Autologous bone marrow transplantation for children with AML in first remission.

In Children's cancer group (CCG) 2891, newly diagnosed patients with AML were randomized between standard and intensive timing induction therapies. Patients in first remission who lacked an HLA matched family donor were randomized between an autologous bone marrow transplantation (ABMT) where marrow was purged with 4 hydroperoxycyclophosphamide and consolidation chemotherapy. One hundred and thirty seven patients received an ABMT. Myeloid and platelet engraftment occurred at a median of 44 and 42 days, respectively. Disease-free survival (DFS), relapse-free survival and overall survival at 8 years post induction were 47% (95% confidence interval (CI): 38-55), 50% (CI: 42-59) and 55% (CI: 46-63), respectively. Multivariate analysis of DFS showed WBC <50 000/microl and having received intensively timed induction therapy were associated with improved DFS. Recipients who received intensive timed induction therapy and whose WBC was less than 50 000/microl had a DFS at 8 years of 62% (CI: 49-73). Conversely, recipients who received intensive timed induction therapy patients whose WBC was > or =50 000/microl had a DFS of 33% (CI: 17-50), P=0.003. The results confirm previous studies that ABMT is effective post remission therapy for pediatric patients with AML in first remission.

Postremission therapy for children with acute myeloid leukemia: the children's cancer group experience in the transplant era.

We reviewed consolidation therapy results and analyzed postremission outcomes for 1464 children less than 21 years old at diagnosis in five consecutive Children's Cancer Group acute myeloid leukemia trials between 1979 and 1996. Children in remission were allocated to allogeneic bone marrow transplantation (BMT) (N=373) in first remission, if a matched family donor was available. Remaining children were assigned consolidation chemotherapy (N=688) or autologous purged BMT (N=217), or withdrew from study before assignment, or with unknown data (N=186). Overall and disease-free survival were superior for children assigned allogeneic transplants. High (>50,000/microl) diagnostic white blood cell (WBC) count was prognostic for inferior outcome, but French-American-British (FAB) subtypes were not. Inv(16) is a favorable karyotypic feature for children in first remission and t(8;21) is not. Allogeneic transplantation benefit was evident in most children, including those with high or low diagnostic WBC count, each FAB subtype, and t(8;21), but was not seen in children with inv(16). Therefore, these data suggest reserving matched related donor allogeneic transplantation for children with inv(16) for second remission, but not those with t(8;21).

Intensively timed induction therapy followed by autologous or allogeneic bone marrow transplantation for children with acute myeloid leukemia or myelodysplastic syndrome: a Childrens Cancer Group pilot study.

PURPOSE: Childrens Cancer Group (CCG) protocol 2861 was designed to test the feasibility of aggressively timed induction therapy followed by autologous or allogeneic bone marrow transplantation (BMT) as the sole postremission therapy for newly diagnosed children with acute myeloid leukemia (AML) or myelodysplastic syndrome (MDS). PATIENTS AND METHODS: Between April 1988 and October 1989, 142 patients were eligible for study. All patients entered received a timing-intensive five-drug induction of dexamethasone, cytarabine (Ara-C), thioguanine, etoposide, and daunorubicin (DCTER) over 4 days with a second cycle administered after 6 days of rest, irrespective of hematologic status at that time. Most patients subsequently received a second two-cycle induction course. Those who achieved remission were eligible for bone marrow ablative therapy with busulfan and cyclophosphamide, followed by 4-hydroperoxy-cyclophosphamide (4-HC)-purged autologous or allogeneic BMT rescue. RESULTS: One hundred eight (76%) patients achieved remission: 19 (13%) died of complications of the leukemia and/or chemotherapy, and 15 (11%) failed to achieve remission. Seventy-four patients subsequently underwent BMT with either autologous (n = 58) or allogeneic (n = 16) rescue. For patients who received autologous rescue with 4-HC-purged grafts, the actuarial disease-free survival (DFS) rate at 3 years from the day of transplant is 51%, compared with 55% for patients who received allogeneic grafts (P = .92). At 3 years, the overall actuarial survival rate for all 142 patients entered on this study is 45%, with an event-free survival (EFS) rate of 37%. Adverse prognostic factors for outcome included an elevated WBC count or the presence of CNS leukemia at the time of AML diagnosis. CONCLUSION: Results suggest that aggressively timed induction therapy followed by marrow ablation and BMT rescue with either autologous or allogeneic grafts for children with newly diagnosed AML or MDS is both feasible and effective.

Comparison of preparative regimens in transplants for children with acute lymphoblastic leukemia.

PURPOSE: Preparative regimens involving total-body irradiation (TBI) produce significant late toxicities in some children who receive bone marrow transplants, including impaired growth and intellectual development. Busulfan is often used as an alternative to TBI, but there are few data regarding its relative efficacy. PATIENTS AND METHODS: We compared outcomes of HLA-identical sibling transplants for acute lymphoblastic leukemia (ALL) in children (< 20 years of age) who received cyclophosphamide plus TBI (CY/TBI) (n = 451) versus those who received busulfan plus cyclophosphamide (Bu/CY) (n = 176) for pretransplant conditioning. Patients received transplants between 1988 and 1995 and their results were reported to the International Bone Marrow Transplant Registry by 144 participating institutions. The CY/TBI and Bu/CY groups did not differ in gender, immune phenotype, leukocyte count at the time of diagnosis, chromosome abnormalities, remission status, or length of initial remission. T-cell depletion was used more frequently in the CY/TBI group; the Bu/CY group included a higher proportion of children who were less than 5 years of age. The median follow-up period was 37 months. RESULTS: The 3-year probabilities of survival were 55% (95% confidence interval [CI], 50% to 60%) with TBI/CY and 40% (95% CI, 32% to 48%) with Bu/CY (univariate P =.003). The 3-year probabilities of leukemia-free survival were 50% (95% CI, 45% to 55%) and 35% (95% CI, 28% to 43%), respectively (univariate P =.005). In a multivariate analysis, the risks of relapse were similar in the two groups (relative risk [RR], 1.30 for Bu/CY v CY/TBI; P =.1). Treatment-related mortality was higher in the Bu/CY group (RR, 1.68; P =.012). Death and treatment failure (relapse or death, inverse of leukemia-free survival) were more frequent in the Bu/CY group (RR, 1. 39; P =.017 for death; RR, 1.42; P =.006 for treatment failure). CONCLUSION: These data indicate superior survival with CY/TBI conditioning, compared with Bu/CY conditioning, for HLA-identical sibling bone marrow transplants in children with ALL.

Morphologic, immunologic, and cytogenetic classification of acute myeloid leukemia and myelodysplastic syndrome in childhood: a report from the Childrens Cancer Group.

The purposes of this report are to reaffirm concordance difficulties with the acute myeloid leukemia (AML) French-American-British (FAB) classification, to present the frequency of previously delineated AML syndromes in pediatric patients and to describe additional characteristic AML profiles utilizing composite morphologic, cytogenetic and immunophenotypic data. Profiles of 124 children with acute myeloid leukemia (AML) and 13 children with myelodysplastic syndrome entered on the Childrens Cancer Group (CCG) pilot study CCG-2861 were examined. Concordance between institutions and reviewers for FAB designation was 65%. Discordance was found principally between M1 and M2, M2 and M4, and M4 and M5. In 49% of marrow specimens, leukemic blasts expressed at least one T lineage-related antigen; 24% expressed the B lineage-related antigen CD19. CDw14 correlated with FAB M4 or M5 morphology and was the only surface antigen associated with a specific FAB subtype. Normal karyotypes were found for 15% of the 75 children with satisfactory karyotype preparations. Recurring aberrations, found in 76% of children, included t(15;17)(q22;q11), t(8;21)(q22;q22), inv(16)(p13q22), rearrangements of band 11q23, t(6;9) (p23;q34), trisomy 8 and monosomy 7. Results from this pilot study and from the current CCG randomized trial correlating morphology, immunophenotyping and cytogenetics, will help to classify AML into unique subgroups with differing clinical consequences or therapy requirements.

The effects of antithymocyte globulin on natural killer cells.

We studied the effects of antithymocyte globulin (ATG) on natural killer (NK) cell activity to evaluate whether ATG may be useful in preventing NK-mediated disorders such as rejection of mismatched bone marrow (BM). We found that four of four lots of ATG eliminated greater than 95% of NK activity against K562. The effect of ATG on NK-mediated suppression of granulocyte-macrophage colony-forming units (CFU-GM) was also studied. ATG at a concentration of 1.0 mg/ml completely abrogated NK-mediated suppression of CFU-GM, as did monoclonal antibody Leu 11b/anti-CD 16 and complement (BRC). In contrast, OKT3/anti-CD3 had no effect on NK-mediated suppression of CFU-GM, suggesting that the effector cells were most likely NK cells rather than T cells. We also studied the effect of gamma interferon (INF) on NK-mediated suppression of CFU-GM and showed that eliminating NK cells with Leu 11b and BRC or ATG did not reduce gamma INF suppression of CFU-GM, suggesting that gamma INF directly suppressed CFU-GM rather than augmenting NK activity. Based on these results we postulate that ATG may have a role in the prevention or treatment of suppression of hematopoiesis thought to be caused by NK cells.

Double fluorochrome analysis of human bone marrow lymphoid cells: studies with terminal transferase and anti-T-cell monoclonal antibodies.

Mature thymus-derived (T) lymphocytes are generally believed to be derived from a bone marrow progenitor cell. Data from studies with animals suggest that the enzyme terminal deoxynucleotidyl transferase (Tdt) is expressed in many T-cell progenitors in bone marrow. In this study we attempted to identify Tdt+ bone marrow cells in man that may be committed to T lineage based on coexpression of Tdt and antigens that have previously been useful in characterization of thymocytes or peripheral-blood T cells. We used a panel of ten monoclonal antibodies against such antigens to analyze Tdt+ bone marrow cells using two-color immunofluorescence. We found that T-cell-associated antigens were not expressed on Tdt+ bone marrow cells and that T cells in bone marrow have a phenotype similar if not identical to peripheral-blood T cells. These results support the hypothesis that many postthymic immunocompetent T cells are found in human marrow. Our results also suggest that if Tdt+ bone marrow cells are committed to T lineage, then the acquisition of mature T-cell-associated antigens is an intrathymic event.

Characterization of thymocytes expressing the common acute lymphoblastic leukemia antigen.

We studied the relationship between CALLA + thymocytes and two known markers of T-lymphocyte differentiation, Tdt and the sheep erythrocyte receptor. Thymocytes were studied using double fluorochrome analysis (with monoclonal anti-CALLA antibody and anti-Tdt) before and after E rosette separation. We found that approx. 4% of unseparated thymocytes were CALLA + and that most CALLA + cells were also Tdt +. After E rosettes separation CALLA + Tdt + cells were found mostly in the ER- fraction (20% of ER- cells) while only 1.0% of ER + cells were CALLA +. The expression of CALLA on ER- Tdt + thymocytes suggests that CALLA may define cells early in T-cell differentiation.

Autoimmune manifestations of the Wiskott-Aldrich syndrome.

OBJECTIVE: To describe and review the autoimmune features and typical manifestations of Wiskott-Aldrich syndrome (WAS). DESIGN: Case series and review of the literature. SETTING: Tertiary care medical center and pediatric referral center. PATIENTS: The presentation, diagnosis, and management of two cases are reported. In addition to the typical features of WAS, the first patient had hemolytic anemia, arthritis, leukocytoclastic vasculitis, and colitis. The second patient had colitis and arthralgias. Detailed review of features and therapeutic options in WAS as exemplified by these two patients are presented. Both patients had bone marrow transplantation, the only definitive treatment for WAS. CONCLUSIONS: WAS has variable clinical and autoimmune manifestations. Diagnosis must be suspected in a boy with small, decreased number of platelets and autoimmune problems or infections. Bone marrow transplantation is the only successful mode of treatment for all aspects of WAS.

Autologous bone marrow transplantation for acute myeloid leukemia in remission or first relapse using monoclonal antibody-purged marrow: results of phase II studies with long-term follow-up.

One hundred and thirty-eight patients with AML underwent ABMT with monoclonal antibody plus complement-purged marrow between August 1984 and March 1997. One hundred and ten patients were in CR (CR1: 23; CR2/3: 87) and 28 were in first relapse (R1) at ABMT. Preparative regimens included busulfan (16 mg/kg) and CY (120 mg/kg) (n = 93), CY (120 mg/kg over 2 days) with TBI (1200 cGy) (n = 35), and busulfan (16 mg/kg) plus etoposide (60 mg/kg) (n = 10). CR1 patients treated with CY/TBI (n = 7) had 3- and 5-year disease-free survival (DFS) rates of 71% and 57%. CR1 patients treated with BU/CY (n = 12), had 3- and 5-year DFS rates of 45%. Three and 5-year DFS for CR2/3 patients treated with CY/TBI (n = 26) was 23%. Three- and 5-year DFS for patients in CR2/3 treated with BU/CY (n = 55) was 31 and 28%. Three- and 5-year DFS for patients in R1 treated with BU/CY (n = 26) was 37%. In multivariate analysis, increased age was associated with greater risk of death and relapse. For CR2/3 patients, the length of CR1 was a significant predictor of DFS. ABMT performed in CR or R1 results in excellent 5-year DFS and OS. The contribution of purging may require a randomized trial comparing purged vs unpurged stem cell infusions.

Cytokine-mobilized allogeneic peripheral blood stem cell transplants in children result in rapid engraftment and a high incidence of chronic GVHD.

Between October 1995 and October 1998, 24 children aged 9 months to 17 years (median 11 years) underwent cytokine-mobilized allogeneic peripheral blood stem cell (PBSC) transplantation for treatment of hematological disorders. All of the transplants were the first allogeneic transplant for the recipient. Twenty patients were transplanted for hematological malignancies (ALL = 8, AML = 6, CML = 4, MDS = 2) and four patients were transplanted for non-malignant disease (thalassemia major = 2, Wiskott-Aldrich syndrome = 1, Kostmann's syndrome = 1). Nineteen donors were HLA-identical siblings, four were HLA-matched or single antigen mismatched parents, and one was a syngeneic transplant. Donors aged 8 to 38 years (median 15 years, 14 donors <18 years) received G-CSF 10 microg/kg/day subcutaneously beginning 4 days before PBSC collection and were submitted to one to three leukapheresis collections. The median CD34+ cell yield was 7.8 x 106 cells/kg recipient body weight. All patients achieved an ANC >0.5 x 109/l after a median of 13 days (range 10-21). Twenty-three patients eventually achieved platelet transfusion independence. One patient died on day 63 without ever achieving platelet transfusion independence. Four patients received platelet transfusions to maintain a platelet count well above 20 x 109/l due to bleeding complications. Of the 19 evaluable patients, the median time to a non-transfused platelet count of 20 x 109/l was 12 days (range 0-44). Ten of 23 at-risk patients developed acute GVHD grades II to IV, with grades III to IV in four patients. Twelve of 19 patients followed for at least 100 days have developed chronic GVHD (extensive = 2, limited = 10) with an actuarial risk of chronic GVHD of 75% at 1 year. The Kaplan-Meier estimate of event-free survival is 65% at 2 years. Four patients died (GVHD = 3, VOD = 1), three patients relapsed, and one patient with thalassemia major had a late graft failure with autologous recovery. Based upon our experience, allogeneic PBSCT is safe for both pediatric donors and recipients and engraftment of neutrophils and platelets is rapid. Bone Marrow Transplantation (2000) 25, 13-18.

Post-transplant lymphoproliferative disorder after autologous peripheral stem cell transplantation in a pediatric patient.

Post-transplant lymphoproliferative disorder (PTLD) is a complication of allogeneic bone marrow transplantation (BMT). Rare cases of PTLD after autologous BMT have been reported only in adults. This case report is the first to describe PTLD in a pediatric patient after autologous peripheral stem cell transplantation (PSCT). This 2-year-old male with stage IV neuroblastoma underwent autologous PSCT. The post-PSCT course was complicated by fever with hematochezia and a lung mass. On day 94 post PSCT, colonoscopy revealed an ulcer due to a PTLD, monomorphic type, B cell phenotype, associated with Epstein-Barr virus. Fine needle aspiration identified the lung mass as neuroblastoma. PTLD can occur in pediatric autologous PSCT recipients, and may occur more frequently in autologous grafts manipulated by T cell depletion or CD34+ cell selection.

A modified method for human bone marrow filtration prior to bone marrow transplantation.

A technique is described for filtering harvested bone marrow using disposable materials, namely a 4 x 4 inch piece of sterile gauze that is gently packed into the barrel of a 60-ml plastic disposable syringe, which is connected directly to a blood collection bag. The filtration of marrow directly into the collection bag eliminates additional filtration steps and therefore may potentially reduce the incidence of inadvertent microbial contamination. In this study we describe this filtering technique and compare it to the method described by Thomas and Storb. Numbers of granulopoietic progenitors (CFU-GM) and erythropoietic progenitors (BFU-E), total white cell counts, percentage of cells positive for the CD3 (OKT3) lymphocyte surface membrane marker, and volume changes were studied following filtration by each method. The two techniques were shown to be comparable in terms of these parameters. Furthermore, when compared with historical controls, this method resulted in a reduced incidence of microbial contamination compared to filtration using successive stainless steel screens.

FK506 (tacrolimus) in the treatment of steroid-resistant acute graft-versus-host disease in children undergoing bone marrow transplantation.

Steroid-resistant graft-versus-host disease (GVHD) is an often lethal complication of bone marrow transplantation (BMT). FK506 (tacrolimus) is a new potent immunosuppressant which has been shown to be superior to conventional immunosuppression in the prevention and treatment of graft rejection in recipients of solid organ transplants. To determine whether FK506 is effective in the treatment of steroid-resistant acute GVHD, 6 children with biopsy-proven severe GVHD were studied. FK506 was administered as intravenous or oral therapy and the dose was adjusted to achieve serum levels between 0.5 and 1.0 microgram/ml by ELISA. Steroid doses were tapered based on clinical grading in each organ. Within 1-2 days, improvement occurred in skin and gut in all patients, and in liver in 3 patients. Toxicity attributable to FK506 was similar to that described in solid organ transplant patients and included neurotoxicity, nephrotoxicity and gastrointestinal effects. While FK506 is effective in the treatment of steroid-resistant acute GVHD, toxicity may limit its use. Further studies evaluating FK506 as GVHD prophylaxis and treatment of less advanced GVHD are needed.

Lymphoid progenitor cells in severe combined immunodeficiency.

In the present study we evaluated the possibility that patients with severe combined immunodeficiency (SCID) might be deficient in lymphoid progenitor cells in bone marrow. Bone marrow from six patients with SCID was studied for the presence of cells expressing antigens associated with the earliest known stages of lymphopoiesis--terminal transferase (Tdt), the common acute lymphocytic leukemia antigen (CALLA), and p24. Four of six patients had detectable Tdt+, CALLA+, and p24+ cells, although they were quantitatively reduced compared to results from normal infant marrow. In two of six patients no bone marrow mononuclear cells expressing any of these markers were detected. These two patients were more lymphopenic than the other four SCID patients. The absence or deficiency of Tdt+, CALLA+, and p24+ bone marrow cells in some patients with SCID (two of six in the present study) is consistent with the lymphopenia seen in these patients and suggests that the underlying defects which result in SCID affect the production of immature as well as more differentiated lymphocytes.

Expression of the CD4 molecule on acute nonlymphocytic leukemia (ANLL) cell lines.

Mature and immature monocytes express the CD4 molecule similar to that expressed on lymphocytes. Since monocytes and cells of myeloid lineage are derived from a common progenitor, we studied a panel of nine myeloid leukemia cell lines for the expression of the CD4 molecule. We found that six of nine myeloid leukemia cell lines (U937, KG1-B, HL-60, THP-1, HEL 92.1.7, KMOE) expressed CD4, the exceptions being the erythroleukemia line K562, myeloblast line KG1-REV, and megakaryocytic line, CHRF-288. SDS-PAGE analysis of HL-60 cells immunoprecipitated with OKT4 showed the presence of a 55 kd molecule similar in weight to that seen on lymphocytes. These data suggest that some myeloid progenitor cells express the CD4 molecule and that the CD4 may have a broader distribution within hematopoietic cells.

Comparison of two immuno-mechanical methods of T-cell depletion of human bone-marrow for prevention of graft-versus-host disease: soybean lectin agglutination and sheep erythrocyte rosette depletion versus triple rosette depletion.

Depletion of T-lymphocytes from HLA-mismatched donor bone marrow can be accomplished by either triple neuraminidase-treated sheep erythrocyte depletion (3En) or soybean agglutination and sheep erythrocyte depletion (SBA/E/En). T-lymphocyte depletion by 3En resulted in higher yields of all marrow precursor phenotypes (studied by a battery of monoclonal antibodies) than did SBA/E/En depletion. While both procedures enriched for some early precursor cells (e.g., HLA-DR and/or terminal deoxynucleotidyl transferase (TdT) positive cells), greater numbers of these cells were available after 3En. Clinical benefits which may be derived from the larger inoculum of T-depleted bone-marrow cells available after 3En treatment remain to be studied.

Absorption and elimination of photodieldrin by Daphania and goldfish.

The alga, Ankistrodesmus spiralis, becomes saturated with photodieldrin in 8 hours. The Daphnia whether contaminated by feeding on the algae saturated with photodieldrin or by directly absorbing this insecticide eliminate about 50 per cent of the absorbed photodieldrin in 4 days which increases up to 70 per cent in 7 days. However, continuous exposure of daphnids to photodieldrin results in increased absorption and accumulation of this insecticide. Transfer of the goldfish contaminated with photodieldrin to clean water results in initial elimination in 24 hours whose rate in is then is then subsequently reduced. These two organisms are thus capable of eliminating their body residues of photodieldrin, the "terminal residue: of dieldrin, if their environment is decontaminated.