Christos Sazeides

ISG15 deficiency is a type I interferonopathy characterized by elevated circulating type I interferon (IFN-I), intracranial calcifications, fibrotic skin lesions, and occasionally inflammatory lung disease. However, the mechanisms driving immune-mediated fibrosis remain poorly understood. In this study, we combined molecular biology approaches with spatial transcriptomics in ex vivo patient samples and in vitro models to characterize a novel loss-of-function ISG15 variant and to elucidate disease-associated molecular pathways. Analysis of patient skin biopsies revealed increased IFN-I–dependent apoptosis, altered macrophage polarization, enhanced epithelial-to-mesenchymal transition (EMT), and myofibroblast activation. In vitro, ISG15 knockout (KO), but not wild-type (WT), epithelial cells were predisposed to EMT following combined TGF-β and IFN-I stimulation. Additionally, ISG15 KO fibroblasts displayed impaired mechanical wound healing and increased IFN-I–induced apoptosis compared with WT cells. Collectively, our findings suggest that chronic IFN-I exposure in ISG15 deficiency disrupts wound repair, skews macrophage activation, and promotes EMT, contributing to fibrosis across multiple organ systems.

Type I interferons (IFN-Is) are cytokines with potent antiviral and inflammatory capacities. IFN-I signaling drives the expression of thousands of IFN-I–stimulated genes (ISGs), whose aggregate function results in the control of viral infections. A few of these ISGs are tasked with negatively regulating the IFN-I response to prevent overt inflammation. ISG15 is a negative regulator whose absence leads to persistent, low-grade elevation of ISG expression and concurrent, often self-resolving, mild autoinflammation. The limited breadth and low-grade persistence of ISGs expressed in ISG15 deficiency are sufficient to confer broad-spectrum antiviral resistance. Inspired by the antiviral state of humans with ISG15 deficiency, we identified a nominal collection of 10 ISGs that recapitulated the broad antiviral potential of the IFN-I system, which typically induces the expression of thousands of ISGs. The expression of this 10-ISG collection in an IFN-I–nonresponsive cell line increased cellular resistance to Zika virus, vesicular stomatitis virus, and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). A lipid nanoparticle–encapsulated messenger RNA (mRNA) formulation of this 10-ISG collection reduced influenza A virus plaque size in samples collected from infected mice when given prophylactically. Moreover, when used collectively and delivered prophylactically, the 10-ISG collection was able to protect hamsters against a lethal SARS-CoV-2 challenge, in contrast with the lack of efficacy when mRNAs were delivered individually. These findings suggest that these 10 ISGs have potential as a broad-spectrum antiviral prophylactic.

Summary: Type I interferons (IFN-I) are cytokines with potent antiviral and inflammatory capacities. IFN-I signaling drives the expression of hundreds of IFN-I stimulated genes (ISGs), whose aggregate function results in the control of viral infection. A few of these ISGs are tasked with negatively regulating the IFN-I response to prevent overt inflammation. ISG15 is a negative regulator whose absence leads to persistent, low-grade elevation of ISG expression and concurrent, self-resolving mild autoinflammation. The limited breadth and low-grade persistence of ISGs expressed in ISG15 deficiency are sufficient to confer broad-spectrum antiviral resistance. Inspired by ISG15 deficiency, we have identified a nominal collection of 10 ISGs that recapitulate the broad antiviral potential of the IFN-I system. The expression of the 10 ISG collection in an IFN-I non-responsive cell line increased cellular resistance to Zika, Vesicular Stomatitis, Influenza A (IAV), and SARS-CoV-2 viruses. A deliverable prophylactic formulation of this syndicate of 10 ISGs significantly inhibited IAV PR8 replication in vivo in mice and protected hamsters against a lethal SARS-CoV-2 challenge, suggesting its potential as a broad-spectrum antiviral against many current and future emerging viral pathogens. One-Sentence Summary: Human inborn error of immunity-guided discovery and development of a broad-spectrum RNA antiviral therapy