Bridging the Species Divide: Transgenic Mice Humanized for Type-I Interferon Response

Abstract

We have generated transgenic mice that harbor humanized type I interferon receptors (IFNARs) enabling the study of type I human interferons (Hu-IFN-Is) in mice. These “HyBNAR” (Hybrid IFNAR) mice encode transgenic variants of IFNAR1 and IFNAR2 with the human extracellular domains being fused to transmembrane and cytoplasmic segments of mouse sequence. B16F1 mouse melanoma cells harboring the HyBNAR construct specifically bound Hu-IFN-Is and were rendered sensitive to Hu-IFN-I stimulated anti-proliferation, STAT1 activation and activation of a prototypical IFN-I response gene (MX2). HyBNAR mice were crossed with a transgenic strain expressing the luciferase reporter gene under the control of the IFN-responsive MX2 promoter (MX2-Luciferase). Both the HyBNAR and HyBNAR/MX2-Luciferase mice were responsive to all Hu-IFN-Is tested, inclusive of IFNα2A, IFNβ, and a human superagonist termed YNSα8. The mice displayed dose-dependent pharmacodynamic responses to Hu-IFN-I injection, as assessed by measuring the expression of IFN-responsive genes. Our studies also demonstrated a weak activation of endogenous mouse interferon response, especially after high dose administration of Hu-IFNs. In sharp contrast to data published for humans, our pharmacodynamic readouts demonstrate a very short-lived IFN-I response in mice, which is not enhanced by sub-cutaneous (SC) injections in comparison to other administration routes. With algometric differences between humans and mice taken into account, the HyBNAR mice provides a convenient non-primate pre-clinical model to advance the study of human IFN-Is.

Enhanced in Vivo Efficacy of a Type I Interferon Superagonist with Extended Plasma Half-life in a Mouse Model of Multiple Sclerosis

“…24. Measurements were made using either an Agilent 7300 real time PCR system (96-well setup) or for some of the studies (see Figs.…”

“…24. In brief, D-luciferin was injected 10 min prior to luminescence measurements for each time point.…”

“…Exchange of the five C-terminal amino acids to those from human IFN␣8 increases binding to IFNAR2 by 15-fold (23). Combination of both sets of mutations led to the variant YNS␣8, which exhibits significantly higher potency than IFN-␤ in cell culture systems (22)(23)(24). To enhance the plasma half-life of this molecule, we appended to the N terminus a PAS polypeptide comprising 600 residues of Pro, Ala, and Ser, which forms a natively disordered, uncharged biopolymer with large hydrodynamic volume, i.e.…”

“…PASylated YNS␣8 Activates a Long Lasting IFN-I Response in Mice-HyBNAR transgenic mice that harbor humanized IFNAR1 and IFNAR2 receptors and that respond sensitively to human IFN-I injection (24) were used to evaluate the in vivo activity of PAS-YNS␣8. These mice were further crossed with a transgenic mouse strain expressing the luciferase reporter under the control of the IFN-I-responsive MX2 promoter (24,32) to follow the course of human IFN-I activation in real time.…”

“…However, in studying the effect of these superagonists in a mouse model, we are faced with a dilemma, because the human and mouse IFNAR sequences have diverged considerably, and cross-species activation of human IFN-I subtypes in mouse cells is weak and nonphysiological. To allow the study of human IFN-I in mice, we have recently generated transgenic mice harboring humanized IFNAR receptors (HyBNAR) that sensitively transduce human IFN-I signaling (24). Here, we examine a human IFN-I superagonist with a greatly improved pharmacological lifespan through PASylation (25), a biological alternative to PEGylation, using the HyBNAR transgenic mouse model.…”