Metformin for treatment of cytopenias in children and young adults with Fanconi anemia.

Fanconi anemia (FA), a genetic DNA repair disorder characterized by marrow failure and cancer susceptibility. In FA mice, metformin improves blood counts and delays tumor development. We conducted a single institution study of metformin in nondiabetic patients with FA to determine feasibility and tolerability of metformin treatment and to assess for improvement in blood counts. Fourteen of 15 patients with at least 1 cytopenia (hemoglobin < 10 g/dL; platelet count < 100 000 cells/µL; or an absolute neutrophil count < 1000 cells/µL) were eligible to receive metformin for 6 months. Median patient age was 9.4 years (range 6.0-26.5 ). Thirteen of 14 subjects (93%) tolerated maximal dosing for age; 1 subject had dose reduction for grade 2 gastrointestinal symptoms. No subjects developed hypoglycemia or metabolic acidosis. No subjects had dose interruptions caused by toxicity, and no grade 3 or higher adverse events attributed to metformin were observed. Hematologic response based on modified Myelodysplastic Syndrome International Working Group criteria was observed in 4 of 13 evaluable patients (30.8%; 90% confidence interval, 11.3-57.3). Median time to response was 84.5 days (range 71-128 days). Responses were noted in neutrophils (n = 3), platelets (n = 1), and red blood cells (n = 1). No subjects met criteria for disease progression or relapse during treatment. Correlative studies explored potential mechanisms of metformin activity in FA. Plasma proteomics showed reduction in inflammatory pathways with metformin. Metformin is safe and tolerable in nondiabetic patients with FA and may provide therapeutic benefit. This trial was registered at as #NCT03398824.

SPRINTing to Innovation: Children's Hospital of Philadelphia's Strategic Approach to Discovering Its Untapped Innovation Potential.

PROBLEM: There is a clear and urgent need for health care innovation in the United States. Hospital employees routinely recognize pain points that affect care delivery and are in a unique position to propose innovative and practical solutions, yet leaders rarely solicit ideas for investment and development from frontline providers and staff, revealing an untapped resource with innovation potential. APPROACH: To address these deficiencies, the Children's Hospital of Philadelphia expanded its innovation infrastructure with the competition-based SPRINT program in 2015. All hospital employees are encouraged to apply with early-stage innovative ideas, and if selected, are provided with business, legal, technical, and scientific project management support to help accelerate their projects toward commercial viability. SPRINT was modeled around 4 core tenets: (1) small, dynamic, and attentive project manager-led teams; (2) low barriers to entry; (3) emphasis on outreach; and (4) fostering innovators. OUTCOMES: Over its first 4 cycles from 2015 to 2018, 271 innovative teams applied to the SPRINT program, which led to support for 30 projects (11% acceptance rate). About a quarter of the projects each year were submitted by physician-led teams (mean 23%), a third by nonphysician clinical providers (mean 33%), and almost half were submitted by employees without direct patient contact (mean 44%). Nurses have emerged as the largest applicant group. Eleven of the SPRINT-supported projects (37%) resulted in commercial endpoints. NEXT STEPS: SPRINT has proven to be an effective model for supporting institution-wide, employee-driven health care innovation, especially among frontline clinical and nonclinical personnel. Critical next steps for the program include a formal cost-benefit analysis and the earlier participation of technology transfer and intellectual property experts to improve the commercialization roadmap for many SPRINT projects.

L-leucine improves anemia and growth in patients with transfusion-dependent Diamond-Blackfan anemia: Results from a multicenter pilot phase I/II study from the Diamond-Blackfan Anemia Registry.

BACKGROUND: Diamond-Blackfan anemia (DBA) is an inherited bone marrow failure syndrome characterized by anemia, short stature, congenital anomalies, and cancer predisposition. Most cases are due to mutations in genes encoding ribosomal proteins (RP) leading to RP haploinsufficiency. Effective treatments for the anemia of DBA include chronic red cell transfusions, long-term corticosteroid therapy, or hematopoietic stem cell transplantation. In a small patient series and in animal models, there have been hematologic responses to L-leucine with amelioration of anemia. The study objectives of this clinical trial were to determine feasibility, safety, and efficacy of L-leucine in transfusion-dependent patients with DBA. PROCEDURE: Patients ≥2 years of age received L-leucine 700 mg/m2 orally three times daily for nine months to determine a hematologic response and any improvement in growth (NCT01362595). RESULTS: This multicenter, phase I/II study enrolled 55 subjects; 43 were evaluable. There were 21 males; the median age at enrollment was 10.4 years (range, 2.5-46.1 years). No significant adverse events were attributable to L-leucine. Two subjects had a complete erythroid response and five had a partial response. Nine of 25, and 11 of 25, subjects experienced a positive weight and height percentile change, respectively, at the end of therapy. CONCLUSIONS: L-leucine is safe, resulted in an erythroid response in 16% of subjects with DBA, and led to an increase in weight and linear growth velocity in 36% and 44% of evaluable subjects, respectively. Further studies will be critical to understand the role of L-leucine in the management of patients with DBA.

Biexponential R2* relaxometry for estimation of liver iron concentration in children: A better fit for high liver iron states.

BACKGROUND: R2* relaxometry's capacity to calculate liver iron concentration (LIC) is limited in patients with severe overload. Hemosiderin increases in these patients, which exhibits a non-monoexponential decay that renders a failed R2* analysis. PURPOSE/HYPOTHESIS: To evaluate a biexponential R2* relaxometry model in children with different ranges of iron overload. STUDY TYPE: Retrospective. POPULATION: In all, 181 children with different conditions associated with iron overload. FIELD STRENGTH/SEQUENCE: 1.5T, T2 *-weighted gradient echo sequence. ASSESSMENT: Bi- and monoexponential R2* relaxometry were measured in the liver using two regions of interest (ROIs) using a nonproprietary software: one encompassing the whole liver parenchyma (ROI-1) and the other only the periphery (ROI-2). These were drawn by a single trained observer. The residuals for each fitting model were estimated. A ratio between the residuals of the mono- and biexponential models was calculated to identify the best fitting model. Patients with 1) residual ratio ≥1.5 and 2) R2*fast ≥R2*slow were considered as having a predominant biexponential behavior. STATISTICAL TESTS: Nonparametric tests, Bland-Altman plots, linear correlation, intraclass correlation coefficient. Patients were divided according to their LIC into stable (n = 23), mild (n = 58), moderate (n = 61), and severe (n = 39). RESULTS: The biexponential model was more suitable for patients with severe iron overload when compared with the other three LIC categories (P < 0.001) for both ROIs. For ROI-1, 37 subjects met criteria for a predominant biexponential behavior. The slow component (5.7%) had a lower fraction than the fast component (94.2%). For ROI-2, 22 subjects met criteria for a predominant biexponential behavior. The slow component (4.7%) had a lower fraction than the fast component (95.2%). The intraobserver variability between both ROIs was excellent. DATA CONCLUSION: The biexponential R2* relaxometry model is more suitable in children with severe iron overload. LEVEL OF EVIDENCE: 3 Technical Efficacy: Stage 1 J. Magn. Reson. Imaging 2019;50:1191-1198.

Protocol optimization for cardiac and liver iron content assessment using MRI: What sequence should I use?

OBJECTIVE: To determine the optimal MRI protocol and sequences for liver and cardiac iron estimation in children. METHODS: We evaluated patients ≤18 years with cardiac and liver MRIs for iron content estimation. Liver T2 was determined by a third-party company. Cardiac and Liver T2* values were measured by an observer. Liver T2* values were calculated using the available liver parenchyma in the cardiac MRI. Linear correlations and Bland-Altman plots were run between liver T2 and T2*, cardiac T2* values; and liver T2* on dedicated cardiac and liver MRIs. RESULTS: 139 patients were included. Mean liver T2 and T2* values were 8.6 ±â€¯5.4 ms and 4.5 ±â€¯4.1 ms, respectively. A strong correlation between liver T2 and T2* values was observed (r = 0.96, p < 0.001) with a bias (+4.1 ms). Mean cardiac bright- and dark-blood T2* values were 26.5 ±â€¯12.9 ms and 27.2 ±â€¯11.9 ms, respectively. Cardiac T2* values showed a strong correlation (r = 0.81, p < 0.001) with a low bias (-1.0 ms). The mean liver T2* on liver and cardiac MRIs were 4.9 ±â€¯4.7 ms and 4.6 ±â€¯3.9 ms, respectively. A strong correlation between T2* values was observed (r = 0.96, p < 0.001) with a small bias (-0.2 ms). CONCLUSION: MRI protocols for iron concentration in the liver and the heart can be simplified to avoid redundant information and reduce scan time. In most patients, a single breath-hold GRE sequence can be used to evaluate the iron concentration in both the liver and heart.

A Standardized Clinical Pathway to Decrease Hospital Admissions Among Febrile Children With Sickle Cell Disease.

BACKGROUND AND OBJECTIVE: Recurrent hospital admissions for patients with sickle cell disease (SCD) are costly and contribute to a low quality of life for patients. We implemented a clinical pathway to safely discharge SCD patients with fever who are evaluated in the emergency department (ED) of a large tertiary care center. METHODS: An interdisciplinary team of ED and hematology physicians, nurses, and an improvement advisor developed a clinical pathway that identified febrile SCD patients at low risk of serious bacterial infection based on historical, clinical, and laboratory criteria who could be discharged from the ED. Phone follow-up was planned through the use of an automated electronic notification that was sent to an established hematology follow-up pool at the time of ED discharge. We conducted two "fake front end" trials in the ED to receive feedback on our process before full implementation. A postpathway implementation quality improvement team monitored discharge rates, phone follow-up rates and adverse events. RESULTS: In the first 9 weeks postpathway implementation, 100 SCD patients were evaluated for fever; 84 (24%) met low-risk criteria and were discharged home. This reduction in admission rate has been maintained throughout the 3 years postimplementation. Successful phone follow-up was achieved in all discharged patients within 24 hours and no adverse events were identified. CONCLUSIONS: Low-risk febrile patients with SCD can be safely discharged from the ED. An automated notification system within the electronic medical record system can facilitate patient follow-up after ED discharge. Future quality improvement efforts aimed to further reduce admissions in this population should target patients with modifiable risk factors for serious bacterial infection.

Red blood cell alloimmunization in transfused patients with bone marrow failure syndromes.

BACKGROUND: Red blood cell (RBC) alloimmunization is a concern for patients who receive multiple or chronic transfusions. Alloimmunization prevalence in transfused patients with bone marrow failure syndrome (BMFS) is unknown. This study aimed to determine physician practice for RBC antigen matching, immunization rates, and antibody specificities in patients with BMFS. STUDY DESIGN AND METHODS: The clinical records of all patients with BMFS seen at the Children's Hospital of Philadelphia between 2001 and 2015 were reviewed. Immunization rate was determined per 100 units transfused. RESULTS: ABO/D, C, E, and K (CEK) RBC matching was requested for 21.8% of patients. A total of 3782 RBC units were transfused to 87 patients, of which 2551 (67.5%) were CEK matched and 1231 (32.5%) were ABO/D only matched. The majority of units transfused to patients on a chronic transfusion regimen were CEK matched (89.6% of 2728 units). No anti-C, -E, or -K antibodies formed in any patient during the 14-year study period. Two alloantibodies and two autoantibodies formed, resulting in a rate of 0.05 alloantibodies and 0.05 autoantibodies per 100 units transfused. The prevalence of alloimmunization was 2.3%. CONCLUSION: The rate and prevalence of RBC alloimmunization were low in patients with BMFS. CEK matching avoided alloimmunization to these antigens in chronically transfused patients.

Disrupted lymphocyte homeostasis in hepatitis-associated acquired aplastic anemia is associated with short telomeres.

Hepatitis-associated aplastic anemia (HAA) is a variant of acquired aplastic anemia (AA) in which immune-mediated bone marrow failure (BMF) develops following an acute episode of seronegative hepatitis. Dyskeratosis congenita (DC) is an inherited BMF syndrome characterized by the presence of short telomeres, mucocutaneous abnormalities, and cancer predisposition. While both conditions may cause BMF and hepatic impairment, therapeutic approaches are distinct, making it imperative to establish the correct diagnosis. In clinical practice, lymphocyte telomere lengths (TL) are used as a first-line screen to rule out inherited telomeropathies before initiating treatment for AA. To evaluate the reliability of TL in the HAA population, we performed a retrospective analysis of TL in 10 consecutively enrolled HAA patients compared to 19 patients with idiopathic AA (IAA). HAA patients had significantly shorter telomeres than IAA patients (P = 0.009), including four patients with TL at or below the 1st percentile for age-matched controls. HAA patients had no clinical features of DC and did not carry disease-causing mutations in known genes associated with inherited telomere disorders. Instead, short TLs were significantly correlated with severe lymphopenia and skewed lymphocyte subsets, features characteristic of HAA. Our results indicate the importance of caution in the interpretation of TL measurements in HAA, because, in this patient population, short telomeres have limited specificity.

Emergence of clonal hematopoiesis in the majority of patients with acquired aplastic anemia.

Acquired aplastic anemia (aAA) is a nonmalignant disease caused by autoimmune destruction of early hematopoietic cells. Clonal hematopoiesis is a late complication, seen in 20-25% of older patients. We hypothesized that clonal hematopoiesis in aAA is a more general phenomenon, which can arise early in disease, even in younger patients. To evaluate clonal hematopoiesis in aAA, we used comparative whole exome sequencing of paired bone marrow and skin samples in 22 patients. We found somatic mutations in 16 patients (72.7%) with a median disease duration of 1 year; of these, 12 (66.7%) were patients with pediatric-onset aAA. Fifty-eight mutations in 51 unique genes were found primarily in pathways of immunity and transcriptional regulation. Most frequently mutated was PIGA, with seven mutations. Only two mutations were in genes recurrently mutated in myelodysplastic syndrome. Two patients had oligoclonal loss of the HLA alleles, linking immune escape to clone emergence. Two patients had activating mutations in key signaling pathways (STAT5B (p.N642H) and CAMK2G (p.T306M)). Our results suggest that clonal hematopoiesis in aAA is common, with two mechanisms emerging-immune escape and increased proliferation. Our findings expand conceptual understanding of this nonneoplastic blood disorder. Future prospective studies of clonal hematopoiesis in aAA will be critical for understanding outcomes and for designing personalized treatment strategies.

Severe hemolytic transfusion reaction due to anti-D in a D+ patient with sickle cell disease.

A 5-year-old male with sickle cell disease presented with pain, dark urine, and fatigue 10 days after a red blood cell (RBC) transfusion. Laboratory evaluation demonstrated severe anemia, blood type O+, and anti-D in the serum. Anti-D in a D+ patient led to RH genotyping, which revealed homozygosity for RHD*DAU4 that encodes partial D antigen. Anti-D in this patient whose RBCs exclusively express partial D caused a delayed hemolytic transfusion reaction after exposure to D+ RBCs. The finding of anti-D in a D+patient should be investigated by molecular methods to help distinguish an alloantibody from an autoantibody.

Acquired aplastic anemia in children.

This article provides a practice-based and concise review of the etiology, diagnosis, and management of acquired aplastic anemia in children. Bone marrow transplantation, immunosuppressive therapy, and supportive care are discussed in detail. The aim is to provide the clinician with a better understanding of the disease and to offer guidelines for the management of children with this uncommon yet serious disorder.

Genome architecture marked by retrotransposons modulates predisposition to DNA methylation in cancer.

Epigenetic silencing plays an important role in cancer development. An attractive hypothesis is that local DNA features may participate in differential predisposition to gene hypermethylation. We found that, compared with methylation-resistant genes, methylation-prone genes have a lower frequency of SINE and LINE retrotransposons near their transcription start site. In several large testing sets, this distribution was highly predictive of promoter methylation. Genome-wide analysis showed that 22% of human genes were predicted to be methylation-prone in cancer; these tended to be genes that are down-regulated in cancer and that function in developmental processes. Moreover, retrotransposon distribution marks a larger fraction of methylation-prone genes compared to Polycomb group protein (PcG) marking in embryonic stem cells; indeed, PcG marking and our predictive model based on retrotransposon frequency appear to be correlated but also complementary. In summary, our data indicate that retrotransposon elements, which are widespread in our genome, are strongly associated with gene promoter DNA methylation in cancer and may in fact play a role in influencing epigenetic regulation in normal and abnormal physiological states.

NUP98-HOX translocations lead to myelodysplastic syndrome in mice and men.

The myelodysplastic syndromes (MDS) are a group of clonal hematopoietic stem cell disorders characterized by ineffective hematopoiesis, peripheral blood cytopenias, dysplasia, and a propensity for transformation to acute myeloid leukemia (AML). A wide spectrum of genetic aberrations has been associated with MDS, including chromosomal translocations involving the NUP98 gene, most commonly leading to fusions of NUP98 with abd-b group HOX genes, including HOXD13. We used vav regulatory elements to direct expression of a NUP98-HOXD13 (NHD13) fusion gene in hematopoietic tissues. NHD13 transgenic mice faithfully recapitulate all the key features of MDS, including peripheral blood cytopenias, bone marrow dysplasia and apoptosis, and transformation to acute leukemia. The MDS that develops in NHD13 transgenic mice is highly lethal; within 14 months, 90% of the mice died of either leukemic transformation or severe anemia and leukopenia due to progressive MDS. These mice provide a preclinical model that can be used for the evaluation of MDS therapy and biology.

A neurologic presentation of familial hemophagocytic lymphohistiocytosis which mimicked septic emboli to the brain.

Familial hemophagocytic lymphohistiocytosis is an inherited deficiency of natural killer cell function and excessive cytokine activity, which predominantly presents in early childhood. The initial symptoms of familial hemophagocytic lymphohistiocytosis are often nonspecific but may be predominantly neurologic. The case presented here describes an 18-month-old boy who initially presented with fever, encephalopathy, and hemiparesis. He had innumerable brain lesions visualized on magnetic resonance imaging scans. An infectious etiology was excluded, and brain, liver, and bone marrow biopsies were nonspecific but consistent with hemophagocytic lymphohistiocytosis. Cells were sent for flow cytometry perforin analysis, which demonstrated defective natural killer cell function. A diagnosis of familial hemophagocytic lymphohistiocytosis was confirmed by mutation analysis and decreased expression of the perforin gene, in the patient and immediate family members. These results showed the patient to be a compound heterozygote for perforin mutations. His case illustrates the potential for a fulminant neurological presentation of familial hemophagocytic lymphohistiocytosis with widespread lesions in the brain.

Retroviral insertional mutagenesis identifies genes that collaborate with NUP98-HOXD13 during leukemic transformation.

The t(2;11)(q31;p15) chromosomal translocation results in a fusion between the NUP98 and HOXD13 genes and has been observed in patients with myelodysplastic syndrome (MDS) or acute myelogenous leukemia. We previously showed that expression of the NUP98-HOXD13 (NHD13) fusion gene in transgenic mice results in an invariably fatal MDS; approximately one third of mice die due to complications of severe pancytopenia, and about two thirds progress to a fatal acute leukemia. In the present study, we used retroviral insertional mutagenesis to identify genes that might collaborate with NHD13 as the MDS transformed to an acute leukemia. Newborn NHD13 transgenic mice and littermate controls were infected with the MOL4070LTR retrovirus. The onset of leukemia was accelerated, suggesting a synergistic effect between the NHD13 transgene and the genes neighboring retroviral insertion events. We identified numerous common insertion sites located near protein-coding genes and confirmed dysregulation of a subset of these by expression analyses. Among these genes were Meis1, a known collaborator of HOX and NUP98-HOX fusion genes, and Mn1, a transcriptional coactivator involved in human leukemia through fusion with the TEL gene. Other putative collaborators included Gata2, Erg, and Epor. Of note, we identified a common insertion site that was >100 kb from the nearest coding gene, but within 20 kb of the miR29a/miR29b1 microRNA locus. Both of these miRNA were up-regulated, demonstrating that retroviral insertional mutagenesis can target miRNA loci as well as protein-coding loci. Our data provide new insights into NHD13-mediated leukemogenesis as well as retroviral insertional mutagenesis mechanisms.