Abstract

During viral infections, type I interferons (IFN) are induced and play a key role in counteracting initial viral spread. Twelve different human IFN subtypes exist that bind the same receptor; however, they elicit unique host responses and display distinct potencies of antiviral activities. Our previous studies on HIV and HBV demonstrated that the clinically used IFN2 is not the most effective one among the IFN subtypes. By sequence modeling, we identified a region in helix B with mainly conserved residues at the outside facing IFNAR1, but variable residues at the inside facing the core of IFN, potentially representing a putative tunable anchor to tune pleiotropic IFN responses. Using site-directed mutagenesis various mutations were introduced into the IFN2b backbone targeting sites which are important for binding to IFNAR1 and IFNAR2, the putative tunable anchor, or outside these three regions. Selected mutations were based on sequence differences to high antiviral subtypes IFN6 and IFN14. Treatment assays against HBV and HIV identified several critical residues for the antiviral activity of IFN mainly in the IFNAR1 binding region. Combined mutations of the IFN2 IFNAR1/2 binding sites or the IFNAR1 binding region plus the putative tunable anchor by those of IFN14 further augmented activation of different downstream signaling cascades providing a molecular correlate for the enhanced antiviral activity. We describe here important functional residues within IFN subtype molecules, which enabled us to design novel and innovative drugs that may have the potential to be used in clinical trials against a variety of different viral infections. ImportanceThe potency of IFN to restrict viruses was already discovered in 1957. However, until today only IFN2 out of the 12 distinct human IFN subtypes has been therapeutically used against chronic viral infections. There is convincing evidence that other IFN subtypes are far more efficient than IFN2 against many viruses. In order to identify critical antiviral residues within the IFN subtype sequence, we designed hybrid molecules based on the IFN2 backbone with individual sequence motifs from the more potent subtypes IFN6 and IFN14. In different antiviral assays with HIV or HBV, residues binding to IFNAR1 as well as combinations of residues in the IFNAR1 binding region, the putative tunable anchor, and residues outside these regions were identified to be crucial for the antiviral activity of IFN. Thus, we designed artificial IFN molecules, based on the clinically approved IFN2 backbone, but with highly improved antiviral activity against several viruses.