Oral citrulline as arginine precursor may be beneficial in sickle cell disease: early phase two results.

L-Arginine may be a conditionally essential amino acid in children and adolescents with sickle cell disease, particularly as required substrate in the arginine-nitric oxide pathway for endogenous nitrovasodilation and vasoprotection. Vasoprotection by arginine is mediated partly by nitric oxide-induced inhibition of endothelial damage and inhibition of adhesion and activation of leukocytes. Activated leukocytes may trigger many of the complications, including vasoocclusive events and intimal hyperplasias. High blood leukocyte counts during steady states in the absence of infection are significant laboratory risk factors for adverse complications. L-Citrulline as precursor amino acid was given orally twice daily in daily doses of approximately 0.1 g/kg in a pilot Phase II clinical trial during steady states in four homozygous sickle cell disease subjects and one sickle cell-hemoglobin C disease patient (ages 10-18). There soon resulted dramatic improvements in symptoms of well-being, raised plasma arginine levels, and reductions in high total leukocyte and high segmented neutrophil counts toward or to within normal limits. Continued L-citrulline supplementation in compliant subjects continued to lessen symptomatology, to maintain plasma arginine concentrations greater than control levels, and to maintain nearly normal total leukocyte and neutrophil counts. Side effects or toxicity from citrulline were not experienced. Oral L-citrulline may portend very useful for palliative therapy in sickle cell disease. Placebo-controlled, long-term trials are now indicated.

Full hematopoietic engraftment after allogeneic bone marrow transplantation without cytoreduction in a child with severe combined immunodeficiency.

Bone marrow transplantation (BMT) for severe combined immunodeficiency (SCID) with human leukocyte antigen (HLA)-identical sibling donors but no pretransplantation cytoreduction results in T-lymphocyte engraftment and correction of immune dysfunction but not in full hematopoietic engraftment. A case of a 17-month-old girl with adenosine deaminase (ADA) deficiency SCID in whom full hematopoietic engraftment developed after BMT from her HLA-identical sister is reported. No myeloablative or immunosuppressive therapy or graft-versus-host disease (GVHD) prophylaxis was given. Mild acute and chronic GVHD developed, her B- and T-cell functions became reconstituted, and she is well almost 11 years after BMT. After BMT, repeated studies demonstrated: (1) Loss of a recipient-specific chromosomal marker in peripheral blood leukocytes (PBLs) and bone marrow, (2) conversion of recipient red blood cell antigens to donor type, (3) conversion of recipient T-cell, B-cell, and granulocyte lineages to donor origin by DNA analysis, and (4) increased ADA activity and metabolic correction in red blood cells and PBLs.

Allogeneic bone marrow transplantation for children with acute leukemia: long-term follow-up of patients prepared with high-dose cytosine arabinoside and fractionated total body irradiation.

High-dose therapy and allogeneic matched sibling bone marrow transplantation (BMT) is considered to be the treatment of choice for children with relapsed acute lymphoblastic leukemia (ALL), or for children with acute myeloid leukemia (AML) in first remission. However, the rate of bone marrow relapse after transplant for either of these diseases remains high. In this study, we assessed the efficacy and toxicity of high-dose cytosine arabinoside and total body irradiation (TBI) followed by allogeneic BMT, for children with acute leukemia or myelodysplastic syndrome (MDS). Sixty-five pediatric patients underwent allogeneic related (n = 57) or unrelated (n = 8) BMT. Twenty-seven were transplanted for ALL in second remission (CR2), and 16 for AML in first remission (CR1). The other 22 were high risk patients: six were transplanted for ALL in third remission (CR3), two for AML in CR2, two for myelodysplastic syndrome (MDS) and 12 for acute leukemia in relapse. Patients were prepared with cytosine arabinoside 3000 mg/m2 per dose twice daily for 6 days followed by 12000 cGy TBI as 200 cGy fractions twice daily for 3 days. Minimum follow-up is 21 months. Five-year event-free survival (EFS) and the actuarial relapse rate is 59 and 14% for patients with ALL in second remission, and 38 and 14+% for patients with AML in first remission. Twelve patients have relapsed (three are alive in remission after testicular or marrow relapse) and 28 have died of other causes. Acute GVHD with or without infection was the cause of death in 11 patients. Ten of the 11 patients who died of acute GVHD were considered at 'high risk' for GVHD (inadequate GVHD prophylaxis, or mismatched family donor or a matched unrelated donor). Toxicities in the immediate post-BMT period included diarrhea, oropharyngeal mucositis and conjunctivitis. Significant late toxicities included short stature, avascular necrosis of bone, and poor school performance (most often in patients who had received prior cranial irradiation). Our conclusions are that high-dose Ara-C and TBI followed by allogeneic bone marrow transplantation is effective therapy for children in second complete remission of their acute leukemia. However, significant late toxicities occur, and it is clear that more effective, less toxic therapies are necessary for these patients.

Bone marrow transplantation for peripheral T-cell lymphoma in children and adolescents.

We report nine children with relapsed (n = 8) or high-risk (n = 1) peripheral T-cell lymphoma (PTCL) who underwent autologous (n = 6) or allogeneic (n = 3) bone marrow transplantation (BMT). These children received transplants as part of a prospective phase I/II study of thioTEPA (TT) and total body irradiation (TBI) with escalating doses of VP-16. The median age of these patients at time of BMT was 6.5 years (range 2.5 years to 14 years). Three were transplanted with active disease after failing salvage chemotherapy. Of the other six, one was transplanted in first complete remission (CR) and five in second or subsequent CR. Of these nine patients, eight are free of disease a median of 25 months after BMT (range, 6 to 48 months), with an estimated 2-year relapse-free survival (RFS) of 89%. Six of these eight patients have been followed for 12 or more months after BMT, and in each their current remission exceeds their longest previous remission duration. The toxicity of the TT/TBI +/- VP-16 regimens was significant but manageable, predominantly consisting of severe mucositis. For a comparison, we reviewed retrospective data on the six additional children and adolescents with PTCL who underwent BMT during the 3-year period preceding this phase I/II study. The median age at BMT of these six patients was 19 years (range 15.5 years to 20 years). These patients were prepared for BMT with a variety of other regimens. One had no response to BMT and the other five relapsed at 1.5 to 5 months after BMT (median, 3 months) with an RFS of 0%. Our data suggest that thioTEPA plus TBI, with or without VP-16, is an effective preparative regimen for BMT for young patients with relapsed or high-stage PTCL and leads to prolonged RFS.

Successful treatment of severe aplastic anemia by bone marrow transplantation from HLA nonidentical family members: preliminary results utilizing cyclophosphamide and 600 cGY fractionated total body irradiation.

We have performed bone marrow transplants on four children with severe aplastic anemia who lacked an human leukocyte antigen (HLA)-identical sibling donor. Patients were prepared with cyclophosphamide and 600 cGy fractionated total body irradiation, and then received marrow from a parent donor mismatched for one (two patients), two (one patient), or three (one patient) HLA antigens. All four patients engrafted. One died early of acute graft-versus-host disease. The three others showed sustained complete hematopoietic reconstitution. Two are alive and hematologically normal 43-87 months after transplant. Both have had acute and chronic graft-versus-host disease (CGVHD), and one of the two remains on immunosuppressive drugs. The fourth died at 48 months after transplant of CGVHD. The previous experience with HLA-incompatible marrow transplants is reviewed, and the rationale for this preparative regimen is discussed. Cyclophosphamide and 600 cGy fractionated total body irradiation is an effective preparative regimen for children with severe aplastic anemia receiving transplants from HLA-nonidentical parental donors, allowing engraftment and full hematologic reconstitution.

Allogeneic transplantation of blood-derived, T cell-depleted hemopoietic stem cells after myeloablative treatment in a patient with acute lymphoblastic leukemia.

This report describes an allogeneic peripheral blood stem cell transplant in a patient who had received marrow ablative therapy. The patient was an 18-year-old white male with acute lymphocytic leukemia in third remission for whom an allogeneic bone marrow transplant was recommended. His HLA-identical sibling preferred to donate peripheral blood stem cells rather than marrow. The donor cells were collected with 10 apheresis procedures and depleted of T lymphocytes to prevent excessive graft-versus-host disease. Nine collections were cryopreserved. The patient received high-dose cytosine arabinoside and 12 Gy of total body irradiation, followed by infusion of all cryopreserved donor cells. A portion of the tenth apheresis product collected on the day of transplant containing 1.8 x 10(9) T lymphocytes was infused without further processing to approximate the number of T lymphocytes given in an allogeneic bone marrow transplant; the remainder was T lymphocyte depleted and infused. More than 1 x 10(9)/l granulocytes were present on day +11. A bone marrow biopsy on day +27 showed trilineage engraftment. Cytogenetic studies demonstrated that the recipient's marrow and peripheral blood were populated exclusively with donor cells. Allogeneic peripheral stem cell transplantation produced an early hematopoietic engraftment. Since the patient died on day +32, sustained engraftment could not be evaluated.

High-dose cytosine arabinoside and fractionated total-body irradiation: an improved preparative regimen for bone marrow transplantation of children with acute lymphoblastic leukemia in remission.

Twenty children with acute lymphoblastic leukemia in second (18 patients) or third (two patients) complete remission after bone marrow relapse received allogeneic bone marrow transplants from histocompatible sibling donors. The preparative regimen for marrow transplantation consisted of 12 doses of 3,000 mg/m2 cytosine arabinoside twice daily for six days followed by 1,200 cGy total-body irradiation (six doses of 200 cGy twice daily for three days). The preparative regimen was well tolerated, and all patients showed marrow engraftment promptly. Twelve patients are alive in complete remission 12+ to 79+ months posttransplant; eight patients are over 48 months posttransplant. Six patients died 1 to 9 months posttransplant of nonleukemic causes: (two each of graft-v-host disease, interstitial pneumonitis, and infection). Two patients developed recurrent leukemia at 15 and 30 months posttransplant. Both have died at 19 and 36 months posttransplant. Life table analysis reveals an actuarial survival and event-free survival rate of 58% and a marrow relapse rate of 17%. These results suggest that high-dose cytosine arabinoside and fractionated total-body irradiation is a relatively nontoxic and highly effective preparative regimen for allogeneic bone marrow transplantation for acute lymphoblastic leukemia that deserves further evaluation.

Barrett's esophagus in three children after antileukemia chemotherapy.

Barrett's esophagus, a columnar metaplasia of the lower esophagus that is usually associated with gastroesophageal reflux (GER), was found in three children on long-term antileukemia chemotherapy. Two of the children had been on a standard acute lymphoblastic leukemia (ALL) maintenance protocol with 2 to 3 years of methotrexate and 6-mercaptopurine administration. The third child received daunorubicin, cytosine arabinoside, and 6-thioguanine for treatment of acute myelogenous leukemia (AML). None of the patients had clinical or pathologic evidence of GER disease. We propose that the Barrett's esophagus in these patients did not result from the usual peptic esophagitis, but rather from chemotherapy-induced esophageal mucosal injury.

Differentiation of presumed sepsis from acute graft-versus-host disease by C-reactive protein and serum total IgE in bone marrow transplant recipients.

Fever after bone marrow transplantation may indicate the onset of bacterial or opportunistic infection, or acute graft-versus-host disease (GVHD). In an attempt to differentiate between infection and GVHD, we prospectively studied 41 bone marrow transplants in 38 patients (24 allogeneic, 17 autologous). Elevation of C-reactive protein (CRP) proved to be a good indicator of disseminated infections. In 40 episodes of documented (11) or presumed (29) sepsis, CRP rose above 5 mg/dl in 38 episodes (95%), and above 10 mg/dl in 32 episodes (80%). The CRP concentration paralleled the clinical course of the infectious episodes. Elevated CRP values were not observed in the 15 episodes of acute GVHD without concurrent infection. High peak values of serum total IgE, ranging from 4-fold to over 4000-fold baseline, were observed posttransplant in 18/22 allogeneic BMT recipients, temporally associated with activation of acute GVHD. IgE was elevated neither in episodes of sepsis without concurrent GVHD, nor in viral or focal bacterial infections. In general, septic infections were characterized by high CRP but low IgE levels. Acute GVHD without concurrent infection was characterized by high IgE but low CRP. We conclude that CRP and serum total IgE utilized together in serial fashion are helpful in distinguishing sepsis from acute GVHD.

Ganglioside GD2 specific monoclonal antibody 3F8: a phase I study in patients with neuroblastoma and malignant melanoma.

The murine IgG3 monoclonal antibody (MoAb) 3F8, specific for the ganglioside GD2, activates human complement, is active in antibody-dependent cell-mediated cytotoxicity (ADCC), and can target specifically to human neuroblastoma in patients with metastatic disease. In a phase I study, 3F8 was administered intravenously (IV) to 17 patients with metastatic GD2 positive neuroblastoma or malignant melanoma at doses of 5, 20, 50, and 100 mg/m2. Serum 3F8 levels achieved were proportional to the dose of 3F8 infused. However, serum antimouse antibody levels did not increase with the amount of 3F8 administered. Toxicities included pain, hypertension, urticaria, and complement depletion. All acute side effects were controllable with symptomatic therapy. No long-term side effects were detected in patients observed for more than 14 months. None of the 17 patients received any antitumor therapy postantibody treatment. Antitumor responses occurred in seven of 17 patients. These ranged from complete clinical remissions to mixed responses. The murine monoclonal antibody (MoAb) 3F8 has clinical utility for the diagnosis and therapy of neuroblastoma and melanoma.

Hemophilic arthropathy: assessment with MR imaging.

Fifteen patients with hemophilia, 14 of whom had hemophilic arthropathy, were examined with magnetic resonance (MR) imaging to determine if it could be used to assess hemophilic arthropathy, especially synovial hypertrophy and the status of the articular cartilage. Thirty-five joints of the appendicular skeleton were imaged. Four joints in two patients were clinically normal. Synovial hypertrophy was detected in 28 joints and appeared as areas of low to intermediate signal intensity on T1- and T2-weighted images, with foci of increased signal intensity on T2-weighted images (presumed to be due to areas of fluid or inflammation) in 16 joints. Abnormal articular cartilage was demonstrated in 26 joints; bone lesions, fluid collections, and joint space narrowing could also be seen. MR imaging appears to be useful in depicting the components of hemophilic arthropathy.

Neutrophil elastase produces 52-kD and 30-kD glucocorticoid receptor fragments in the cytosol of human leukemia cells.

Characterization of glucocorticoid receptors in leukemia cells is important to understand mechanisms of glucocorticoid resistance but has been impeded by receptor fragmentation in cytosol extracts. We recently found that formation of 52- and 30-kilodalton (kD) glucocorticoid receptor fragments in cytosol of leukemia cells is due to proteolysis and is blocked by diisopropylfluorophosphate (DFP). In the present study, we identify a 28-kD serine protease in cytosol of leukemia cells that binds [3H]DFP and correlates with the formation of 52- and 30-kD receptor fragments. This protease is immunoprecipitated by antiserum to neutrophil elastase. Limited digestion of [3H]dexamethasone-21-mesylate-labeled receptors by purified neutrophil elastase produces 52- and 30-kD receptor fragments. Receptor fragmentation in the cytosol of leukemia cells in inhibited by methoxysuccinyl-alanyl-alanyl-prolyl-valyl-chloromethylketone, a highly specific inhibitor of neutrophil elastase. The addition of as few as 5% neutrophils to a lymphoid cell suspension provides sufficient elastase to produce receptor fragmentation. Our findings indicate that neutrophil elastase is responsible for receptor fragmentation in the cytosol of leukemia cells. The neutrophil elastase may be endogenous to the leukemia cells or may come from neutrophils that contaminate leukemia cell suspensions.

Diagnostic imaging of human neuroblastoma with radiolabeled antibody.

In a previous study, the authors showed that iodine-131 labeled monoclonal antibody (Mab 3F8) could be used to image human neuroblastoma xenografts in mice with excellent tumor-to-tissue ratios. In this study they report their experience with six patients scanned with radiolabeled 3F8. There was strong accumulation of the labeled antibody in viable tumor, but no significant uptake was noted in normal brain, liver, spleen, or adrenal glands. Tumor-to-nontumor activity ratios varied but were approximately 10:1-20:1. This ratio yields good contrast for visualization. Time-activity curves show that radioactivity levels in normal tissue have a half-time of about 40 hours, whereas tumor tissues show a half-time of about 60 hours. Significant gastric secretion of free iodine demonstrated that the Mab was being deiodinated. Calculated radiation doses indicate that tumors receive at least ten times the dose to other tissues. The results indicate that Mab 3F8 has clinical potential for both imaging and therapy of human neuroblastomas.

Detection of neuroblastoma cells in bone marrow using GD2 specific monoclonal antibodies.

Three murine monoclonal antibodies (Mab) against a cell surface antigen, the disialoganglioside GD2, were investigated for detecting bone marrow metastasis in patients with neuroblastoma (NB) by indirect immunofluorescence (IF). As few as 0.01% NB cells could be detected. No Mab reactivity was found in 60 non-NB marrows. Thirty-five marrows from patients with stage III and stage IV NB at diagnosis were examined during the course of their disease. Tumor involvement was found in 74% by Mab, 55% by the two-layer soft agar clonogenic assay (CA), and 27% by conventional histochemical stains. All marrows containing NB histologically were positive for tumor by Mab; 71% were also positive by CA. Of histologically negative marrows, 63% were Mab positive, the majority (78%) of which were also positive by CA. All Mab nonreactive samples were negative histologically and by CA. We conclude that these antibodies are highly sensitive and specific in detecting NB metastasis in bone marrow.

Abdominal neuroblastoma: magnetic resonance imaging and tissue characterization.

Magnetic resonance imaging was performed in eight children with abdominal neuroblastomas. Five patients had serial examinations after diagnosis, and 15 computed tomography (CT) scans were available for comparison. MR imaging was as precise as CT in detecting the presence or absence of liver involvement and more efficient in determining the relationship of tumor to vascular structures. However, it did not differentiate tumor from normal kidney as accurately as CT. Inversion recovery (IR) images yielded the greatest soft-tissue contrast resolution between liver and tumor. T1 values of tumor were much higher than those of normal liver but overlapped those of normal kidneys. T2 values of tumor were significantly higher than those of the liver and were slightly, but not significantly, lower than those of the kidneys. Either T1- or T2- weighted pulse sequences should, therefore, provide adequate liver-tumor differentiation, but more heavily T2- weighted images appear necessary to distinguish between tumor and kidney. T1 values of tumors usually decreased with rapid tumor regression, while T2 values changed independently. The ability to quantitate tissue parameters helps in selecting appropriate imaging sequences and may be of use in following the progress of tumors.