Matched Sibling Bone Marrow Transplantation Research Articles

Reduced toxicity matched sibling bone marrow transplant results in excellent outcomes for severe congenital neutropenia.

Severe congenital neutropenia (SCN) is caused by germline mutations, most commonly in ELANE, impacting neutrophil maturation and leading to high risk of life-threatening infections. Most patients with ELANE-mutant SCN can achieve safe neutrophil counts with chronic Granulocyte-Colony Stimulating Factor (G-CSF). However, up to 10% of patients have neutropenia refractory to G-CSF and require allogeneic stem cell transplant. Traditional conditioning for these patients includes busulfan and cyclophosphamide which is associated with significant toxicities. We present five patients with SCN without myeloid malignancy transplanted using a reduced toxicity regimen of busulfan, fludarabine and thymoglobulin. 5 pediatric patients with SCN underwent matched sibling donor bone marrow transplant (MSD-BMT) between 2014-2022 on or per CHP14BT057 (NCT02928991), a prospective, single center trial testing elimination of cyclophosphamide from conditioning in pediatric patients with single lineage inherited BMF syndromes. All patients had MSDs and no evidence of MDS. Conditioning consisted of PK-adjusted busulfan, fludarabine, and thymoglobulin, with calcineurin inhibitor and mycophenolate mofetil GVHD prophylaxis. With median follow-up of 48.4 months, overall and event-free survival were 100%. There was no acute GVHD and one instance of chronic limited GVHD. Patients exhibited >95% donor myeloid chimerism at 5 years post-BMT. Two patients experienced CMV reactivation without end-organ disease, and no other viral reactivation or significant infections occurred. MSD-BMT with reduced toxicity myeloablation for SCN provides excellent outcomes while minimizing toxicity. These data suggest that busulfan, fludarabine, and ATG can be considered an efficacious, low-toxicity standard of care regimen for patients with SCN undergoing MSD-BMT.

Chronic granulomatous disease identified in the evaluation of atypical Kawasaki disease in an infant

An 8-week-old full-term male was admitted after 5 days of fever and cough. He was previously admitted at 3 weeks of age for a Klebsiella pneumoniae urinary tract infection (UTI) and treated with 10 days of antibiotics. Blood and cerebrospinal fluid cultures, serum inflammatory markers, and respiratory viral testing obtained at that time were negative. During the current admission, urine cultures grew Klebsiella oxytoca and he was again treated for a UTI.His fevers persisted and on day 8 of illness he met criteria for atypical Kawasaki disease with elevated inflammatory markers, elevated liver enzymes, thrombocytosis, leukocytosis, and anemia. Prior to initiating IVIG, the patient underwent abdominal ultrasound to rule out occult infection, revealing multiple liver and spleen abscesses. Aspirated fluid grew Serratia marcescens and he was started on cefepime. Additional imaging of the chest, abdomen, and pelvis revealed pulmonary nodules. Evaluation of these nodules demonstrated no additional mycobacterial, fungal, or bacterial infections.Chronic granulomatous disease (CGD) was suspected based on the patient’s age, multi-organ abscesses with Serratia marcescens, and parental consanguinity (first cousins). Neutrophil oxidative burst testing showed absent ROS production. Genetic testing demonstrated a homozygous pathogenic variant in NCF2 (exon 3 deletion), confirming the diagnosis of autosomal-recessive CGD. He received high-dose methylprednisolone with a prolonged taper to promote antibiotic penetration of his CGD-associated liver abscesses. Repeat imaging showed interval improvement and the patient began prophylactic itraconazole and trimethoprim-sulfamethoxazole.Familial testing revealed heterozygous carrier status of the same NCF2 variant in both parents and the patient’s two sisters. At 10 months of age, the patient underwent a 10/10 matched sibling bone marrow transplant. Chimerism studies showed rapid neutrophil engraftment with delayed T cell engraftment. As of follow up at 12 months of age, he has appropriate post-transplant weight gain, no infectious complications, and no evidence of GVHD.Initial presentation in patients with CGD and liver/spleen abscesses can be insidious with isolated fever and elevated inflammatory markers. Such patients can readily meet criteria for atypical Kawasaki disease, highlighting the importance of maintaining a broad differential diagnosis for fever of unknown origin with a high index of suspicion for primary immunodeficiency.

Reduced-Intensity Conditioning Is Successful in Matched Sibling Bone Marrow Transplantation As a Cure for Sickle Cell Disease

Introduction: Sickle cell disease (SCD) is the most common genetic disease in this country, affecting 1 in every 72,000 individuals in the United States, with sickle cell trait found in 1 in 12 African-Americans. Without curative therapy, disease related mortality is as high as 14% in adolescents and young adults, with a decreased life expectancy compared to individuals without SCD. Hematopoietic stem cell transplantation (HSCT) is the only established cure, and matched sibling HCST have been performed for 30 years with great success. Unfortunately, only about 14% of SCD patients have a matched sibling. Most SCD HSCT protocols use myeloablative (MAB) conditioning in matched sibling transplants with average overall survival (OS) and disease-free survival (DFS) of 90% and 95% respectively, but with potential long-term complications including infertility following its use. Reduced Intensity Conditioning (RIC) has been used in young patients with malignancies with better tolerance and fewer complications. In an attempt to minimize risk and maximize cure, our study examined the use of a RIC regimen for both matched sibling and alternative donor HSCT in SCD patients ages 2-30.Primary Objective: To determine DFS after matched sibling or unrelated transplant after a conditioning regimen consisting of Alemtuzumab, Fludarabine, and Melphalan.Methods: The Institutional Review Board of Hackensack University Medical Center approved this study for human subjects. Strict stopping rules were established for graft rejection and death. Eligibility criteria included a diagnosis of SCD for patients receiving matched sibling donor HSCTs, but did not require any specific complications. For alternative donor HSCTs, eligibility criteria included specific complications associated with severe disease, and required a 9-10/10 unrelated donor or 5-6/6 UCB in the donor registry. All patients underwent partial exchange transfusions for a goal of HbS <45% at the start of conditioning. Conditioning initially consisted of Alemtuzumab beginning on d -22 followed by Fludarabine and Melphalan. However, Alemtuzumab was moved up to d -16 after 7 patients were enrolled to reduce graft vs host disease (GVHD). Due to increasing host chimerisms in 3 patients, the Alemtuzumab was moved back to d-22 after an additional 7 patients were enrolled. Post-HSCT, GVHD prophylaxis consisted of Tacrolimus (trough goal of 10-20) through d+100, mycophenolate mofetil through d+45, and steroids (for unrelated donors) through d+28. Patients were monitored closely for signs of infection with weekly PCRs for adenovirus, CMV and EBV. Donor chimerism by STR and HbS percentages by electropheresis were assessed at d+30, d+60, d+100, 1 year and 2 years post HSCT.Results: A total of 20 patients (17 related, 3 unrelated) were enrolled, with age at transplant ranging from 2.25 to 25.4 years. 3 patients were initially consented to the related arm, but then were found to be ineligible due to organ dysfunction or compliance issues. OS and DFS in patients with related donors were 100% and 94%, with one patient suffering secondary graft rejection. The OS and DFS in patients with unrelated donors were 66% and 33%, with one patient dying from GVHD complications and a second patient suffering secondary graft rejection. The unrelated patient with graft failure did receive a second HSCT off study, but died due to GVHD complications. Mean neutrophil engraftment occurred at d+14 and d+11 and platelet engraftment occurred at d+40 and d+26 for related and unrelated donors, respectively. The incidence of acute GVHD stage II-IV was 35% (7/20 patients), occurring in 6/17 (35%) related and 1/3 (33%) unrelated BMT patients. Chronic GVHD was noted in 20% of patients (4/20), occurring in 3/17 (18%) related and 1/3 (33%) unrelated BMT patients. Chimerisms ranged from 20-100% with HbS% of 0-62%.Conclusions : Our results show that a RIC regimen is well-tolerated with successful engraftment in patients undergoing matched sibling HSCT. However, the complications of GVHD & graft rejection in unrelated transplants greatly affected the DFS. The results are promising for continued use of this regimen for related HSCT even in SCD patients with minimal complications, but other alternatives need to be developed for unrelated HSCT. DisclosuresNo relevant conflicts of interest to declare.

Impact of Splenomegaly on Outcomes of Fully Matched Sibling Bone Marrow Transplant Among Children with Low Risk Thalassemia

Objective:Severe thalassemia syndromes (ST) are the most common non-infectious life-threatening disease in South Asia. ST is highly curable by BMT but rejection may occur in 5 to 30% of cases. In a limited resource setting appropriate case selection for BMT and aggressive down staging pre-transplant are crucial in order to achieve optimal outcomes. Though hepatomegaly > 2 cm has clearly been proven to be associated with poor transplant outcomes whereas the role of splenomegaly as an independent risk factor for rejection in low risk thalassemia has not been clarified.Methods:We analyzed a total of 87 consecutive first bone marrow transplants carried out between August 2013 and Dec 2016 after conditioning with busulfan (14 mg/kg oral, not adjusted to serum levels), cyclophosphamide (200 mg/kg) and anti-thymocyte globulin (Genzyme 4 mg/kg or Fresenius 16 mg/kg on days -12 to -10). 74 patients received cyclosporine and methotrexate, and 13 patients received cyclosporine and mycophenolate as GVHD/rejection prophylaxis. G-CSF-primed bone marrow (5 μg/kg/dose twice daily for 3 to 5 days prior to harvest) was the source of hematopoietic stem cells in all cases. All patients were considered low risk based on liver size < 2 cm from costal margin and age less than 15 years (median 6.9 years, range 1.1 to 14.5). All sibling couples were HLA-compatible.Results:Of the 87 low risk transplants 12 children rejected (13.8%) of which 5 were primary and 7 were secondary graft failures. Out of the total 87 patients 11 patients had significant splenomegaly at transplant (spleen > 3 cm below costal margin and/or z score > 3 spleen size measured by ultrasound adjusted for height). Four of the rejections (1 primary and 3 secondary) occurred in children with significant splenomegaly which is statistically significant (log rank p=0.02) (figure 1). Splenomegaly also seems to increase treatment-related mortality albeit not statistically significant (log rank p=0.17). Out of the 7 children who had treatment related mortality 2 had splenomegaly. One of them died as a consequence of primary graft failure and the other due to a large intracranial bleed. We could not detect any difference in other variables potentially associated with rejection risk such as gender mismatch, serum ferritin, ABO mismatch, mean cell dose (median 6.75; range 2.6 to 29.44 x 108 nucleated cells /kg) and immunosuppressive therapy between splenomegaly and no splenomegaly groups. In the latter group 41% of children received hydroxyurea pre-BMT compared to 81% in the splenomegaly group (P= 0.01) which in fact should have helped to reduce rejection rates in the splenomegaly group. By multivariate analysis significant splenomegaly (p= 0.0049) seems to be more relevant than gender mismatch in terms of rejection risk (p = 0.0379), while cell dose (p=0.9546) was not a significant factor.Conclusion:Splenomegaly may increase rejection risk in low risk thalassemia transplants transplanted with ATG-BuCy. Strategies to reduce spleen size prior to transplant should be considered. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Moyamoya Syndrome in Children with Sickle Cell Disease: A 10-Year Single-Center Experience

▪Background: Moyamoya syndrome (MMS) is a relatively uncommon vascular complication of sickle cell disease (SCD) characterized by progressive stenosis of the supraclinoid carotid arteries and development of typical collaterals. MMS confers a significant lifetime risk of ischemic stroke, and frequently results in multiple cerebrovascular events leading to poor outcomes. Management of MMS in this population is challenging, and to date no clinical trials have evaluated the long-term neurologic outcome of treatment with chronic transfusion versus surgical intervention [Encephaloduroarteriosynangiosis (EDAS) or Encephalomyoarteriosynangiosis (EMAS)]. Determination of efficacy and safety of various interventions and information about long-term progression could guide treatment decisions and potentially support the establishment of treatment guidelines. We report clinical and radiographic characteristics and neurological outcomes in children with SCD and MMS treated at our center over the last 10 years.Methods: We conducted a retrospective chart review of all patients with SCD who were diagnosed with MMS at our center from 2006 to 2016. Cases were identified utilizing radiology data mining software (MONTAGETM Search and Analytics). The radiological images and reports of potential MMS cases were independently reviewed by a hematologist, a radiologist and a neurologist to confirm the diagnosis of MMS. Relevant clinical data were collected and summarized using descriptive statistics.Results: During the study period, 542 unique patients with SCD required neuroimaging at our institution. Indications for neuroimaging included concerns for stroke, abnormal TCD, headaches or follow up imaging. Out of the 542 patients, 23 patients (4%) [18 males] were diagnosed with MMS. The median age at time of diagnosis of MMS was 6 years [Interquartile range (IQR) 4-8]. The genotype was SS in 20 patients, S-beta0 thalassemia in 2, and SC in 1. Of the 23 patients with MMS, 16 patients (70%) were diagnosed with MMS after initially presenting with a clinically overt ischemic stroke, while 7 patients (30%) were diagnosed following an abnormal or conditional surveillance Transcranial Doppler (TCD) study. MMS was bilateral in 16 patients (70%) and unilateral in the remaining 7 patients (30%). Prior to diagnosis, baseline laboratory values were (median and IQR): hemoglobin (Hb) 7.5 g/dL [IQR 6.7-8]; absolute reticulocyte count 333 k/mL [IQR 227-414]; white blood cell count 14.6 k/mL [IQR 12-18]; platelet count 371 k/mL [IQR 282-424]; total bilirubin 3.25 mg/dL [ IQR 2-4.45]; lactate dehydrogenase 522 u/L [IQR 426-862]; and HbF 6.6% [IQR 3.4-17.1]. Of the 16 patients who presented with clinically overt ischemic stroke, 11 patients (69%) experienced recurrent ischemic events (mean 2 strokes/patient). After diagnosis of MMS, 19/23 patients (82%) were started on chronic monthly transfusions aimed at maintaining HbS level below 30%. Nine patients (39%) were treated with indirect surgical revascularization procedure performed at a median of 17 months [IQR3.5-36] after MMS diagnosis. Two patients received matched sibling bone marrow transplantation. Two patients died after suffering a massive stroke.Conclusion: Children with SCD who develop MMS are at significant risk of recurrent strokes and poor neurologic outcomes. Optimal management remains unclear but should include consideration for urgent surgical revascularization procedures. The most effective treatment as well as the optimal time for revascularization procedure should be addressed by a large multi-center trial. Future studies should also evaluate mechanisms underlying MMS. DisclosuresNo relevant conflicts of interest to declare.

Outcomes of allogeneic transplantation for hemoglobin Bart's hydrops fetalis syndrome in Hong Kong.

Hemoglobin Bart's hydrops fetalis syndrome (BHFS) was once considered a fatal condition universally. Medical advances over the past three decades have resulted in increasing numbers of BHFS survivors. This retrospective review summarized local territory-wide experience and outcomes of BHFS patients who received allogeneic hematopoietic stem cell transplantation (HSCT) in Hong Kong. All BHFS patients who underwent allogeneic HSCT in Hong Kong, either in one of the two former pediatric transplant centers (Queen Mary Hospital and Prince of Wales Hospital) on or before 2019 or in the single territory-wide pediatric transplant center (Hong Kong Children's Hospital) since 2019, from January 1, 1996, till December 31, 2020, were included. Basic demographic data, perinatal history, transplant details, long-term outcomes, and morbidities were reviewed. Total five allogeneic HSCT were performed in two males and three females at a median age of 22months, which include one 8/8 matched-sibling bone marrow transplant, one 5/6 matched-sibling cord blood transplant with HLA-DR antigenic mismatch, two 12/12 matched-unrelated peripheral blood stem cell transplant (PBSCT), and one haploidentical PBSCT with TCRαβ/CD45RA depletion from maternal donor. Neutrophil and platelet engrafted (>20×109 /L) at a median of 15 and 22days, respectively. All achieved near full donor chimerism at 1month. All patients survived and remained transfusion-independent without significant morbidities with median follow-up duration of 10years. To conclude, local data demonstrated favorable outcome of allogeneic HSCT for BHFS patients, but sample number is small. Non-directive approach in counseling and international collaboration is recommended.

45. Clinical utility of NGS hemeonc panel in a pediatric patient (pancytopenia concerning for primary myelofibrosis) with KRAS mutation

Application of next-generation sequencing (NGS) panels is usually limited to diagnosis, prognosis, and development of personalized treatment strategies/targeted therapies for patients with myeloid malignancies e.g., Myelodysplastic syndrome (MDS), myeloproliferative neoplasms (MPN), and acute myelogenous leukemia (AML). Here we present a case of 3 year old male that presented with pancytopenia and splenomegaly. His initial bone marrow revealed mild to moderate fibrosis (normocellular; no increase in blasts). A repeat bone marrow biopsy demonstrated myelofibrosis and platelet dysplasia. The physician was concerned that this pediatric patient has primary myelofibrosis and requested the NGS Hemeonc panel testing to be performed on the patient. Library preparation was performed on the DNA extracted from the peripheral blood using Illumina's TruSeq Custom Amplicon Technology. NGS was performed with Illumina's MiSeq platform, using a panel targeting 148 exons within 30 genes most commonly implicated in the development and/or persistence of MDS, MPN, and AML (ABL1, ASXL1, CBL, CEBPA, CSF3R, DNMT3A, ETV6, EZH2, FLT3, GATA2, IDH1, IDH2, JAK2, JAK3, KIT, KRAS, MPL, NPM1, NRAS, PHF6, PTPN11, RUNX1, SETBP1, SF3B1, SH2B3, SRSF2, TET2, TP53, U2AF1, WT1). NGS analysis revealed a KRAS mutation (c.37G > T, p.G13C) in this patient. A matched sibling bone marrow transplant was performed following myeloablative preparation on this patient and the post stem cell transplant specimen (peripheral blood) was sent in 2017 for further analysis of the previously detected KRAS mutation (p.G13C) using NGS Hemeonc panel. The patient was found to be negative for the same KRAS mutation on the post stem cell specimen indicating the clinical utility of the NGS Hemeonc panel.

Low-cost matched sibling bone marrow transplant for standard-risk thalassemia in a limited-resource setting

Thalassemias are the most common inherited genetic disorder in India and a major public health burden with bone marrow transplant (BMT) considered the only established curative therapy. We describe outcomes for patients (n = 71) with standard-risk thalassemia (liver size < 2 cm and age <15 years), receiving BMT in 6 low-cost start up BMT centers in the Indian sub-continent from August-2013 to July-2016. Patients received HLA-matched sibling donor unmanipulated BMT. Conditioning was with busulfan (14 mg/kg oral total over days −10 to −7, serum levels not measured), cyclophosphamide (200 mg/kg total over days −5 to −2) and anti-thymocyte globulin (Genzyme 4 mg/kg or Fresenius 16 mg/kg over days −12 to −10). Kaplan Meier survival analysis revealed thalassemia free survival (TFS) of 83% with overall survival (OS) of 93% with a median follow up of 17.5 months (IQR 13.4–22.6). Twelve percent of the patients rejected, 12% had severe GVHD (7% acute grade 3–4 GVHD and 4% had moderate/severe chronic GVHD). All 3 patients with acute GVHD Grade 3 are off immunosuppression, and those with chronic GVHD are well with Lansky score >80 at the last follow up. 5 patients (7%) died, mortality related to transplant. Enough data existed for 2 centers in India (36/71 transplants) to analyze overall costs from admission up to one-year post-BMT which revealed a median cost of Rs 7,30,445 ($11519) [Range Rs 4,52,821–10,32,842 ($ 7079–16147)]. In conclusion, children with thalassemia in resource limited settings can achieve good outcomes with BMT at a reasonable cost.

Using Haploidentical (haplo) Donors and High-Dose Post-Transplant Cyclophosphamide (PTCy) for Refractory Severe Aplastic Anemia (SAA)

SAA is a life-threatening hematopoietic stem cell disorder that is treated with either bone marrow transplantation (BMT) or immunosuppressive therapy (IST). BMT is the treatment of choice for young patients with a matched sibling donor. Those with acquired SAA lacking a matched sibling donor and older adults (> 40yrs) generally receive IST because of the high toxicity of BMT in these patients. Roughly two-thirds will respond but up to 40% of patients eventually relapse or acquire a secondary clonal disease such as paroxysmal nocturnal hemoglobinuria (PNH) or myelodysplastic syndromes. Thus, a major challenge in treating SAA is the management of patients who are have relapsed after or are refractory to IST or who have acquired a secondary clonal disorder. BMT is the only curative therapy for patients with inherited SAA but problems with graft failure, graft-versus-host disease (GVHD), and lack of matched unrelated donors especially for ethnic minorities have limited the success of alternative donor BMT, especially for adult patients. PTCy after HLA-haplo BMT has been shown to facilitate engraftment and yield rates of GVHD comparable to those of matched sibling BMT in hematologic malignancies. We hypothesized that PTCy would improve outcomes for patients with refractory SAA by reducing the risk of GVHD and expanding the donor pool to include haplo related and mismatched unrelated donors (URD).Patients received an allogeneic BMT (alloBMT) for relapsed or refractory SAA. Relapsed/ refractory disease was defined as having failed previous course of IST. In the majority of the cohort, haplo donors were used due to lack of availability of suitable and timely URDs. Nine patients underwent haploBMT. A mismatched URD was used in one case of no suitably available haplo donor given the inherited nature of her disorder. The nonmyeloablative preparatory regimen included antithymocyte globulin (0.5 mg/kg day-9 and 2 mg/kg on days -8,-7), fludarabine (30mg/m2/day on days - to -2), low dose CY (14.5 mg/kg on days -4, -5 ) and total body irradiation (200cGy day -1). After the unmanipulated marrow graft was infused on day 0, cyclophosphamide 50mg/kg/day IV on days +3,+4 post-transplant was administered. All patients received tacrolimus and mycophenolate mofetil (MMF) beginning day+5 through 35 (MMF) and day +5 through 1 year (tacrolimus) for additional graft versus host disease (GVHD) prophylaxis. Neutrophil engraftment was defined as an ANC >1.0 x109/L measured for three consecutive measurements on different days. Red cell engraftment was defined as days from last red blood cell transfusion. Platelet engraftment was defined as a platelet count >50,000 for 7 days without transfusion.Ten patients were treated according to this IRB approved protocol. Eight patients had documented acquired disease and received grafts from 5/10 related donors. Six had failed ATG-containing regimens previously and two relapsed after high dose cyclophosphamide as IST. Two patients had inherited syndromes: one with Diamond Blackfan who received a graft from 5/10 related donor and one with telomeres less than the first percentile in length with a presumed familial syndrome who received a 9/10 unrelated graft. Median follow-up time is 17.5 months (range 5-49). The median age was 34 years (range 17-54) with 6 patients greater than age 30 years, and 50% were males. Median time to neutrophil engraftment was 18 days (range 16-24). Median time to red cell engraftment was 25 days (range 16-48). Median time to platelet engraftment was 27.5 days (range 22-108). At the time of BMT, six patients had PNH clones, and all were eliminated. All patients are alive and well, fully engrafted with 100% donor chimerism in blood and marrow. Two patients had grade 1-2 skin acute GVHD. These two also had mild chronic GVHD of the skin/mouth requiring systemic steroids. One patient was able to come off all immunosuppression by 15 months and the other by 17 months.Nonmyeloablative alloBMT with PTCy appears promising in patients with refractory acquired and inherited SAA with acceptable rates of engraftment, eradication of pre-existing clonal diseases, and expansion of the donor pool. Given these promising results, this approach is being utilized in the next BMTCTN national trial for acquired refractory SAA. We believe this approach could also be considered as upfront therapy for SAA, and such a trial is in development at Hopkins DisclosuresBrodsky:Alexion Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees.

From pre to post-BMT: autoimmunity and vasculitis in wiskott-aldrich syndrome

Background Wiskott-Aldrich Syndrome (WAS) is an X-linked primary immunodeficiency associated with recurrent infections, eczema, and small platelets. Patients are at risk of autoimmune complications which represent a more severe form of the WAS disease spectrum. Case history We described a 4-year-old boy who was diagnosed with WAS, in the context of mucosal bleeding and severe thrombocytopenia. The thrombocytopenia did not respond to intravenous immunoglobulins and steroids. Several months post-diagnosis, he developed left lower lobe pneumococcal pneumonia, arthritis and a vasculitic rash which was leukocytoclastic on histology. This was followed by a rise in his IgG levels (peaking at 46 g/L), abnormal liver function tests which were predominantly cholestatic, and persistently low C4 levels. Liver biopsy showed vasculitis and autoimmune cholangitis. His anti-C1q autoantibodies and rheumatoid factor were also significantly elevated. He was treated with a combination of anti-CD20 monoclonal antibody and steroids, resulting in a significant improvement in his clinical symptoms and laboratory markers. He underwent a matched sibling bone marrow transplant (BMT) and is currently well 45 days post-transplant. Discussion In conclusion, vasculitis in WAS requires prompt treatment due to the risk of severe morbidity and is a consideration for BMT.

A novel homozygous mutation in recombination activating gene 2 in 2 relatives with different clinical phenotypes: Omenn syndrome and hyper-IgM syndrome

A novel homozygous mutation in recombination activating gene 2 in 2 relatives with different clinical phenotypes: Omenn syndrome and hyper-IgM syndrome

Successful engraftment of donor marrow after allogeneic hematopoietic cell transplantation in autosomal-recessive hyper-IgE syndrome caused by dedicator of cytokinesis 8 deficiency

A child with homozygous partial deletion of the DOCK8 gene showed characteristic clinical findings of autosomal recessive hyper-IgE syndrome and full donor chimerism early after matched sibling bone marrow transplantation.

Blood and marrow transplantation for sickle cell disease: overcoming barriers to success.

Sickle cell disease (SCD) is a common health problem in the United States; yet, the only curative therapy, a bone marrow transplant (BMT), is seldom applied. The objective of this report is to review the most recent clinical trials involving blood and BMT for SCD and to discuss novel approaches to overcome the many barriers to successful use of BMT for SCD. In select patients, disease-free and overall survival is greater than 80% following matched sibling BMT for SCD. Unfortunately, most patients with SCD do not have a suitable human lymphocyte antigen-matched sibling donor. In an attempt to expand the donor pool, several groups are beginning to explore the use of alternative sources of stem cells such as haploidentical donors and umbilical cord cell blood. The curative potential of BMT in SCD is irrefutable, with outstanding results in children following a myeloablative conditioning regimen and a matched sibling donor transplant. Well tolerated and effective application of alternative sources of stem cells for BMT in SCD could greatly increase the cure rate for this devastating disease.

Time‐course and risk factors of hypothyroidism following allogeneic hematopoietic stem cell transplantation (HSCT) in children conditioned with fractionated total body irradiation

Hypothyroidism is a complication of hematopoietic stem cell transplantation (HSCT)1 that has been studied primarily in children after matched sibling bone marrow transplantation. Unrelated donors and umbilical cord blood (CB) are now used in transplantation, and hypothyroidism in these populations is undocumented. Children who underwent fractionated total body irradiation (fTBI) and received bone marrow or umbilical cord blood transplantation from siblings or unrelated donors were examined for hypothyroidism over time. All children who received fTBI prior to allogeneic HSCT at The Children's Hospital (TCH), Denver between 1994 and 2002, and survived >1 year were included (N = 41 eligible; N = 33 met inclusion criteria). Probability of hypothyroidism was estimated by the Kaplan-Meier method and groups were compared using Cox's proportional hazards regression model. Hypothyroidism developed in 52% of the children at a median time of 1.9 years (range 0.7-8.3 years). Primary compensated hypothyroidism was diagnosed in 94% of those affected. Children who received unrelated donor HSCT were 8.4 times more likely to develop hypothyroidism than those receiving matched sibling HSCT. Recipients of CB were 4.6 times more likely to have hypothyroidism than recipients of bone marrow. There is a significant risk of developing hypothyroidism following fTBI, and allogeneic HSCT, that appears greater in recipients of unrelated donor HSCT and CB transplantation. Most of our patients who received BMT had sibling donors and the majority of CB transplants were from unrelated donors. It is difficult to determine which association is related to hypothyroidism, but the combination is significant.

Capitalizing on Mendelian randomization to assess the effects of treatments.

The stage of disease and general fitness of patients usually influences the treatments they receive. Because of this, tests of treatments and treatment comparisons need to ensure that treatment comparison groups are made up of patients who are as similar as possible. Without this assurance, any differences in the progress of patients receiving different treatments cannot confidently be ascribed to differential effects of treatments: they may simply reflect differences in the characteristics of patients receiving the different treatments. This is the rationale underlying random allocation (randomization) to treatment groups in randomized trials—to ensure that whether treatment is given or withheld is unrelated to factors influencing the prognoses of patients. Sometimes, for one or more of a variety of reasons, the organization of randomized controlled trials (RCTs) presents challenges that are judged likely to be very difficult to overcome. One such example relates to the need to assess the effects of bone marrow transplantation in the treatment of acute myeloid leukaemia in children. Randomized comparisons of treatment with and without bone marrow transplantation in this form of leukaemia have been judged unlikely to be achieved successfully, yet it is clearly important to obtain reliable, unbiased estimates of the effects of transplantation—wanted and unwanted—because the disadvantages may outweigh any advantages of this invasive treatment. In 1991, Richard Gray and Keith Wheatley reported an ingenious method for obtaining unbiased estimates of the effects of bone marrow transplantation without conducting a traditional randomized trial.1 They pointed out that unbiased comparisons could be made between child patients who had a genetically compatible sibling, and so in principle could receive a matched sibling bone marrow transplant, with other child patients who had no genetically compatible sibling, and so were incapable of receiving a matched sibling bone marrow transplant. Because having or not having a genetically compatible sibling is a matter of chance—it is determined by random assortment of genes at the time of gamete formation and conception—this situation produces what is effectively a randomized comparison. Whether a child with leukaemia belongs to the group with genetically compatible siblings or the group without such siblings will not be related to potential confounding factors such as disease stage and general fitness at the time of diagnosis. In a form of ‘intention-to-treat analysis’, Gray and Wheatley noted that an unbiased comparison meant comparing all the patients with a genetically compatible sibling with all the patients without such a sibling, regardless of whether or not every patient with a potential donor actually received a transplant (Figure 1).1 Figure 1 Intention to treat analysis of children with leukaemia Gray and Wheatley appear to have been the first authors to refer to this particular way of capitalizing on chance events in nature to create unbiased comparison groups to assess the effects of a treatment. They called it ‘Mendelian randomization’. Several further studies have now been carried out using their design,2-4 including studies to assess the effects of treatment for acute lymphoblastic leukaemia.5-7 Some of these studies have confirmed that like is being compared with like in comparison groups defined in this way.3-6 They have also shown that there are differences in prognostic factors between groups defined by the treatment they received; differences that would confound a conventional observational analysis comparing different treatments.4-6 The basic design outlined in Figure 1 might be improved by taking into account the number of siblings each patient has.8 Patients with more siblings have a greater chance of having a genetically compatible donor, and therefore groups defined by having a compatible donor will differ according to average number of siblings and thus will differ by factors that may be related to prognosis. Indeed, a later study found that the number of siblings could itself be related to survival.9 The study by Gray and Wheatley (Gray, personal communication) and another more recent study4 applied an analysis restricted to patients with at least one sibling, but exact stratification or matching on number of siblings might be a more robust approach.

The Molecular Characteristics of T-Cell Immune Reconstitution in Leukemia Patients after Allogeneic Hematopoietic Stem Cell Transplantation.

The clinical and experimental results indicated that the T cell immune reconstitution was slow after hematopoietic stem cell transplantation. This study was To analyze the molecular characteristics of T lymphocytic clones during immune reconstitution in leukemia patients after allogeneic hematopoietic stem cell transplantation(Allo-HCST)by investigating complementarity determining region (CDR3) repertoires of T cell receptor β chain variable region (TCRVB) and made it possible to understand the relationship between the express of TCRVB and immune reconstitution. Reverse transcriptase-polymerase chain reaction(RT-PCR) amplified 24 subfamily genes of TCRVB from peripheral blood lymphocytes (PBLs) of twenty-four patients with leukemia underwent three kinds of Allo-HSCT(haploidentical bone marrow transplantation, matched sibling bone marrow transplantation and matched-unrelated PBSCT), five normal donors as control. The PCR products were further analyzed by genescane to evaluate the clonality of VB subfamily. The monoclonal bands which associated with GVHD and CMV infection were obtained through denaturation polyacrylamide gel electrophoresis and sequenced. Compared the sequences of TCRVB CDR3 gene repertoire with other sequences associated with GVHD or CMV infection which had been reported. +2~+19months after transplantation, for nine patients among them which underwent haploidentical bone marrow transplantation, there were 6~14 VB subfamilies expressed with 33% 15 VB subfamilies expressed, which 45%of them were polyclones. In the 5 patients of matched-unrelated PBSCT, there were 10~15 VB families expressed, with 45% the polyclone expression. For ten patients underwent matched sibling bone marrow transplantation, 10~16 VB subfamilies expressed and more than 48% were polyclones. Monoclones and oligoclones existed in 24 VB subfamilies, no common monoclone VB subfamilies expressed. According to the usage of VB family and polyclonal expression, Immune reconstitution in patients which underwent haploidentical BMT was later than that of other two groups (p<0.05), there were no different between other two groups (p>0.05). After Allo-HSCT, two patients detected TCRVB in +2m and +3m and found that it has tendency of adding to use VB subfamilies and increasing to express CDR3 polymorphism. Using biology-information tool, compared 23 TCRVB CDR3 molecules which related to GVHD and CMV infection and found that, different cases in the same VB subfamilies may share similarity in amino acid motif, while in different VB subfamilies none of clones appeared to share the same amino acid motif. In 1.5 years after AHSCT, the usage of TCRVB subfamilies still restricted. T-cell immune reconstitution in patients which underwent haploidentical BMT was later than that of other two groups. TCRVB CDR3 molecules which related to GVHD and CMV infection showed that different cases in the same VB subfamilies may share similarity in amino acid motif, while in different VB subfamilies none of clones appeared to share the same amino acid motif.

Acute Adult Leukemia (ATL) Successfully Treated with Full Matched Sibling Bone Marrow Transplantation and Haplo-Identical (3 Locus Mis-Matched) Peripheral Stem Cell Transplantation.

[Back ground] HTLV-1 known as the 1st recognized human retrovirus infection described 1979 is endemic to some regions of the world, especially southwest Japan, Caribbean and others. Acute ATL type among the HTLV-1 associated leukemia/lymphoma has the poorest prognosis after various chemotherapies that are easily mutated chemo-resistantly. We hereby present 2 acute ATL cases receiving stem cell transplantation. [Case 1] The patient was a 21 years-old female. Induction therapy like ALL setting and then CHOP 14 for acute ATL induced to CR. The patient had HTLV-1 negative full matched 31 years-old brother. We performed bone marrow transplantation at day 90 from the beginning of initial chemotherapy. Typical conditioning regimens and immunotherapies consisted of CY (60mg/kg×3) +TBI (12Gy) and tacrolimus+steroid respectively. The engraftment was day 14 with no aGVHD of grade 2 to 4 observed till day 100. HTLV-1 viral load by RT -PCR was not detected at day 40 after BMT although HTLV-1 antibody and Western blotting analysis were still positive. [Case 2] The patient was a 32 years-old male. He was admitted our emergency room due to hypercalcemia (Ca 26.5mg/dl).CHOP and other chemotherapies were performed for a critical life saving situation. However, the control of his hypercalcemia was difficult even under intensive chemotherapy with steroid, elcatonin and bisphosphonates. His metastatic pulmonary calcification and arthralgia due to hypercalcemia were progressive on bone scintigram and CAT scan. The Japan Marrow Donor Program failed to provide an adequate donor. We decided to initiate peripheral stem cell transplantation (PBSCT) from his haplo-identical (3 locus mis-matched) HTLV-1 positive 68 years-old mother with informed consent from the patient and his family, as there was no life expectancy from usual chemotherapeutic treatment only. We performed PBSCT at day 90. The conditioning regimen and immunotherapy consisted of fludarabine phosphate (25mg×5) +L-PAM (240mg) +TBI (4Gy) and tacrolimus +steroid +short term MTX from the beginning of initial chemotherapy. At day 16 from PBSCT, Hypercalcemia and metastatic calcification as well as leukemic status improved dramatically. Engraftment and complete chimera were observed on day 11 and day 26 respectively with no aGVHD till day100. Although immunotherapy induced CMV infection was severe, the infection improved by using ganciclovir and foscarnet.[Results] The former case showed positive for results the successful eradication of HTLV-1. The latter case is still positive for HTLV-1 on PCR. However, this case showed CR in acute ATL with neither hypercalcemia nor its related complications.[Conclusion] In our limited studies, the modality for stem cell transplantation in acute ATL to cure or control leukemic cells in young patients has strong potential, even in a haplo-identical setting.

Unrelated Donor Umbilical Cord Blood Transplant Following Intrauterine Transfusions for Treatment of Alpha Thalassemia Major.

Homozygous α0-thalassemia (deletion of all 4 α-globin genes) results in Hb Bart's (γ4 tetramers). The high oxygen affinity of Hgb Bart's leads to severe hypoxia inutero with resulting profound edema (hydrops) and congestive heart failure. Almost universally fetal death occurs prior to diagnosis, therefore not allowing the opportunity for treatment. Advancement of maternal-fetal medicine and neonatal ultrasound has led to the possibility of in utero diagnosis and treatment with inrauterine transfusions. Also, cases of premature infants surviving and undergoing chronic transfusion therapy have been reported. Hematopoietic stem cell transplant (HSCT) has become increasingly used in the cure of beta thalassemia, and provides the potential to cure a0thalassemia as well. To date, 3 reported cases of a0thalassemia have been successfully treated with HSCT. Two children underwent matched sibling bone marrow transplant at 20 and 21 months of age and one underwent a 5/6 HLA matched sibling umbilical cord blood transplant (UCBT). One child had received intrauterine transfusions. We report a case of an infant with a0thalassemia successfully treated with intrauterine transfusion therapy followed by unrelated donor UCBT. The child's mother, of Cambodian descent, presented to the obstetrician at 23 weeks with concerns of decreased fetal movement. Fetal ultrasound revealed a thickened placenta and hydrops fetalis. Based on the family's nationality and history of previous fetal loss, the suspicion of a0thalassemia was raised. Umbilical cord blood sampling revealed a hypochromic, microcytic anemia with target cells. The hemoglobin electrophoresis demonstrated Hgb Barts and Portland. Subsequent genotyping confirmed deletion of all 4 α-globin genes. Options presented to the family included termination of pregnancy as well as the option of intrauterine transfusion followed by either chronic transfusion therapy with iron chelation or the possibility of HSCT. The patient received three intrauterine transfusions prior to delivery at 32 weeks gestation. A poor physical profile, non-reassuring heart rate and breech position led to the premature delivery. Apgars were 1 at one minute, 6 at five minutes, and 9 at ten minutes. The baby was admitted to the NICU and required mechanical ventilation for two days. The hospital course was relatively uneventful and included red blood cell transfusion. After discharge, he was maintained with intermittent transfusions until 6 months of age. Following informed consent, he was conditioned for transplant with busulfan, cytoan and rabbit ATG. Since the infant lacked an HLA matched sibling, he received a 5/6 HLA matched unrelated donor umbilical cord blood unit delivering 11.8 × 107 nucleated cells/kg. Neutrophil engraftment (ANC>500) was achieved on day + 15. FK506 and methotrexate 5mg/m2 on days 1, 3 and 6 was utilized for GVHD prophylaxis. His course was complicated by moderate venoocclusive disease, small subdural hemorrhage, and CMV and adenovirus viremia. He has not had any evidence of graft versus host disease. Initial chimerism (RFLP) showed approximately 63% donor derived cells, and subsequent testing showed increasing chimerism and 100% T-cell engraftment. He clinically is doing very well post transplant. This case demonstrates that intrauterine transfusion followed by unrelated donor UCBT is feasible for the treatment of a0thalassemia.

Calcitonin A Genotype Is Associated with Risk of Acute Graft-Versus-Host Disease Following Allogeneic Bone Marrow Transplantation for Children with Acute Myeloid Leukemia in First Remission.

Background: Procalcitonin (PCT) is a 13 kD 116-amino acid prohormone. Elevated serum levels of procalcitonin have been noted in patients with bacterial infections such as septicemia, pneumonia and immune-mediated conditions like graft-versus-host disease (GvHD) and organ rejection. In this study, we sought to examine if a single base pair polymorphism (a T to C transition) at position −624 of the calcitonin A gene (CALCA) was associated with risk of acute GvHD.Subjects: 113 children (median age 10 y; range 0.38 – 19.0 y) with de novo acute myeloid leukemia (AML) treated on recent Children's Oncology Group protocols 2891, 2941 or 2961. All of these patients received matched sibling bone marrow transplant (BMT) in first clinical remission. Ablative conditioning consisted of busulfan and cyclophosphamide. All patients received methotrexate until day 100 for GvHD prophylaxis.Methods: All clinical DNA specimens were subjected to whole genome amplification. Genotyping was performed using the 5′-nuclease (TaqMan) assay on the ABI 7700 DNA analyzer (Applied Biosystems).Results: Observed CALCA -624 T>C genotypes in 113 patients were: TT = 52 (46%), TC = 43 (38%) and CC = 18 (16%); T allele = 73 (65%) and C allele = 40 (35%). There were no significant differences in patient characteristics (age, gender, race, study 2891/2941/2961, white blood count at diagnosis, FAB type, or cytogenetics) between the TT vs. combined TC or CC groups. Overall, acute GvHD grade I–IV (Seattle criteria) was observed in 55/113 patients (49%). Nineteen out of 52 (37%) with TT, while 21 out of 43 (49%) with TC, and 15 out of 18 (83%) with CC CALCA genotypes developed acute GvHD. Alternatively, acute GvHD was observed in 37% patients with common TT genotype and in 59% patients with variant TC or CC genotype (p=0.023). The odds ratio for developing acute GvHD (grade I – IV) for children with TC or CC genotypes was 2.5 (CI 1.17, 5.33) compared with those with TT genotype. The difference in observed acute GvHD remained significant when the analyses were limited to more uniform subgroups: 54 patients on single trial, 2961 only (p=0.028) and 42 whites only on 2961 trial (p=0.028). Severe acute GvHD (grade III – IV) was seen in 6/52 (12%) patients with TT vs. 6/61 (10%) in those with TC or CC genotypes; this difference was not statistically significant (p =0.771). There were no significant differences in day +100 transplant related deaths, engraftment, relapse rate or overall survival. Plans: Association between CALCA genotypes and infectious complications will be also analyzed when clinical data abstraction is completed. CALCA −1752 C>G and 3740 A>G genotyping is in progress and clinical correlative analyses will be repeated at genotype and haplotype levels.

Complete Remission Following Clofarabine Treatment in Refractory Juvenile Myelomonocytic Leukemia

Juvenile myelomonocytic leukemia (JMML) is the most common myeloproliferative/myelodysplastic disorder seen in children. The treatment of choice, allogeneic stem cell transplantation, provides the only known cure for the disease, but relapse after transplant is common. The authors describe a 5-year-old boy diagnosed at age 34 months with JMML that evolved to acute myeloid leukemia. Initial treatment consisted of fludarabine and cis-retinoic acid therapy, followed by a matched sibling bone marrow transplant. After a relapse, he received a second transplant from the same donor, using peripheral blood stem cells, followed by repeated donor leukocyte infusions. After the second relapse, he received the farnesyltransferase inhibitor R115777 (tipifarnib, Zarnestra), but the leukemia persisted. When bone marrow blasts numbered 60% of the mononuclear cells, he received single-agent clofarabine induction (52 mg/m/d) for 5 days. After three courses, he attained a remission marrow with 5% blasts and disappearance of the 5q- and 9q- cytogenetic abnormalities.