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Fanconi anemia is an inherited disease characterized by congenital malformations, pancytopenia, cancer predisposition, and sensitivity to cross-linking agents. The molecular diagnosis of Fanconi anemia is relatively complex for several aspects including genetic heterogeneity with mutations in at least 16 different genes. In this paper, we report the mutations identified in 100 unrelated probands enrolled into the National Network of the Italian Association of Pediatric Hematoly and Oncology. In approximately half of these cases, mutational screening was carried out after retroviral complementation analyses or protein analysis. In the other half, the analysis was performed on the most frequently mutated genes or using a next generation sequencing approach. We identified 108 distinct variants of the FANCA, FANCG, FANCC, FANCD2, and FANCB genes in 85, 9, 3, 2, and 1 families, respectively. Despite the relatively high number of private mutations, 45 of which are novel Fanconi anemia alleles, 26% of the FANCA alleles are due to 5 distinct mutations. Most of the mutations are large genomic deletions and nonsense or frameshift mutations, although we identified a series of missense mutations, whose pathogenetic role was not always certain. The molecular diagnosis of Fanconi anemia is still a tiered procedure that requires identifying candidate genes to avoid useless sequencing. Introduction of next generation sequencing strategies will greatly improve the diagnostic process, allowing a rapid analysis of all the genes.

Epidemiologic investigation of invasive fungal diseases (IFDs) in allogeneic hematopoietic stem cell transplantation (allo-HSCT) may be useful to identify subpopulations who might benefit from targeted treatment strategies. The Gruppo Italiano Trapianto Midollo Osseo (GITMO) prospectively registered data on 1858 consecutive patients undergoing allo-HSCT between 2008 and 2010. Logistic regression analysis was performed to identify risk factors for proven/probable IFD (PP-IFD) during the early (days 0 to 40), late (days 41 to 100), and very late (days 101 to 365) phases after allo-HSCT and to evaluate the impact of PP-IFDs on 1-year overall survival. The cumulative incidence of PP-IFDs was 5.1% at 40 days, 6.7% at 100 days, and 8.8% at 12 months post-transplantation. Multivariate analysis identified the following variables as associated with PP-IFDs: transplant from an unrelated volunteer donor or cord blood, active acute leukemia at the time of transplantation, and an IFD before transplantation in the early phase; transplant from an unrelated volunteer donor or cord blood and grade II-IV acute graft-versus-host disease (GVHD) in the late phase; and grade II-IV acute GVHD and extensive chronic GVHD in the very late phase. The risk for PP-IFD was significantly higher when acute GVHD was followed by chronic GVHD and when acute GVHD occurred in patients undergoing transplantation with grafts from other than matched related donors. The presence of PP-IFD was an independent factor in long-term survival (hazard ratio, 2.90; 95% confidence interval, 2.32 to 3.62; P

We have conducted a long-term prospective study of children undergoing bone marrow transplantation (BMT) to assess morbidity and mortality for liver disease. One hundred eleven consecutive children were enrolled between June 1985 and June 1995 and were followed-up for a median of 5.5 years after BMT. Before transplant 48/111 children (43%) had abnormal alanine aminotransferase (ALT), none were HBsAg+ and 4/111 were anti-HCV+. After BMT 4/111 patients (3.6%) died of liver failure. No relationship was found between pretransplant hepatitis B (HBV) or C (HCV) infection or elevated transaminases and development of severe liver damage. Eighty-two out of one hundred and eleven patients (74%) had abnormalities of ALT after BMT, transient (n = 54) or persistent (n = 28). None developed clinical signs or symptoms of end stage liver disease or of cirrhosis during follow-up. No significant difference in prevalence of liver disease, was found between children with normal or abnormal ALT at BMT (relative risk [RR] = 1.04). HCV infection could be implicated in the etiology of chronic liver disease in 14/28 patients; 2 other patients were found infected by HBV alone (1 case) or combined with HCV (1 case). In the remaining 12 the etiology of chronic liver disease could not be defined. Posttransplant hepatitis B occurred in 4/111 children (3.6%), including a recipient from a donor who had been previously vaccinated against HBV, while no patient who had been vaccinated developed hepatitis B. The rate of posttransplant seroconversion to anti-HCV was 15%.

Immunosuppressive therapy (IST) with antithymocyte globulin and cyclosporin A (CyA) is the standard treatment for children with acquired aplastic anaemia (AAA) lacking a matched donor. Survival rates of more than 80% at 5 years are achieved, but the response is drug-dependent in 15-25% of cases. This study, of 42 consecutive children with AAA treated with IST, assessed the incidence of CyA-dependence, CyA and granulocyte colony-stimulating factor (G-CSF) tapering schedules and the impact of drug accumulation on progression to myelodysplasia/acute myeloid leukaemia (MDS/AML). Overall survival was 83% at 10 years. CyA-dependence without a predictive marker was observed in 18% of responders. Probability of discontinuing CyA was 60.5% at 10 years; a slow CyA tapering schedule was performed in 84% of patients; the cumulative incidence of relapse was 16% at 10 years. Relapse risk was significantly associated with rapid CyA discontinuation: 60% compared to 7.6% in the slow tapering group (P = 0.001). Cumulative incidence of MDS/AML was 8% at 10 years, with a significant correlation with both G-CSF cumulative dose and second IST. This long-term follow-up of children with AAA shows that IST with a slow CyA tapering course is an effective treatment with a low-relapse rate in these cases.

A new live-trap (named LOT trap) for small mammals is described herein. This trap is easily built and is considerably cheaper than commercial traps. The trap body is made of a PVC 8x8 cm pipe, with a tin door and a transparent plexiglas back door. The efficacy of this trap was tested in an experimental field study, and the results are presented and discussed. Capture rates were significantly different between types of trap, with 81.6% of captured rodents for LOT and 18.4% of rodents for multiple-capture traps.

Acquired aplastic anemia (AA) is a rare heterogeneous disease characterized by pancytopenia and hypoplastic bone marrow. The incidence is 2-3/million inhabitants/year, in Europe, but higher in East Asia. Survival in severe aplastic anemia (SAA) has markedly improved in the past 2 decades because of advances in hematopoietic stem cell transplantation, immunosuppressive and biologic drugs, and supportive care. In SAA hematopoietic stem cell transplant (HSCT) from a matched sibling donor (MSD) is the treatment of choice. If a MSD is not available, the options include immunosuppressive therapy (IST) or unrelated donor HSCT. The objective of this guideline is to provide healthcare professionals with clear guidance on the diagnosis and management of pediatric patients with AA. A preliminary, evidence-based document issued by a group of pediatric hematologists was discussed, modified and approved during a series of "Consensus Conferences" according to procedures previously validated by the AIEOP Board. The guidelines highlight the importance of referring pediatric patients with AA to pediatric centers with long experience in diagnosis, differential diagnosis, management, supportive care and follow-up of AA.

We have analyzed 1448 patients with acquired aplastic anemia grafted between 2005 and 2009, and compared outcome of identical sibling (n=940) versus unrelated donor (n=508) transplants. When compared to the latter, sibling transplants were less likely to be performed beyond 180 days from diagnosis (39% vs. 85%), to have a cytomegalovirus negative donor/recipient status (15% vs. 23%), to receive antithymocyte globulin in the conditioning (52% vs. 61%), and more frequently received marrow as a stem cell source (60% vs. 52%). Unrelated donor grafts had significantly more acute grade II-IV (25% vs. 13%) and significantly more chronic graft-versus-host disease (26% vs. 14%). In multivariate analysis, the risk of death of unrelated donor grafts was higher, but not significantly higher, compared to a sibling donor (P=0.16). The strongest negative predictor of survival was the use of peripheral blood as a stem cell source (P

We have previously shown that hemopoietic stem cell transplant (HSCT) recipients can be stratified on day+7 as having low, intermediate or a high risk of transplant-related mortality (TRM). With the aim of reducing TRM and GVHD, intermediate and high-risk patients (n = 170) were randomized to receive anti-thymocyte globulin (ATG, thymoglobuline) on day+7 (n = 84) or no treatment (n = 86) (controls). There was a reduction of TRM from 35% in controls to 29% in ATG patients (P = 0.3), of acute GVHD III-IV from 15 to 5% (P = 0.02) and of chronic GVHD from 26 to 11% (P = 0.03); survival was comparable. The predictive value of the day+7 score on TRM was confirmed for controls (19 vs 42% for intermediate vs high risk, respectively, P = 0.03), whereas ATG abrogated this predictive effect (29 vs 29%). ATG reduced GVHD (P = 0.006) in high-risk patients, but not in patients with an intermediate risk. In conclusion, we confirm that TRM can be predicted on the basis of day+7 laboratory values, after alternative donor HSCT; in high-, but not intermediate-risk patients, the administration of ATG on day+7 reduces GVHD. These results may represent a platform for risk-adapted post transplant immune modulation. Bone Marrow Transplantation (2010) 45, 385-391; doi:10.1038/bmt.2009.151; published online 6 July 2009

In view of the lack of guidance on the use of irradiated blood products (IBP) in patients with aplastic anaemia (AA) receiving immunosuppressive therapy (IST) with antithymocyte globulin (ATG) and ciclosporin, we conducted the first survey from European and USA centres to determine current practice on this issue. The routine use of pre-storage leucocyte-depleted blood products has reduced the risk of allosensitization in haematological malignancies, although data is lacking in AA [The Trial to reduce Alloimmunization to Platelets study group, 1997; Marsh et al, 2009]. Transfusion-associated graft-versus-host disease (TA-GVHD) is a rare but invariably fatal complication following transfusion of viable lymphocyte-containing blood products. It is abolished by the use of IBP in at-risk patients. For haematological patients, the routine use of IBP is recommended for all autologous and allogeneic haemopoietic stem cell tranplantation (HSCT) recipients, patients with congenital immune deficiency, Hodgkin lymphoma, and patients receiving purine analogues. In Germany, Non-Hodgkin lymphoma is also an indication for IBP [British Committee for Standards in Haematology (BCSH) Blood Transfusion Task Force, 1996; Williamson et al, 2007; Agbaht et al, 2007; The Board of the German Medical Association on the recommendation of the Scientific Advisory Board 2009]. Treatments with either ATG or Alemtuzumab (anti-CD52 monoclonal antibody) are also under consideration as an indication by the BCSH (J. Treleaven, Royal Marsden Hosptial, Sutton, UK, personal communication, 2009). The rationales for considering giving only IBP to patients with AA are (1) to prevent TA-GVHD during ATG treatment and (2) to help reduce allo-sensitization, which occurs frequently in AA patients (Killick et al, 1997; Laundy et al, 2004). Animal data show that irradiation of all red cell and platelet transfusions before HSCT reduces the risk of sensitization to minor histocompatibility antigens and the risk of graft rejection after dog leucocyte antigen-identical marrow grafts [Bean et al, 1994]. It is likely, but unproven, that the routine clinical use of leucocyte-depleted blood products has helped to reduce this risk. Leucodepletion should also reduce the chances of a recipient developing TA-GVHD. There were 13 cases of TA-GVHD reported to Serious Hazards of Blood Transfusion (SHOT) in the UK between 1996 and 2005, and of these, 11 occurred after transfusion of non-leucocyte depleted products and 2 after transfusion of leucocyte-depleted products; there have been no reported cases since 2001 [Williamson et al, 2007]. We were not aware of any published cases of TA-GVHD following ATG treatment for haematological disorders. We therefore performed an email survey of 12 European Group for Blood and Marrow Transplantation (EBMT) centres and two USA centres in 2008. Questionnaires were sent to lead representatives who were also representatives on the EBMT Severe Aplastic Anaemia Working Party (EBMT SAAWP) from the following countries: UK, Switzerland, Germany, Italy, Sweden, Netherlands, Italy and France. Two large centres in the USA were also included, The National Institutes of Health and Medical College of Wisconsin, on account of their specialist interest in AA. Both adult and paediatric centres were included in the survey. The questionnaire comprised the following questions: (1) Do you routinely give only IBP after ATG? (2) If so, how long for? (3) Which blood products do you irradiate? (4) What is/are your reason(s) for giving IBP? (5) Have you ever seen a case of TA-GVHD after ATG? (6) Do you have the same policy for children and (7) Any other comments to add? The results are summarized in Figure 1. Twelve of 14 centres (85%) routinely only give IBP after ATG. Of these, 6 of 12 centres indicated that they give IBP to all AA patients regardless of treatment. The reason for not giving IBP at 2 centres was lack of published evidence. There was no consensus on how long to continue IBP after IST. Two of 12 centres gave IBP indefinitely, 5 until patients became transfusion independent, two for 6 months, two for 12 months and one until the lymphocyte count was > 1.5 × 109/1. All of the 12 centres gave IBP to avoid TA-GVHD after ATG. Two centres had a universal IBP policy to avoid TA-GVHD for all haematology/oncology patients, to avoid possible errors of not giving IBP to patients known to be at risk. Only three centres indicated that reduction in allosensitization was another indication for giving IBP. Seven of 10 centres irradiated both red cell and platelet transfusions, and three irradiated red cells, platelets and fresh frozen plasma. Eight centres managed both children and adults with AA, and all reported the same policy for children and adults. A patient with severe AA was reported to have developed probable TA-GVHD 20 years ago after treatment with ATG. A further case was reported in a liver transplant recipient more than 10 years ago. No further details on these two cases of TA-GVHD were available. Figure 1 Results of survey on the use of irradiated blood products in aplastic anaemia patients treated with immunosuppressive therapy. The risk of TA-GVHD post-ATG is not known but appears to be low. This may reflect irradiation practices, and the use of leucodepleted blood products, or it may be truly low. However, horse ATG (Lymphoglobuline) was routinely used as treatment for AA, but was withdrawn from the market in 2007. Rabbit ATG is more immunosuppressive than horse ATG. It causes a more prolonged lymphopenia, higher peak Epstein-Barr virus (EBV) viral levels post-ATG for AA than horse ATG [Scheinberg et al, 2007], more EBV post-transplant lymphoproliferative disease after HSCT [Scheinberg et al, 2007]. Therefore, there is concern that rabbit ATG might increase the risk of TA-GVHD compared with horse ATG. Therefore, based on background information we have on the beneficial effect of IBP, and on the results of this survey, we propose that patients with AA should receive IBP during and after ATG treatment. This policy should probably be continued for at least as long as patients are receiving IST, such as ciclosporin.

We have analyzed 2,002 patients grafted in Europe between 1976 and 1998 from an identical twin (n = 34), from an HLA-identical sibling (n = 1,699) or from an alternative donor (n = 269), which included unrelated and family mismatched donors. The proportions of patients surviving in these three groups are, respectively, 91, 66 and 37%: major causes of failure were acute graft-versus host disease (GvHD) (11%), infection (12%), pneumonitis (4%), rejection (4%). In multivariate Cox analysis, factors predicting outcome were patient’s age (p < 0.0001), donor type (p < 0.0001), interval between diagnosis and bone marrow transplantation (BMT) (p < 0.0005), year of BMT (p = 0.0005) and female donor for a male recipient (p = 0.02). Patients were then divided in two groups according to the year of BMT: up to or after 1990. The overall death rate dropped from 43 to 24% (p < 0.00001). Improvements were seen mostly for grafts from identical siblings (from 54 to 75%, p < 0.0001), and less so for alternative-donor grafts (from 28 to 35%; p = 0.07). Major changes have occurred in the BMT protocol: decreasing use of radiotherapy in the conditioning regimen (from 35 to 24%; p < 0.0001) and increasing use of cyclosporin (with or without methotrexate) for GvHD prophylaxis (from 70 to 98%; p < 0.0001). In conclusion, the outcome of allogeneic BMT for patients with severe aplastic anemia has considerably improved over the past two decades: young patients, grafted early after diagnosis from an identical sibling, have currently an over 80% chance of long-term survival. Transplants from twins are very successful as well. The risk of complications with alternative donor transplants is still high.

The safety and efficacy of reduced-intensity conditioning (RIC) followed by allogeneic stem cell transplantation (SCT) for relapsed lymphomas remains unresolved. We conducted a prospective, multicentered, phase II trial. A total of 170 relapsed/refractory lymphomas received a RIC regimen followed by SCT from sibling donors. The primary study end point was non-relapse mortality (NRM). Histologies were non-Hodgkin's lymphomas (NHL) (indolent (LG-NHL), n=63; aggressive (HG-NHL), n=61; mantle cell lymphoma (MCL), n=14) and Hodgkin's disease (HD, n=32). Median follow-up was 33 months (range, 12-82). The results show that frequencies were as follows: cumulative NRM at 3 years, 14%; acute and chronic graft-versus-host disease (GVHD) 35 and 52%, respectively; 3-year overall survival (OS), 69% for LG-NHL, 69% for HG-NHL, 45% for MCL and 32% for HD (P=0.058); and 3-year relapse incidence, 29, 31, 35 and 81%, respectively (P

Background. Severe liver disease, including fulminant hepatic failure and venoocclusive disease can occur after bone marrow transplantation (BMT). The aim of our study was to assess risk factors for veno occlusive disease and severe liver disease occurring within 6 months from BMT. Methods. A total of 193 consecutive patients from 15 BMT Centers were prospectively enrolled between January and June 1995. Data on donors and recipients before and after transplant were collected and included age, gender, alanine aminotransferase (ALT), hepatitis B (HBV), and hepatitis C virus (HCV) markers, hematological disease, status and type of BMT, conditioning regimen and graft versus host disease prophylaxis. Statistical analysis included univariate descriptive and multivariate analysis based on logistic regression on major end-points. Results. Forty-three of 193 patients died during the study period, and liver disease was the main cause of death (13 of 43, 30%). Incidence of severe veno occlusive disease was 8%, fulminant hepatic failure 0.5% and 12% of cases had ALT >500 U/L (normal ≤42 U/L). A de novo HBV or HCV infection occurred in 3.2 and 7% of patients respectively. Predictive risk factors for life-threatening liver disease were: unrelated donors (relative risk=5.8, confidence interval=1.7-19.8) and abnormal BMT donor ALT (relative risk=6.3, confidence interval=1.5-25.5). Conclusions. This study indicates that ongoing or previous infection with HBV or HCV in donor or recipient is not an absolute contraindication for BMT. However, abnormal ALT levels in BMT donors were a significant predictor of potentially lethal liver complications. The occurrence of de novo HBV or HCV infection did not correlate with severity of liver disease observed in the first 6 months posttransplant. These findings should be carefully evaluated before disregarding HBV or HCV positive siblings with normal transaminase levels in favor of unrelated donors.

About 30% of patients with severe aplastic anaemia (SAA) unresponsive to one course of immunosuppressive (IS) therapy with antithymocyte or antilymphocyte globulin can achieve complete or partial remission after a second IS treatment. Among various second-line treatments, rabbit ATG (r-ATG) could represent a safe and effective alternative to horse ALG (h-ALG). In a multicentre study, 30 patients with SAA (17 males and 13 females, median age 21 years, range 2-67) not responding to a first course with h-ALG plus cyclosporin (CyA) and granulocyte colony stimulating factor (G-CSF), were given a second course using r-ATG (3.5 mg/kg/d for 5 d), CyA (5 mg/kg orally from day 1 to 180) and G-CSF (5 microg/kg subcutaneously from day 1 to 90). The median interval between first and second treatment was 151 d (range 58-361 d). No relevant side-effects were observed, but one patient died early during treatment because of sepsis. Overall response, defined as transfusion independence, was achieved in 23/30 (77%) patients after a median time of 95 d (range 14-377). Nine patients (30%) achieved complete remission (neutrophils/=2.0 x 109/l, haemoglobin/=11 g/dl and platelets/=100 x 109/l). The overall survival rate was 93% with a median follow-up of 914 d (range 121-2278). So far, no patient has relapsed. Female gender was significantly associated with a poorer likelihood to respond (P = 0.0006). These data suggest that r-ATG is a safe and effective alternative to h-ALG for SAA patients unresponsive to first-line IS treatment.

We report the results of the first prospective randomized multicenter study of immunosuppressive treatment in patients with previously untreated nonsevere aplastic anemia (AA) as defined by a neutrophil count of at least 0.5 × 109/L and transfusion dependence. Patients were randomized to receive cyclosporin (CSA) alone or the combination of horse antithymocyte globulin ([ATG] Lymphoglobuline; Merieux, Lyon, France) and CSA. The endpoint of the study was the hematologic response at 6 months. One hundred fifteen patients were randomized and assessable with a median follow-up period of 36 months; 61 received CSA and 54 ATG and CSA. In the CSA group, the percentage of complete and partial responders was 23% and 23%, respectively, for an overall response rate of 46%. A significantly higher overall response rate of 74% was found in the ATG and CSA group, with 57% complete and 17% partial responders (P = .02). Compared with CSA alone, the combination of ATG and CSA resulted in a significantly higher median hemoglobin level and platelet count at 6 months. Fewer patients required a second course of treatment before 6 months due to a nonresponse. In the CSA group, 15 of 61 (25%) patients required a course of ATG before 6 months because of disease progression, compared with only 3 of 54 (6%) in the ATG and CSA group. The survival probabilities for the two groups were comparable, 93% (CSA group) and 91% (ATG and CSA group), but at 180 days, the prevalence of patients surviving free of transfusions, which excluded patients requiring second treatment because of nonresponse, death, disease progression, or relapse, was 67% in the CSA group and 90% in the ATG and CSA group (P = .001). We conclude that the combination of ATG and CSA is superior to CSA alone in terms of the hematologic response, the quality of response, and early mortality, and a second course of immunosuppression is less frequently required.

Fanconi anemia (FA) is an autosomal recessive disease characterized by pancitopenia, congenital malformations, predisposition to cancers and chromosomal instability. We report the clinical and molecular features of a patient initially identified as a potential FA case only because of chemotherapy toxicity during the treatment of a T-lineage acute lymphoblastic leukemia (ALL). Cells from this patient showed a moderate chromosomal instability, increasing sensitivity to DNA crosslinking agents but normal response to ionizing radiation. The analysis of FA proteins demonstrated a marked reduction of FANCD2 (>95%), but normal levels of FANCA or FANCG. Interestingly, this defect was associated with a homozygous missense mutation of FANCD2, resulting in a novel amino-acid substitution (Leu153Ser) at residue Leu153, which is highly conserved through evolution. The FANCD2(L153S) protein, whose reduced expression was not due to impaired transcription, was detected also in its monoubiquitinated form in the nucleus, suggesting that the mutation does not affect post-translation modifications or subcellular localization but rather the stability of FANCD2. Therefore, the hypomorphic Leu153Ser mutation represents the first example of a FANCD2 defect that might promote clonal progression of tumors, such as T-ALL, and severe chemotherapy toxicity in patients without any clinical manifestations typical of FA.

Fluorescence in situ hybridization and comparative genomic hybridization characterized 6p rearrangements in eight primary and in 10 secondary myeloid disorders (including one patient with Fanconi anemia) and found different molecular lesions in each group. In primary disorders, 6p abnormalities, isolated in six patients, were highly heterogeneous with different breakpoints along the 6p arm. Reciprocal translocations were found in seven. In the 10 patients with secondary acute myeloid leukemia/myelodysplastic syndrome (AML/MDS), the short arm of chromosome 6 was involved in unbalanced translocations in 7. The other three patients showed full or partial trisomy of the 6p arm, that is, i(6)(p10) (one patient) and dup(6)(p) (two patients). In 5/7 patients with unbalanced translocations, DNA sequences were overrepresented at band 6p21 as either cryptic duplications (three patients) or cryptic low-copy gains (two patients). In the eight patients with cytogenetic or cryptic 6p gains, we identified a common overrepresented region extending for 5-6 megabases from the TNF gene to the ETV-7 gene. 6p abnormalities were isolated karyotype changes in four patients. Consequently, in secondary AML/MDS, we hypothesize that 6p gains are major pathogenetic events arising from acquired and/or congenital genomic instability.

Purpose Older age and a previously failed autologous stem-cell transplantation (SCT) are poor prognostic factors for patients receiving myeloablative conditioning and allogeneic SCT. Reduced-intensity conditioning (RIC) regimens achieved a significant reduction of treatment-related mortality, but the influence of previously described risk factors on the outcome of this novel transplantation strategy have not been fully analyzed yet. Patients and Methods One hundred fifty patients with advanced hematologic malignancies received a RIC regimen containing thiotepa (10 mg/kg), fludarabine (60 mg/m2), and cyclophosphamide (60 mg/kg), followed by an allogeneic transplantation from an HLA-identical sibling donor. Patients were divided into two cohorts according to age; 90 patients were younger than 55 years, and 60 patients were 55 years old or older. The other pretransplantation characteristics were fairly balanced. Results Actuarial 5-year nonrelapse mortality (NRM) rate was not statistically different between the groups (13% in the younger group and 19% in the older group). By univariate and multivariate analysis, NRM was significantly higher in older patients who previously experienced failure with an autograft. The occurrence of grade 3 to 4 acute graft-versus-host disease (GVHD) or extensive chronic GVHD was associated with a higher NRM in both age cohorts. Overall survival (OS) was not statistically different between the younger (66%) and older groups (61%). By multivariate analysis, refractory disease was associated with a worse OS irrespective of age group. Conclusion RIC transplantations show a rather low NRM, and age ≥ 55 years per se cannot be considered a risk factor anymore. The timing of transplantation and novel strategies for the prevention of severe GVHD could further improve patient outcome.

Previous studies suggested a link between the use of G-CSF and increased incidence of myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) after immunosuppressive therapy (IST) for severe aplastic anemia (SAA). This European survey included 840 patients who received a first-line IST with (43%) or without (57%) G-CSF. The incidences of MDS/AML in patients who did or did not receive G-CSF were 10.9% and 5.8%, respectively. A significantly higher hazard (1.9) of MDS/AML was associated with use of G-CSF. Relapse of aplastic anemia was not associated with a worse outcome in patients who did not receive G-CSF as first therapy, whereas relapse was associated with a significantly worse outcome in those patients who received G-CSF. These results emphasize the necessity of the current European randomized trial comparing IST with or without G-CSF and to alert physicians that adding G-CSF to IST is currently not standard treatment for SAA.

The aim of this study was to ascertain prevalence and natural history of hepatitis C virus (HCV) infection in a large cohort of patients cured of childhood leukemia who had been followed prospectively for liver disease for at least 10 years since chemotherapy withdrawal: 114 consecutive patients entered the study. Liver function tests and ultrasonography were used to assess presence of liver disease. Patients were tested for antibody to HCV and for serum HCV-RNA at the end of chemotherapy and at the end of follow-up. At chemotherapy withdrawal, 56 patients (49%) were HCV-RNA positive, often without detectable anti-HCV, and in these cases, transaminase levels were more elevated during (P = .08) and after (P = .04) chemotherapy compared with HCV-RNA negative cases. Patients were then followed-up 13 to 27 years (mean, 17) after chemotherapy withdrawal. During this period, 38 initially anti-HCV negative patients seroconverted to anti-HCV and 17 initially anti-HCV positive cases lost reactivity. Forty patients were persistently HCV-RNA positive in serum, while 16 initially viremic patients became HCV-RNA negative during follow-up. At the end of the observation period, a persistent transaminase elevation was detected only in four HCV-RNA positive and anti-HCV positive cases, while no patient developed signs or symptoms of decompensated liver disease. Thus, hepatitis C was a frequent finding in long-term survivors after chemotherapy. It was associated with an atypical serologic profile and did not cause severe liver impairment over a period of 13 to 27 years.

Hepatitis C virus (HCV)-infection is common in persons with leukemia. Blood transfusions are the typical source and HCV-related chronic hepatitis a common outcome. Development of HCV-infection and -related hepatitis raises important questions about subsequent leukemia treatment including the natural history of the infection and need for treatment modification. Although the natural history of HCV-infection and -related hepatitis in this setting is unknown, data from normal persons with HCV-infection suggest that short-term survival is not likely to be decreased in persons with leukemia and these complications. In contrast, long-term survival may decrease because of a high rate of chronic hepatitis, cirrhosis, and possibly hepatocellular carcinoma. There are no convincing data that HCV-infection or -related hepatitis or alterations in anti-leukemia drug dose or schedule prompted by abnormal liver function tests, alter leukemia outcome. Consequently, it is uncertain whether drug doses and/or schedule should be modified in persons with leukemia and HCV-infection or -related hepatitis. Short-term outcome of blood cell and bone marrow transplants is also unaffected by HCV-infection or -related hepatitis.

Allogeneic hematopoietic stem cell transplantation (HSCT) represents the only curative treatment for patients with myelodysplastic syndrome (MDS), but involves non-negligible morbidity and mortality. Registry studies have shown that advanced disease stage at transplantation is associated with inferior overall survival. To define the optimal timing of allogeneic HSCT, we carried out a decision analysis by studying 660 patients who received best supportive care and 449 subjects who underwent transplantation. Risk assessment was based on both the International Prognostic Scoring System (IPSS) and the World Health Organization classification-based Prognostic Scoring System (WPSS). We used a continuous-time multistate Markov model to describe the natural history of disease and evaluate the effect of allogeneic HSCT on survival. This model estimated life expectancy from diagnosis according to treatment policy at different risk stages. Relative to supportive care, estimated life expectancy increased when transplantation was delayed from the initial stages until progression to intermediate-1 IPSS-risk or to intermediate WPSS-risk stage, and then decreased for higher risks. Modeling decision analysis on WPSS versus IPSS allowed better estimation of the optimal timing of transplantation. These observations indicate that allogeneic HSCT offers optimal survival benefits when the procedure is performed before MDS patients progress to advanced disease stages. Am. J. Hematol. 88:581–588, 2013. © 2013 Wiley Periodicals, Inc.

Hepatitis B virus (HBV)-infected individuals are occasionally used as donors for bone marrow transplantation (BMT). We studied the rate of HBV infection and the clinical expression of the associated liver disease in patients receiving marrow from HBsAg+ donors. We performed a retrospective survey in 14 BMT units in Italy in which all BMTs performed between 1984 and 1994 were reviewed and those involving HBsAg+ donors were identified. Donors and recipients were analyzed for HBV markers and liver disease. A total of 24 of 2,586 patients (0.9%) had received an HBsAg+ marrow. HBsAg became detectable in 22% of pre-BMT HBsAg-patients, but only 5.5% became chronic HBsAg carriers. Antigenemia developed more frequently in anti-HBs-compared with anti-HBs+ patients independently of passive prophylaxis with hyperimmune anti-HBs Ig, although the difference was not significant. Severe liver failure with death occurred in 21% of patients, which was a value greater than that generally observed after BMT in our units (3.7%). Patients with an anti-HBe+ donor had higher frequency of liver failure (28% v 0%) and alanine aminotransferase peaks as compared with those of patients with an HBeAg+ donor. Liver failure was not observed in anti-HBs+ recipients. The use of HBsAg+ donors, particularly if anti-HBe+, increases the risk of severe liver disease in BMT recipients. Anti-HBs positivity may prevent severe liver damage.

This review summarises serologic profiles and clinical features of HCV infection in children with leukemia. The diagnosis of HCV infection is currently based on the detection in serum of antiviral antibodies and HCV-RNA. Studies on leukemic children, however, have clearly shown that only HCV-RNA testing correctly identifies HCV infection, as specific antibodies become detectable in serum only after chemotherapy withdrawal in almost all cases. Viraemia instead appears early in the course of leukemia, and infected patients become carriers. The pattern of liver disease is rather homogeneous. Early onset, persisting ALT elevation during chemotherapy often with drastic reduction during high-dose chemotherapy, followed by sharp exacerbations of liver cell necrosis. ALT normalise after chemotherapy withdrawal in most cases, despite persisting viraemia. As regards to the ultimate prognosis of liver disease in these children, we have observed that, among a series of 119 patients followed for at least 10 years off-therapy, none has developed clinical decompensated liver disease and only 6% still has abnormal ALT levels. On the other hand, the need for prolonged follow-up of children surviving leukemia with chronic HCV infection is emphasized by the fact that the natural history of HCV infection has not been fully clarified and that additional late sequelae are likely to occur.

Bone marrow has been shown to be superior to peripheral blood, as a stem cell source, in young patients (20 years of age) with acquired aplastic anemia undergoing a matched sibling transplant. The aim of this study was to test whether this currently also holds true for older patients with acquired aplastic anemia.We analyzed 1886 patients with acquired aplastic anemia who received a first transplant from a human leukocyte antigen identical sibling between 1999 and 2009, with either bone marrow (n=1163) or peripheral blood (n=723) as the source of stem cells.In multivariate Cox analysis negative predictors for survival were: patient's age over 20 years (RR 2.0, P0.0001), an interval between diagnosis and transplantation of more than 114 days (RR 1.3, P=0.006), no anti-thymocyte globulin in the conditioning (RR 1.6, P=0.0001), a conditioning regimen other than cyclophosphamide (RR=1.3, P=0.008) and the use of peripheral blood as the source of stem cells (RR 1.6, P0.00001). The survival advantage for recipients of bone marrow rather than peripheral blood was statistically significant in patients aged 1-19 years (90% versus 76% P0.00001) as well as in patients aged over 20 years (74% versus 64%, P=0.001). The advantage for recipients of bone marrow over peripheral blood was maintained above the age of 50 years (69% versus 39%, P=0.01). Acute and chronic graft-versus-host disease were more frequent in peripheral blood transplants. Major causes of death were graft-versus-host disease (2% versus 6% in bone marrow and peripheral blood recipients, respectively), infections (6% versus 13%), and graft rejection (1.5% versus 2.5%).This study shows that bone marrow should be the preferred stem cell source for matched sibling transplants in acquired aplastic anemia, in patients of all age groups.

This prospective study was initiated in 1993 with the aim to study late effects and responses to antiviral therapy in a cohort of hepatitis C virus (HCV)-infected patients. A total of 195 patients were included from 12 centers. In all, 134 patients had undergone allogeneic and 61 autologous hematopoietic SCT (HSCT). The median follow-up from HSCT is currently 16.8 years and the maximum 27.2 years. Overall 33 of 195 patients have died of which 6 died from liver complications. The survival probability was 81.6% and the cumulative incidence for death in liver complications was 6.1% at 20 years after HSCT. The cumulative incidence of severe liver complications (death from liver failure, cirrhosis and liver transplantation) was 11.7% at 20 years after HSCT. In all, 85 patients have been treated with IFN; 42 in combination with ribavirin. The sustained response rate was 40%. The rates of severe side effects were comparable to other patient populations and no patient developed significant exacerbations of GVHD. Patients receiving antiviral therapy had a trend toward a decreased risk of severe liver complications (odds ratio=0.33; P=0.058). HCV infection is associated with morbidity and mortality in long-term survivors after HSCT. Antiviral therapy can be given safely and might reduce the risk for severe complications.

The pattern of hepatitis C virus (HCV) serum markers and liver disease was investigated in 11 leukemic children showing anti-HCV reactivity at least once during long-term observation to define the role of HCV infection and the behavior of HCV serologic markers in this patient cohort. Antibodies to HCV by first- and second-generation enzyme-linked immunosorbent assay (ELISA) and by second-generation (four antigens) recombinant immunoblotting assay (RIBA) and HCV-RNA by nested polymerase chain reaction (PCR) were serially examined in serum. Liver disease was defined according to transaminase levels. Seven of 11 patients were found HCV-RNA positive during chemotherapy and after blood transfusion, 3 of 11 became viremic during follow-up, and 1 of 11 was always HCV-RNA negative. Seroconversion to anti-HCV positivity by second-generation ELISA occurred in all the HCV-RNA positive children either during or after chemotherapy. Alanine aminotransferase (ALT) levels were elevated in all the HCV-RNA positive patients during antileukemic treatment and normalized in seven of them after therapy withdrawal, despite persisting viremia. These results indicate that HCV- RNA testing by polymerase chain reaction is required to correctly identify HCV infection in patients with leukemia while on chemotherapy. Viremia did not correlate with ALT levels and anti-HCV patterns.

134 patients with acquired aplastic anaemia (AA) were given HALG 15 mg/kg/d for 5 d and methylprednisolone for 1 month, and randomized to receive (n = 69) or not (n = 65) oxymetholone 2 mg/kg/d p.o. daily for 4 months. Early mortality (120 d) was comparable in the two arms 12/69 (17%) and 11/65 (17%), and correlated with the severity of the disease (39%, 10% and 6% respectively in patients with neutrophil counts (PMN)0.2, 0.2-0.5,0.5 x 10(9)/l). The response rate at 120 d was significantly greater in patients receiving androgens (56% v 40%; P = 0.04); it was 68% v 48% (P = 0.02) in patients surviving 120 d, and 78% v 27% (P = 0.03) in females with PMN less than 0.5 x 10(9)/l. In a multivariate Cox analysis on patients with less than 0.5 x 10(9)/l PMN, the probability of responding without androgens was reduced compared to the androgen treatment arm (P = 0.05). Survival was comparable in the two groups (71% v 65%). It was superior (74% v 50%), but not significantly (P = 0.1) in females with PMNor = 0.5 x 10(9)/l receiving androgens. Side-effects, including biochemical abnormalities and virilization, could be controlled and were reversible. In conclusion, the addition of androgens to HALG and methylprednisolone as first line treatment of aplastic anaemia significantly improves the response rate at 4 months, particularly in females with low neutrophil counts, although there is no significant effect on short-term survival. The reversible side-effects warrant the use of androgens as an adjunct to the first course of ALG in females with severe AA.