Interferonopathies in Brazil: The Pivotal Role of the Interferon Signature for Diagnosis and for the Genetic Sequencing in the CNE3i

Interferonopathies are rare autoinflammatory disorders driven by a constitutive upregulation of the type I interferon (IFN-I) pathway, leading to chronic activation of this cytokine. Despite advances, direct IFN-I measurement is challenging, making diagnosis reliant on clinical suspicion and genetic testing, which often yields inconclusive results due to a lack of established clinical criteria and the limited number of genes described as related to interferonopathies. Indirect methods, such as the “IFN signature” (expression of interferon-stimulated genes—ISGs), are used, but many patients still lack a confirmed genetic diagnosis. To this end, we aim to evaluate the clinical utility of the interferon signature as a diagnostic screening tool and to integrate it with genetic analysis in a cohort of Brazilian patients with suspected interferonopathy. We conducted a cross-sectional study within the Centro nacional de Erros Inatos da Imunidade e Imunodesregulação (CNE3i) (Certificate of Presentation for Ethical Consideration [CAAE]: 73174223.1.1001.0068). Patients with clinical suspicion of interferonopathy underwent peripheral blood collection for peripheral blood mononuclear cell (PBMC) isolation and RNA/DNA extraction. The IFN-I signature was assessed via quantitative PCR (qPCR) of a 6-gene panel (IFI27, ISG15, IFI44L, RSAD2, IFIT1, and SIGLEC1), and the IS (interferon score) was determined. All patients underwent exome sequencing (NovaSeqX), and Illumina EMEDGENE was used for secondary analysis. To date, 73 patients have been evaluated. The IS confirmed IFN-1 pathway activation (resulted positive) in 43/73 cases (59%). Among those with IS-positive patients, 27 (62%) remain without a definitive genetic diagnosis (Figure 1). Targeted reanalysis searching for genes involved in nucleic acid metabolism from exome data in 11 cases with positive IS identified potentially relevant variants of uncertain significance (VUS) in genes associated with interferonopathy (RNASEH2A, ADAR1, SAMHD1, PNPT1, and ATM). The interferon signature proved to be a highly effective front-line screening test, accurately stratifying 59% of clinically suspected cases. Its high positivity rate among undiagnosed patients (62%) underscores its critical role in confirming the pathophysiological mechanism and prioritizing cases for advanced genomic investigation. The IS significantly aids in differentiating interferonopathies from other syndromes, reducing diagnostic delay, and providing a functional biomarker for future therapeutic monitoring. FINEP funding: 0956/24; FAPESP funding: 2023/09965-0; Instituto de Investigação em Imunologia funding CNPQ/MCTI: 408685/2024-7.Figure 1.Interferon signature in suspected interferonopathies at the CNE3i (Centro Nacional de Erros Inatos da Imunidade e Imunodesregulação) in Brazil. (1) The number of samples received and the main clinical phenotypes. (2) The samples that have already been analyzed, along with the mean interferon score and the mean level of IFN gene expression. (3) The number of individuals who have already been sequenced and the top three diagnoses identified at the CNE3i. IS, interferon score; AGS, Aicardi-Goutières syndrome; PRAAS, proteasome-related autoinflammatory diseases; COPA, coatomer subunit alpha Protein; SAVI, STING-associated vasculopathy with infant onset; SPENCD, spondyloenchondrodysplasia with immune dysregulation.