT remission lasted longer than any previous one. Actuarial 2- and 3-year disease-free and overall survival of patients in second and third CR was 25% (+/- 9%) and 18% (+/- 9%), and 29% (+/- 9%) and 23% (+/- 9%), respectively. ABMT avoids the problems of graft-versus-host disease and of finding suitable donors for allogeneic marrow transplantation.     

Circulating granulocyte colony-stimulating factor (G-CSF) levels after allogeneic and autologous bone marrow transplantation: endogenous G-CSF production correlates with myeloid engraftment.

Myeloid engraftment after bone marrow transplantation (BMT) is influenced by a number of variables, including cytoreductive chemoradiotherapy, genetic disparity, number of reinfused committed myeloid progenitor cells, healthy microenvironment, and the presence of hematopoietic growth factors. Granulocyte colony-stimulating factor (G-CSF) stimulates proliferation of myeloid progenitor cells and enhances myeloid engraftment after BMT. We investigated the temporal relationship between endogenous G-CSF production and myeloid engraftment in both children and adults after allogeneic (ALLO) and autologous (AUTO) BMT. Circulating endogenous G-CSF levels ranged between 0 and 2552 pg/mL. The correlation coefficient between circulating serum G-CSF levels and the peripheral absolute neutrophil count (ANC) was r = -.875 (P less than .001). The endogenous serum G-CSF level was highest during the first week after BMT, when the ANC was less than or equal to 200/microL (699 +/- 82.3 pg/mL) (P less than .001). Both children and adults demonstrated a similar inverse relationship between circulating G-CSF level and degree of neutropenia. One patient failed to engraft after AUTO BMT and also failed to generate any endogenous G-CSF production. Lastly, once the serum G-CSF level decreased to less than 200 pg/mL, a mean of 6.1 +/- 0.9 days elapsed before the ANC was greater than or equal to 500/microL for 2 consecutive days. This study demonstrates that endogenous G-CSF production is associated with myeloid engraftment in both children and adults after AUTO and ALLO BMT and that the rate of increase and decrease in endogenous G-CSF may be predictive of either failure to engraft or duration of neutropenia.     

Evaluation of hematopoiesis by granulocyte elastase after hematopoietic stem cell transplantation

To evaluate hematopoietic restoration after hematopoietic stem cell transplantation (HSCT), we sequentially monitored the post-HSCT level of granulocyte elastase (GE), a sensitive parameter of qualitative and quantitative changes in granulopoiesis. We compared it with routinely used hematopoietic recovery indices, such as leukocyte and reticulocyte count. We also compared levels of GE in the clinical administration of different colony stimulating factors (granulocyte-colony stimulating factor (G-CSF) versus macrophage-colony stimulating factor (M-CSF) and in different types of hematopoietic stem cell transplantation (allogeneic bone marrow transplantation (BMT) versus autologus peripheral blood stem cell transplantation (PBSCT)). Days to first increase of GE after HSCT were 3.0 and 2.3 days earlier than increase of leukocytes in allo-BMT and auto-PBSCT, respectively. Recovery of highly fluorescent reticulocytes and monocytes were later than recovery of GE. These results indicated that granulopoiesis after transplantation started before the increase in peripheral leucocyte count, and that GE was the earliest indicator of hematopoietic recovery. On the basis of GE level, M-CSF had the same stimulating effect on granulopoiesis as G-CSF. The nadir of GE in PBSCT was significantly higher than that in BMT, indicating continuous granulopoiesis in PBSCT. From these results, we concluded that measurements of GE can be used for the clinical evaluation of myelosuppression by different conditioning regimens as well as of granulopoiesis induced by various cytokines.     

Allogeneic bone marrow transplantation versus chemotherapy in high-risk childhood acute lymphoblastic leukaemia in first remission

We compared the outcome of children with high-risk acute lymphoblastic leukaemia (HR-ALL) in first complete remission (first CR) treated with chemotherapy (CHEMO) or with allogeneic bone marrow transplantation (BMT) in a multicentre study.   All children treated by the Italian Paediatric Haematology Oncology Association for HR-ALL in first CR between 1986 and 1994 were eligible for the study. 30 children were given BMT at a median of 4 months from first CR, with preparative regimens including total-body irradiation ( n  =25/30). 130 matched controls for BMT patients were identified among 397 HR-ALL CHEMO patients. Matching on main prognostic factors and duration of first CR was adopted to control the selection and time-to-transplant biases. The comparative analysis was based on the results of a stratified Cox model. The estimated hazard ratios of BMT versus CHEMO at 6 months, 1 year and 2 years after CR were 1.38 (CI 0.59–3.24), 0.69 (CI 0.27–1.77) and 0.35 (CI 0.06–1{\raise 5mu ..91), with an overall non-significant difference between the two groups ( P  = 0.34). With a median follow-up of 4 years, the disease-free survival was 58.5% (SE 9.3) in the BMT group and 47.7% (SE 4.8) in the CHEMO group, at 4 years from CR. Non-leukaemic death occurred in 4% of CHEMO and 10% of BMT patients. In the BMT group the estimated cumulative incidence of relapse at 1.5 years from CR was 31.5% (SE 8.8) and did not change thereafter, whereas in the CHEMO group the corresponding figure was 29.2% (SE 4.1) and the incidence continued to increase thereafter (48.2% (SE 4.8) at 4 years from CR).   The results of this study suggest that, with respect to the CHEMO group, the higher risk of early failure in the BMT group is outweighed by the lower risk of relapse after 1 year. Results prompt the need for a prospective study, in order to demonstrate the likely advantage of BMT in HR childhood ALL in first CR.     

Highly fluorescent reticulocyte count predicts haemopoietic recovery after immunosuppression for severe aplastic anaemia

Highly fluorescent reticulocyte (HFR) counts are the most reliable and sensitive index of haemopoietic recovery after bone marrow or peripheral blood stem cell transplantation. We report the behaviour of HFRs during haemopoietic recovery in two patients who were affected by severe aplastic anaemia (SAA) and treated with horse antithymocyte globulin (ATG), cyclosporin A (CsA) and granulocyte colony-stimulating factor (G-CSF). A HFR value > 5% of the total reticulocyte count, a reticulocyte count > 30 x 10(9)/l, and a polymorphonuclear (PMN) count > 0.5 x 10(9)/l were found after 9 and 8, 20 and 46, and 16 and 22 days, respectively, after the end of ATG. HFR recovery to > 5% anticipated the rise of PMN > 0.5 x 10(9)/l by at least 7 and 14 days, respectively. Thus, HFR evaluation could be used as a reliable and early marker of response to immunosuppression in severe aplastic anaemia.