HLA-matched Bone Marrow Transplantation Research Articles

Encephalopathy After Bone Marrow Transplantation

Encephalopathy After Bone Marrow Transplantation

Survival Difference of AML Patients Receiving HLA Matched Sibling Allogeneic Bone Marrow Transplants (ABMT) Correlates with HLA-Cw Ligand Groups for Killer Immunoglobulin-Like Receptors (KIRs).

The reactivity of NK cells and some T cell populations is regulated by the interaction of KIRs with target cell HLA class I molecules. KIR interactions have been suggested to influence outcomes of haploidentical and HLA-identical allogeneic hematopoietic stem cell transplants, particularly for AML patients. We analyzed the KIR ligand phenotypes of 60 AML patients who received HLA-identical sibling myeloablative ABMT from 4/9/97-11/5/03. The median age was 45 yrs (range 8–62). At the time of transplant 24 patients (40%) were in CR. All patients received a busulfan/cyclophosphamide-based preparative regimen and all received bone marrow (T-cell replete) as their stem cell source. Patient HLA KIR ligands were categorized as: 1) HLA-Cw groups C1/C1, C2/C2, C1/C2; 2) HLA-Bw4 (+ or −); 3) HLA-A3 or A11 (+ or −) (as reviewed in Farag et al Blood 100:1935, 2002). Recursive partitioning analysis for post-transplant time-related outcomes (acute GVHD, grade 3/4 acute GVHD, chronic GVHD, extensive chronic GVHD, freedom from relapse and survival) was performed for each KIR ligand group. Patients with C1/C1 or C2/C2 (n=26) had improved survival compared to the C1/C2 group (n=34) (median survival 43.5 months vs. 5.8 months, respectively, p=0.018), as shown below: [Display omitted] This survival difference was associated with more relapse in the C1/C2 group (p=0.048), but not with incidence or severity of acute or chronic GVHD, age, infection or pretransplant disease status. There were 26/34 (76%) deaths in the C1/C2 group with 15 (58%) due to relapse as compared to 13/26 (50%) deaths in the C1/C1 + C2/C2 group where 4/13 (31%) were due to relapse. The median follow up of survivors was 36.3 mos (range 7.8–72.4 mos). No significant differences in outcomes were observed when patients were analyzed for the presence or absence of HLA-Bw4 or A3/11. The majority of patients had KIR genotyping performed for those KIRs with established HLA ligands. Among those tested there were no cases in which the donor did not have at least one inhibitory KIR gene specific for a Cw ligand present in the patient or donor. This may suggest that KIR expression at the cellular level rather than KIR genotype alone should be investigated. In conclusion, AML patients undergoing matched sibling donor ABMT who were heterozygous for HLA-Cw KIR ligand groups (C1/C2) had reduced survival compared to patients homozygous for these groups. The higher relapse rate observed in the heterozygous ligand group may suggest a less effective graft-versus-leukemia (GVL) response. Since C1/C2 heterozygotes have a greater opportunity to engage inhibitory KIRs than do C1 or C2 homozygotes, they may more effectively inhibit KIR-positive NK and T cell populations involved in GVL responses.

Quinine as a multidrug resistance inhibitor: a phase 3 multicentric randomized study in adult de novo acute myelogenous leukemia

AbstractBased on our previous demonstration that quinine could be used clinically to reverse P-glycoprotein–mediated resistance, we designed a multicenter, randomized trial aiming to determine whether quinine would improve the survival of adult patients (15-60 years old) with de novo acute myelogenous leukemia (AML). These patients randomly received (n = 213) or did not receive (n = 212) a 30 mg/kg/day continuous intravenous infusion of quinine in combination with induction chemotherapy combining idarubicine and cytarabine and, depending on bone marrow examination at day 20, an additional course of cytarabine and mitoxantrone. The mean steady-state quinine concentration was 7.8 mg/L and the mean multidrug resistance reversing activity of serum was 1.96. Complete remission (CR) was obtained in 344 patients (80.9%) without significant influence of quinine. Of the patients in complete remission, 82 were assigned to receive HLA-matched bone marrow transplants, whereas 262 were assigned to 2 courses of intensive consolidation chemotherapy, with or without quinine, depending on initial randomization. The 4-year actuarial overall survival (OS) of the 425 eligible patients was 42.0% ± 2.5%, without significant influence of quinine. Of 160 patients who could be studied, 54 demonstrated rhodamine 123 efflux. In these patients, quinine significantly improved the CR rate from 12 of 25 (48.0%) to 24 of 29 (82.8%) (P= .01). However, there was no significant difference in OS. Neithermdr1gene nor P-glycoprotein expression influenced the outcome. We conclude that quinine does not improve the survival of adult patients with de novo AML, even though it improves CR rate in a small subgroup of patients defined by rhodamine 123 efflux.

Impact of prophylactic donor leukocyte infusions on mixed chimerism, graft-versus-host disease, and antitumor response in patients with advanced hematologic malignancies treated with nonmyeloablative conditioning and allogeneic bone marrow transplantation

In an attempt to capture graft-versus-tumor effects without graft-versus-host disease (GVHD), the authors initiated a trial of nonmyeloablative allogeneic bone marrow transplantation (BMT) in patients with advanced hematologic malignancies, with the majority of patients having chemotherapy-refractory disease. Forty-two patients received an HLA-matched related donor BMT after a cyclophosphamide and antithymocyte globulin-based conditioning that also included thymic irradiation for patients who had not received prior mediastinal radiotherapy. Prophylactic donor leukocyte infusion (pDLI) at a dose of 1 × 10 7 CD3 + cells per kilogram were given beginning 5 weeks post-BMT to 16 patients with mixed chimerism (MC) but without GVHD, whereas 26 patients did not receive pDLI, either because of GVHD or early relapse. Twelve of 16 patients (75%) receiving pDLI had T cell chimerism at the time of pDLI ≥40%. These patients, by day 100 post-BMT, either converted to full donor chimerism (FDC) (n = 10) or had an increase in or stable donor chimerism (n = 2) after pDLI. Four of 4 patients whose T cell chimerism was ≤20% at the time of pDLI, lost the graft. In contrast, only 5 of 18 evaluable patients (28%) not receiving a pDLI converted to FDC by day 100 post-BMT, 7 maintained MC, and 10 of an evaluable 22 lost the graft. Patients who had undergone a previous autologous stem cell transplant had a higher rate of conversion to FDC (69% v 31%) and higher incidence of GVHD (69% v 34%) compared with those who did not have a previous autologous SCT. Eleven of 16 patients (69%) who received a pDLI achieved a remission with 50% 1-year progression-free survival rate and 44% 3-year overall survival rate. Nineteen of 42 patients (45%) had ≥grade II acute GVHD, including 12 after BMT and 7 after pDLI. Approximately one third of patients, after having initial MC, eventually lost their donor graft. The authors conclude that (1) pDLI has the potential to convert MC to FDC; (2) sustained remissions can be achieved in patients with chemorefractory hematologic malignancies who receive a pDLI, albeit with a significant risk of acute GVHD; and (3) the degree of donor T cell chimerism at the time of pDLI is predictive of the fate of MC, ie, donor T cell chimerism ≥40% or ≤20% at the time of pDLI correlates with conversion of MC or loss of the graft, respectively.

Prospective Trial of Chemotherapy and Donor Leukocyte Infusions for Relapse of Advanced Myeloid Malignancies After Allogeneic Stem-Cell Transplantation

Patients with advanced myeloid malignancies who experience relapse after allogeneic bone marrow transplantation (BMT) have a poor prognosis. Long-term survival after chemotherapy alone, second myeloablative transplant, or donor leukocyte infusions (DLIs) alone is unusual. DLIs may have minimal effectiveness in advanced disease because adequate cellular responses are not able to develop in the presence of bulky, fast-growing disease. A chemotherapy strategy was used to debulk disease before administration of granulocyte colony-stimulating factor (G-CSF)-primed DLIs. Sixty-five patients experiencing hematologic relapse of myeloid malignancy after HLA-matched sibling BMT were prospectively treated with cytarabine-based chemotherapy, then G-CSF-primed DLIs. No prophylactic immunosuppression was provided. Twenty-seven of 57 assessable patients experienced a complete response. Graft-versus-host disease (GVHD) was observed in 56% of the patients. Treatment-related mortality was 23%. Overall survival at 2 years for the entire cohort was 19%. Patients with a complete response were more likely to survive, with 1- and 2-year survival rates of 51% and 41%, respectively, with a median follow-up of more than 2 years. The 1-year survival for nonresponders was 5%. A posttransplant remission lasting more than 6 months before relapse was associated with a higher likelihood of response. GVHD was not required for durable remission. Salvage treatment with chemotherapy before DLI can help some patients with advanced myeloid relapse and is not dependent on GVHD. Patients with short remissions after BMT are unlikely to benefit from this approach, and the approach is associated with significant treatment-related mortality. Modifications of this approach or entirely different approaches will be required for most patients with this difficult clinical problem.

Prospective Trial of Chemotherapy and Donor Leukocyte Infusions for Relapse of Advanced Myeloid Malignancies After Allogeneic Stem-Cell Transplantation

PURPOSE: Patients with advanced myeloid malignancies who experience relapse after allogeneic bone marrow transplantation (BMT) have a poor prognosis. Long-term survival after chemotherapy alone, second myeloablative transplant, or donor leukocyte infusions (DLIs) alone is unusual. DLIs may have minimal effectiveness in advanced disease because adequate cellular responses are not able to develop in the presence of bulky, fast-growing disease. A chemotherapy strategy was used to debulk disease before administration of granulocyte colony-stimulating factor (G-CSF)–primed DLIs. PATIENTS AND METHODS: Sixty-five patients experiencing hematologic relapse of myeloid malignancy after HLA-matched sibling BMT were prospectively treated with cytarabine-based chemotherapy, then G-CSF–primed DLIs. No prophylactic immunosuppression was provided. RESULTS: Twenty-seven of 57 assessable patients experienced a complete response. Graft-versus-host disease (GVHD) was observed in 56% of the patients. Treatment-related mortalit...

Pulmonary Function Changes in Long-Term Survivors of Chronic Myelogenous Leukemia After Allogeneic Bone Marrow Transplantation: A Taiwan Experience

Pulmonary function in 42 patients with chronic myelogenous leukemia (CML) was tested before and after HLA-matched (39 related, 3 unrelated) allogeneic bone marrow transplantation (BMT) between 1985 and 1999. Pulmonary function tests (PFTs) including ventilatory capacity, lung volumes, and diffusion capacity for carbon monoxide (DLCO) were performed before and 3, 6, 12, and 24 months after BMT, and every 12 months thereafter. Possible pre- and post-BMT risk factors were evaluated for their influence on pulmonary function. Patients were divided into two groups according to their survival duration for more than 12 months or not.Pretransplant PFTs were essentially normal except for mild reduction in DLCO values in the short-term survival group. Overall pulmonary function changes revealed persistent and significant decrease of forced vital capacity (FVC) and DLCO values after BMT. The DLCO values reached abnormal levels (<80%) and showed a trend of incomplete recovery. Decrease of forced expiratory volume in the first second (FEV1) and vital capacity were also noted but the FEV1/FVC ratio remained within normal limits after BMT. Transient fall of total lung capacity after BMT was noted. However, its values did not reach abnormal levels such as to cause restrictive ventilatory impairment. Possible risk factors including gender, smoking, bronchiolitis obliterans, acute and chronic graft-versus-host disease (GVHD) were found to have significant influences on posttransplant pulmonary function changes by multiple regression analysis. Most patients except those who developed bronchiolitis obliterans were clinically asymptomatic. Development of bronchiolitis obliterans was the most important factor to cause both clinical symptoms and impaired pulmonary function.In summary, pulmonary function changes before and after HLA-matched allogeneic BMT in long-term survivors of CML only showed modest dysfunction. The primary negative presentation with the development of oxygenation defect had no clinical significance in most patients. The influences on the impairment of pulmonary function were multifactorial.

BK VIRUS-ASSOCIATED FATAL RENAL FAILURE FOLLOWING LATE-ONSET HEMORRHAGIC CYSTITIS IN AN UNRELATED BONE MARROW TRANSPLANTATION

The human polyomavirus BK (BKV)-associated hemorrhagic cystitis (HC) has been a frequent and seldom life-threatening complication after bone marrow transplantation (BMT). The authors report a male with myelodysplastic syndrome, who developed BKV-associated late-onset HC 12 days after HLA-matched unrelated BMT. His urine contained epithelial cells with intranuclear inclusion bodies suggestive of BKV infection and was positive for BKV in polymerase chain reaction. He did not respond to any treatment for HC. In addition, he developed BKV-associated acute renal failure on day 26, followed by hepatic veno-occlusive disease on day 42. This is the first case in which BKV may be associated with fatal progressive renal failure.

Minor H antigens: genes and peptides.

In this review, we describe the evidence from which the existence of non-MHC histocompatibility (H) antigens was deduced, the clinical setting of bone marrow transplantation in which they are important targets for T-cell responses, and the current understanding of their molecular identity. We list the peptide epitopes of the human and murine minor H antigens now identified at the molecular level, their MHC restriction molecules and the genes encoding them. Identification of the peptide epitopes allows T-cell responses to these antigens following transplantation of MHC-matched, minor H-mismatched tissues to be enumerated using tetramers and elispot assays. This will facilitate analysis of correlations with host-versus-graft (HVG), graft-versus-host (GVH) and graft-versus-leukaemia (GVL) reactions in vivo. The potential to use minor H peptides to modulate in vivo responses to minor H antigens is discussed. Factors controlling immunodominance of T-cell responses to one or a few of many potential minor H antigens remain to be elucidated but are important for making predictions of in vivo HVG, GVH and GVL responses and tailoring therapy after HLA-matched bone marrow transplantation and donor lymphocyte infusion.

ARE THE FUNCTIONAL ASSAYS USEFUL IN PREDICTING GRAFT-VERSUS-HOST DISEASE IN HLA-MATCHED SIBLING BONE MARROW TRANSPLANTATION?

A reply to the letter from Affaticati et al. There is no doubt that the frequency analysis of helper T lymphocyte precursors (HTLp) and cytotoxic T lymphocyte precursors (CTLp) has great impact on the detection of alloreactivity. However, the debate on the clinical values of these functional assays for the prediction of graft-versus-host disease (GVHD) occurrence after bone marrow transplantation (BMT) has carried on for the last decade. One can easily find several reports in the literature either for (1–4) or against (5–9) the usefulness of these functional assays for predicting GVHD in HLA-matched sibling BMT. Although our observation (9) showed no significant correlation between the HTLp/CTLp frequencies and the incidence of acute GVHD after HLA-matched sibling BMT, there are other reports showing the contrary (5,7). Due to the lack of standard protocols for these functional assays, there are considerable variations in the methodology used by different investigators, which makes it difficult, sometimes impossible, to compare or correlate data from different research groups. For a rigorous examination on the association between the frequencies of HTLp/CTLp and the incidence of GVHD, it is crucial to establish standard protocols and to study a sufficient number of transplant recipients with homogeneity of disease and uniformity of conditioning and GVHD prophylaxis. The effectiveness of the immunosuppressive regimen after transplantation and the precision of the HLA matching also need to be taken into consideration. The discordance in one or more of these factors could be attributable to the controversy regarding the usefulness of HTLp/CTLp frequency estimation for GVHD prediction. During the last few years, the sensitivity of these assays has been improved by eliminating antigen-presenting cells from the responder population and/or T cells from the stimulator population (10,11). However, the complexity of the assay procedure makes these protocols impracticable for routine clinical use, and researchers are still searching for a better method. The combined limiting dilution assay (LDA) used in our study was established by Bouma (12) et al. and has proven to be as sensitive as single LDA to detect alloreactive HTLp and CTLp frequencies in HLA-mismatched settings. Our observation (9) has demonstrated that this combined LDA may not be ideal in HLA-identical sibling settings. The possible reasons have been discussed in detail in our previous articles (8,9). Although Affaticati et al. (13), using a single LDA, showed a positive correlation of CTLp frequency with clinical acute GVHD grades II–IV, the statistical significance remained marginal (P =0.04), with a third of patients who did not develop severe GVHD demonstrating high frequencies. It may well be that the positive CTLp frequencies detected in both publications by Affaticati et al. and ourselves were against minor histocompatibility antigens. Some of these responses detected in cohorts of patients developing no or low-grade GVHD may, however, not be relevant or strong enough in the in vivo clinical setting (in the presence of prophylaxis regimens for example) to give rise to GVHD. However, in our view, CTLp frequency estimation remains an inconclusive predictive indicator of GVHD occurrence and severity in the HLA-matched sibling setting, mainly due to overlapping results of high frequency within cohorts of patients demonstrating either no or low-grade GVHD (grades 0–1). Anne Dickinson Xiao-Nong Wang

Survival after transplantation of unrelated donor umbilical cord blood is comparable to that of human leukocyte antigen–matched unrelated donor bone marrow: results of a matched-pair analysis

AbstractUmbilical cord blood (UCB) is being increasingly used for hematopoietic stem cell transplantation and has been associated with a reduced incidence of severe graft-versus-host disease (GVHD). To further investigate the relative merits of unrelated donor UCB versus bone marrow (BM), a matched-pair analysis comparing the outcomes of recipients of 0 to 3 human leukocyte antigen (HLA)–mismatched UCB and HLA-A, B, DRB1-matched BM was performed. UCB patients, who received cyclosporine (CSA) and methylprednisolone (MP), were matched for age, diagnosis, and disease stage with BM patients, who received either methotrexate (MTX) and CSA (26 pairs) or T-cell depletion (TCD) and CSA/MP (31 pairs). Patients were predominantly children (median age, 5 years) undergoing transplantation for malignancy, storage diseases, BM failure, and immunodeficiency syndromes between 1991 and 1999. Although neutrophil recovery was significantly slower after UCB transplantation, the probability of donor-derived engraftment at day 45 was 88% in UCB versus 96% in BM-MTX recipients (P = .41) and 85% in UCB versus 90% in BM-TCD recipients (P = .32), respectively. Platelet recovery was similar in UCB versus BM pairs. Furthermore, incidences of acute and chronic GVHD were similar in UCB and BM recipients, with 53% of UCB versus 41% of BM-MTX recipients alive (P = .40) and 52% of UCB versus 56% of BM-TCD recipients alive at 2 years (P > .80), respectively. These data suggest that despite increased HLA disparity, probabilities of engraftment, GVHD, and survival after UCB transplantation are comparable to those observed after HLA-matched BM transplantation. Therefore, UCB should be considered an acceptable alternative to HLA-matched BM for pediatric patients.

Procalcitonin does not discriminate infection from inflammation after allogeneic bone marrow transplantation.

Procalcitonin (PCT) is an early marker of bacterial infection but little is known about its value in neutropenic allogeneic bone marrow transplant (BMT) recipients. We collected plasma from 12 recipients of T-cell-depleted HLA-matched related BMT recipients who had been treated preemptively with meropenem from the day after BMT for at least 15 days. PCT and C-reactive protein (CRP) concentrations were determined on BMT days 1, 5, 8, 12, and 15, and their relationship to inflammatory events (IE), including mucositis, microbiologically and clinically defined infections, acute graft-versus-host disease (GVDH), and unexplained fever, was then determined. The PCT concentrations were all low and never exceeded 4 microg/liter, unlike CRP concentrations, which spanned the full range up to 350 mg/liter. All patients had mucositis, and there was no significant difference between PCT concentrations associated with mucositis alone and those associated with an additional IE on BMT days 1 to 12. However, on BMT day 15, the mean concentrations of PCT were 0.37 +/- 0.05 microg/liter for the 10 patients that had an additional IE, compared with 0.11 +/- 0.03 microg/liter for the 2 patients with mucositis only (P = 0.012), and GVHD rather than infection was involved in six cases. PCT was also not a sensitive marker of gram-positive bacteremia or pulmonary aspergillosis. Thus, PCT is of little value in discriminating infections from other inflammatory complications that occur following allogeneic BMT.

T-cell frequency analysis does not predict the incidence of graft-versus-host disease in HLA-matched sibling bone marrow transplantation.

Graft-versus-host disease (GVHD) is a major and sometimes fatal complication of allogeneic bone marrow transplantation (BMT). The prediction of GVHD remains an important issue in preventing morbidity and mortality after allogeneic BMT. In the past 10 years, there has been great interest in using the frequency analysis of alloreactive helper and cytotoxic T lymphocyte precursors (HTLp and CTLp) to detect recipient-specific alloreactivity and thus predict GVHD in HLA-matched related and unrelated BMT. However, the results remain controversial. The intention of the present study was to investigate whether alloreactive HTLp and CTLp frequencies measured in donor peripheral blood before BMT would be a useful predictor for the occurrence of acute GVHD after HLA-matched sibling BMT. A combined limiting dilution assay was used to determine alloreactive HTLp and CTLp frequencies for 42 HLA-matched sibling patient/donor pairs. The pretransplantation host-reactive HTLp and CTLp frequencies were then correlated with post-transplantation clinical outcomes of acute GVHD. The association between HTLp/CTLp frequencies and the incidence of acute GVHD was determined using the Fisher's exact test. The mean values of HTLp and CTLp frequencies for this cohort of HLA-matched sibling patient/donor pairs were 1:321,322 (range 1:71,000 to 1:1000,000) and 1:195,260 (range 1:3,717 to 1:1000,000), respectively. Acute GVHD (> or =II) was observed in one of four patients with high (>1:100,000) HTLp frequencies and 20 of 36 patients with low (<1:100,000) HTLp frequencies. Similarly, 6 of 10 patients with high (>1: 100,000) CTLp frequencies and 14 of 29 patients with low (<1:100,000) CTLp frequencies developed acute GVHD (> or =II). The overall correlation between hostreactive HTLp/CTLp frequencies and the incidence of acute GVHD in this cohort of patients was 42.5% and 53.9%, respectively. There was no significant difference in the incidence of acute GVHD between the patients with either high or low host-reactive HTLp/ CTLp frequencies (P=0.331 and 0.716, respectively). The data were also analyzed separately for the adult patient group based on GVHD prophylaxis with either cyclosporine alone or the combination of cyclosporine and methotrexate. Within these two prophylaxis groups, neither HTLp nor CTLp frequencies correlated with acute GVHD. Host-reactive HTLp and CTLp frequency analysis did not provide informative prediction for the occurrence of acute GVHD after HLA-matched sibling BMT.

Changes in the periodontal status of patients undergoing bone marrow transplantation.

Patients receiving an HLA-matched bone marrow transplant (BMT) from a relative or unrelated donor undergo a permanent alteration of their immune system, followed by a prolonged period of immunodeficiency. This study aimed to examine alterations in the periodontal status of patients over 6 months post-bone marrow transplantation. Thirty-seven patients scheduled for bone marrow transplantation participated in this study. One calibrated examiner carried out periodontal examinations (clinical and radiographic) immediately prior to and at 3 and 6 months after transplantation. All patients followed an intense oral care program. Subgingival plaque samples were analyzed by enzyme-linked immunosorbent assay (ELISA) for the presence of Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans, and Prevotella intermedia. Data were subjected to statistical analyses to determine the relationships between the frequency distribution of the radiographic and clinical variables over time. Gains in clinical attachment level (CAL) of > or =2 mm at 4 or more sites from baseline to 6 months post-BMT were noted in 9/16 patients (56%), while 6/16 (38%) patients experienced a loss of CAL > or =2 mm at 4 or more sites in the same period. At a site level, 4.8% of sites exhibited a gain in CAL > or =2 mm between baseline and 3 months post-BMT while 2.3% of sites showed a loss of CAL > or =2 mm in the same period. From baseline to 6 months, a gain in CAL of > or =2 mm was recorded at 3.1% of sites, and 2.4% of sites experienced a loss of > or =2 mm. A significant improvement in the gingival index occurred between all sequential time periods when assessed at a site level. At a patient level, 11/18 (61%) patients showed a significant change in gingival index between baseline and 3 months and 10/16 (63%) between baseline and 6 months. There was no significant relationship between clinical changes and the prevalence of the periodontal pathogens at the various time periods. An improvement in periodontal health was recorded between baseline and 6 months post-transplantation. Most of the improvement in periodontal status was noted in the first 3 months after BMT, with a slight decline in periodontal health between 3 and 6 months post-transplant. No significant alteration was noted in the prevalence of periodontal pathogens during the study period.

SOMNOLENCE SYNDROME IN A CHILD FOLLOWING 1200-cGy TOTAL BODY IRRADIATION IN AN UNRELATED BONE MARROW TRANSPLANTATION

Neurological complications may occur following intensive chemotherapy and hematopoietic cell transplantation. Postirradiation somnolence syndrome has been observed in children with acute lymphoblastic leukemia who received central nervous system preventive therapy with 1800-2400 cGy cranial irradiation. The authors report a 16-year-old boy with chronic myelogenous leukemia in chronic phase, who developed symptoms compatible with thes omnolence syndrome (SS) 6 weeks following HLA-matched unrelated bone marrow transplantation (BMT). The preparative regimen consisted of 1200 cGy total body irradiation (TBI), cytosine arabinoside and cyclophosphamide. The patient developed lethargy and low-grade fever, with intermittent rhythmical delta activity in electroencephalograph. Here covered spontaneously without specific therapy 3 weeks after developing symptoms. This is the first report describing that as low as 1200 cGy TBI can induce SS in a child. After allogeneic BMT, some patients develop neurological symptoms. The authors suggest that somnolence syndrome should be included in differential diagnosis in these patients.

Use of a skin explant model for predicting GVHD in HLA-matched bone marrow transplants - effect of GVHD prophylaxis.

Over the last decade we have successfully evaluated the use of a human skin explant assay for predicting acute GVHD in HLA-matched sibling transplants. In the present study, we modified GVHD prophylaxis on an individual patient basis depending on the GVHD outcome predicted by the skin explant model. We have summarised our previous data describing how the skin explant assay results correctly predict GVHD occurrence and severity in 45/56 patients (80%); P< 0.0001, chi2 19.97, df = 1. In a further cohort of 19 patients, all were predicted to develop grade II or above GVHD. These patients were given increased GVHD prophylaxis with the addition of methotrexate and a significant reduction in the expected incidence of GVHD was observed (P = 0.02; chi2 7.7, df = 1; Fisher exact test P = 0.04). The results from these studies suggest that modifying GVHD prophylaxis, based on skin explant assay results, may reduce the expected incidence and severity of GVHD. We suggest that the technique might be used for selective GVHD prophylaxis in T cell non-depleted HLA matched sibling transplants.

Bone Marrow Transplantation in a Child with Hemophagocytic Lymphohistiocytosis Using a Less Toxic Conditioning Regimen

Background Hemophagocytic lymphohistiocytosis (HLH) is a rare non-neoplastic, frequently fatal disease of childhood. HLA-matched bone marrow transplantation (BMT) can bring about long-term remission and an eventual cure. Methods We report on the beneficial effect of BMT in a 2-month-old male using a less intensive conditioning regimen. The regimen included busulfan at 4 mg/kg/day (total dose 16 mg/kg), etoposide at 300 mg/m 2/day (total dose 900 mg/m 2), and cyclophosphamide at 50 mg/kg/day (total dose 150 mg/kg). Prophylaxis for graft-vs.-host disease included methotrexate and cyclosporine. Results An absolute neutrophil count of 500 μL was noticed on +day 12 (engraftment day). At present, i.e., 400 days after the procedure, the patient is asymptomatic, his physical examination is normal, and a slightly increased level of gamma-glutamyl-transpeptidase (GGT) and alkaline phosphatase are the only laboratory abnormalities. Conclusions In this case, the conditioning regimen was adequate for the eradication of the disease and allowed persistent engraftment without significant toxicity. The results in our patient suggest that a less toxic regimen is feasible and permits rapid engraftment without compromising the effectiveness of chemotherapy.

Retrovirus-mediated WASP gene transfer corrects defective actin polymerization in B cell lines from Wiskott-Aldrich syndrome patients carrying 'null' mutations.

Boys affected with Wiskott-Aldrich syndrome (WAS) present with variable association of thrombocytopenia, eczema and immune deficiency. If untreated, WAS patients may succumb to intracerebral hemorrhages, severe infections or malignancies. Allogeneic bone marrow transplantation (BMT) can cure all aspects of the disease, but HLA-identical donors are not available to all patients and mismatched BMTs are unfortunately associated with high mortality and morbidity. The good success of HLA-matched BMT, however, makes WAS a potential candidate for hematopoietic stem cell gene therapy. WAS patients carry mutations of the Wiskott-Aldrich syndrome protein gene encoding WASP, a 502-amino acid proline-rich protein with demonstrated involvement in the organization of the actin cytoskeleton. To verify the feasibility of genetic correction for this disease, the WASP cDNA was expressed in EBV-immortalized B cell lines obtained from WAS patients using a retroviral vector. Transduced WAS cells showed levels of WASP expression similar to those found in cells from normal donors, without detectable effects on viability or growth characteristics. In addition, retrovirus-mediated expression of WASP led to improvement of cytoplasmic F-actin expression and formation of F-actin-positive microvilli, a process shown to be defective in untransduced WAS cell lines. These preliminary results indicate a potential use for retrovirus-mediated gene transfer as therapy for WAS.

A human minor histocompatibility antigen specific for B cell acute lymphoblastic leukemia.

Human minor histocompatibility antigens (mHags) play an important role in the induction of cytotoxic T lymphocyte (CTL) reactivity against leukemia after human histocompatibility leukocyte antigen (HLA)-identical allogeneic bone marrow transplantation (BMT). As most mHags are not leukemia specific but are also expressed by normal tissues, antileukemia reactivity is often associated with life-threatening graft-versus-host disease (GVHD). Here, we describe a novel mHag, HB-1, that elicits donor-derived CTL reactivity in a B cell acute lymphoblastic leukemia (B-ALL) patient treated by HLA-matched BMT. We identified the gene encoding the antigenic peptide recognized by HB-1-specific CTLs. Interestingly, expression of the HB-1 gene was only observed in B-ALL cells and Epstein-Barr virus-transformed B cells. The HB-1 gene-encoded peptide EEKRGSLHVW is recognized by the CTL in association with HLA-B44. Further analysis reveals that a polymorphism in the HB-1 gene generates a single amino acid exchange from His to Tyr at position 8 within this peptide. This amino acid substitution is critical for recognition by HB-1-specific CTLs. The restricted expression of the polymorphic HB-1 Ag by B-ALL cells and the ability to generate HB-1-specific CTLs in vitro using peptide-loaded dendritic cells offer novel opportunities to specifically target the immune system against B-ALL without the risk of evoking GVHD.

Selective CD4+ T-cell depletion does not prevent graft-versus-host disease.

Donor-derived CD4+ T cells may play a role in the development of graft-versus-host disease (GVHD) and graft-versus-leukemia reaction after allogeneic bone marrow transplantation (BMT). Therefore, we evaluated the effect of CD4+ T-cell depletion on GVHD and graft-versus-leukemia reaction after HLA-matched BMT. CD4 depletion was performed using anti-CD4 monoclonal antibodies and immunomagnetic beads, initially in small-scale experiments on bone marrow and granulocyte colony-stimulating factor-mobilized peripheral blood apheresis products. The result was elimination of the CD4+ T cells from both sources (0% and 2+/-1.4% CD4+ cells, respectively). Subsequently, we used this technique for large-scale negative selection of CD4+ T cells from bone marrow grafts of four consenting leukemic patients in relapse (ALL-3, ANLL-1) (M-3, F-1). The large-scale CD4+ T-cell depletion resulted in >98% (n=4) elimination of CD4+ cells. The resulting population included 17.7+/-4.6% CD3+ T cells, 8.9+/-2.5% CD8+ T cells, 0.1+/-0.1% CD16+ natural killer cells, and 2.3+/-3.2% CD34+ hematopoietic progenitor cells. Patients were transplanted with 2.84+/-1.31 x 10(8) viable cells/kg. They received cyclosporine starting on day -1 as GVHD prophylaxis. Engraftment was fast with a white blood cell count of >1 x 10(9)/L on day 13.2+/-0.5, an absolute neutrophil count of >0.5 x 10(9)/L on day 13.8+/-0.5, and a platelet count of >25 x 10(9)/L on day 26.5+/-6.8. Immunological reconstitution was normal, and peripheral blood phenotyping 3 weeks after BMT disclosed 49.0+/-5.0% CD3, 14.3+/-12.4% CD4, and 59.5+/-7.8% CD8 T cells in addition to 17.0+/-3.0% CD16+ and 9.0+/-3.0% CD56 natural killer cells. Three out of four patients developed very early grade IV GVHD beginning on day 12 (10-13) and died 2-4 months after BMT. One patient is alive and well with a follow-up of 36 months. We conclude that selective CD4 T-cell depletion does not prevent GVHD.