Hemalatha G. Rangarajan

Emergence of T cells that recognize nonpolymorphic antigens during graft-versus- host disease.

Chronic GVHD is a major cause of morbidity and mortality in allogeneic stem cell transplantation recipients and typically develops from antecedent acute GVHD. In contrast to acute GVHD, chronic GVHD has much broader tissue involvement and clinical manifestations that bear striking similarity to what is observed in autoimmune diseases. How autoimmunity arises out of alloimmunity has been a longstanding unresolved issue. To address this question, in the present study, we performed a comprehensive analysis of the clonotypic T-cell response using complementary murine models that simulate what occurs during the transition from acute to chronic GVHD. These studies revealed repertoire skewing and the presence of high-frequency clonotypes that had undergone significant in vivo expansion, indicating that GVHD-associated autoimmunity was characterized by antigen-driven expansion of a limited number of T-cell clones. Furthermore, we observed that T cells with identical TCRβ CDR3 nucleotide sequences were capable of recognizing donor and host antigens, providing evidence that the loss of self-tolerance during acute GVHD leads to the emergence of self-reactive donor T cells that are capable of recognizing nonpolymorphic tissue or commensally derived antigens. These data provide a mechanistic framework for how autoimmunity develops within the context of preexisting GVHD and provide additional insight into the pathophysiology of chronic GVHD.

The use of novel Therakos™ Cellex® for extracorporeal photopheresis in treatment of graft‐versus‐host disease in paediatric patients

Extracorporeal photopheresis (ECP) is an established second line treatment option for graft‐versus‐host disease (GVHD) post‐haematopoietic progenitor cell transplant. At our centre, the Therakos™ Cellex® has replaced the Therakos™ UVAR‐XTS™ machine for ECP since 2009. We reviewed the records of 385 procedures using the Therakos™ Cellex® for safety and tolerability. Nine patients underwent ECP for GVHD. The median age was 13·5 years (range 3·7–24) and weight was 49·2 kg (range 18·5–86·3). The mean duration per procedure was 106 min (range 60–205). Fifteen (3·9%) procedures were cancelled and 10 (2·6%) were delayed, with central venous line (CVL) issues being the most frequent problem. With the use of prophylactic tissue plasminogen activator, fewer CVL‐related occlusions were observed (4·7% vs. 2·3%). There was one episode of a CVL‐associated thrombosis and one episode of delayed bleeding. There were four episodes of viral reactivation, four CVL‐associated infections (1142 catheter days) and one episode of systemic inflammatory response syndrome. No patient experienced symptomatic hypotension. This is the first report outlining the safety and tolerability of the Therakos™ Cellex® device for ECP in children and young adults.

Outcomes of matched sibling donor hematopoietic stem cell transplantation for severe sickle cell disease with myeloablative conditioning and intermediate‐dose of rabbit anti‐thymocyte globulin

Allogeneic hematopoietic stem cell transplantation (HSCT) is a curative therapy for sickle cell disease (SCD) in children. Despite excellent outcomes of matched sibling donor (MSD) HSCT, there is still 5–10% chance of rejection and transplant related mortality (TRM) with 12–23% incidence of graft versus host disease (GVHD). We postulated that an intermediate dose of rabbit anti‐thymocyte globulin (r‐ATG, 10 mg/kg cumulative) would be effective in preventing both rejection and GVHD.

Anicteric veno-occlusive disease after hematopoietic stem cell transplantation in children.

Hepatic veno-occlusive disease (VOD) or sinusoidal obstruction syndrome is a potentially fatal complication following hematopoietic stem cell transplantation (HSCT) and occurs in ~ 30% of the children undergoing HSCT. Two diagnostic criteria have been used for the diagnosis of VOD, the Baltimore criteria by Jones et al. comprising hyperbilirubinemia of42mg/dL and any two of the following: weight gain of 45% from baseline, hepatomegaly or ascites. The diagnosis of VOD based on the Seattle criteria by McDonald et al. can be made if any two of the following are present by day +20 after HSCT: hyperbilirubinemia (42mg/dL), ascites or weight gain of 42% from baseline, or hepatomegaly. Prompt diagnosis and early intervention is crucial as patients with severe VOD may develop multi-organ failure, requiring intensive care and carry a high mortality of up to 84%. Yet there is no consensus on grading severity of VOD. Although liver biopsy is helpful for confirming the diagnosis, in the setting of coagulopathy and thrombocytopenia, it is usually avoided. Ultrasonography has been used in the diagnosis of VOD; in the study by Lassau et al. on multivariate analysis, two ultrasound criteria (splenomegaly and ascites) and one doppler criterion (flow recorded in paraumbilical vein) were correlated with the severity of VOD (P = 0.0001). Demonstration of reversal of portal venous blood flow by ultrasonography can be helpful, but is often a late finding indicating advanced disease. The Baltimore criteria are more often used in adults and appear to be more stringent than the Seattle criteria; as patients without hyperbilirubinemia are excluded. In contrast, the Seattle criteria may lead to over diagnosis, as some patients may meet the diagnostic criteria earlier in the course of the disease. Higher serum bilirubin levels have been associated with increased severity of VOD and worse outcomes. Bearman et al. used total serum bilirubin level and percentage weight gain to create risk curves that would predict the likelihood of developing severe VOD. Total serum bilirubin is often used as a marker for response to treatment with defibrotide, as discussed in studies by Ho et al. and Richardson et al. However some patients with VOD may not have hyperbilirubinemia but still have all the other features consistent with VOD. In a recent study by Myers et al., five patients had a bilirubin of ⩽ 2mg/dL at the time of diagnosis of VOD and reversal of portal venous flow (PVF) by ultrasonography; two of these continued to have a bilirubin of o2mg/dL. There have been few studies focusing specifically on anicteric VOD. The primary aim of this study is to assess the clinical course and outcomes of patients with anicteric VOD after HSCT in children. A retrospective review of the medical charts of all patients diagnosed with VOD while undergoing HSCT from 1992 through June 2014 at Nationwide Children’s Hospital, Columbus, OH, USA was performed. Both Seattle and Baltimore criteria were retrospectively applied to each patient diagnosed and treated as VOD. Patients who were diagnosed with VOD but maintained a total serum bilirubin level ⩽ 2mg/dL during the entire course of treatment for VOD were defined as having ‘Anicteric VOD’. The level of supportive care during treatment for VOD was defined as follows: ‘Level 1’—general floor level management (that is, diuretics, pain control, fluid restriction, transfusion and so on), ‘Level 2’: paediatric intensive care unit level support for ⩽ 2 body systems (that is, mechanical ventilation for respiratory failure, vasopressors for heart failure and so on) and ‘Level 3’: paediatric intensive care unit level support ⩾ 3 body systems (that is, mechanical ventilation+vasopressors+dialysis and so on). Standard descriptive statistical methods were used for analysis. Statistical analysis was done using SAS 9.3 statistical software (SAS Institute, Cary, NC, USA). Categorical variables were compared between VOD groups using a Fisher’s exact test. Continuous variables were compared using a nonparametric twosided Wilcoxon rank-sum test. P-values o0.05 were considered significant. Acquisition of patient data was approved by the Institutional Review Board at Nationwide Children’s Hospital. Thirty patients were diagnosed with VOD during the study period and patient demographics and characteristics are presented in Tables 1 and 2. One patient with undocumented bilirubin level was excluded from the study. The remaining 29 patients were included in the final analysis. Nine of 29 (31%) had anicteric VOD and 20/29 (69%) had VOD with icterus. The majority (n= 26) of the patients received myeloablative conditioning for malignant (n= 24) or non-malignant (n= 5) diseases. There was no difference in the characteristics of patients with anicteric VOD versus icteric VOD with regard to percentage weight gain, hepatomegaly, abdominal pain, demonstration of PVF reversal, duration of treatment or outcome. Ascites was seen in 5 of the 9 (55.6%) and 19 of 20 (95%) cases with anicteric and icteric VOD, respectively. The median level of supportive care was level 2 for the icteric VOD group as compared with level 1 for the anicteric group (P= 0.007). While analysing ultrasonography results for the two groups, of the nine patients with anicteric VOD, data were missing in four (all recovered from VOD), one showed reversal of PVF and recovered, four patients had no reversal of PVF (two recovered and two died). In the icteric VOD group, 12 showed reversal of PVF (five recovered and seven died), five had no reversal of PVF (one recovered and four died) and three had missing data (two recovered and one died). Overall, six patients with no reversal of PVF died from VOD (two with anicteric VOD and four with icteric VOD). Seven cases were diagnosed with VOD by the Seattle criteria at a median of day +15 after transplant. However, curative treatment with defibrotide was delayed by 1–11 days for lack of hyperbilirubinemia; and two of these never developed hyperbilirubinemia. Curative anticoagulation therapy was given in 1/9 (11%) cases with anicteric VOD versus 9/20 (45%) of the icteric VOD. Fourteen (14) of the 29 patients died (48%) from complications of VOD, 2 with anicteric and 12 with icteric VOD died. Although there have been numerous studies characterizing VOD and its treatments, few studies have specifically focused on anicteric VOD. There seems to be a poor understanding and awareness of anicteric VOD as a diagnosis relative to icteric VOD. As patients with anicteric VOD do not meet the Baltimore criteria, the diagnosis of VOD and its treatment may be delayed if only the Baltimore criteria are followed. It is well known that delay in the initiation of supportive care and curative treatment may result in Bone Marrow Transplantation (2016) 51, 135–137

Hepatoblastoma and prune belly syndrome: a potential association

Prune belly syndrome (PBS) is a congenital anomaly characterized by the clinical triad of lax abdominal musculature, bilateral cryptorchidism, and abnormalities of the kidney and urinary tract. Previous reports of malignancy in patients with PBS have been limited to germ cell tumors. Hepatoblastoma (HBL) is the most common hepatic malignancy of childhood, affecting approximately 100 children each year in the USA. We describe a set of 4 pediatric patients with PBS and HBL. All individuals were born after 2002. These subjects lacked genetic, natal, or environmental factors known to confer risk of HBL. The occurrence of PBS and HBL in these patients constitutes a novel potential association.

Clinical risks and healthcare utilization of hematopoietic cell transplantation for sickle cell disease in the USA using merged databases.

Advances in allogeneic hematopoietic cell transplantation for sickle cell disease have improved outcomes, but there is limited analysis of healthcare utilization in this setting. We hypothesized that, compared to late transplantation, early transplantation (at age <10 years) improves outcomes and decreases healthcare utilization. We performed a retrospective study of children transplanted for sickle cell disease in the USA during 2000–2013 using two large databases. Univariate and Cox models were used to estimate associations of demographics, sickle cell disease severity, and transplant-related variables with mortality and chronic graft-versus-host disease, while Wilcoxon, Kruskal-Wallis, or linear trend tests were applied for the estimates of healthcare utilization. Among 161 patients with a 2-year overall survival rate of 90% (95% confidence interval [CI] 85–95%) mortality was significantly higher in those who underwent late transplantation versus early (hazard ratio (HR) 21, 95% CI 2.8–160.8, P=0.003) and unrelated compared to matched sibling donor transplantation (HR 5.9, 95% CI 1.7–20.2, P=0.005). Chronic graftversus host disease was significantly more frequent among those translanted late (HR 1.9, 95% CI 1.0–3.5, P=0.034) and those who received an unrelated graft (HR 2.5, 95% CI 1.2–5.4; P=0.017). Merged data for 176 patients showed that the median total adjusted transplant cost per patient was

Safety Profile of Good Manufacturing Practice Manufactured Interferon γ‐Primed Mesenchymal Stem/Stromal Cells for Clinical Trials

467,747 (range:

Severe transplant-associated thrombotic microangiopathy in patients with hemoglobinopathies

344,029–

Venous Thromboembolism in Pediatric Hematopoietic Cell Transplant: a Multicenter Cohort Study

799,219). Healthcare utilization was lower among recipients of matched sibling donor grafts and those with low severity disease compared to those with other types of donor and disease severity types (P<0.001 and P=0.022, respectively); no association was demonstrated with late transplantation (P=0.775). Among patients with 2-year pre- and post-transplant data (n=41), early transplantation was associated with significant reductions in admissions (P<0.001), length of stay (P<0.001), and cost (P=0.008). Early transplant outcomes need to be studied prospectively in young children without severe disease and an available matched sibling to provide conclusive evidence for the superiority of this approach. Reduced post-transplant healthcare utilization inpatient care indicates that transplantation may provide a sustained decrease in healthcare costs over time.

Treatment of recurrent CNS disease post‐bone marrow transplant in familial HLH

Mesenchymal stem/stromal cells (MSCs) are widely studied by both academia and industry for a broad array of clinical indications. The collective body of data provides compelling evidence of the clinical safety of MSC therapy. However, generally accepted proof of therapeutic efficacy has not yet been reported. In an effort to generate a more effective therapeutic cell product, investigators are focused on modifying MSC processing protocols to enhance the intrinsic biologic activity. Here, we report a Good Manufacturing Practice‐compliant two‐step MSC manufacturing protocol to generate MSCs or interferon γ (IFNγ) primed MSCs which allows freshly expanded cells to be infused in patients on a predetermined schedule. This protocol eliminates the need to infuse cryopreserved, just thawed cells which may reduce the immune modulatory activity. Moreover, using (IFNγ) as a prototypic cytokine, we demonstrate the feasibility of priming the cells with any biologic agent. We then characterized MSCs and IFNγ primed MSCs prepared with our protocol, by karyotype, in vitro potential for malignant transformation, biodistribution, effect on engraftment of transplanted hematopoietic cells, and in vivo toxicity in immune deficient mice including a complete post‐mortem examination. We found no evidence of toxicity attributable to the MSC or IFNγ primed MSCs. Our data suggest that the clinical risk of infusing MSCs or IFNγ primed MSCs produced by our two‐step protocol is not greater than MSCs currently in practice. While actual proof of safety requires phase I clinical trials, our data support the use of either cell product in new clinical studies. Stem Cells Translational Medicine 2017;6:1868–1879