Pluripotent cell-based phenotypic dissection identifies a high-frequency somatic NLRC4 mutation as a cause of autoinflammation

Objective To elucidate the genetic background of a patient with neonatal-onset multisystem inflammatory disease(NOMID) who does not carry any NLRP3 mutation. MethodsA Japanese male diagnosed as NOMID was recruited. The patient had no NLRP3 mutation even as low frequency mosaicism. We performed whole exome sequencing(WES) of the patient and his parents. Induced pluripotent stem cells(iPSCs) were established from the fibroblasts of the patient. iPSCs were then differentiated into monocytic lineage to evaluate the cytokine profile. Results We established multiple iPSC clones from an NOMID patient and incidentally found that the phenotype of monocytes from iPSC clones were heterogeneous, and could be grouped into “diseased” and “normal” phenotype. Because each iPSC clone was derived from a single somatic cell, we hypothesized the patient had somatic mosaicismof an IL-1β-related gene. WES of both representative iPSC clones and patient's blood identified a novel heterozygous NLRC4mutation, p.T177A (c.529A>G), as a specific mutation in “diseased” iPSC clones. Knockout of the NLRC4gene using CRISPR/ Cas9system in a mutant iPSC clone abrogated the pathogenic phenotype. Conclusion We concluded the patient as having somatic mosaicismof a novel NLRC4mutation. To our knowledge, this is the first case showing somatic NLRC4mutation causes autoinflammatory symptoms compatible to NOMID. The present study demonstrates the significance of prospective genetic screeningcombined with iPSC-based phenotypic dissection for individualized diagnoses. This article is protected by copyright. All rights reserved.