Bone Marrow Transplantation Conditioning Research Articles

Effect of antithymocyte globulin source on outcomes of bone marrow transplantation for severe aplastic anemia.

For treatment of severe aplastic anemia, immunosuppressive therapy with horse antithymocyte globulin results in superior response and survival compared with rabbit antithymocyte globulin. This relative benefit may be different in the setting of transplantation as rabbit antithymocyte globulin results in more profound immunosuppression. We analyzed 833 severe aplastic anemia transplants between 2008 and 2013 using human leukocyte antigen (HLA)-matched siblings (n=546) or unrelated donors (n=287) who received antithymocyte globulin as part of their conditioning regimen and bone marrow graft. There were no differences in hematopoietic recovery by type of antithymocyte globulin. Among recipients of HLA-matched sibling transplants, day 100 incidence of acute (17% versus 6%, P<0.001) and chronic (20% versus 9%, P<0.001) graft-versus-host disease were higher with horse compared to rabbit antithymocyte globulin. There were no differences in 3-year overall survival, 87% and 92%, P=0.76, respectively. Among recipients of unrelated donor transplants, acute graft-versus-host disease was also higher with horse compared to rabbit antithymocyte globulin (42% versus 23%, P<0.001) but not chronic graft-versus-host disease (38% versus 32%, P=0.35). Survival was lower with horse antithymocyte globulin after unrelated donor transplantation, 75% versus 83%, P=0.02. These data support the use of rabbit antithymocyte globulin for bone marrow transplant conditioning for severe aplastic anemia.

Combining Flow Cytometry and Molecular Assessment Improves the Prognostic Value of Pre-Transplant Minimal Residual Disease in Acute Myeloid Leukemia

BACKGROUND AND AIMSAllogeneic bone marrow transplantation (BMT) offers the greatest chance of cure for high-risk acute myeloid leukemia (AML) patients. Persistence of disease or high levels of pre BMT minimal residual disease (MRD) have been reported to predict relapse risk after BMT. Multicolor flow cytometry (MFC) is the most common tool to evaluate MRD, whereas WT1 gene expression is the most widely applicable and standardized molecular MRD marker in AML. The aim of this study was to analyze the role of pre-BMT combined MFC-molecular MRD assessment as predictor for the post-transplant relapse risk.MATERIALS AND METHODSWe retrospectively analyzed the outcome of 224 consecutive AML patients receiving allo-BMT in 1st or 2nd CR. Pre-BMT marrow samples were analysed for WT1 expression and MFC as MRD evaluation. Median age at transplant was 44 years. Disease phase was CR1 in 161 (72%) and CR2 in 63 patients (28%). BMT conditioning wasmyeloablative in 163 (73%), whereas 61 patients (27%) received reduced intensity conditioning. Stem cell source was HLA-identical sibling in 79 (35%),haploidentical in 59 (27%) and alternative donor in 86 (38%).Median follow-up was 64 months (95% CI 50.3 - 77.3 months). Cumulative Incidence of Relapse (CI of relapse) at various time-points was calculated in competing risk analysis by takingin account non relapse mortality as competing event. Overall Survival (OS) was calculated from the time of BMT until death by any cause or last follow-up.A positive MFC MRD was defined by the presence of no less than 25 clustered leukemic cells/105 total events (threshold of 2.5x10-4 residual leukemic cells) at four-color flow-cytometry. WT1 copy number/Abl copy number 500x104 was used as cut-off value for abnormal WT1 expression.RESULTSRelapse occurred in 62 patients (27.7%). Three-year estimate of CI of relapse was 25.9% (median not reached). The CI of relapse was significantly affected by occurrence of acute GVHD (lower for grade ≥2, p <0.05) and MRD status before BMT, measured with any method (p <0.001 for WT1-based MRD, p<0.03 for MFC based MRD, p<0.0001 for combined MRD). In multivariate analysis the combined MRD evaluation and the onset ofaGVHDwere the only independent predictor of CI of relapse (p <0.001 and 0.04, respectively, Table I).Specifically, patients with double positive molecular and MFC MRD had a very poor outcome, with a 3-year CI of relapse of 51.9% (Figure 1). On the contrary, patients with negative combined MRD had a very low CI of relapse, regardless of the conditioning regimen received, whereas a trend toward benefit frommyeloablativeconditioning was observed in patients with MFC positive/WT1 negative MRD and double positive patients.Patients with negative MFC-MRD undergoing BMT in 1st or 2nd CR had a very similar outcome (3-year CI of relapse 8.3 and 5.6%, respectively). Similarly, MFC positive/WT1 negative MRD patients had a 3-year CI of relapse of 17.3 and 18.% for CR1 and CR2 patients, respectively. Patients who were MRD positive by both methods undergoing BMT in CR1 or CR2 had a similar, very high CI of relapse (Figure 2).MRD evaluation was also a strong predictor of long term survival: 3- years OS was 73.5% for MFC negative and 36.7% for double WT1 and MFC MRD positive patients, respectively, (p <0.001). Multivariate OS analysis showed that conditioning intensity and combined MRD evaluation significantly influenced OS duration (p <0.005 and <0.001, respectively)CONCLUSIONSPre transplant MRD evaluation by combined molecular and MFC assessment on bone marrow samples is a reliable predictor of relapse risk. Patients with both negative pre-BMT MRD markers have a significantly lower CI of relapse, while patients with both positive MRD markers display an higher risk of relapse. Given the high prognostic value, combined molecular/MFC pre-transplant MRD status may be taken in account when considering the most appropriate stem cell source or conditioning for each patient. Furthermore, identifying patient in morphological CR at BMT but with a very high risk of relapse, such as double positive patients, could open the way to apply pre-emptive therapeutic strategies to prevent AML relapse, from donor lymphocyte infusion to other innovative approaches. [Display omitted] [Display omitted] [Display omitted] DisclosuresGobbi:Janssen: Consultancy, Honoraria; Novartis: Consultancy, Research Funding; Celgene: Consultancy; Roche: Honoraria; Mundipharma: Consultancy, Research Funding; Takeda: Consultancy; Gilead: Honoraria.

A Depleting Anti-CD45 Monoclonal Antibody as Isolated Conditioning for Bone Marrow Transplantation in the Rat.

ObjectiveA monoclonal antibody (mAb) against the leukocyte common antigen CD45 (RT7 in rats) could facilitate bone marrow transplantation (BMT). This study in rats evaluates a depletive rat anti-RT7a mAb as isolated tool for BMT conditioning without using irradiation or any chemotherapeutic / immunosuppressive agent.MethodsThe model used a CD45 di-allelic polymorphism (RT7a/RT7b). The anti-RT7a mAb was intravenously administered to LEW.1W rats (RT1uRT7a) at 5, 10 and 15 mg/kg. 1x108 BM cells of MHC syngeneic (RT1u), MHC disparate (RT1l) or MHC haploidentical (RT1u/l) donors were transplanted. All BM donor strains carried the RT7b allele so that their CD45+ cells were not affected by the anti-RT7a mAb. Recipients were monitored for reconstitution and donor-chimerism in blood leukocytes.ResultsmAb dosages of 5 or 10 mg/kg were myelosuppressive, whereas 15 mg/kg was myeloablative. Multi-lineage donor-chimerism at day 100 indicated engraftment of MHC syngeneic BM after any used mAb dosage (5 mg/kg: 46+/-7%; 10 mg/kg: 62+/-5%; 15 mg/kg: 80+/-4%). MHC disparate BM resulted in autologous reconstitution after conditioning by 10 mg/kg of the mAb and caused transient chimerism ending up in death associated with aplasia after conditioning by 15 mg/kg of the mAb. MHC haploidentical BM (F1 to parental) engrafted only after conditioning by 15 mg/kg (chimerism at day 100: 78+/-7%). Abandonment of α/β TCR+ cell depletion from BM grafts impaired the engraftment process after conditioning using 15 mg/kg of the mAb in the MHC syngeneic setting (2 of 6 recipients failed to engraft) and the MHC haploidentical setting (3 of 6 recipients failed).ConclusionThis depletive anti-RT7a mAb is myelosuppressive and conditions for engraftment of MHC syngeneic BM. The mAb also facilitates engraftment of MHC haploidentical BM, if a myeloablative dose is used. RT7b expressing, BM-seeded α/β TCR+ cells seem to impair the engraftment process after myeloablative mAb conditioning.

Targeted busulfan and fludarabine-based conditioning for bone marrow transplantation in chronic granulomatous disease.

Chronic granulomatous disease (CGD) is a primary immunodeficiency disease caused by impaired phagocytic function. Hematopoietic stem cell transplantation (HSCT) is a definitive cure for CGD; however, the use of HSCT is limited because of associated problems, including transplantation-related mortality and engraftment failure. We report a case of a patient with CGD who underwent successful HSCT following a targeted busulfan and fludarabine reduced-toxicity myeloablative conditioning. Intravenous busulfan was administered once daily for 4 consecutive days (days –8 to –5), and the target area under the curve was 75,000 µg·hr/L. Fludarabine (40 mg/m2) was administered once daily for 6 consecutive days from days –8 to –3. Antithymocyte globulin (2.5 mg/kg/day) was administered from days –4 to –2. The patient underwent successful engraftment and did not have any severe toxicity related to the transplantation. Conditioning with a targeted busulfan and fludarabine regimen could provide a better outcome for HSCT in CGD, with close regulation of the busulfan dose.

Long-term outcomes of granulocyte colony-stimulating factor-combined conditioning in allogeneic hematopoietic stem cell transplantation from HLA-identical family donors for myeloid malignancies

Dormant leukemia cells, which might escape the cytotoxic effect of conditioning before hematopoietic stem cell transplantation (HSCT), could be induced to enter the cell cycle by granulocyte colony-stimulating factor (G-CSF) and become more susceptible to the cell-cycle-specific agent cytarabine arabinoside (Ara-C). Based on this effect, we have utilized G-CSF-combined high-dose Ara-C in myeloablative conditioning for allogeneic bone marrow or peripheral blood stem cell transplantation from HLA-identical family donors since 1988. We report on the long-term outcomes of allogeneic HSCT using a conditioning regimen of 12Gy total body irradiation and G-CSF-combined high-dose Ara-C in 89 adult patients with acute myeloid leukemia, chronic myelogenous leukemia, or myelodysplastic syndrome. With a median follow-up of 135 months, the probability of overall survival and cumulative incidence of relapse at 5 years were 67.8% and 19.4%, respectively. In multivariate analysis, disease status at HSCT was associated with survival and relapse. These data demonstrate that G-CSF-combined myeloablative conditioning could be safely and effectively used for patients with myeloid malignancies.

Preventive measures for cyclophosphamide-related hemorrhagic cystitis in blood and bone marrow transplantation: an Italian multicenter retrospective study.

Hemorrhagic cystitis (HC) is a troublesome and potentially life-threatening complication of bone marrow transplantation (BMT). HC can appear within a few hours after chemotherapy or after weeks or months. Early-onset HC (EOHC) is usually associated with the conditioning regimen. The main aim of this study was to describe the incidence of EOHC in patients undergoing BMT regimens including high-dose cyclophosphamide (CY) and the effects of the main preventive measures adopted in Italian nursing practice. The authors retrospectively analyzed the clinical records of 158 Italian patients who underwent BMT from 2006-2008. Thirty-one patients (19.6%) developed EOHC. One hundred and forty-seven patients (93%) given high-dose CY were treated with hyperhydration combined with 2-mercaptoethane sulphonate (mesna) and diuresis alkalinization, and only 51 (32.3%) patients were preventively catheterized and received continuous bladder irrigation (CBI). By univariate analysis, prophylactic urethral catheterization and CBI did not decrease EOHC incidence questioning if these measures were to be routinely recommended. Previous studies showed increased discomfort and urinary tract infection in catheterized patients; therefore, nurses may fulfill an important role in balancing the benefits and harms of preventive catheterization and CBI in patients who received BMT conditioning including high-dose CY.

Intestinal Helminth Colonization Regulates Lethal Graft Versus Host Disease and Preserves Graft Versus Tumor in Bone Marrow Transplanted Mice

Introduction.Graft versus host disease (GVHD) is a frequent and severe complication of allogeneic bone marrow transplantation (BMT). Novel and less toxic treatment regimens are needed to suppress GVHD and preserve graft versus tumor (GVT). Self-limited intestinal helminth colonization stimulates host regulatory T cell (Treg) subsets. The intestine is a prime organ in GVHD generation, with Tregs playing a pivotal role in controlling GVHD. Treg-mediated suppression of GVHD preserves GVT. We hypothesized that self-limited intestinal colonization with helminths protects BMT mice from lethal acute GVHD and preserves the GVT.Methods. Three weeks after administering 3rd stage adult Heligmosomoides polygyrus larvae into 5-6 week old male WT Balb/C recipient mice (H2d), we initiated H2b→H2d MHC I/II mismatch model of acute GVHD by total body irradiation (TBI; 850 cGy) and administration of total splenic T cells and T cell depleted (TCD) bone marrow (BM) cells from male C57BL/6 WT (MHC: H2b) donors into uninfected control and helminth-infected male wild-type (WT) Balb/C (MHC: H2d) bone marrow recipients. In certain experiments, we used donor T cells from TGFβ RII dominant negative (DN) (TGFβ RII DN) mice (MHC: H2b) whose T cells are unresponsive to TGFβ-mediated immune regulation due to over-expression of a truncated TGFβ receptor II. Th1 (IFNγ, TNFα) cytokine generation was analyzed by ELISA. Splenic and mesenteric lymph node (MLN) cells were stained for H2b, H2d, T cell surface markers and FoxP3. Tissues were analyzed for GVHD-related inflammation. GVT was assessed in uninfected and helminth-infected BMT recipients by administration of luciferase expressing A20 leukemia/lymphoma cells (A20-luc; H2d) and IVIS imaging. Statistical and survival difference between groups was determined by Student’s t-test and Kaplan Meier curves, respectively.Results. Helminths increased ~3-fold CD4 T cell and FoxP3+ Treg TGFβ generation (p<0.01 btw uninfected and helminth-infected). Helminths suppressed donor T cell Th1 cytokine production (p<0.05 btw control and helminth-infected mice (N=3)) and regulated clinical as well as histopathological (colon and lung) GVHD score (p<0.05 between control and helminth-infected, for each parameter from multiple experiments). Helminths promoted the survival of recipient T cells after TBI (Spleen control: 1.4±0.9 x 104 vs. helminth-infected: 11.0±6.9 x 104 CD3+ T cells (N=8; p<0.05); MLN control: 0.09±0.09 x 104 vs. helminth-infected: 6.0±5.2 x 104 (N=8; p<0.001)). Surviving T cells in H. polygyrus-colonized mice were enriched for FoxP3+ Tregs (spleen: 1.16±0.6 x 104 and MLN: 0.65 ±0.56 x 104 FoxP3+ Tregs). Recipient T cells enriched for Tregs survived during GVHD, as analyzed 6 days after BMT (3.3±1.1% of splenic and 15.9±3.5% of MLN host T cells being FoxP3+ Tregs). The number of recipient T cells in uninfected BMT mice was low for further Treg analysis. Helminths increased the number of GVHD-regulating donor-derived H2b+ FoxP3+ Tregs (Spleen: 66±32 x 103 (control) vs. 206±73 x 103 (helminth-infected) Tregs per mouse (p<0.05; N=6); MLN: 1.6 ±1.1 x 103 (control) vs. 4.9±2.0 x 103(helminth-infected) Tregs per mouse (p<0.05; N=4). Helminth infection was associated with >15 weeks survival of 40% of recipients, while all uninfected BMT recipient mice died of GVHD (p<0.01 between uninfected (N=10) and helminth-infected (N=10) WT recipients). Helminths did not promote the survival of recipients, when T cells unresponsive to TGFβ were used as donor lymphocytes. Helminths preserved the GVT in BMT mice transferred A20-luc. All uninfected and helminth-infected TCD BM transferred recipients died within few weeks displaying significant luciferase activity, while no luciferase signal was evident at day 70 in surviving (5/9) helminth-infected mice administered TCD BM and splenic T cells along with A20-luc. Uninfected mice (9/9) transferred splenic T cells, TCD BM and A20-luc died of GVHD without showing luciferase activity.Conclusions. Our results show that (1) helminths stimulate TGFβ producing host T cells and FoxP3+ Tregs that survive BMT conditioning with TBI; (2) helminths regulate GVHD by inducing donor Tregs and in a TGFβ dependent manner; (3) helminths preserve GVT. With a safe side-effect profile, helminths or helminth products have the potential to be investigated as an alternative strategy to prevent or treat GVHD. DisclosuresNo relevant conflicts of interest to declare.

Chemokine treatment rescues profound T-lineage progenitor homing defect after bone marrow transplant conditioning in mice

AbstractDevelopment of T cells in the thymus requires continuous importation of T-lineage progenitors from the bone marrow via the circulation. Following bone marrow transplant, recovery of a normal peripheral T-cell pool depends on production of naïve T cells in the thymus; however, delivery of progenitors to the thymus limits T-lineage reconstitution. Here, we examine homing of intravenously delivered progenitors to the thymus following irradiation and bone marrow reconstitution. Surprisingly, following host conditioning by irradiation, we find that homing of lymphoid-primed multipotent progenitors and common lymphoid progenitors to the thymus decreases more than 10-fold relative to unirradiated mice. The reduction in thymic homing in irradiated mice is accompanied by a significant reduction in CCL25, an important chemokine ligand for thymic homing. We show that pretreatment of bone marrow progenitors with CCL25 and CCL21 corrects the defect in thymic homing after irradiation and promotes thymic reconstitution. These data suggest new therapeutic approaches to promote T-cell regeneration.

Cytostatic conditioning in experimental allogeneic bone marrow transplantation: Busulfan causes less early gastrointestinal toxicity but Treosulfan results in improved immune reconstitution

Background: Acute graft versus host disease (aGVHD) after allogeneic bone marrow transplantation (allo-BMT) is associated with significant morbidity and mortality. We evaluated the impact of the conditioning regimen on aGVHD comparing Treosulfan (Treo) and Busulfan (Bu) with total body irradiation (TBI).Methods: Using a haploidentical murine model, B6D2F1 mice conditioned with Bu (100 mg/kg)/Fludarabine (Flu, 500 mg/kg) or Treo (6000 mg/kg)/Flu (500 mg/kg) or TBI with 14 Gy received bone marrow cells and splenocytes (20 × 106) from either syngeneic (B6D2F1) or allogeneic (C57BL/6N) donors in order to analyze aGVHD outcome.Results: Conditioning with Bu/Flu or Treo/Flu resulted in significantly reduced aGVHD severity and improved survival (p < 0.05) after allo-BMT compared to TBI. On day 5 after allo-BMT, the organ damages of Bu/Flu conditioned animals were significantly reduced in association with diminished expression of tumor necrosis factor in serum compared to Treo/Flu. Interestingly, the early toxicity of Treo/Flu did not result in significantly higher aGVHD severity; furthermore, a significantly improved immune reconstitution of B220-positive B cells was observed at day 42 after Treo/Flu conditioning compared to Bu/Flu.Conclusion: Conditioning with Treo/Flu or Bu/Flu results in decreased aGVHD severity compared to TBI. Moreover, Treo/Flu was associated with improved immune reconstitution despite the early toxicity.

DI-066 Analysis of the profile of candidates for bone marrow transplant and respective conditioning chemotherapy

BackgroundBone marrow transplant (BMT) involves intravenous transference of haematopoietic stem cells (HSC) from a donor to a recipient after the administration of chemotherapy, possibly in association with radiotherapy. The HSC...

16S rRNA Gene Pyrosequencing Reveals Shift in Patient Faecal Microbiota During High-Dose Chemotherapy as Conditioning Regimen for Bone Marrow Transplantation

Gastrointestinal disturbances are a side-effect frequently associated with haematological malignancies due to the intensive cytotoxic treatment given in connection with bone marrow transplantation (BMT). However, intestinal microbiota changes during chemotherapy remain poorly described, probably due to the use of culture-based and low-resolution molecular methods in previous studies. The objective of our study was to apply a next generation DNA sequencing technology to analyse chemotherapy-induced changes in faecal microbiota. We included eight patients with non-Hodgkin's lymphoma undergoing one course of BMT conditioning chemotherapy. We collected a prechemotherapy faecal sample, the day before chemotherapy was initiated, and a postchemotherapy sample, collected 1 week after the initiation of chemotherapy. Total DNA was extracted from faecal samples, denaturing high-performance liquid chromatography based on amplification of the V6 to V8 region of the 16S ribosomal RNA (rRNA) gene, and 454-pyrosequencing of the 16 S rRNA gene, using PCR primers targeting the V5 and V6 hypervariable 16S rRNA gene regions were performed. Raw sequence data were screened, trimmed, and filtered using the QIIME pipeline. We observed a steep reduction in alpha diversity and significant differences in the composition of the intestinal microbiota in response to chemotherapy. Chemotherapy was associated with a drastic drop in Faecalibacterium and accompanied by an increase of Escherichia. The chemotherapy-induced shift in the intestinal microbiota could induce severe side effects in immunocompromised cancer patients. Our study is a first step in identifying patients at risk for gastrointestinal disturbances and to promote strategies to prevent this drastic shift in intestinal microbiota.

How does bone marrow transplantation affect ovarian function and fertility?

This article sets out to review the literature on gonadal toxicity of bone marrow transplantation (BMT) in women, and preventive and curative actions available for girls and women facing BMT and infertility. The real incidence of ovarian damage after BMT is difficult to assess. Series reporting ovarian function and pregnancies after BMT vary greatly in terms of endpoints, number of patients included, age at and indications for BMT, conditioning regimens and duration of follow-up. The risk of premature ovarian failure (POF) is very high, however, and increases with age and in case of BMT conditioning treatment with total body irradiation. Embryo, oocyte and ovarian cortex cryopreservation are effective fertility preservation methods that should be discussed with all girls and young women requiring BMT. Despite a lack of literature on long-term follow-up of fertility after BMT in large populations, it is clear that the risk of ovarian damage and POF is very high. Fertility preservation options should therefore be discussed with all patients having to undergo BMT.

Strategy for bone marrow transplantation in eculizumab-treated paroxysmal nocturnal hemoglobinuria

Although the recent introduction of eculizumab has had a significant impact on the management of paroxysmal nocturnal hemoglobinuria (PNH), bone marrow transplantation (BMT) remains the only therapeutic option for patients who develop severe aplasia in the clinical course of PNH. However, information regarding BMT for eculizumab-treated PNH patients is scarce, and two major points-the optimal duration of eculizumab therapy, and the optimal BMT conditioning regimen-remain unclear. Here, we describe the clinical course of a PNH patient who was successfully treated with unrelated reduced-intensity BMT. Eculizumab was discontinued 2weeks prior to the initiation of the conditioning regimen, which consisted of fludarabine 180mg/m(2), cyclophosphamide 100mg/kg, rabbit anti-thymocyte globulin 2mg/kg, and TBI 3Gy. Complete donor chimerism was rapidly achieved in association with a rapid decrease in the proportion of PNH erythrocytes. The patient became transfusion-free immediately after BMT, and had no recurrence of hemolysis. The present case suggests that discontinuation of eculizumab before BMT and the use of a highly lymphoablative conditioning regimen may act as a successful treatment strategy in BMT for PNH. Further studies are warranted to evaluate the efficacy and safety of this treatment strategy.

Antithymocyte Globulin-induced Acute Lung Injury During Transplantation for Aplastic Anemia

A 10-year-old boy with acquired, very severe aplastic anemia developed acute lung injury after the administration of equine antithymocyte globulin, during conditioning for allogenic bone marrow transplantation. Limited cases of antithymocyte globulin-induced acute lung injury have been described in adults. The respiratory worsening was sudden and required mechanical ventilation. The clinical course was complicated by sepsis with Escherichia coli, vancomycin-resistant enterococci, and Stenotrophomonas maltophilia. Implications for treatment are discussed and earlier literature is reviewed.

Rapid Recovery of CD4 Counts Following Low Dose Alemtuzumab Conditioning During HLA Matched Related Bone Marrow Transplant for Children with Haemoglobinopathies

AbstractAbstract 2335CMV viraemia and disease remains a major cause of morbidity and mortality following allogeneic bone marrow transplantation (BMT). We investigated the impact of low dose alemtuzumab conditioning in paediatric matched sibling allogeneic BMT for haemoglobinopathy on immune reconstitution (early and late CD3/4+, CD3/8+, CD3-/CD56+ cell recovery) and subsequent viral infections in 18 consecutive transplants (14 β thalassaemia major and 4 sickle cell anaemia). A secondary aim was to investigate the impact of immune reconstitution on graft versus host disease but due to the extremely low incidence of acute and chronic graft versus host disease with this protocol (acute grade II to IV 11.6% and chronic 4.6%) we were unable to assess this. Transplant protocol: following hypertransfusion to suppress endogenous haemopoiesis, patients were conditioned with oral busulfan 14 mg/kg (days -9 to -6), cyclophosphamide 200 mg/kg (days -5 to -2) and alemtuzumab 0.3 mg/kg (days -8 to -6). Graft versus host disease prophylaxis was provided with ciclosporin for six months and methotrexate 10 mg/m2 on days +4 and +7. All transplants used BM as source of stem cells. CMV PCR surveillance was undertaken twice weekly and CMV reactivation was treated with intravenous foscarnet or ganciclovir, followed by secondary prophylaxis with valganciclovir. Routine peripheral blood immunophenotyping of lymphocyte subsets was performed at 60-, 90-, 120-, 150- and 180- days post-BMT. Results:the median age of transplant recipients was 7 years (range 2 – 17 years) and median follow up 254 days (range 203–378). The median CD3/CD4+ cell count rose above 200 cells/μl by day 180 and above 300 cells/μl by day 254 following the withdrawal of immune suppression at six months (fig 1A), the median CD8+ cell count increased earlier, to >200 cells/μl by day 60 and 300 cells/μl by day 180 (fig 1B) and the median CD56+ cell count reached 200 cells/μl by day 60 and plateaued thereafter (fig 1C). Overall, CMV reactivation occurred in 13 patients (72.2%) after a median interval of 35 days following BM infusion (range day-1 to day +72): 10/13 patients (76.9%) in whom both recipient and donor were CMV positive (CMV +/+); 1/3 patients in whom recipient was CMV negative and donor CMV positive (CMV -/+); and in 1/1 who was CMV positive with a CMV negative donor (CMV +/−). Significantly there was no clinical CMV disease in any patient and with the use of oral valganciclovir there were no subsequent CMV reactivations. CMV reactivation occurred in patients with lower CD4+ cell counts at day 60 (67 cells/μl) compared with the CMV-unaffected group (a median CD4+ count 298 cells/μl; p= 0.0262). Four patients experienced other viral infections: HHV6 infection requiring foscarnet on day +85 (n=1), urinary BK virus not requiring treatment (n=2) and parainfluenza pneumonitis requiring ribavarin on day +12 (n=1). Conclusion:Low dose alemtuzumab conditioning is associated with rapid CD4+ cell recovery (CD4+ >200 cells/μl by day +180) in children undergoing allogeneic matched related donor BMT for haemoglobinopathy. Although there is a high frequency of CMV reactivation (72.2%) in patients with low CD4+ counts at day 60 (median CD4+ count 67 cells/μl), no clinical CMV disease was seen and subsequent reactivations of CMV were prevented by the use of valganciclovir secondary prophylaxis whilst awaiting full immune reconstitution. [Display omitted] [Display omitted] [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Autologous Peripheral Blood Stem Cell Transplantation in Children with Refractory or Relapsed Lymphoma: Results of Children’s Oncology Group Study A5962

This prospective study was designed to determine the safety and efficacy of cyclophosphamide, BCNU, and etoposide (CBV) conditioning and autologous peripheral blood stem cell transplant (PBSCT) in children with relapsed or refractory Hodgkin and non-Hodgkin lymphoma (HL and NHL). Patients achieving complete remission (CR) or partial remission (PR) after 2 to 4 courses of reinduction underwent a granulocyte-colony stimulating factor (G-CSF) mobilized PBSC apheresis with a target collection dose of 5 × 10⁶ CD34(+)/kg. Those eligible to proceed received autologous PBSCT after CBV (7200 mg/m², 450-300 mg/m², 2400 mg/m²). Forty-three of 69 patients (30/39 HL, 13/30 NHL) achieved a CR/PR after reinduction. Thirty-eight patients (28 HL, 10 NHL) underwent PBSCT. All initial 6 patients who received BCNU at 450 mg/m² experienced grade III or IV pulmonary toxicity compared to none of the subsequent 32 receiving 300 mg/m² (P < .0001). The probability of overall survival (OS) at 3 years for all patients is 51% and for transplanted patients is 64%. The 3-year event-free survival (EFS) is 38% (45% for HL; 30% NHL). The 3-year EFS in transplanted patients is 66% (65% HL; 70% NHL). Initial duration of remission of ≥12 versus <12 months was associated with a significant increase in OS (3 years OS 70% versus 34%) (P = .003). BCNU at 300 mg/m(2) in a CBV regimen prior to PBSCT is well tolerated in relapsed or refractory pediatric lymphoma patients. A short duration (<12 months) of initial remission is associated with a poorer prognosis. Last, a high percentage of patients achieving a CR/PR after reinduction therapy can be salvaged with CBV and autologlous PBSCT.

Role of reduced intensity conditioning in T-cell and B-cell immune reconstitution after HLA-identical bone marrow transplantation in ADA-SCID

The treatment of choice for severe combined immunodeficiency is bone marrow transplantation from an HLA-identical donor sibling without conditioning. However, this may result in low donor stem cell chimerism, leading to reduced long-term immune reconstitution. We compared engraftment, metabolic, and T-cell and B-cell immune reconstitution of HLA-identical sibling bone marrow transplantation performed in 2 severe combined immunodeficiency infants with adenosine deaminase deficiency from the same family treated with or without a reduced intensity conditioning regimen (busulfan/fludarabine). Only the patient who received conditioning showed a stable mixed chimerism in all lineages, including bone marrow myeloid and B cells. The use of conditioning resulted in higher thymus-derived naïve T cells and T-cell receptor excision circles, normalization of the T-cell repertoire, and faster and complete B-cell and metabolic reconstitution. These results suggest the utility of exploring the use of reduced intensity conditioning in bone marrow transplantation from HLA-identical donor in severe combined immunodeficiency to improve long-term immune reconstitution.

Growth Velocity in Pediatric Bone Marrow Transplantation: Significance of Donor Type and Treatment Factors

Children who have undergone bone marrow transplantation (BMT) often have decreased growth. Growth is a multifactorial process, and the factors that influence growth after BMT are not completely understood. The authors hypothesized that donor type may be a factor influencing growth. Sixty-five children and adolescents who underwent BMT (32 related matched, 33 unrelated matched) were evaluated. Growth velocity (height standard deviation) was assessed prior to and 2 years following BMT. The results indicated that children and adolescents who underwent unrelated matched transplants had lower growth velocity (P < .059) than those with related matched transplants. Those who received the standard conditioning regimen that included total body irradiation (TBI) had a significantly lower growth velocity (P < .045) than those with chemotherapy-only regimens. Significant correlates of growth velocity included younger age at BMT and pre-BMT growth velocity. Thus, donor type, age at BMT, prior treatment, and BMT conditioning regimens that include TBI may all affect growth post-BMT. Careful monitoring of growth velocity is required for patients who have received an unrelated donor BMT.

High-dose mitoxantrone and etoposide conditioning in autologous bone marrow transplantation for relapsed Hodgkin's disease.

We administered high-dose mitoxantrone in combination with etoposide to 6 patients with relapsed Hodgkin's disease as the conditioning regimen for autologous bone marrow transplantation. This regimen was well tolerated and no significant cardiotoxicity was observed. Responses of the Hodgkin's disease to this therapy were favourable but short-lived. Serial measurements of the serum levels of mitoxantrone suggested an open 3-compartment model of drug distribution. The rapid early phase of drug distribution was followed by an intermediate phase and a slow terminal drug-elimination phase. However, mitoxantrone was still detected in the serum of all patients 7 days after the last dose of mitoxantrone and on the day of bone marrow re-infusion. The clinical significance of such findings is unclear but they may suggest a need for the use of other anthracycline-related cytotoxic agents for the conditioning in autologous bone marrow transplantation.

Cold agglutinin disease (CAD)with autoimmune haemolytic anaemia: a case report of a coronary artery disease patient

Cold agglutinin disease (CAD) with autoimmune haemolytic anemia is characterized by the production of harmful cold autoantibodies associated with increased red cell destruction during exposure to cold. The treatment of CAD is very difficult and a great effort is required to obtain therapeutic success. Cyclophosphamide is a potent immunosuppressive agent which is widely used in all bone marrow transplantation conditioning regimens for patients with acquired severe aplastic anemia. In this report, we describe the case of a coronary artery disease patient with severe CAD, but without lymphoproliferative disease, in which general measures and immunosuppressive therapies were adopted, there by avoiding blood transfusions.