Plasma cell-free RNA signatures of inflammatory syndromes in children.

Inflammatory syndromes, including those caused by infection, are a major cause of hospital admissions among children and are often misdiagnosed because of a lack of advanced molecular diagnostic tools. In this study, we explored the utility of circulating cell-free RNA (cfRNA) in plasma as an analyte for the differential diagnosis and characterization of pediatric inflammatory syndromes. We profiled cfRNA in 370 plasma samples from pediatric patients with a range of inflammatory conditions, including Kawasaki disease (KD), multisystem inflammatory syndrome in children (MIS-C), viral infections, and bacterial infections. We developed machine learning models based on these cfRNA profiles, which effectively differentiated KD from MIS-C-two conditions presenting with overlapping symptoms-with high performance [test area under the curve = 0.98]. We further extended this methodology into a multiclass machine learning framework that achieved 80% accuracy in distinguishing among KD, MIS-C, viral, and bacterial infections. We further demonstrated that cfRNA profiles can be used to quantify injury to specific tissues and organs, including the liver, heart, endothelium, nervous system, and the upper respiratory tract. Overall, this study identified cfRNA as a versatile analyte for the differential diagnosis and characterization of a wide range of pediatric inflammatory syndromes.

Treatment of Multisystem Inflammatory Syndrome in Children.

BACKGROUND: Evidence is urgently needed to support treatment decisions for children with multisystem inflammatory syndrome (MIS-C) associated with severe acute respiratory syndrome coronavirus 2. METHODS: We performed an international observational cohort study of clinical and outcome data regarding suspected MIS-C that had been uploaded by physicians onto a Web-based database. We used inverse-probability weighting and generalized linear models to evaluate intravenous immune globulin (IVIG) as a reference, as compared with IVIG plus glucocorticoids and glucocorticoids alone. There were two primary outcomes: the first was a composite of inotropic support or mechanical ventilation by day 2 or later or death; the second was a reduction in disease severity on an ordinal scale by day 2. Secondary outcomes included treatment escalation and the time until a reduction in organ failure and inflammation. RESULTS: Data were available regarding the course of treatment for 614 children from 32 countries from June 2020 through February 2021; 490 met the World Health Organization criteria for MIS-C. Of the 614 children with suspected MIS-C, 246 received primary treatment with IVIG alone, 208 with IVIG plus glucocorticoids, and 99 with glucocorticoids alone; 22 children received other treatment combinations, including biologic agents, and 39 received no immunomodulatory therapy. Receipt of inotropic or ventilatory support or death occurred in 56 patients who received IVIG plus glucocorticoids (adjusted odds ratio for the comparison with IVIG alone, 0.77; 95% confidence interval [CI], 0.33 to 1.82) and in 17 patients who received glucocorticoids alone (adjusted odds ratio, 0.54; 95% CI, 0.22 to 1.33). The adjusted odds ratios for a reduction in disease severity were similar in the two groups, as compared with IVIG alone (0.90 for IVIG plus glucocorticoids and 0.93 for glucocorticoids alone). The time until a reduction in disease severity was similar in the three groups. CONCLUSIONS: We found no evidence that recovery from MIS-C differed after primary treatment with IVIG alone, IVIG plus glucocorticoids, or glucocorticoids alone, although significant differences may emerge as more data accrue. (Funded by the European Union's Horizon 2020 Program and others; BATS ISRCTN number, ISRCTN69546370.).

Intravenous immunoglobulin resistance in Kawasaki disease patients: prediction using clinical data.

BACKGROUND: About 10-20% of Kawasaki disease (KD) patients are resistant to the initial infusion of intravenous immunoglobin (IVIG). The aim of this study was to assess whether IVIG resistance in KD patients could be predicted using standard clinical and laboratory features. METHODS: Data were from two cohorts: a Korean cohort of 7101 KD patients from 2015 to 2017 and a cohort of 649 KD patients from San Diego enrolled from 1998 to 2021. Features included laboratory values, the worst Z-score from the initial echocardiogram or during hospitalization, and the five clinical KD signs at presentation. RESULTS: Five machine learning models achieved a maximum median AUC of 0.711 [IQR: 0.706-0.72] in the Korean cohort and 0.696 [IQR: 0.609-0.722] in the San Diego cohort during stratified 10-fold cross-validation using significant laboratory features identified from univariate analysis. Adding the Z-score, KD clinical signs, or both did not considerably improve the median AUC in either cohort. CONCLUSIONS: Using commonly measured clinical laboratory data alone or in conjunction with echocardiographic findings and clinical features is not sufficient to predict IVIG resistance. Further attempts to predict IVIG resistance will need to incorporate additional data such as transcriptomics, proteomics, and genetics to achieve meaningful predictive utility. IMPACT: We demonstrated that laboratory, echocardiographic, and clinical findings cannot predict intravenous immunoglobin (IVIG) resistance to a clinically meaningful extent using machine learning in a homogenous Asian or ethnically diverse population of patients with Kawasaki disease (KD). Visualizing these features using uniform manifold approximation and projection (UMAP) is an important step to evaluate predictive utility in a qualitative manner. Further attempts to predict IVIG resistance in KD patients will need to incorporate novel biomarkers or other specialized features such as genetic differences or transcriptomics to be clinically useful.

Risk Factors Associated with Intensive Care Admission in Children with Severe Acute Respiratory Syndrome Coronavirus 2-Related Multisystem Inflammatory Syndrome (MIS-C) in Latin America: A Multicenter Observational Study of the REKAMLATINA Network.

Background: Multisystem inflammatory syndrome in children (MIS-C) associated with coronavirus disease 2019 varies widely in its presentation and severity, with low mortality in high-income countries. In this study in 16 Latin American countries, we sought to characterize patients with MIS-C in the pediatric intensive care unit (PICU) compared with those hospitalized on the general wards and analyze the factors associated with severity, outcomes, and treatment received. Study Design: An observational ambispective cohort study was conducted including children 1 month to 18 years old in 84 hospitals from the REKAMLATINA network from January 2020 to June 2022. Results: A total of 1239 children with MIS-C were included. The median age was 6.5 years (IQR 2.5-10.1). Eighty-four percent (1043/1239) were previously healthy. Forty-eight percent (590/1239) were admitted to the PICU. These patients had more myocardial dysfunction (20% vs 4%; P < 0.01) with no difference in the frequency of coronary abnormalities (P = 0.77) when compared to general ward subjects. Of the children in the PICU, 83.4% (494/589) required vasoactive drugs, and 43.4% (256/589) invasive mechanical ventilation, due to respiratory failure and pneumonia (57% vs 32%; P = 0.01). On multivariate analysis, the factors associated with the need for PICU transfer were age over 6 years (aOR 1.76 95% CI 1.25-2.49), shock (aOR 7.06 95% CI 5.14-9.80), seizures (aOR 2.44 95% CI 1.14-5.36), thrombocytopenia (aOR 2.43 95% CI 1.77-3.34), elevated C-reactive protein (aOR 1.89 95% CI 1.29-2.79), and chest x-ray abnormalities (aOR 2.29 95% CI 1.67-3.13). The overall mortality was 4.8%. Conclusions: Children with MIS-C who have the highest risk of being admitted to a PICU in Latin American countries are those over age six, with shock, seizures, a more robust inflammatory response, and chest x-ray abnormalities. The mortality rate is five times greater when compared with high-income countries, despite a high proportion of patients receiving adequate treatment.

Characterization of SARS-CoV-2 and common cold coronavirus-specific T-cell responses in MIS-C and Kawasaki disease children.

The immunopathogenesis of multisystem inflammatory syndrome (MIS-C) in children that may follow exposure to SARS-CoV-2 is incompletely understood. Here, we studied SARS-CoV-2-specific T cells in MIS-C, Kawasaki disease (KD), and SARS-CoV-2 convalescent controls using peptide pools derived from SARS-CoV-2 spike or nonspike proteins, and common cold coronaviruses (CCC). Coordinated CD4+ and CD8+ SARS-CoV-2-specific T cells were detected in five MIS-C subjects with cross-reactivity to CCC. CD4+ and CD8+ T-cell responses alone were documented in three and one subjects, respectively. T-cell specificities in MIS-C did not correlate with disease severity and were similar to SARS-CoV-2 convalescent controls. T-cell memory and cross-reactivity to CCC in MIS-C and SARS-CoV-2 convalescent controls were also similar. The chemokine receptor CCR6, but not CCR9, was highly expressed on SARS-CoV-2-specific CD4+ but not on CD8+ T cells. Only two of 10 KD subjects showed a T-cell response to CCC. Enumeration of myeloid APCs revealed low cell precursors in MIS-C subjects compared to KD. In summary, children with MIS-C mount a normal T-cell response to SARS-CoV-2 with no apparent relationship to antecedent CCC exposure. Low numbers of tolerogenic myeloid DCs may impair their anti-inflammatory response.

Clinical Presentation and Outcomes of Kawasaki Disease in Children from Latin America: A Multicenter Observational Study from the REKAMLATINA Network.

OBJECTIVES: To describe the clinical presentation, management, and outcomes of Kawasaki disease (KD) in Latin America and to evaluate early prognostic indicators of coronary artery aneurysm (CAA). STUDY DESIGN: An observational KD registry-based study was conducted in 64 participating pediatric centers across 19 Latin American countries retrospectively between January 1, 2009, and December 31, 2013, and prospectively from June 1, 2014, to May 31, 2017. Demographic and initial clinical and laboratory data were collected. Logistic regression incorporating clinical factors and maximum coronary artery z-score at initial presentation (between 10 days before and 5 days after intravenous immunoglobulin [IVIG]) was used to develop a prognostic model for CAA during follow-up (>5 days after IVIG). RESULTS: Of 1853 patients with KD, delayed admission (>10 days after fever onset) occurred in 16%, 25% had incomplete KD, and 11% were resistant to IVIG. Among 671 subjects with reported coronary artery z-score during follow-up (median: 79 days; IQR: 36, 186), 21% had CAA, including 4% with giant aneurysms. A simple prognostic model utilizing only a maximum coronary artery z-score ≥2.5 at initial presentation was optimal to predict CAA during follow-up (area under the curve: 0.84; 95% CI: 0.80, 0.88). CONCLUSION: From our Latin American population, coronary artery z-score ≥2.5 at initial presentation was the most important prognostic factor preceding CAA during follow-up. These results highlight the importance of early echocardiography during the initial presentation of KD.

Bridging a diagnostic Kawasaki disease classifier from a microarray platform to a qRT-PCR assay.

BACKGROUND: Kawasaki disease (KD) is a systemic vasculitis that mainly affects children under 5 years of age. Up to 30% of patients develop coronary artery abnormalities, which are reduced with early treatment. Timely diagnosis of KD is challenging but may become more straightforward with the recent discovery of a whole-blood host response classifier that discriminates KD patients from patients with other febrile conditions. Here, we bridged this microarray-based classifier to a clinically applicable quantitative reverse transcription-polymerase chain reaction (qRT-PCR) assay: the Kawasaki Disease Gene Expression Profiling (KiDs-GEP) classifier. METHODS: We designed and optimized a qRT-PCR assay and applied it to a subset of samples previously used for the classifier discovery to reweight the original classifier. RESULTS: The performance of the KiDs-GEP classifier was comparable to the original classifier with a cross-validated area under the ROC curve of 0.964 [95% CI: 0.924-1.00] vs 0.992 [95% CI: 0.978-1.00], respectively. Both classifiers demonstrated similar trends over various disease conditions, with the clearest distinction between individuals diagnosed with KD vs viral infections. CONCLUSION: We successfully bridged the microarray-based classifier into the KiDs-GEP classifier, a more rapid and more cost-efficient qRT-PCR assay, bringing a diagnostic test for KD closer to the hospital clinical laboratory. IMPACT: A diagnostic test is needed for Kawasaki disease and is currently not available. We describe the development of a One-Step multiplex qRT-PCR assay and the subsequent modification (i.e., bridging) of the microarray-based host response classifier previously described by Wright et al. The bridged KiDs-GEP classifier performs well in discriminating Kawasaki disease patients from febrile controls. This host response clinical test for Kawasaki disease can be adapted to the hospital clinical laboratory.

Cardiovascular toxicities with pediatric tyrosine kinase inhibitor therapy: An analysis of adverse events reported to the Food and Drug Administration.

We sought to examine cardiovascular toxicities associated with tyrosine kinase inhibitors in pediatrics. We examined 1624 pediatric adverse events with imatinib, dasatinib, sorafenib, pazopanib, crizotinib, and ruxolitinib reported to the Food and Drug Administration between January 1, 2015, and August 14, 2020. There were 102 cardiovascular event reports. Hypertension was the most commonly reported cardiovascular event and was most frequently associated with sorafenib and pazopanib. The presence of infection increased the reporting odds of cardiovascular events overall and specifically cardiac arrest, heart failure, and hypertension. These data provide early insight into cardiovascular toxicities with tyrosine kinase inhibitor use in pediatrics.

Addressing sequential and concurrent treatment regimens in a small n sequential, multiple assignment, randomized trial (snSMART) in the MISTIC study.

Multisystem Inflammatory Syndrome in children (MIS-C) is a rare and novel pediatric complication linked to COVID-19 exposure, which was first identified in April 2020. A small n, Sequential, Multiple Assignment, Randomized Trial (snSMART) was applied to the Multisystem Inflammatory Syndrome Therapies in Children Comparative Effectiveness Study (MISTIC) to efficiently evaluate multiple competing treatments. In the MISTIC snSMART study, participants are randomized to one of three interventions (steroids, infliximab or anakinra), and potentially re-randomized to the remaining two treatments depending on their response to the first randomized treatment. However, given the novelty and urgency of the MIS-C disease, in addition to patient welfare concerns, treatments were not always administered sequentially, but allowed to be administered concurrently if deemed medically necessary. We propose a pragmatic modification to the original snSMART design to address the analysis of concurrent versus sequential treatments in the MISTIC study. A modified Bayesian joint stage model is developed that can distinguish a concurrent treatment effect from a sequential treatment effect. A simulation study is conducted to demonstrate the improved accuracy and efficiency of the primary aim to estimate the first stage treatments' response rates and the secondary aim to estimate the combined first and second stage treatments' responses in the proposed model compared to the standard snSMART Bayesian joint stage model. We observed that the modified model has improved efficiency in terms of bias and rMSE under large sample size settings.

Endothelial Cell Response in Kawasaki Disease and Multisystem Inflammatory Syndrome in Children.

Although Kawasaki disease (KD) and multisystem inflammatory syndrome in children (MIS-C) share some clinical manifestations, their cardiovascular outcomes are different, and this may be reflected at the level of the endothelial cell (EC). We performed RNA-seq on cultured ECs incubated with pre-treatment sera from KD (n = 5), MIS-C (n = 7), and healthy controls (n = 3). We conducted a weighted gene co-expression network analysis (WGCNA) using 935 transcripts differentially expressed between MIS-C and KD using relaxed filtering (unadjusted p < 0.05, >1.1-fold difference). We found seven gene modules in MIS-C, annotated as an increased TNFα/NFκB pathway, decreased EC homeostasis, anti-inflammation and immune response, translation, and glucocorticoid responsive genes and endothelial-mesenchymal transition (EndoMT). To further understand the difference in the EC response between MIS-C and KD, stringent filtering was applied to identify 41 differentially expressed genes (DEGs) between MIS-C and KD (adjusted p < 0.05, >2-fold-difference). Again, in MIS-C, NFκB pathway genes, including nine pro-survival genes, were upregulated. The expression levels were higher in the genes influencing autophagy (UBD, EBI3, and SQSTM1). Other DEGs also supported the finding by WGCNA. Compared to KD, ECs in MIS-C had increased pro-survival transcripts but reduced transcripts related to EndoMT and EC homeostasis. These differences in the EC response may influence the different cardiovascular outcomes in these two diseases.