Mechanisms underlying genetic susceptibility to multisystem inflammatory syndrome in children (MIS-C)

Abstract

Background: Multisystem inflammatory syndrome in children (MIS-C) is a pediatric complication of severe acute respiratory syndrome coronavirus 2 infection that is characterized by multiorgan inflammation and frequently by cardiovascular dysfunction. It occurs predominantly in otherwise healthy children. We previously reported haploinsufficiency of suppressor of cytokine signaling 1 (SOCS1), a negative regulator of type I and II interferons, as a genetic risk factor for MIS-C.

Objectives: We aimed to identify additional genetic mechanisms underlying susceptibility to severe acute respiratory syndrome coronavirus 2-associated MIS-C.

Methods: In a single-center, prospective cohort study, whole exome sequencing was performed on patients with MIS-C. The impact of candidate variants was tested by using patients' PBMCs obtained at least 7 months after recovery.

Results: We enrolled 18 patients with MIS-C (median age = 8 years; interquartile range = 5-12.25 years), of whom 89% had no conditions other than obesity. In 2 boys with no significant infection history, we identified and validated hemizygous deleterious defects in XIAP, encoding X-linked inhibitor of apoptosis, and CYBB, encoding cytochrome b-245, beta subunit. Including the previously reported SOCS1 haploinsufficiency, a genetic diagnosis was identified in 3 of 18 patients (17%). In contrast to patients with mild COVID-19, patients with defects in SOCS1, XIAP, or CYBB exhibit an inflammatory immune cell transcriptome with enrichment of differentially expressed genes in pathways downstream of IL-18, oncostatin M, and nuclear factor κB, even after recovery.

Conclusions: Although inflammatory disorders are rare in the general population, our cohort of patients with MIS-C was enriched for monogenic susceptibility to inflammation. Our results support the use of next-generation sequencing in previously healthy children who develop MIS-C.

Keywords: COVID-19; MIS-C; Multisystem inflammatory syndrome in children; SARS-CoV-2; whole exome sequencing.

Graphical abstract

Fig 1

Genetic risk factors for MIS-C. A, Schematic of XIAP with structural modeling of the ubiquitin-associated (UBA) domain identifies a new hydrogen bond (indicated by the dotted red line) formed between the S421N mutant found in patient 1 and alanine 417. B, Flow cytometric quantification of XIAP protein expression in CD14⁺ monocytes from a control (Ctrl) and the patient from 2 experiments with 3 controls. C, Quantification of HLA-DR⁺TNF-α⁺ monocytes, gated on CD14⁺ cells, after stimulation with 200 ng/mL of muramyl dipeptide or LPS for 2.5 hours in 2 experiments with 5 controls. D, IL-1β secretion after stimulation of PBMCs from 4 controls and patient 1 with indicated stimuli, pooled from 2 experiments. E, TNF-α secretion after stimulation of PBMCs from 4 controls and patient 1, pooled from 2 experiments. F, Schematic of CYBB. G, Quantification of the neutrophil oxidative burst in the presence and absence of stimulation with phorbol 12–myristate 13–acetate, pooled from 3 independent experiments with 13 controls. max, Maximum; MFI, mean fluorescence intensity; n.s., not significant; NOI, neutrophil oxidative index; stim, stimulated; unstim, unstimulated.

Fig 2

A, Transcriptome analysis of unstimulated PBMCs from patients 1 and 2, as well as those from the patient with SOCS1 haploinsufficiency whom we previously described, compared with PBMCs from 4 otherwise healthy individuals who had previously had mild COVID-19. PBMCs were collected at least 7 months after recovery, at which time the individuals had returned to their baseline state of health. ∗P < .05; ∗∗P < .01 by the Mann-Whitney test. B, Uniform Manifold Approximation and Projection (UMAP) plots depict transcriptional clusters generated by single-cell RNA sequencing of PBMCs from a control and a patient with active MIS-C but no identifiable genetic diagnosis. This research sample was taken early in the patient’s course; she had received 1 dose of IVIG but subsequently required additional doses of IVIG and methylprednisolone before clinical improvement occurred. Ingenuity Pathway Analysis of differentially expressed genes indicates upregulation of signaling pathways downstream of type I and/or II interferons, STAT1, IRF3, and IRF7. mono, Monocyte; NF-κB, nuclear factor κB; NK, natural killer.