The incidence of ultra-rare bleeding coagulation disorders is unknown but is critically important to understand the need for specialized health care services and linkage of individuals to treatment. However, the ability to identify and characterize these disorders may be difficult based upon the absence of specialized testing and the presence of more than one condition, which may impact bleeding phenotype. To better understand the nature and frequency of ultra-rare bleeding coagulation disorders in the US, the American Thrombosis and Hemostasis Network (ATHN) initiated ATHN 10, a collaborative effort within its network of 140 ATHN-affiliated hemophilia treatment centers (HTCs), to perform whole gene sequencing and collection of detailed phenotypic data, including bleeding scores on enrolled patients with one of 31 identified ultra-rare bleeding coagulation disorders. Thus far, 1,225 individuals with an established ultra-rare bleeding coagulation disorder have been sequenced for their expected factor deficiency in a panel that includes their targeted gene along with 29 other clotting factor and selected platelet protein genes to determine the presence of novel variants and possibility of more than one disorder impacting the bleeding phenotype. Over 20,720 whole gene sequences have been analyzed in ATHN 10 with 176 novel variants being identified. In 31% of individuals, an additional mutation was found in a second gene not suspected in the original diagnosis. Some of the secondary variants, resulted in a decreased production of the second affected factor, which possibly influenced the bleeding phenotype more than expected for identified deficiency alone. These secondary deficiencies, or modifying variants, were surprisingly common and found in participants across 8 regions in the US Hemophilia Treatment Center Network, with unique clusters identified. Although most of the 176 novel variants, not previously described in the literature, were identified in single individuals, there were 6 novel variants on 5 different genes that were found in multiple individuals. The variants F10: R445H, ITGB3: 615-1G>C, and F7: G406D were found in individuals at different HTCs. Variants F7: G432S, FGB: 719-9_719-2delCATTTGCA, and FGG: T80fs*7 were found in three multigenerational family studies at the same HTC. By leveraging the infrastructure at ATHN-affiliated HTCs and the laboratory services at the Center for Inherited Blood Disorders (CIBD)/Hematology Advanced Diagnostic Laboratory (CAP/CLIA Certified), the ATHN 10 project reduced the barriers associated with genetic testing and has expanded knowledge of the incidence of new pathogenic variants at reduced cost, allowing HTCs to provide more specific care and genetic counseling. In addition, whole gene sequencing has confirmed the role of certain genetic hot spots in clotting factor genes as well as the role of splice site variants in determining factor activity levels and bleeding phenotypic variance. This information not only helps patients and physicians, but also provides researchers with valuable resources to further our understanding of the pathophysiology of factor function and deficiencies.

Background and Significance: Exercise intolerance and dyspnea on exertion (DOE) are frequent after pulmonary embolism (PE) despite anticoagulation and may impact quality of life. There is little research that fully explains why this occurs, nor are there specific treatments or guidelines for PE complications in children or young adults. The FUVID Program (NCT04583878) will examine the impact of 3 mechanisms underlying exercise intolerance and DOE: cardiac (right ventricular maladaptation and its coupling with the pulmonary circulation) dysfunction, pulmonary limitations secondary to persistent vascular obstruction, and skeletal muscle metabolic abnormalities on exercise intolerance and DOE after PE. Study Design and Methods: This prospective, multi-center, observational study is enrolling previously healthy patients ages 8-21 years with acute, first-episode PE with or without DVT. A positive control group with no PE but with physical activity restrictions mimicking a deconditioning effect as patients with PE (cohort 1) and control group with no prescribed physical activity restrictions and otherwise considered healthy (cohort 2) will be enrolled. Patients will be enrolled within 8 weeks of diagnosis of PE from 13 enrollment and 25 recruitment sites across the US. Due to the complexity of the cutting-edge metabolic and functional tools only available at UTSW (namely, MR spectroscopy at 7 Telsa to study in-vivo muscle metabolic function and perfusion and exercise cardiac MRI), research assessments will occur centrally at 3- and 12-months post-diagnosis. Exercise capacity will be defined as peak oxygen uptake (VO2), expressed as a percent predicted based on ideal body weight during cardiopulmonary exercise testing. The primary comparison will be between participants with and without exercise intolerance. Key secondary subgroup analyses will include: sex (male/female), PE category (low risk/non-low-risk), and thrombolysis (yes/no). Results/Discussion: We have enrolled 67 PE participants (planned=80) from participating enrollment sites and 30 case-controls (planned=30). We propose that 1) the right ventricle will be unable to overcome afterload placed on it during exercise, preventing optimal coupling with pulmonary circulation, 2) arterial desaturation during maximal cycling exercise combined with dysfunctional breathing of deconditioning will provoke greater DOE and intolerance, and 3) depletion and recovery of muscle phosphocreatine in-vivo will predict exercise intolerance. Our results will provide critically informative data to phenotype pediatric post-PE syndrome accurately and allow investigation of cardiac, vascular, and musculoskeletal mechanisms associated with post-PE syndrome assessed during exercise.

With increasing early and upfront use of rituximab and caplacizumab in the modern management of immune-mediated thrombotic thrombocytopenic purpura (iTTP), the risk of refractory disease is expected to decline. However, despite the use of adequate initial therapy, a small subset of patients develop a refractory disease which is difficult to manage. Bortezomib has come to be known as a safe and effective treatment option for refractory iTTP, but its use in children is limited. Here, we describe the case of an adolescent patient with refractory iTTP who had a satisfactory and sustained response to the use of bortezomib.

Primary erythrocytoses (PE) include germline gain of function mutations of erythropoietin (EPO) receptor (EPOR) gene and polycythemia vera (PV), a clonal disease due to acquired somatic JAK2 mutations. Both these PEs have intrinsically abnormal early erythroid progenitors (BFU-E) characterized by their in vitro EPO hypersensitive growth (EPORmutations), or even EPO independent growth (the hallmark of PV); both have low plasma EPO (Chapter 58, Williams Hematology, 10e). Secondary erythrocytoses (SE) have intrinsically normal BFU-Es but are caused by increased circulating erythropoiesis stimulators, chiefly EPO. In contrast, congenital Chuvash erythrocytosis is characterized by augmentation of hypoxia inducible factors (HIFs) causing high EPO and hypersensitive BFU-E's EPO responses; thus, it has features of both PE and SE. We report a 7-year-old normoxic girl with history of symptomatic erythrocytosis since birth of unknown etiology, with hemoglobin of 22 g/dL at birth, consistently elevated EPO at 50-75 mU/mL, and normal hemoglobin oxygen affinity. Her BFU-E grew without EPO yet no somatic PV defining mutations were found. A targeted analysis of previously described erythrocytoses genes was negative. Whole genome sequencing, performed under the auspices of Undiagnosed Disease Network, identified a heterozygous missense variant (p. Lys769Leu) of CMTR1(Cap Methyltransferase 1) gene, not listed in the Genome Aggregation Database (gnomAD). Cap methylation on mRNA protects mRNA from being degraded and promotes RNA translation by recruiting translation initiation factors (PMID:30312682). The variant was present in the proband but not in her asymptomatic mother (father was not available for the analysis). This variant was found in leukocytes and in nail DNA, indicating germline origin. Transcriptional X-chromosome inactivation assay (PMID:18641369) revealed polyclonal granulocytes and platelets, confirming that the erythrocytosis is caused by germline mutation, not by somatic mutation-driven clonal hematopoiesis. Because of her high EPO, we hypothesized that CMTR1K769L variant might cause high HIF activity. HIF-mediated gene regulation is tissue- and stage of differentiation-specific. Expression of HIF target genes SLC29A1, VEGFA, EDN1, and LDHA was upregulated in granulocytes while in platelets all target genes except SLC29A1 were upregulated. To test if CMTR1 regulates HIF transcriptional activity, 1) we correlated CMTR1 transcript levels with HIF target gene expression levels in granulocytes and platelets. CMTR1 transcript levels were positively correlated with expression levels of HIF target genes. 2) we tested if CMTR1 overexpression and knockdown alters HIF target gene expression. In erythroid cell lines (K562 and HEL), overexpressed CMTR1 increased SLC2A1 and VEGFA (HIF target genes transcripts) while knockdown of CMTR1 downregulated their expression. This demonstrates that CMTR1 positively regulates HIF transcriptional activity, suggesting that CMTR1K769L accounts for increased plasma EPO and for EPO mediated feature of SE. As erythroid progenitors express endogenous erythropoietin (PMID 9573013) which contributes to their differentiation and proliferation (PMID 10903342), we evaluated if CMTR1K769L also promotes endogenous erythroid EPO accounting for EPO independent growth of CMTR1K769L mutated BFU-Es and thus provides potential mechanism as a feature of PEfor the erythrocytosis. The proband's endogenous EPO transcript levels in in vitro expanded erythroid cells at day 4, when most of cells are erythroid progenitors, were 10 times higher than the mother's see Figure. We present a new cause of erythrocytosis associated with the novel germline mutation of principal epigenetic modifier of RNA, CMTR1 (PMID:33922187), associated with high EPO (SE). Her BFU-E grew without extrinsic EPO (PE) likely due to previously unobserved stimulation of EPO production in erythroid progenitors. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Background:A rise in hospital-acquired venous thromboembolism (HA-VTE) in children has led to increased awareness regarding VTE prophylaxis and risk assessment. Despite no consensus exists regarding these practices in pediatrics.Objective:To describe common practices in VTE prophylaxis, VTE risk assessment models, and anticoagulation dosing strategies in pediatric hospitals that are members of the Children's Hospital Acquired Thrombosis (CHAT) Consortium.Methods:An electronic survey of 44 questions evaluating practices surrounding pediatric HA-VTE risk assessment and prevention was distributed between August 9, 2021, and August 30, 2021, to the primary investigators from the 32 institutions within the CHAT Consortium.Results:The survey response rate was 100% (n = 32). In total, 85% (n = 27) of the institutions assess HA-VTE, but only 63% (n = 20) have formal hospital guidelines. Within the institutions with formal guidelines, 100% (n = 20) include acute systemic inflammation or infection and presence of a central venous catheter (CVC) as risk factors for VTE. Pharmacologic prophylaxis is prescribed at 87% (28) of institutions, with enoxaparin being the most frequent (96%, n = 27). Variability in responses persisted regarding risk factors, risk assessment, thromboprophylaxis, dosing of prophylactic anticoagulation or anticoagulant drug monitoring. A majority of providers were comfortable providing thromboprophylaxis across all age groups. In addition, the global coronavirus disease 2019 increased the providers' use of prophylactic anticoagulation 78% (n = 25).Conclusion:Practices among institutions are variable in regard to use of HA-VTE prophylaxis, risk assessment, or guideline implementation, highlighting the need for further research and a validated risk assessment model through groups like the CHAT Consortium.

Trisomy 18 is a well-documented cause of spontaneous abortions. When live births do occur, median survival time has been estimated between two and fourteen days. This severely shortened life expectancy is attributed to potentially lethal congenital malformations involving the heart, central nervous system, and gastrointestinal system, among others. Infrequently, affected children survive beyond a year and evidence suggests this may become more common due to increased willingness to intervene in the neonatal period and the success of surgical corrections. This case report documents how a six-year-old female with trisomy 18 has challenged provider expectations and gives perspective in an emerging treatment gray-zone.

Post-Pulmonary Embolism impairment (PPEI), also called Post-PE syndrome is being increasingly recognized as a long-term sequelae of pulmonary embolism (PE) in adults, but data regarding its prevalence in pediatrics is unknown. PPEI likely represents a spectrum of severity, with the most severe manifestation being chronic thromboembolic pulmonary hypertension (CTEPH) affecting a small proportion of individuals, and with many individuals with less severe PPEI suffering from reduced quality of life and functional limitations. The pathophysiology is poorly understood but involves residual pulmonary artery thrombi leading to right ventricular damage and abnormal gas exchange. The prevalence, risk factors and a consensus definition in the pediatric and adolescent populations is unknown, as are optimum treatment strategies. In addition to physical sequelae, psychological consequences of PE are significant and also under recognized.

Multisystem inflammatory syndrome of children (MIS-C) continues to be a highly concerning diagnosis in those recently infected with SARS-CoV-2. The diagnosis of MIS-C cases will likely become even more challenging as vaccine uptake and natural immunity in previously infected persons leads to lower circulating rates of SARS-CoV2 infection and will make cases sporadic. Febrile children presenting with cardiac dysfunction, symptoms overlapping Kawasaki disease or significant gastrointestinal complaints warrant a thorough screen in emergency departments, urgent care centers, and outpatient pediatric or family medicine practices. An increased index of suspicion and discussion regarding higher level of care (transferring to pediatric tertiary care centers or to intensive care) continues to be recommended. Herein we outline a broad approach with a multidisciplinary team for those meeting the case definition and believe such an approach is crucial for successful outcomes.

Low molecular weight heparin (LMWH) remains the most commonly prescribed pediatric anticoagulant. There is debate whether LMWH anti‐Xa assays with or without exogenous antithrombin (AT) best reflect anticoagulation effect, and how much discrepancy exists between assay types.

Introduction: Obesity is associated with endothelial dysfunction, hemostatic and fibrinolytic disturbances. The relationship between obesity and elevated Von Willebrand Factor (VWF) is complex and not fully elucidated. There is a significant knowledge gap regarding the impact of BMI on VWF levels. Given the proinflammatory effect associated with abdominal obesity, we hypothesized that there would be an increased prevalence of obesity among individuals with Low VWF (LVWF) compared to Type 1 Von Willebrand Disease (T1VWD) in the ATHN (American Thrombosis and Hemostasis Network) dataset. Methods: A retrospective review of de-identified patients included in the ATHN dataset as of March 2018 was performed. The dataset was queried for all patients with a diagnosis of "T1VWD," who were over 18 years of age when labs were drawn and when BMI was recorded, who had VWF Ristocetin cofactor (RCO) levels <50 IU/dL and who had BMI entered within 24 months of the date of lab entry. Subjects were categorized with VWF RCO ≤30% as T1VWD and 30-50% as LVWF. We used the NIH definitions for BMI (BMI<18.5, underweight; 18.5-24.9, normal; 25-29.9, overweight; 30-39.9, obese; BMI >40, extremely obese). Results: Of the 6939 patients with T1VWD in the ATHN dataset, 4754 patients had VWF RCO <50%, 1019 were above the age of 18, resulting in 548 evaluable subjects with BMI and laboratory metrics. There were 186 patients in the T1VWD cohort, and 362 patients in the LVWF cohort, with a Female:Male ratio >3:1 (Table 1). BMI was treated as a continuous measurement and on bivariate analysis there was not a statistically significant difference (p=0.593), with mean BMI 28.2 (17.2-52) in T1VWD and 28.6 (15.3-55.4) in LVWF. Prevalence of obesity (BMI ≥30) was not significantly different between cohorts (T1VWF 32% vs. LVWF 36%, p=0.345, Table 1). The prevalence of obesity by age (18-39, 40-59, >60 years) was similar among both cohorts with the exception of a larger proportion of obese individuals over the age of 60 in the LVWF cohort (63% vs 25%). Mean FVIII level for LVWF cohort was significantly higher compared to that for T1VWD cohort (80% vs. 53%; p<0.001). In addition, extremely obese patients had an elevated mean FVIII level compared to overweight patients (81% vs. 60%; p=0.041, Table 2). Among individuals with BMI≥30, there were increased rates for Black race (p=0.013), and Medicaid and Medicare rates (p=0.028) when compared to non-obese individuals (Table 3). While rates of obesity are known to vary regionally, no conclusions could be drawn as there was disproportionate geographic clustering in states with well-established hemophilia treatment centers. Conclusions: Our analysis identified that 34.8% of adults categorized as VWD in the ATHN data set are obese, with similar prevalence among T1VWD and LVWF. This finding, coupled with associated race, ethnic and socioeconomic risk factors can help prioritize prevention and weight management as a critical component of the comprehensive care model. Increasing degree of obesity may be associated with elevated FVIII, and should be studied prospectively in larger cohorts and the potential impact on cardiovascular risk. Limitations of this analysis include incomplete laboratory data, lack of longitudinal laboratory data, unknown potential confounders including pregnancy or medication effect, and non-uniform geographic distribution of patients. Further research is needed to evaluate the impact of obesity on bleeding phenotype, bleeding related complications, and management practices, as well as the effects of weight change on VWF and FVIII levels. Disclosures Schaefer: Siemens: Research Funding; Stago: Research Funding.

Introduction: Enoxaparin (low molecular weight heparin, LMWH) is the most commonly prescribed anticoagulant for pediatric patients with venous thromboembolism (VTE) (Goldenberg et al. 2015). The gold-standard LMWH activity (anti-Xa activity) assays differ in whether they add exogenous antithrombin (AT) or dextran sulfate (Ignjatovic et al. 2007). Adding AT would "standardize" results in patients with low antithrombin, such as infants and asparaginase-treated patients (Mitchell et al 2010); however, questions remain about which assay best reflects the patient's anticoagulation effect and the degree of discrepancy between assays. We assessed LMWH activity in residual plasma samples from a cohort of anticoagulated pediatric acute lymphoblastic leukemia and lymphoma (ALL) patients, with history of VTE and variable AT levels, on four platforms (two instruments and their kits +/- exogenous AT). Methods: We analyzed 60 de-identified residual plasma samples from 12 anticoagulated ALL patients (2-19 years. Mean 13.75 yr) who had AT levels obtained for clinical care. All consented to the IRB-approved Oncology Tissue Repository. LMWH activity was assessed on Siemens and Stago instruments using their recommended kits that did or did not contain exogenous AT (Table 1), according to manufacturer recommendations, by experienced laboratorians. Results: Results were interpretable on 236/240 with 4 rejected for lipemia. Mean AT activity was 80 (42-138 ng/ml, lab normal >81%). Correlation was acceptable for the published kit ranges of LMWH activity when comparing kits +AT (Berichrom® to Stachrom®, r=0.82, p<0.0001), -AT (Innovance® to STA®-Liquid Anti-Xa: r=0.93, p<0.0001), and within the same manufacturer (Berichrom® to Innovance®, r=0.92, p<0.0001, Stachrom® to STA®-Liquid Anti-Xa r=0.98, p<0.0001) (Table 2, Figure 1, Figure 2). Comparing +AT or not by manufacturer, there was a nonsignificant trend to higher LMWH activity result with +AT kits. When AT levels were <70 ng/ml (n=19, mean 56 ng/dl) there was a trend to underestimate LMWH activity when AT was not added; this reached significance when the Stago methods were compared (Table 2.) Conclusions: There is acceptable correlation of LMWH activity using kits with or without exogenous AT in anticoagulated pediatric plasma samples; however, in low AT samples, LMWH activity trends lower in platforms without exogenous AT. Use of residual samples precluded conclusions about the clinical impact of this difference; however, there were a few instances where subtherapeutic activity in the -AT assay was in the therapeutic range (0.5-1 activity units) in the +AT assay. Clinicians should know whether exogenous AT is used in their own laboratory's assay and understand that added AT could overestimate the LMWH effect in patients with low AT levels. Disclosures Schaefer: Stago: Research Funding; Siemens: Research Funding.

Pediatric RCC is a rare pediatric neoplasm and is distinctly different compared to adult RCC, often demonstrating translocation morphology evidenced by unique histopathological features and TFE3 or TFEB nuclear expression. We report three cases of pediatric TFE3 positive RCC (TFE3-RCC) occurring in the setting of chronic kidney disease and long-term pharmacological immunosuppression, including two cases that developed in the native kidney following kidney transplantation. Together, these cases suggest that the kidney microenvironment in combination with immune dysregulation is likely contributing factors in the pathogenesis of some pediatric RCC, warranting further study. Long-term post-transplant surveillance may warrant screening for RCC.