Allogeneic bone marrow transplantation-mediated transfer of specific immunity against Toxocara canis associated with excessive IgE.

A girl with myelodysplastic syndrome (RAEB-T) received HLA-identical bone marrow from her younger brother after myeloablative treatment with busulfan and cyclophosphamide. After bone marrow transplantation, fever, exanthema, pruritus, and a pulmonary infiltrate were treated symptomatically. Bacterial cultures remained negative. Leukocyte engraftment began on day 10, and all blood cell populations proved to be of donor origin on FISH analysis. Increasing IgE levels (21 000 U/ml) on day 14 after BMT, positive RAST, specific IgG-antibodies, and missing Toxocara (T.) canis antigens in the recipient indicated donor-derived seroconversion. Before BMT, the recipient had been negative for T. canis in routine parasitological screening, and the donor proved to be positive for T. canis antibody by ELISA. This report suggests that the transfer of IgE immunity in the absence of detectable antigens may be responsible for IgE-mediated symptoms consistent with toxocara infection and confirms the need for parasite screening in donor medical examinations.

A single centre experience with allogeneic stem cell transplantation for severe aplastic anaemia in childhood.

BACKGROUND: Severe aplastic anaemia (SAA) is a rare disorder which has a fatal course when allogeneic stem cell transplantation (SCT) or an immunosuppressive regimen is not applied. Stem cell replacement is the only curative approach for these patients but it is limited by the availability of a compatible donor. PATIENTS: Between 1982 and 1993, 18 children (15 boys, 3 girls) with SAA and HLA identical, MLC negative donors underwent SCT in our institution. SAA was preceded by viral infection in 8 patients (3x hepatitis, 1x measles, 1x herpes simplex infection and 3x viral upper respiratory tract infections). It was drug-associated in one and idiopathic in the 9 others. The median age at diagnosis was 9.7 years (range, 2 months to 16 years). Pretreatments included corticosteroids in 11/18 patients, androgens in 4 patients in addition, two had received cyclosporin A (CSA). One patient progressing from Diamond- Blackfan anaemia to SAA had multiple immunosuppressive treatment courses over 7 years before his grand-uncle was identified as donor while 4 patients had no treatment prior to SCT. METHODS: Early SCT (within 90 days after diagnosis) was performed in 9/18 patients and the median interval between diagnosis and SCT was 2.6 months (range, 0.5 to 7 years). The stem cell source was the bone marrow (BM) of a syngeneic twin in 2 patients, the BM (13 patients) or the cord blood (1 patient) of a sibling whilst it was BM from a HLA-phenotypical family donor (1 father, 1 grand-uncle) in two patients. Cyclophosphamide 50 mg/kg on 4 consecutive days was given as preparative regimen to 16 patients but not to the two syngeneic twins. Rejection prophylaxis included total lymphoid irradiation in 5/16 patients while in the other 11 patients donor buffy coat cells were given on days +1 to +4. The syngeneic twins had no need for either approach. Patients received a median number of 3.7 x 10(8)/kg nucleated cells (range, 2.6 to 6.7). Prophylaxis of graft versus host disease (GVHD) was carried out with MTX alone (n = 12), with CSA alone (n = 2) or with both (n = 4). All patients received standard supportive care. RESULTS: The overall survival is 89% at the median observation time of 100 months. The median time to reach 500 granulocytes was 24 days (range, 15 to 40). Median time to become transfusion independent after BMT was 30 days for platelets (range, 2 to 111) and was 28 days for packed red blood cells (range, 6 to 128). Acute GVHD was observed in 10/18 patients and involved only skin in 6 patients, skin and liver or gut in two patients and all 3 organs in another two patients. Seven of 10 patients had grade 1 to 2 a GVHD toxicity, whereas 3 patients experienced grade 3 to 4 acute GVHD. Chronic GVHD developed in 5 patients. Acute transplant related mortality was 5.5%. Cause of death was persisting non engraftment till day +180 after 2 transplant procedures in a boy with previous platelet transfusions from his mother. Late mortality occurred in 2 patients: one chronic GVHD associated haemorrhage 20 months after SCT and one chronic GVHD associated septicaemia 10 years after SCT. CONCLUSION: Although this report reflects patients data accumulated over 15 years, results compare favourably with more recent survival data. Acute and late transplant related toxicity was low in patients undergoing early transplantation with adequate prior supportive care. This data confirms that SCT still should be the first treatment choice if an HLA identical sibling is available.

Influence of fractionated total body irradiation on mucosal toxicity in intensified conditioning regimens for autologous bone marrow transplantation in pediatric cancer patients.

From April 1988 to March 1991 28 children with generalized solid tumors (N = 15) or hematologic malignancies (N = 13) received intensified myelotoxic regimens followed by autologous stem cell rescue (ABMT). These intensified regimens consisted of 12 Gy fractionated total body irradiation (FTBI) and 2 (or 3) cytotoxic drugs (group A, n = 19) or a combination of 3 cytotoxic drugs (group B, n = 9). FTBI-containing regimens produced more severe mucositis > = WHO grade 3 (p = 0.01) and a longer duration of severe mucositis. The mucositis had a median duration of 8 days (range 0-28) in group A compared with median 0 days (range 0-7) in group B (p < 0.01). Acute renal and liver toxicity were not different. The probability of overall survival at day +100 was 89% in all patients. In terms of long-term survival FTBI containing regimen did not prove superior: 5 out of 19 patients in group A and 6 out of 9 patients in group B have been survivors for a minimum of 3 years. In conclusion, severe gastrointestinal toxicity of such intensive regimens is avoidable if FTBI is omitted.

Tolerance of high doses of amphotericin B by infusion of a liposomal formulation in children with cancer.

Conventional amphotericin B (Amph-B) is the drug of choice for treating systemic fungal infections. Recently, a new formulation has become available, encapsulated in liposomes (Amph-lip). This new form of administration was developed in order to lower the acute side effects and to offer the possibility of administering high doses of amphotericin B. Experience with Amph-lip is limited, especially in children. We treated four children with documented systemic fungal infections with Amph-lip and administered it empirically to 12 children. Fifteen of these 16 children were severely granulocytopenic oncologic patients. One 3-month-old baby suffered from systemic candidiasis. Amph-lip was preferred to conventional Amph-B in children with organ dysfunction developing as a consequence of conventional chemotherapy or bone marrow transplantation, after failure of conventional Amph-B to improve a fungal infection, and after adverse drug reactions had occurred. The daily doses of Amph-lip ranged from 1 to 6 mg/kg (median 3 mg/kg), the cumulative doses from 13 to 311 mg/kg (median 75 mg/kg). Acute adverse reactions or organ function abnormalities attributable to Amph-lip did not occur in 402 administrations. Amph-lip has proven to be well tolerated by children in terms of acute toxicity and in the long term. Although large cumulative doses were given, organ function abnormalities attributable to Amph-lip doses were not detected in any of ten long-term survivors over a median observation time of 36 months (range 30-44 months). Amph-lip appears to be a promising alternative antifungal treatment, especially for patients with impaired organ function, when high doses of amphotericin B are necessary.

Treatment of infant leukemia with busulfan, cyclophosphamide +/- etoposide and bone marrow transplantation.

Infants with acute leukemia have a poor chance of being cured by conventional chemotherapy. We therefore treated cases of infant leukemia with high dose chemotherapy followed by bone marrow transplantation (BMT). Six suffered from acute leukemia and one from refractory anemia with excess of blasts (RAEB-t). The conditioning regimen consisted of busulfan (BU) and cyclophosphamide (CY), and was intensified by adding etoposide (VP) in four cases. At the time of BMT the children were 4, 5, 12, 13, 13, 14, and 20 months old. Three children were autografted, three received HLA-identical marrow from a sibling donor, and one child received matched unrelated donor marrow. All five children who were grafted in complete (CR) or partial remission (PR) are alive and well in CR 7, 13, 24, 37, and 46 months after allogeneic (two patients) or autologous (three patients) BMT, and 13, 17, 29, 42, and 53 months after initial diagnosis. The child with RAEB-t and the one transplanted in second chemotherapy-resistant relapse of acute non-lymphoblastic leukemia relapsed at 7 and 17 months respectively. The chemotherapy regimen was well tolerated. BU-CY-VP is a promising alternative treatment to regimens including total body irradiation for very young children suffering from acute leukemia.

Recovery kinetics after chemotherapy and circulating mononuclear cells expressing the CD34 antigen in pediatric cancer patients.

Hematopoietic progenitor cells collected from the peripheral blood are capable of restoring hematopoiesis after myeloablative therapy. The numbers of circulating mononuclear cells expressing the CD34 antigen were calculated and the colony-forming capacity was determined in 26 blood samples, which were drawn during rapid rise of leukocytes after chemotherapy cycles that were followed by aplasia. Culture assay after 14 days revealed a median 507 (210-2029) myeloid progenitors (CFU-GEMM/GM) per 10(5) nucleated cells (NC) in 13 CD34-positive samples, and only a median 76 (9-224) in 13 CD34-negative ones (p less than 0.001). Median 343 (175-2450) erythroid burst-forming units (BFU-E) per 10(5) NC were detected in the CD34-positive samples, whereas only 72 (10-315) per 10(5) NC were found in the negative ones (p less than 0.01). The percentage of CD34-positive cells clearly correlated with the growth of CFU-GEMM/GM and BFU-E (p less than 0.01). The content of CD34-positive cells in circulation was determined within 120 min by FACS analysis and predicted colony-forming capacity of circulating mononuclear cells. These observations will help to select the optimal individual days for leukaphereses.

Conventional vs. liposomal amphotericin B in immunosuppressed children.

Invasive fungal infections, mostly caused by Candida and Aspergillus species, are a major cause of early morbidity and mortality in immunocompromised children. The treatment of choice for systemic fungal infections is still the early intravenous administration of amphotericin B. However, conventional AMB therapy is often limited by severe side effects such as fever, chills, bronchospasm, and nephrotoxicity. In recent reports liposomal AMB (AmBisome) was shown to be effective in the treatment of severe systemic fungal infections. So far, clinical experience with AmBisome in children is still anecdotal and no comparative study is yet available. In the following we report on 11 immunosuppressed children who were treated with conventional or liposomal AMB for longer than 3 weeks.

Limited tolerance of intensified conditioning regimens in children receiving methotrexate/cyclosporin A for graft-versus-host disease prophylaxis.

Twenty-one patients with a median age of 9 years (0.5-19) underwent intensified myeloblative therapy: 1800 mg/m2 etoposide (VP) was added to 120 mg/kg cyclophosphamide (CY) and 12 Gy fractionated total body irradiation (FTBI) or 12-16 mg/kg busulfan (BU) for treatment of acute lymphoblastic leukemia (11 patients), acute myeloid leukemia (8 patients), non-Hodgkin's lymphoma (1 patient), or myelodysplastic syndrome (1 patient). Severe liver toxicity occurred in 5 of 7 children (71%) receiving short-term methotrexate (MTX) and additional cyclosporin A (CSA) for prophylaxis of graft-versus-host disease (GVHD). Three of them died of subsequent acute renal failure on days 8, 13, and 34. In contrast, acute severe organ toxicity occurred in only 1 of 14 children (7%) receiving the same intensified regimens who were autografted (7 pts) or received MTX alone for GVHD prophylaxis (7 pts). These observations suggest that GVHD prophylaxis with MTX and CSA may adversely influence the tolerance of intensified antileukemic regimens in children.

Unusually long survival after autografting in second partial remission of translocation t(4;11) acute infant leukemia.

About 70% of children with acute lymphoblastic leukemia may be cured by conventional chemotherapy. The prognosis is considerably worse in infant leukemia with a translocation t(4;11). We report an infant with a diagnosis of cytochemically undifferentiated acute hybrid leukemia (pre pre B-ALL coexpressing one myelomonocytic marker) and t(4;11). Initial clinical presentation and the course of the disease were typical for t(4;11) acute leukemia. After an early hematologic relapse intensive chemotherapy resulted only in a second partial remission 7 months after initial diagnosis. Subsequent bone marrow transplantation with 16 mg/kg busulfan and 200 mg/kg cyclophosphamide followed by the infusion of autologous purged bone marrow resulted in a continuous second remission which has lasted 46 months so far.

Is treatment intensification by adding etoposide and carboplatin to fractionated total body irradiation and melphalan acceptable in children with solid tumors with respect to toxicity?

Conventional chemotherapy results in high mortality rates in patients with solid tumors involving the bones or the bone marrow. High dose melphalan (MEL) with or without total body irradiation followed by bone marrow transplantation (BMT) has prolonged survival, but curative potential has remained disappointing. In order to improve survival 20 children with generalized or relapsed solid tumors (neuroblastoma, peripheral neuroectodermal tumor, Ewing's sarcoma, rhabdomyosarcoma) underwent autologous (n = 16) or allogeneic (n = 4) BMT. The myeloablative regimen consisted of 12 Gy fractionated total body irradiation (FTBI) and high dose MEL. In 12 of these patients this regimen was intensified by giving 60 mg/kg etoposide (1800 mg/m2 VP), and 1.5 g/m2 carboplatin (CBDCA) was added in seven of these 12 patients. The intensification of FTBI and MEL by adding VP and CBDCA was followed by acceptable toxicity. Acute liver toxicity in 15/20 patients (75%) and acute renal toxicity in 17/20 patients (85%) did not exceed WHO grade 1. The use of the conditioning regimen FTBI-MEL-VP-CBDCA during first chemotherapy response is a promising approach in treatment of children suffering from generalized solid tumors.

Amphotericin B serum levels in pediatric bone marrow transplant recipients.

We studied amphotericin B (AMB) serum levels (n = 590) in 41 pediatric patients, who underwent allogeneic (21) or autologous (20) bone marrow transplantation (BMT). All patients received AMB orally as part of a total gut decontamination; 30/41 patients (73%) had AMB i.v. either for prophylaxis or therapy of fungal infections. Rapid initial dose escalation of AMB and the infusion over 1 h only were well tolerated by the children. Serum level monitoring allowed AMB long-term treatment safely to be administered in children suffering from transplantation-related complications (veno-occlusive disease of the liver, graft-versus-host disease of the liver). An h.p.l.c. method was used for monitoring AMB serum trough levels to avoid levels exceeding 2 mg/l. One lethal fungal infection was observed in 41 pediatric BMT recipients (2.4%). Rapidly increasing doses of AMB at start of therapy and drug monitoring by h.p.l.c. might help to reduce fungal mortality and renal toxicity by a dose sparing effect in BMT recipients.

[Autologous bone marrow transplantation in children with acute leukemia or non-Hodgkin's lymphoma]. / Autologe Knochenmarktransplantation bei Kindern mit akuter Leukämie oder Non-Hodgkin-Lymphom.

More than 95% of children with acute lymphoblastic leukemia (ALL) achieve complete remission with conventional chemotherapy and 60 to 70% are cured. In contrast, only 70 to 80% of children with acute myeloid leukemia (AML) achieve remission and 30 to 40% are cured. In acute leukemia, ALL and AML, high initial white blood cell counts, poor initial response to therapy, certain structural chromosomal abnormalities, and diagnosis under 1 year of age indicate an increased risk of relapse. Prognosis is poor in children, who relapse during treatment. If no bone marrow donor is available, high dose chemo(radio)therapy and autologous stem cell rescue can be applied in these children to consolidate second remission. Furthermore, autologous bone marrow transplantation may be used to consolidate first remission of individual children, who suffer from acute leukemia with high risk criteria. 19 children with a median age of 8 years (0.5 to 19 years) underwent high dose chemo(radio)therapy and autologous bone marrow and/or blood derived stem cell rescue. 11 suffered from ALL, 4 had AML, and 4 suffered from Non Hodgkin's lymphoma (NHL). 7/13 children (54%), who were in first or second remission after early relapse, are alive in continued remission for median 24 months (9 to 44) after autografting. Children in more advanced stage of disease had no benefit from high dose therapy and autologous stem cell rescue.

High-dose melphalan, etoposide +/- carboplatin (MEC) combined with 12-gray fractionated total-body irradiation in children with generalized solid tumors.

Long-term disease-free survival is poor in patients with primary generalized or relapsed solid tumors. High-dose chemoradiotherapy with stem cell rescue improved survival, but more effective protocols are needed. From January 1988 to November 1988, we treated 7 patients (median age, 9 years; range, 3-23 years) with an intensified treatment program. They received 12-Gy fractionated, total-body irradiation (FTBI). High-dose chemotherapy (MEC) consisted of melphalan (120-140 mg/m2 Mel) and etoposide (40-60 mg/kg VP-16) with or without carboplatin (1.5 g/m2 CBDCA). Although we combined 12-Gy FTBI with Mel, VP-16, +/- CBDCA in doses used previously for high-dose single-agent chemotherapy, the extramedullary toxicity of FTBI with ME(C) was tolerable. Two of the four patients who were grafted without delay after good initial chemotherapy response are still alive in continued complete remission 30 and 33 months, respectively, after initial diagnosis. Early application of FTBI and ME(C) during first chemotherapy response might improve outcome in patients with primarily generalized solid tumors.

Capillary leak syndrome during low dose granulocyte-macrophage colony-stimulating factor (rh GM-CSF) treatment of a patient in a continuous febrile state.

A 20 years old man with peripheral primitive neuroectodermal tumor involving the bone marrow received 12 Gy fractionated total body irradiation, 140 mg/m2 melphalan, 1800 mg/m2 etoposide, and 1500 mg/m2 carboplatin for consolidation of first remission. Thereafter, 250 micrograms/m2/day recombinant human granulocyte-macrophage colony-stimulating factor (rh GM-CSF) (Behring Werke) were administered as continuous infusion 4 days after infusion of autologous bone marrow and peripheral stem cells to accelerate granulocyte reconstitution for control of a continued febrile state. The clinical picture of capillary leak syndrome developed with weight gain, pleural effusions and peripheral edema. The patient's condition stabilized after discontinuation of rh GM-CSF. Eight days later he died of invasive aspergillosis. The clinical course of our patient suggests a potentially fatal toxic effect of rh GM-CSF, even in low dose, in the setting of septicemia or fungemia.

Autoantibodies in chronic GVHD: high prevalence of antinucleolar antibodies.

Sera from 32 bone marrow allograft recipients were screened for the presence of autoantibodies 4-61 months post-graft. Sera from 12 of 19 patients with extensive chronic graft-versus-host disease (c-GVHD) stained the nucleolar region strongly in immunofluorescence, indicating the presence of specific antinucleolar antibodies. In contrast, none of three patients with limited and none of 10 patients without c-GVHD had antinucleolar antibodies. Antibodies reacting with nuclear constituents other than nucleoli were found in five of the 12 antinucleolar positive patients. The appearance of antinucleolar antibodies coincided with early clinical symptoms of c-GVHD. We conclude that the appearance of antinucleolar antibodies after bone marrow transplantation is specific for patients with extensive c-GVHD. Furthermore, the development of extensive c-GVHD is paralleled by the emergence of these antinucleolar antibodies.

The median daily increment of leukocytes during hematopoietic recovery reflects the myeloid progenitor cell yield during leukapheresis in children.

Twenty-two consecutive patients with a median age of 11 years (range 3-23) underwent 103 leukaphereses for collection of peripheral stem cells. They suffered from various solid tumors and hematologic malignancies. Our aim was to collect at least 2.5 x 10(4) myeloid committed stem cells (CFU-GM) per kg body weight. This stem cell number has reliably resulted in rapid and sustained hematopoietic reconstitution after autografting in our institution. Peripheral stem cell mobilization was induced by myelosuppressive therapy alone. If the stem cell collections were started at the first platelet rise and were continued during the following 8 days, the stem cell yield of one procedure was high - median 1.02 x 10(4) CFU-GM/kg body weight (range 0.02-13.3). In 69 such 'well-timed' collections, a median of only 4 leukaphereses (range 2-6) were needed to collect high numbers of CFU-GM. The yield from leukaphereses carried out at any other time was unsatisfactory. In 34 'random' collections, a median of only 0.27 x 10(4) CFU-GM/kg body weight (range 0.01-1.11) was obtained in one procedure. In 69 well-timed collections the CFU-GM yield depended on the absolute number of circulating leukocytes (r = 0.369, p = 0.002) and mononuclear cells (r = 0.476, p less than 0.001). The median daily increment of leukocytes to the day of apheresis proved to be an excellent parameter for the prediction of the yield and could be used to select the best days for leukaphereses (r = 0.403, p = 0.002). These findings may help to make the collection of blood-derived stem cells more efficient in pediatric patients.

Autologous rescue with blood-derived stem cells in a child with non-Hodgkin's lymphoma.

Autologous blood-derived stem cells were used for stem-cell rescue in a 5-year-old boy with chemotherapy-resistant B-Non-Hodgkin's lymphoma (B-NHL) involving bone marrow. The high dose chemoradiotherapy was carried out 5 months after initial diagnosis during partial remission. The preparative regimen consisted of 12 Gy fractionated, total-body irradiation (FTBI) before 60 mg/kg etoposide. There were 22.96 X 10(4)/kg body weight myeloid precursor cells, granulocyte-macrophage committed stem cells (CFU-GM) collected by intermittent blood flow separation with a Haemonetics 30R in two cytaphereses and stored in liquid nitrogen. Also 11, 82 X 10(4) CFU-GM/kg body weight were recovered and transfused after thawing. Rapid hematopoietic reconstitution ensued: Erythroid precursors were detected on day 9, 1 X 10(9)/L leucocytes were counted on day 11, and 0.5 X 10(9)/L granulocytes on day 13, respectively. The patient required 3 single-donor platelet transfusions, the last one on day 10. On day 17, 100 X 10(9)/L platelets were reached. A bone marrow aspirate on the same day showed good trilineage regeneration. The patient remained in complete remission 7 months after autografting with a normal stem cell content of the bone marrow and in the peripheral blood. On day 226, after stem cell infusion, a bone marrow relapse occurred.

The median rise of leucocytes predicts the optimal days for leukapheresis in children.

22 consecutive patients with a median age of 11 years (range 3-23) underwent 103 leucaphereses after a chemotherapy induced aplasia. They suffered from various solid tumors and hematological malignancies. The stem cell yield of 69 aphereses which were done during the 8 days following the first platelet rise, was median 1.02 x 10(4) myeloid committed stem cells (CFU-GM)/kg/apheresis (range 0.02-13.3), whereas only 0.27 x 10(4) CFU-GM/kg/apheresis (range 0.01-1.11) were obtained in 34 "random" collections (p less than .05). The CFU-GM yield of 69 "well timed" collections depended on the absolute number of circulating leucocytes (p = .002) and mononuclear cells (p less than .001). The median daily increment of leucocytes was a reliable predictor of the stem cell yield (p = .002) and could be used to select the optimal days for leucaphereses during the well timed period. By careful timing alone high stem cell numbers could be collected in various malignancies without using exogenous growth factors.