Background/Purpose

IFNα is emerging as a clinically validated target in SLE yet it is currently unclear if other type I IFNs are contributing to the IFN signature present in many SLE patients. In this study we analyzed SLE patient sera and plasma for the presence of IFNω protein and compared the biological effects imparted by recombinant IFNα and IFNω on human cells. We further examined the effects of blocking IFNα alone versus dual blockade of IFNα and ω using SLE patient-derived IFN stimuli in vitro.

Methods

SLE patient sera and plasma were analyzed for the presence of IFNα and IFNω using a multiplex ELISA. SLE sera or conditioned media from cells exposed to SLE patient immune complexes were utilized as stimuli in an ISRE reporter gene assay (RGA) with selectively neutralizing mAbs to IFNα or IFNω or isotype control. To examine the effects of IFNω treatment, PBMCs from 6 healthy human donors were treated with either recombinant IFNαA or ω and gene and protein expression were analyzed by microarray and, Luminex or ELISA, respectively. To assess the individual contribution of IFNα and IFNω on the IFN signature, unstimulated SLE whole blood from patients having an elevated IFN signature was treated with neutralizing mAbs to IFNα, IFNω or the combination of both and qPCR analysis was performed. A fully-human monoclonal antibody targeting IFNω and multiple subtypes of IFNα was developed and tested for its ability to neutralize IFNα and IFNω-induced IP-10 release in whole blood.

Results

IFNω and IFNα were found to be elevated in the plasma and serum of a subset of SLE patients. IFNω protein was also detected in conditioned media from SLE patient immune complex-stimulated PBMCs. Combined blockade of IFNα and IFNω resulted in further suppression of IFN activity in comparison to IFNα blockade alone using these endogenous preparations of type I IFN. Microarray data from IFN-treated PBMCs indicated that 99.25% of genes modulated by IFNαA treatment versus untreated control were modulated by IFNω at 24h. IFNω exhibited indistinguishable qualitative gene expression responses as compared to IFNαA-treated cells using a 21 gene IFN signature. IFNαA and IFNω treatment induced TLR7, IP-10 and BLyS gene expression. IFNω-mediated BLyS and IP-10 induction was confirmed at the protein level. To determine the impact of various IFN inhibitors on the IFN signature present in SLE donor whole blood, neutralizing antibodies targeting IFNα, IFNω, or the combination of both were added to unstimulated blood. Gene expression analysis by qPCR indicated that combined blockade of IFNα and IFNω resulted in greater reduction of multiple IFN-inducible genes than IFNα blockade alone. We further demonstrate a fully-human monoclonal antibody capable of dose-dependently inhibiting IP-10 release induced by both IFNω and IFNα.

Conclusion

IFNω antagonism enhanced the ability of IFNα antagonists to suppress IFN activity in SLE patient sera, SLE immune complex-induced preparations of IFN and the IFN signature in SLE patient whole blood in vitro.  IFN signatures induced by recombinant IFNω and IFNα were found to be indistinguishable and our current data lends compelling support to the hypothesis that IFNω may contribute to the total type I IFN activity and signature present in some SLE patients.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/elevation-and-functional-activity-of-interferon-omega-in-human-systemic-lupus-erythematosus/