ALBERT

Description: Epstein Barr Virus (EBV) associated Lymphoproliferative Disease (LPD) is a complication of allogeneic haemopoietic stem cell tranplantation (HSCT). In certain groups (congenital immunodeficiency, unrelated and mismatched donor transplants, T cell depletion) the risk may be as high as 25% with significant morbidity and mortality. Strategies to predict such illlness and allow early intervention have therefore assumed importance. We have routinely screened peripheral blood of paediatric, transplanted patients by quantitiative PCR. We report the results of such analysis of 28 successive patients and the EBV serial quantitation in 4 patients with EBV LPD. The median age at time of transplant was 6.5 years. 17 patients received an unrelated donor transplant and one patient received a haplo-identical transplant. The remainder (n=9) received a matched family donor transplant. 23 patients received either Alemtuzumab (n=19) or ATG (n=4). 13 patients had leukaemia, 5 had mucopolysaccharide syndrome, 4 for congential immune deficiency and 6 for non malignant haeamtological conditions. 9 (32%) patients showed no evidence of EBV reactivation using serial PCR monitoring. 10 patients had low level EBV reactivation as defined with a PCR log[copy number] 6, whilst the highest level without disease was 5.2). All 4 patients responded to therapy for EBV LPD, with a combination of rituximab, withdrawal of immune suppression or administration of donor lymphocytes - DLI). At higher EBV levels the quantitative PCR had increasing positive predictive value for clinical LPD. We therefore conclude that EBV serial quantitative PCR is useful in discriminating those who will develop LPD from those that will not. Our data suggest that it is possible to use EBV PCR quantitation further to discriminate asymptomatic EBV reactivation that will resolve without therapy from EBV reactivation that will require intervention. This prevents over exposure of patients to treatments (rituximab, DLI, immune suppression withdrawal) with significant associated toxicity (prolonged B cell aplasia, graft versus host disease).

Description: Bristol has previously published extensively on the use of Campath antibodies during the unrelated donor cell transplantation of children with relapsed or otherwise high risk Acute Lymphoblastic Leukaemia (A.L.L.). These former reports (Br J Haematol1996;94(3):574–8; Blood1999; 94(7) 2236–46) concerned the use of Campath IG (a rat monoclonal antibody directed at CD52) given pre-transplant to the patients by intravenous infusion during conditioning therapy that also included cyclophosphamide (120 mgs/kg) and fractionated total body irradiation (1440 Gy). During this treatment the unrelated donor marrow was T cell depleted in vitro using Campath IM (also monoclonal and of rat origin). The results of such treatment demonstrated results comparable with HLA-matched sibling BMT, with low levels of acute graft versus host disease. In this communication transplant centres in 3 UK hospitals (Bristol, Great Ormond Street, London and Manchester) report their combined experience of using Campath IH (Alemtuzumab, a humanised monoclonal antibody - also directed at CD52) in the conditioning therapy of children with relapsed or otherwise high risk A.L.L.. The conditioning was otherwise again with Cyclophosphamide and fractionated total body irradiation. In this protocol a T cell replete graft was given and there was no in vitro T cell depletion of the unrelated donor marrow. We report 34 successive patients where follow up for at least 12 months was available. The median age was 6.5 years and 28 patients received fully matched grafts (matched at HLA-A, -B, -C, -DRB1 and DQB1). The remaining patients received a graft that was mismatched at one of these class I loci. All patients engrafted and the median time to neutrophil count sustained above 0.5x10e9 / l was 20 days (range 12–58 days). Only tow patients experienced grade III acute GvHD and no patient experienced grade IV acute GvHD. 22 patients survive at a median follow up of 23.8 months including 4 of the patients who received a mismatched graft. There were only 3 deaths in the first 100 days following transplant (2 due to disseminated, invasive adenovirus and one to aspergillus). Beyond 100 days 3 further patients have died of adenovirus, 3 have relapsed, 2 have died of chronic GvHD and one died of a pulmonary haemorrhage of uncertain aetiology. We therefore conclude the combination of Aletuzumab with cyclophosphamide and TBI is an effective and safe conditioning therapy for children with relapsed A.L.L. All patients engraft and there are low rates of acute and chronic GvHD.

Description: Blood loss during corrective surgery for scoliosis and the requirement for homologous blood is an important issue in paediatric spinal surgery. This is particularly pertinent in cases of scoliosis secondary to underlying neuromuscular disease where anaemia may be less well tolerated, both intra-operatively and post-operatively, due to multiple organ dysfunction. Despite the introduction of techniques to minimise the necessity to transfuse homologous blood (pre-deposit autologous donation, cell salvage and isovolaemic haemodilution), it is often required. This brings with it the potential risks of allo-immunization and infective complications. In the UK homologous blood is an increasingly precious resource following the recent exclusion of donors who have themselves received blood products in the last ten years in view of the potential risk of new variant CJD. Aprotinin, a serine proteinase inhibitor (now produced by recombinant technology) with antifibrinolytic properties and the ability to preserve platelet function has been shown to reduce blood loss in major cardiac surgery and liver transplantation. Its effectiveness in the orthopaedic setting has generated more varied reports, however in the setting of spinal surgery for idiopathic scoliosis it has recently been shown to reduce intra-operative blood loss and the requirement for homologous transfusion. We report here our experience with the use of aprotinin in a group of paediatric patients undergoing surgery for scoliosis secondary to an underlying neuromuscular disorder and its effect on intra-operative blood loss. 33 successive patients referred to our institution were allocated to receive aprotinin (n=18) or not (n=15). For those who received the drug a test dose of 50,000KIU was followed by a loading dose of 10,000KIU/kg over 30 minutes. A maintenance infusion was set up at 5,000KIU/hour for the duration of the procedure. The demographics of the two groups were not significantly different. The median age of those children receiving aprotinin was 12 years (7–15) and those who did not 13 years (7–17), [p=0.09]. The median weight of the children receiving aprotinin was 40kg (25–55) and those who did not 38.5kg (21–55), [p=0.74]. The procedures undertaken were posterior spinal fusion (n=26), anterior and posterior fusion (n=3), insertion of growing rods (n=2) and correction of kyphoscoliosis (n=2). All procedures were performed by one of three consultants. The operative time in the two groups was not significantly different, 6.5 hours (4.1–8.9) in those who received aprotinin and 5.7 (4.4–7) in those who did not [p=0.54]. There was however a significant reduction in blood loss in the group who received aprotinin, with an average reduction of 51.6%. Median losses for those who received aprotinin were 1380ml (380ml–2380ml) and 2849ml (1047ml–4651ml) for those who did not [p=0.01]. No adverse events in the form of allergic reactions were observed in the children who received the drug. We conclude that aptotinin is a safe and effective pharmacological intervention which can be used in spinal surgery to reduce operative blood loss. This should translate into a reduced requirement for homologous blood although this was not determined in this study. Evaluation of this secondary endpoint is ongoing.

Description: EBV-driven PCNSL is a rare form of B-cell non-Hodgkin’s lymphoma (NHL) seen in immuno-compromised (particularly AIDs) patients. The prognosis is gloomy and most patients die of their disease as radiotherapy and chemotherapy are usually ineffective in inducing durable remission. We report successful treatment of an 8 years old girl who presented with multifocal EBV-driven B-cell PCNSL with adoptive immunotherapy. Due to rarity of the disease particularly in children, she was investigated for and found to have an underlying primary immunodeficiency associated with defective immunity to herpes viruses. Her underlying immune defect did not fit with any of the currently well characterized congenital immune deficiency syndromes, but as also demonstrated in one of her two brothers, it seemed to have been inherited in an autosomal recessive fashion. Both were persistently lymphopenic (0.3–1.1 X 109/L), with proportionate reduction of T, B, and NK cells. Immunoglobulin classes were normal, as were specific antibody responses to tetanus. Mitogen responses to PHA, serum IgG2 levels and post-vaccination responses to Hib and Pneumococcus were low in both children. Antibody response to EBV was abnormal in both children with development of IgG antibodies to the capsular antigen (VCA) but not the nuclear antigen (EBNA). She was initially treated with conventional chemotherapy (intrathecal as well as systemic high dose Cytarabine and Methrotrexate), Rituximab, anti-virals and immunoglobulins. Neurological deterioration gradually progressed despite four weeks of such treatment. She eventually became bed ridden with quadreparesis and significant bulbar palsy requiring intubation for airway protection. In view of conventional treatment failure, immunotherapy with ex-vivo expanded EBV-specific cytotoxic T-lymphocytes (CTL) from a partially HLA matched donor was initiated. These were given weekly for seven consecutive weeks. There were no untoward effects. She made a remarkable neurological recovery within few weeks and became fully mobile in four months when she received non-myeloablative stem cell transplantation. Post transplant peripheral EBV reactivation (presenting as infectious mononucleosis-like disease without neurological manifestations) was promptly controlled with Rituximab and further doses of allogeneic EBV-CTLs. The child is now well five months post transplantation with evidence of immune reconstitution and remains in complete remission. GVHD prophylaxis has now been totally withdrawn. This is the first report of the successful use of EBV-specific cytotoxic T-lymphocytes in the treatment of primary CNS lymphoma in an immunocompromised patient. It offers new hope in a condition, which until now has been associated with a bleak outlook. The clinical response to immunotherapy is a clear demonstration that although the migration of resting blood lymphocytes across the inter-endothelial tight junctions forming the blood-brain-barrier may be largely restricted in physiological conditions, the migration of activated T lymphocytes into the brain in this clinical setting is unimpeded.