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Treosulfan is an immuno-suppressive and myeloablative alkylating agent that has been introduced as a conditioning agent in stem cell transplantation (SCT). Most studies have been performed in adult patients with malignancy where a low incidence of regimen-related toxicity has been reported. We report the use of treosulfan in 32 consecutive children undergoing SCT for non-malignant disease. Patients received a total treosulfan dose of 36 or 42 g/m(2)/patient given in three daily, divided doses. A range of other conditioning agents and serotherapy was administered to patients who underwent family donor SCT (n = 11), or unrelated donor SCT (n = 21). One patient (3%) died early. Transplant morbidity was limited and mucositis was only mild. Dermatological toxicity was frequent but mild. Twenty-eight patients (87.5%) established donor cell engraftment. In 25 patients (78%) there was adequate, stable donor engraftment. Four patients have required additional transplant procedures to maintain adequate donor-derived haemopoiesis. Twenty-seven patients (84%) survive with a median follow up of 417 d. There were four late deaths due to progression of the underlying disease, graft-versus-host disease or infection. Treosulfan-based conditioning regimens achieve excellent engraftment with reduced regimen-related toxicity in children with non-malignant disease at high risk for both regimen-related toxicity and graft failure.

Infective diarrhoea is common among allogeneic stem cell transplant (SCT) recipients, frequently caused by viruses and may be difficult to differentiate from acute graft-versus-host disease (GVHD). Viral pathogens may directly or indirectly impact upon transplant-related mortality. Rotavirus is one of the most common causes of diarrhoea worldwide, but one of the least studied causes of diarrhoea post SCT. In this retrospective study we describe 21 cases of confirmed rotavirus infection in allogeneic SCT recipients. Most of these cases may occur in clusters during the winter and spring period. Symptoms of rotaviral infection were diarrhoea (95%), vomiting (62%), abdominal pain (38%), weight loss and loss of appetite in 38 and 29% of the cases, respectively. Possible extraintestinal manifestations of rotavirus infection were observed. The duration of the symptoms in this series ranged from 4 days to 4 months with median of 15 days. Patients with rotavirus infection were invariably lymphopenic and/or on immunosuppression for GVHD. Of the patients diagnosed with rotavirus, 86% required hospitalisation. In 57% of the cases, other viral pathogens were isolated near to the rotavirus infection period. Rotavirus infection is an important cause of prolonged diarrhoea post SCT, causing significant morbidity and frequently requiring hospitalisation.

In this study, we retrospectively analysed the clinical features, risk factors and outcome of 22 patients with thrombotic thrombocytopenic purpura (TTP) occurring after allogeneic stem cell transplantation. All but two of these patients received stem cells from unrelated donors (UDs), two-thirds were female, three-quarters were adults and leukaemia was the major reason for transplant. The incidence of TTP was 20 out of 332 patients (6%) with UD transplants and two out of 104 recipients (2%) of matched sibling allografts (P = 0.16). In order to ascertain basic demographic risk factors for the development of TTP, we compared the 22 patients with 434 patients who did not develop TTP. Compared with patients who did not develop TTP, patients with TTP were nearly three times older (P < 0.001) and were more than twice as likely to be female (P = 0.001). Because > 90% of patients were recipients of UD marrow, we then compared the 20 UD-bone marrow transplantation (BMT) patients with 60 randomly selected UD-BMT patients who did not develop TTP. On univariate analysis, age and female gender were again significant risk factors, as was grade II-IV acute graft-versus-host disease (GvHD) (P = 0.002), and there was a trend towards an association with chronic GvHD (P = 0.083). However, after logistic regression analysis, only age and sex remained significant (P < 0.001 and 0.009 respectively). We report an 86% mortality with only three survivors out of 22 patients, and one of these remains thrombocytopenic and red cell transfusion dependent, possibly in part because of graft hypoplasia. Six out of 17 patients responded to plasmapheresis, but the majority of them ultimately succumbed because of TTP, often in association with GvHD or fungal infection.

We conducted a retrospective review of the clinical features and outcome of adenovirus infection in 572 consecutive patients transplanted in a single centre over a 10 year period. One hundred patients (17%) had a total of 105 episodes of adenovirus infection diagnosed at a median of 18 days post transplant (range 2-150 days). The incidence was higher in children than adults (21% vs 9%, P < 0.001) and in unrelated donor vs matched sibling donor transplants (26% vs 9%, P < 0.001). Diarrhoea and fever were the most common presenting features. Reflecting these symptoms, the most common site of isolation was the stool. Serotypes 1, 2 and 7 were the most frequently seen (total of 41/68 or 60% of evaluable cases). In six patients (6%) adenovirus infection was the direct cause of death occurring at a median of 72 days post transplant (range 18-365 days). Five of these six patients had pulmonary involvement and four had associated graft-versus-host disease (GVHD). Three further patients were considered to have severe adenoviral disease (total incidence 9%). Isolation of virus from multiple sites correlated with a poor outcome (P < 0.001). Comorbid viral infection was common in this group with 50% of all patients having other viruses isolated (predominantly polyoma virus and cytomegalovirus). We conclude that adenovirus is commonly isolated after bone marrow transplant and is a cause of significant morbidity but was a rare cause of mortality (6/572 = 1%) in our patient group as a whole. The relative infrequency of severe infection will make it difficult for the transplant physician to decide which patients should receive experimental antiviral drugs such as ribavirin and cidofovir or immunomodulatory therapy with donor white cell infusions.

The biological activities of CD8+ that co-express CD57 remain poorly defined. It is unclear whether all CD8+ cells have the potential to become CD57+ or whether they represent a unique subset with distinct functions. Several studies have reported the association between elevated numbers of CD8+CD57+ and a wide range of clinical disorders such as viral reactivation of human cytomegalovirus (HCMV). In this study, we have investigated the relationship between viral reactivation and the effect of diminished interleukin (IL)-2 production. Using CD8+ cells isolated from patients at various times after allogeneic transplants and in vitro models of HCMV infection, we determined their combined effect on CD8+CD57+. Our results show that high numbers of CD8+CD57+ correlated with diminished killing of HCMV-infected targets. In addition, we showed a synergistic effect between IL-2 and HCMV in the expansion of CD8+CD57+ cells. Furthermore, these cells after anti-CD3 stimulation did not produce tumour necrosis factor (TNF)-alpha or interferon (IFN)-gamma. Interestingly, IL-10 production was elevated in several patients which appeared to be associated with the time from transplant.

The outcome for 39 patients with acute myeloid leukemia (AML) in remission who had CAMPATH 1M T cell-depleted unrelated donor bone marrow transplantations (BMTs) is described. Conditioning was mainly with cyclophosphamide (120 mg/kg) and total body irradiation (TBI) (14.4 Gy), but 5 patients received busulfan in place of TBI and 200 mg/kg cyclophosphamide. All patients received cyclosporin, and short-course methotrexate was given to recipients of mismatched grafts. The patient population was predominantly pediatric (median age, 10 years), but one third of the patients was aged 15 years or above. Twenty-five patients were in second complete remission (CR2), and 14 had high-risk CR1 disease (primarily failed remission induction or antecedent myelodysplastic syndrome, often with complex cytogenetic abnormalities). Both recipient and donor were cytomegalovirus seronegative in 15 of 37 cases (38%); 51% of patients were matched for HLA class I and II. Grade II to IV acute graft-versus-host disease (GVHD) occurred in 24% of patients; chronic GVHD occurred in 5 of 31 evaluable patients (16%), 4 extensive and 1 limited. Relapse occurred in 5 cases (13%); 1 of these 5 patients survives, 24 months after a second unrelated donor transplantation. Two of these relapses were associated with secondary graft failure (incidence rate, 5%). All patients engrafted primarily. Severe viral infection was the major transplant-associated complication, with 12 episodes in 9 patients, 5 of them lethal. Twenty-five patients survive at a median follow-up of 44 months (range, 2-102 months), with estimated actuarial overall and disease-free survival rates at 44 months of 61% (SE 8%) and 57% (SE 8%), respectively. Nineteen patients are more than 2 years post-BMT and may be cured. The functional status of long-term survivors is excellent, with 19 of 21 patients who survive 6 months or more in full-time employment or full-time students. These encouraging results suggest that in patients lacking a sibling donor, unrelated donor BMT for AML in remission achieves survival figures as good as or better than those reported on patients with autologous stem cell transplantation, and that T-cell depletion of grafts is associated with a low relapse rate and an excellent functional status. However, only a randomized study comparing unrelated donor BMT and auto-grafting will resolve which of these treatment strategies is better for patients with AML.

The efficacy of allografting in acute lymphoblastic leukemia (ALL) is heavily influenced by remission status at the time of transplant. Using polymerase chain reaction (PCR)-based minimal residual disease (MRD) analysis, we have investigated retrospectively the impact of submicroscopic leukemia on outcome in 64 patients receiving allogeneic bone marrow transplantation (BMT) for childhood ALL. Remission BM specimens were taken 6 to 81 days (median, 23) before transplant. All patients received similar conditioning therapy; 50 received grafts from unrelated donors and 14 from related donors. Nineteen patients were transplanted in first complete remission (CR1) and 45 in second or subsequent CR. MRD was analyzed by PCR of Ig or T-cell receptor delta or gamma rearrangements, electrophoresis, and allele-specific oligoprobing. Samples were rated high-level positive (clonal band evident after electrophoresis; sensitivity 10(-2) to 10(-3)), low-level positive (MRD detected only after oligoprobing; sensitivity 10(-3) to 10(-5)), or negative. Excluding 8 patients transplanted in CR2 for isolated extramedullary relapse (all MRD-), MRD was detected at high level in 12 patients, low level in 11, and was undetectable in 33. Two-year event-free survival for these groups was 0%, 36%, and 73%, respectively (P <.001). Follow-up in patients remaining in continuing remission is 20 to 96 months (median, 35). These results suggest that MRD analysis could be used routinely in this setting. This would allow identification of patients with resistant leukemia (who may benefit from innovative BMT protocols) and of those with more responsive disease (who may be candidates for randomized trials of BMT versus modern intensive relapse chemotherapy).

Detection of residual disease after completion of therapy or following bone marrow transplantation (BMT) in patients with acute promyelocytic leukaemia (APL) predicts relapse and is associated with a poor prognosis. Here we describe the successful treatment of residual disease post-transplant in APL using prolonged all-trans retinoic acid (ATRA) therapy in two children in whom autologous BMT (ABMT) had been performed in second complete remission (CR). ATRA treatment was well tolerated and found to be beneficial despite its prior use as a component of the initial induction protocol. ATRA therapy post-transplant led to molecular remission as determined by fluorescence in situ hybridization (FISH) as well as reverse transcriptase-polymerase chain reaction (RT-PCR) analyses and remission now exceeds 3.5 years in both patients. Overall, this study not only demonstrates that ATRA may successfully salvage APL patients with residual disease post-transplant, but also suggests a potential role for retinoids post-consolidation as a means of eliminating residual disease which could be beneficial even in patients previously exposed to ATRA as a component of the induction protocol.

We have analysed the behaviour of minimal residual disease (MRD) after allogeneic bone marrow transplantation (allo-BMT) in 71 children with acute lymphoblastic leukaemia (ALL). The method relied on PCR of IgH, TCRdelta and/or TCRgamma gene rearrangements followed by electrophoretic size resolution and allele-specific oligoprobing. Patients were similarly conditioned; 55 received marrow from unrelated donors and 16 from related donors. MRD was assessed at various time-points up to 24 months after BMT. Three children were not evaluable due to transplant-related mortality. MRD was detected in 28/32 patients (88%) who relapsed post-BMT; 16 were positive at all times and 12 were initially negative but became positive at a median of 3 months (range 1.5-11) prior to relapse. In contrast, only eight of 36 (22%) patients who remained in continuing complete remission (CCR) (median follow-up 43 months, range 20-94) showed MRD at any time after BMT (P<0.0001). In these eight patients MRD was found up to 9 months after transplant and at low levels (0.01-0.001%). All eight (median follow-up 39 months, range 24-87) had at least two MRD-negative samples tested subsequently and five of the eight had evidence of grade I-II acute graft-versus-host disease (GvHD), raising the possibility of a graft-versus-leukaemia effect. In general, any evidence of MRD after allo-BMT is a poor prognostic sign. However, if immunotherapy were to be targeted towards patients with evidence of persisting MRD after BMT, the method described would expose only a small proportion of patients to unnecessary additional toxicity.

Only 30% of children have a matched family donor (MFD). Alternative donors can be found from volunteer unrelated donor (UD) panels, and almost every child has a haploidentical parental donor (Haplo). The outcome of alternative donor BMT has previously been inferior due to increased graft rejection and GVHD. The recent outcome of 121 consecutive children undergoing MFD, UD, and Haplo BMT between 1994 and 1997 was analysed. [table in text] Preparative regimens for MFD/UD/Haplo BMT respectively included TBI in 30%/36%/11%, chemotherapy only in 59%/64%/89%, Campath 1G or none n 14%/ 0%/0%. The balance of GVHD and rejection was addressed by T-cell depletion (Campath 1M) in 2/71 MFD BMTs, T-cell depletion with addback of 5 x 10(4) T cells/kg on day zero in 33/41 UD BMTs, and T cell depletion of purified mobilised peripheral blood CD34+ cells and bone marrow in 9/9 Haplo BMTs. Stable engraftment was achieved in 68/71, and 35/41 (one patient after 2nd BMT) and 7/9 MFD, UD, and Haplo BMTs respectively. Median follow up, survival, disease free survival and, death rates for MFD/UD/Haplo BMT were 12/12/12 months, 74%/77%/78%, 69%/67%/67%, and 27%/23%/23% respectively. Within a wide range of paediatric diseases the outcome of alternative donor BMT now parallels that of MFD BMT.

Purpose: Few patients with Philadelphia-positive acute lymphoblastic leukemia (Ph-positive ALL) have been cured by chemotherapy alone. Registry figures show that 38% of patients who have a matched-sibling bone marrow transplant (BMT) are disease-free 2 years after transplant, but the majority of patients lack a sibling donor. Most modern ALL protocols recommend unrelated donor (UD) BMT for patients with Ph-positive ALL in first complete remission (CR1), but the outcome of this is unknown., Patients and Methods: We report the results of 15 children and adolescents who had a T-cell depleted UD-BMT for Ph-positive ALL. Thirteen of 15 had been previously treated on United Kingdom ALL protocols. Nine were in CR1 and six had more advanced disease. Eleven donor recipient pairs were matched at HLA-A, HLA-B, HLA-DR, and HLA-DQ, and four were mismatched at one or two HLA loci., Results: The incidence of greater than grade I acute and chronic graft-versus-host disease (GVHD) was low (13% and 8%, respectively). Six patients have relapsed and seven patients survive at a median of 21 months post-BMT; six of seven are disease free. All seven survivors are in full-time education or work. The 2-year overall and disease-free survivals are 44% +/- 13% and 37% +/- 13% (+/- SE). None of four patients who had mismatched donors survived, but seven of 11 matched recipients survive (P < .05)., Conclusion: UD-BMT can produce prolonged disease-free survival in young patients with Ph-positive ALL who otherwise would have an extremely poor outlook.

The accurate and rapid determination of the origin of haemopoietic cells may provide valuable information as to the aetiology of, and most appropriate therapy for, leucopenia following allogeneic bone marrow or peripheral blood stem cell transplantation. We describe an approach to the analysis of chimaerism post bone marrow transplantation (BMT) based on the immunomagnetic capture of white cells combined with microsatellite polymerase chain reaction (PCR) and resolution of products by polyacrylamide gel electrophoresis (PAGE). This non-isotopic method enables the chimaeric status to be determined from as little as 1.0 ml of profoundly leucopenic peripheral blood (WBC < or =0.1 x 10(9)/l) and has been applicable to all donor/recipient pairs tested so far. Results are available within 6 h of blood sampling and lineage-specific chimaerism is possible. Furthermore, blood transfusions do not interfere with the analysis.

Allogeneic sibling bone marrow transplantation (BMT) is the recommended treatment for relapsed childhood acute lymphoblastic leukaemia (ALL), but appropriate donors are only available in 30% of cases. Unfortunately, BMT from unrelated donors (UD) has been associated with high rates of severe graft-versus-host disease (GvHD) and transplant-related mortality (TRM). In an attempt to improve outcome in UD-BMT we have assessed the impact of T-cell depletion using CAMPATH-1 (anti-CD52) monoclonal antibodies in 50 consecutively referred patients with relapsed ALL in second remission. All were previously treated according to MRC protocols UKALL X and XI, and then given chemotherapy on MRC R1 from relapse until UD-BMT, 19 patients had relapsed on and 31 off therapy. Patients and donors were fully matched at HLA-A, -B, -DR and -DQ loci in 29 cases and mismatched in 21 (four mismatched for more than one antigen). Pre-transplant conditioning comprised CAMPATH-1G, cyclophosphamide and total body irradiation. Bone marrow was T-cell depleted in vitro using CAMPATH-1 antibodies. Additional GvHD prophylaxis consisted of cyclosporin A (42 cases), cyclosporin plus methotrexate (four) or none (four). 47 patients engrafted. The incidence of acute GvHD was very low: two patients with grade II disease in the matched group, four with grade II-IV in the mismatched group. Only four patients have chronic GvHD. The actuarial event-free survival (EFS) at 2 years is 53%, with no significant difference between the matched and mismatched group. Further leukaemic relapse was the most important cause of failure. These results are similar to the most favourable published reports for HLA-matched sibling BMT in relapsed ALL.

Juvenile chronic myeloid leukaemia (JCML) is a rare haematological condition of childhood curable only by bone marrow transplantation (BMT). We report our experience using matched and mismatched unrelated donor BMT for JCML in five patients. Although the procedure is hazardous in terms of toxicity and relapse, two patients are alive and disease-free 28 and 49 months post BMT.

Summary:CAMPATH-1H (C-1H) is widely used in vivo and / or in vitro for T cell depletion in hematopoietic SCT. This humanised monoclonal antibody is specific for CD52, a marker coexpressed on the majority of human lymphocytes with CD48 and other glycosylphosphatidyl-inositol (GPI) anchored proteins. We detected CD52 / CD48 dual expression on >99% of CD3+ lymphocytes from normal individuals and all 15 post-SCT patients whose transplants did not utilise C-1H. By contrast, 23 / 26 patients with transplants involving C-1H (in vivo, in vitro or both) exhibited populations lacking CD52 expression that accounted for 49.7% (4.2–86.2%) of the CD3+ lymphocytes (median and range) in samples evaluated at a median of 2 months post-SCT. Most CD52− cells also lacked CD48 expression. These GPI− T cells were of either donor or mixed donor / recipient origin. They were predominant in the early months after SCT at times of profound lymphopenia and inversely correlated with the recovery of the absolute lymphocyte count (r= − 0.663, P<0.0001). The presence of CD52− cells has been correlated previously with clinical outcome after CAMPATH therapy for both malignant and nonmalignant diseases.Bone Marrow Transplantation (2005) 36, 237–244. doi:10.1038/sj.bmt.1705049; published online 13 June 2005 [ABSTRACT FROM AUTHOR]

Leukemia (2003) 17, 247–251. doi:10.1038/sj.leu.2402759 [ABSTRACT FROM AUTHOR]

The biological activities of CD8+ that co-express CD57 remain poorly defined. It is unclear whether all CD8+ cells have the potential to become CD57+ or whether they represent a unique subset with distinct functions. Several studies have reported the association between elevated numbers of CD8+CD57+ and a wide range of clinical disorders such as viral reactivation of human cytomegalovirus (HCMV). In this study, we have investigated the relationship between viral reactivation and the effect of diminished interleukin (IL)-2 production. Using CD8+ cells isolated from patients at various times after allogeneic transplants and in vitro models of HCMV infection, we determined their combined effect on CD8+CD57+. Our results show that high numbers of CD8+CD57+ correlated with diminished killing of HCMV-infected targets. In addition, we showed a synergistic effect between IL-2 and HCMV in the expansion of CD8+CD57+ cells. Furthermore, these cells after anti-CD3 stimulation did not produce tumour necrosis factor (TNF)-α or interferon (IFN)-γ. Interestingly, IL-10 production was elevated in several patients which appeared to be associated with the time from transplant. [ABSTRACT FROM AUTHOR]

A correction to the article "Successful outcome of allo-SCT in high-risk pediatric AML using chemotherapy-only conditioning and post transplant immunotherapy," which was published on July 14, 2008 is presented.