Tandem high-dose chemotherapy with thiotepa and busulfan-melphalan and autologous stem cell transplantation in very high-risk neuroblastoma patients.

High-risk neuroblastoma is characterised by poor long-term survival, especially for very high-risk (VHR) patients (poor response of metastases after induction therapy). We report the results of an intensified high-dose chemotherapy (HDC) strategy to improve the prognosis of VHR patients. This strategy was based on tandem HDC with thiotepa and busulfan-melphalan (Bu-Mel) followed by autologous stem cell transplantation (ASCT). All data were prospectively recorded in the Gustave Roussy Paediatric ASCT database. From April 2004 to August 2011, 26 patients were eligible for tandem HDC. The median age at diagnosis was 4.4 years (1-15.9). All patients had metastatic disease. MYCN was amplified in 5/26 tumours. Despite the cumulative toxicity of alkylating agents, the toxicity of the intensified HDC strategy was manageable. Thiotepa-related toxicity was mainly digestive, whereas sinusoidal obstruction syndrome was the main toxicity observed after Bu-Mel. The 3-year event-free survival of this cohort was 37.3% (21.3-56.7). This strategy will be compared with combined (131)I-mIBG/Bu-Mel in the upcoming SIOPEN VHR Neuroblastoma Protocol.

Ancestim (r-metHuSCF) plus filgrastim and/or chemotherapy for mobilization of blood progenitors in 513 poorly mobilizing cancer patients: the French compassionate experience.

Ancestim (r-MetHuSCF) is available in France for compassionate use in patients who are candidates for high-dose chemotherapy and autologous transplantation, and who failed in previous attempts at mobilization and collection. We report here data from 513 adult patients who benefited from this program, between January 1998 and July 2007. Given with systematic premedication, ancestim was generally well tolerated, although severe but not life-threatening adverse events were reported in 12 individuals. Overall, a graft was obtained or completed for 235 patients (46%). The median number of collected CD34+ cells was 3.00 × 10(6)/kg (range: 0.03-39.50). The target threshold of 2 × 10(6) CD34+ cells/kg was reached in 161 patients (31%). Factors associated with collection were diagnosis of myeloma, no previous autologous transplant, no more than one previous failed attempt and a mobilization regimen including cytotoxic agents. A total of 207 patients (40%) proceeded to high-dose chemotherapy and autologous transplantation. The median time to reach 0.5 × 10(9)/L neutrophils and 20 × 10(9)/L platelets was 12 (6-40) and 13 (0-31) days, respectively. We conclude that a combination of ancestim with filgrastim successfully mobilized CD34+ cells in peripheral blood, and allowed adequate collection in preparation for autologous transplantation in approximately one-third of poorly mobilizing patients.

[Quality control of defrosted cord blood units: results from an inter-laboratory study]. / Contrôle de qualité des greffons de sang placentaire décongelés : résultats d'une étude multicentrique.

PURPOSE: Today, haematopoietic stem cell graft from placental blood concerns more than 15 % of allogeneic grafts. An inter-laboratory study of the quality control of defrosted cord blood units has been coordinated by the French society for cell and tissue bioengineering (SFBCT), with the cord blood bank of Bourgogne Franche-Comté and controlled by the French health products safety agency (Afssaps). The aim of this study is to ensure the inter-laboratory reproducibility of the quality controls practised by the banks during defrosting. The cellular outputs were analyzed according to the defrosting techniques, according to the method used in flow cytometry: single-platform (SP) versus double-platform (DP), or the product nature, i.e. in total blood or miniaturized. METHODS: Forty-two units of placental blood (USP), which were out of range were provided for defrosting to 14 participating sites. USP were defrosted and controlled according to the procedures of each bank. Once the USP is defrosted, a part of the product was controlled by the site and the other part by Afssaps. Following controls were carried out: numeration of the total nucleated cells (TNC) and of CD34+ cells (made by a SP method in Afssaps) and functional assay. RESULTS: Concerning TNC, the defrosting sites obtained a cellular output of 94 %+/-28 in day 0 compared with an output of 72 %+/-24 in Afssaps showing a rather good stability of the USP transmitted with an average deviation of 23 %+/-22. The freezing process with or without reduction of volume does not affect this variation. Concerning the numeration of CD34+ cells, the average deviation between the participating sites and Afssaps was 29 %+/-23 compared with 21 %+/-16 for the sites using a SP method against 47 %+/-25 for those using a DP method. The CD34+ outputs are equal to 82 % +/- 60 in day 0 for the participating sites against 52 %+/-20 for Afssaps. For the sites using a DP method, it is stressed that this output is particularly high with a rate of 126 %+/-90 (n=15) whereas it is 62 %+/-20 (n=32) for the sites using a SP method. CONCLUSION: These results underline a good stability of viable CD34+ cells and a greater reliability of the SP methods for the CD34+ cell numeration for these defrosted USP. Lastly, the results of the functional assay regarding the average clonogenicities (equal to 15 %) reinforce the conclusions on the quality of the defrosted products.

CD34(+) progenitors are reproducibly recovered in thawed umbilical grafts, and positively influence haematopoietic reconstitution after transplantation.

Cord blood (CB) units are increasingly used for allogeneic transplantation. Cell dose, a major factor for CB selection, is evaluated before freezing by each CB bank, using various techniques. This may introduce variability and affect the prediction of cell recovery after thawing, or haematopoietic reconstitution. Forty-two children were transplanted at the same institution with unrelated CB units. All units were thawed and evaluated at the same cell therapy facility, using standard procedures. We investigated: (i) factors that affect cell loss after thawing, and (ii) the importance of CD34(+) cell doses. Prefreeze and post-thaw CD34(+) cell doses were statistically correlated, thus suggesting that variability in numeration techniques used by different CB banks does not compromise the biological and clinical value of these figures. CD34(+) cell recovery appeared to be correlated with the absolute number of CD34(+) cells per frozen bag. Infused CD34(+) is the cell dose that better correlates with platelet reconstitution delay; in addition, when using a quartile comparison, haematopoietic recovery appeared to be related with prefreeze and post-thaw CD34(+) cell doses. We conclude that enumeration of CD34(+) cells in CB units is of biological significance, and may help select CB units and identify patients at risk of delayed recovery.

Enhanced activation of B cells in a granulocyte colony-stimulating factor-mobilized peripheral blood stem cell graft.

In a randomized study that compared human leucocyte antigen-identical allogeneic granulocyte colony-stimulating factor (G-CSF)-mobilized peripheral blood stem cell (PBSC) versus bone marrow (BM) transplantation, the expression of activation markers, CD23, CD25 and CD45RO by B cells, was compared in blood before and after G-CSF mobilization and in PBSC versus BM grafts. The fractions of CD23+ and CD25+ B cells were higher in PBSC than in BM grafts. Moreover, we observed a G-CSF-induced increase in B-cell fractions in blood as well as in PBSC grafts when compared with BM grafts. Such an enhanced B-cell activation could contribute to the accelerated kinetics of immuno-haematological reconstitution, the occurrence of acute haemolysis in the ABO minor incompatibility setting, as well as the increased incidence of chronic graft-versus-host disease observed after PBSC transplantation.

Allogeneic peripheral blood stem cell transplantation results in less alteration of early T cell compartment homeostasis than bone marrow transplantation.

Since low T cell counts evaluated 1 month after allogeneic bone marrow transplantation (BMT) are associated with an increased risk of leukemia relapse (Powles et al., Blood 1998; 91: 3481-3486), we compared, in a randomized multicentric clinical study, the peripheral blood cells obtained 30 days after allogeneic BMT vs allogeneic G-CSF-mobilized peripheral blood stem cell transplantation (BCT) in an HLA-identical setting. T cell counts were higher 30 days after BCT (718+/-142 cells/microl, n = 20) than after BMT (271+/-53 cells/microl, n = 26, P = 0.006). However, T cells were less activated after BCT than after BMT, as demonstrated by a lower expression level of CD25 and a lower percentage of HLA-DR+ and CD95+ T cells. Furthermore, CD4+, CD8+ and CD45RA+ post-BCT T cell counts correlated with the number of cells infused with the PBSC graft, while such a correlation was not observed between post-BMT counts and BM graft cell numbers, suggesting that the intensity of post-transplant peripheral lymphoid expansion and/or deletion differed between BCT and BMT. A comparison of the input of T cells expressing different CD45 isoforms with the post-transplant cell recovery further confirmed that, within the CD4+ T cell subset, post-transplant expansions occurred at a higher level after BMT than after BCT, affecting mainly the CD4+ CD45RO+ subset. Altogether, our data demonstrate for the first time in a randomized setting that homeostasis of the T cell pool is less altered early after BCT than after BMT. This may have a strong impact on the graft-versus-leukemia (GVL) effect and subsequent relapse rate.

Influence of the hematopoietic stem cell source on early immunohematologic reconstitution after allogeneic transplantation.

Several acute hemolysis episodes, sometimes lethal, have been recently described after transplantation of allogeneic peripheral blood hematopoietic stem cells (PBHSCs). Hemolysis resulted from the production of donor-derived antibodies (Abs) directed at ABO antigens (Ags) present on recipient red blood cells (RBCs). A multicenter randomized phase III clinical study comparing allogeneic PBHSC transplantation (PBHSCT) versus bone marrow hematopoietic stem cell transplantation (BMHSCT) has been conducted in France. In the course of this study, serum anti-A and/or anti-B Ab titers were compared before the conditioning regimen and on day +30 after transplantation in 49 consecutive evaluable PBHSCT (n = 21) or BMHSCT (n = 28) recipients. PBHSCT resulted in a higher frequency of increased anti-A and/or anti-B Ab titers 30 days after transplantation as compared to BMHSCT: 8 (38%) of 21 versus 3 (11%) of 28 (P =.04). In PBHSCT recipients, increased titers were observed mostly after receiving a minor ABO mismatch transplant: 5 of 7 versus 3 of 14 in the absence of any minor ABO mismatch (P =.05), whereas this was not the case after BMHSCT: 1 of 8 versus 2 of 20. Anti-A and/or anti-B serum Abs detectable at day +30 after PBHSCT were always directed against A and/or B Ags absent both on donor and recipient RBCs. Finally, 3 of 21 PBHSCT versus 0 of 28 BMHSCT recipients developed anti-allogeneic RBC Abs other than ABO (P =.07). Overall, the data strongly suggest that immunohematologic reconstitution differs significantly after granulocyte colony-stimulating factor-mobilized PBHSCT when compared to BMHSCT. Such a difference could contribute to the acute hemolysis described after PBHSCT as well as to distinct alloreactivity after PBHSCT.

Effect of granulocyte colony-stimulating factor mobilization on phenotypical and functional properties of immune cells.

Some phenotypic and functional properties of lymphocytes from bone marrow or peripheral blood stem cell donors were compared in a randomized study. Lymphocyte subsets were analyzed by immunocytometry in blood harvested from bone marrow donors (n = 27) and from peripheral blood stem cell donors before and after granulocyte colony-stimulating factor mobilization (n = 23) and in bone marrow and peripheral blood stem cell grafts. Granulocyte colony-stimulating factor mobilization increased the blood T and B, but not NK, lymphocyte counts. All lymphocyte counts were approximately 10-fold higher in peripheral blood stem cell grafts than in bone marrow grafts. Analysis of CD25, CD95, HLA-DR, and CD45RA expression shows that T-cell activation level was lower after granulocyte colony-stimulating factor mobilization. Similarly, granulocyte colony-stimulating factor reduced by twofold to threefold the percentage of interferon-gamma, interleukin-2, and tumor necrosis factor-alpha-secreting cells within the NK, NK-T, and T-cell subsets and severely impaired the potential for interferon-gamma production at the single-cell level. mRNA levels of both type 1 (interferon-gamma, interleukin-2) and type 2 (interleukin-4, interleukin-13) cytokines were approximately 10-fold lower in peripheral blood stem cell grafts than in bone marrow grafts. This reduced potential of cytokine production was not associated with a preferential mobilization of so-called "suppressive" cells (CD3+CD4-CD8-, CD3+CD8+CD56+, or CD3+TCRVA24+CD161+), nor with a modulation of killer cell receptors CD161, NKB1, and CD94 expression by NK, NK-T, or T cells. Our data demonstrate in a randomized setting that quantitative as well as qualitative differences exist between a bone marrow and a peripheral blood stem cell graft, whose ability to produce type 1 and type 2 cytokines is impaired.

Implementation of a specific approval process for blood-components prescription.

We implemented a systematic computer-assisted validation process for transfusion prescriptions to improve transfusion safety. Assessment of this new approach indicates good adoption of validated transfusion guidelines and a reduction of exposure to blood products and overall costs.

Consolidation with a busulfan-containing regimen followed by stem cell transplantation in infants with poor prognosis stage 4 neuroblastoma.

Although infants with stage 4 neuroblastoma (NB) usually have a good prognosis, metastatic relapses after 1 year of age and amplification of the N-myc oncogene are established poor prognostic factors. In order to improve the survival of patients with such high-risk factors, we performed consolidation with a busulfan (600 mg/m2)-melphalan (140 mg/m2)-containing regimen followed by autologous stem cell transplantation (SCT). From 1986 to 1998, 12 patients were treated according to this strategy. Their median age at diagnosis was 9 months (1-11). Consolidation was performed after a metastatic relapse in five children, because of persistent bone metastases in one and as first-line consolidation in six patients whose tumor exhibited N-myc amplification. The 5-year EFS rate is 64. 5% (36-85%) with a median follow-up of 92 months (20-126). One toxicity-related death occurred in a very heavily pretreated patient. Hepatic veno-occlusive disease was the major side-effect that occurred in nine of 12 children. This busulfan-melphalan combination appears to dramatically improve the prognosis of these high-risk infants with metastatic NB. Given its high toxicity, indications for this consolidation must be restricted to high-risk infants and a lower dose of busulfan (480 mg/m2) is recommended in children weighing less than 10 kg. Bone Marrow Transplantation (2000) 25, 937-942.

Allogeneic peripheral blood hematopoietic stem cell transplantation: guidelines for red blood cell immuno-hematological assessment and transfusion practice.Société Française de Greffe de Moelle.

Allogeneic peripheral blood hematopoietic stem cell transplantation (PBSCT) is presently being evaluated in a French randomized study comparing peripheral blood vs bone marrow. Cases of potentially lethal acute hemolysis have recently been reported after allogeneic PBSCT in the presence of a 'minor' ABO incompatibility. Patients were frequently transfused with recipient-compatible and donor-incompatible RBC and usually did not receive methotrexate in addition to cyclosporin A for graft-versus-host disease (GVHD) prophylaxis. In order to homogenize immuno-hematological (IH) assessment and transfusion practices within our protocol, we made proposals to 25 allo-transplant French centers on the following aspects: pre-inclusion IH assessment, IH exclusion criteria, transfusion rules, post-transplant IH surveillance and treatment of hemolysis. Analysis of responses to our proposals led to the elaboration of guidelines which were approved and implemented by the French Bone Marrow Transplantation Society (SFGM). Pre-inclusion IH testing includes mandatory detection and titration of anti-RBC allo-Ab, as well as titration of anti-A and anti-B Ab. The presence in the donor of an anti-A (group A or AB recipients), anti-B (group B or AB recipients) Ab with a titer >1/32 or the presence of allo-Ab against Rh, Kell, Fya, Fyb, Jka, Jkb, Ss Ag present on recipient RBC is an exclusion criterion for the protocol. ABO and RhD compatibility of RBC blood products with both HSC donor and recipient is mandatory. A similar compatibility is also required for Rh (other than D) and Kell Ag. If not possible, compatibility of RBC blood products with the HSC donor is mandatory. Lastly, guidelines regarding post-transplantation IH follow-up as well as acute hemolysis treatment have been elaborated. The implementation of these guidelines should contribute to enhancing the quality of transfusion practice after PBSCT. Such an approach will be applied to other aspects of transfusion medicine in the setting of HSC transplantation. Bone Marrow Transplantation(2000) 25, 507-512.

[Perioperative autotransfusion with salvage blood in cancer surgery] . / Autotransfusion peropératoire par récupération de sang épanché en chirurgie carcinologique.

OBJECTIVES: Intraoperative blood cells salvage using a Cell Saver technique is controversial in oncologic surgery because tumor cells could be aspirated and reinfused to the patient. The goal of this review was to discuss the risk associated with this technique, and the way to minimize it. DATA SOURCES: A review of the literature has been made by questioning PubMed site (http://nbci.nlm.nih.gov) on the period of 1968 to 2000. The key words were: intraoperative blood salvage, blood transfusion, autologous, cancer. Cases reports have been excluded. STUDY SELECTION: Tumor cells aspirated and reinfused have been numbered in both experimental and clinical studies. In clinical studies, the outcome after intraoperative cells salvage/reinfusion has been compared to published data or historical groups of allogeneic transfusion, all in non randomized studies. DATA SYNTHESIS: Both experimental and clinical studies confirmed the presence of cancer cells in the blood either aspirated or reinfused. However, six clinical studies with limited number of patients did not show metastatic spread associated with Cell Saver. The addition of leukocyte filters reduces greatly this quantity of cancer cells. Irradiation of the pack did not destroy tumor cells but blocked their proliferative capacity. In the other hand, some infiltrative tumors were shown to have permanent cancer cells seeding, quantitatively superior to the seeding observed when a Cell Saver is used. CONCLUSION: It seems reasonable to use the Cell Saver in oncologic surgery, if possible with a leukocyte filter, not only in case of unexpected major bleeding (consensus), but also in programmed cases with high risk of huge hemorrhage.

Prognostic factors in metastatic neuroblastoma in patients over 1 year of age treated with high-dose chemotherapy and stem cell transplantation: a multivariate analysis in 218 patients treated in a single institution.

The purpose of this paper is to study prognostic factors in neuroblastoma patients treated with high-dose chemotherapy and hematopoietic stem cell transplantation. Two hundred and eighteen children over 1 year of age and treated for stage 4 neuroblastoma were enrolled in this study. The median age at diagnosis was 39 months, the sex ratio 1.5 and 84% of patients had an abdominal primary tumor. Skeletal disease was detected in 79% of cases and bone marrow involvement in 93%. N-myc oncogene amplification was present in 27% of the patients studied. The probability of event-free survival at 5 years post-diagnosis was 29% in this series. Three major favorable prognostic factors were significant and independent in the multivariate analysis: age under 2 years at diagnosis (P<0.01), absence of bone marrow metastases at diagnosis (P<0.04) and the high-dose conditioning regimen containing busulfanmelphalan combination (P = 0.001). The quality of response to conventional primary chemotherapy was close to significance (P = 0.053). We conclude that factors related to the patient (age) and extent of disease are predictive of outcome in patients with neuroblastoma treated with conventional chemotherapy followed by surgical excision of the primary and consolidation with high-dose chemotherapy. They should be taken into account in future prospective studies. Moreover, the type of conditioning regimen appears to be the most important prognostic factor. This should encourage new investigations into innovative drug combinations.

[Erythrocyte transfusion practices after allogeneic hematopoietic stem cell transplantation]. / Pratiques transfusionnelles érythrocytaires après greffe de cellules souches hématopoiétiques allogéniques.

Allogeneic peripheral blood hematopoietic stem cell transplantation is being evaluated in a randomized French study comparing the use of peripheral blood stem cells vs. bone marrow graft stem cells. In order to standardize immunohematological (IH) assessment and transfusion practices within our protocol, we made suggestions to 25 allo-transplantation French centers on the following elements: pre-inclusion IH assessment, IH exclusion criteria, transfusion rules, post-transplantation IH surveillance and treatment of hemolysis. The analysis of their responses to our suggestions led us to elaborate recommendations which were approved and implemented by the French Bone Marrow Transplantation Society (SFGM). These recommendations concern the transfusion practice in the general framework of allogeneic hematopoietic stem cell transplantation and can therefore be considered as referential.

Phase II study of high-dose thiotepa and hematopoietic stem cell transplantation in children with solid tumors.

From 1987 to 1995, 22 children with refractory solid tumors entered a phase II study of high-dose thiotepa (HDT) (900 mg/m2) followed by stem cell transplantation (SCT) in the Pediatrics Department of the Institut Gustave Roussy. Tumor types were rhabdomyosarcoma (eight), osteosarcoma (seven), neuroblastoma (three), Ewing's sarcoma (three) and Burkitt's lymphoma (one). Before HDT, all had been extensively treated with conventional chemotherapy, surgical resection of the primary tumor (13/22) and of metastases (6/22), and radiotherapy of the primary tumor in three patients. All had measurable disease, at the site of the primary tumor (3 patients), of the metastases (9 patients) or both (10 patients). Toxicity from the HDT was severe but acceptable. No toxicity-related death occurred. The median duration of neutropenia and thrombocytopenia was 18 days (5-37) and 30 days (7-377), respectively. Septicemia was documented in four patients. Severe diarrhea was observed in seven patients. Mild hepatic toxicity occurred 18 times. No CR and 11/22 PR were documented: osteosarcoma 4/7, rhabdomyosarcoma 4/8, Ewing's sarcoma 2/3; 1/1 Burkitt's lymphoma progressed. We conclude that at a dose of 900 mg/m2 followed by SCT support in these heavily pretreated children, the main toxicity induced by thiotepa was digestive. The response rate observed, especially in sarcoma, is particularly encouraging. Thiotepa should be further evaluated in HDC regimens either in combination with other alkylating agents or in rapidly cycled courses of HDC with SCT.

[Blood transfusions in cancer patients]. / Transfusions sanguines chez les malades cancéreux.

In this review, we develop four topics on the relationship between blood transfusion and cancer. First, the rationale for not allowing blood donations from patients with infiltrating tumors is presented. Second, the different possibilities for autotransfusions in cancer patients are discussed. Predeposited autotransfusions are rarely possible in these patients, in addition to the high cost. The usefulness of another method, intraoperative autologous transfusion with blood saved from the surgical field, is not well established. Our third topic concerns the effect of transfusion on cancer induction. In some cases, the risk of cancer is higher after allogenic transfusion resulting from a mechanism involving alterations of the immune function. Finally, the relationship between transfusion and cancer recurrence is controversial. In spite of numerous studies attempting to elucidate this relationship, no final conclusion can be drawn at the present time. What is sure, is that patients requiring blood transfusion have a higher risk of recurrence than patients who do not need transfusion.

Should vancomycin be used empirically in febrile patients with prolonged and profound neutropenia? Results of a randomized trial.

OBJECTIVES: We conducted a randomized trial with ceftazidine alone or associated with amikacin or vancomycin to investigate the efficacy of the daily 3 g dosage of ceftazidime and the efficacy of monotherapy with ceftazidime and to determine if vancomycin should be added empirically. METHODS: Patient inclusion criteria were: age over 10 years, therapeutically-induced neutropenia and fever for at least three hours above 38.5 degrees C in absence of a clear non-infectious aetiology. Patients were randomized into three groups: group C, ceftazidime alone 3 g/day; group CA, ceftazidime 3 g/day plus amikacin 15 mg/kg/day; or group C, ceftazidime 3 g/day plus vancomycin 1.5 g/day. RESULTS: Results from one hundred and two episodes of fever were analyzed. The underlying diseases were haematological malignancies (89 patients) and solid tumours (13 patients). The median duration of neutropenia (< 0.5 x 10(9) PMN/L) was 18 days and the minimum duration of 7 days. The main criterion for the analysis of efficacy was the onset of a major infectious event, i.e. death related to documented or suspected infection and any infectious event considered life-threatening or hindering future treatment of the underlying disease. Eight (22%) patients in group C developed major infectious events compared with four (13%) in group CA and none in group CV (p < 0.01). Major infectious events were mainly due to Gram-positive organisms, particularly Streptococcus species. CONCLUSION: We conclude that: 1) ceftazidime alone and in association with amikacin is effective in preventing Gram-negative major infectious events; and 2) vancomycin should not be added only when a Gram-positive infection is documented, but used empirically.