Serum albumin-binding domain antibodies (AlbudAbs) have previously been shown to greatly extend the serum half-life of the interleukin-1 receptor antagonist IL-1ra. We have subsequently extended this approach to look at the in vitro activity, in vivo efficacy and pharmacokinetics of an agonist molecule, interferon (IFN)-alpha2b, fused to an AlbudAb. Here we describe this molecule and show that in this format AlbudAb half-life extension technology displays significant advantages in comparison with other methods of half-life extension, in particular genetic fusion to serum albumin. When compared directly IFN-alpha2b fused to an Albudab shows higher potency, increased serum half-life and greater efficacy than human serum albumin fused to IFN-alpha2b. AlbudAbs are therefore an ideal platform technology for creation of therapeutics with agonist activity and long serum half-lives.

Interferon alpha (IFNα) is used for the treatment of hepatitis C infection and whilst efficacious it is associated with multiple adverse events including reduced leukocyte, erythrocyte, and platelet counts, fatigue, and depression. These events are most likely caused by systemic exposure to interferon. We therefore hypothesise that targeting the therapeutic directly to the intended site of action in the liver would reduce exposure in blood and peripheral tissue and hence improve the safety and tolerability of IFNα therapy. We genetically fused IFN to a domain antibody (dAb) specific to a hepatocyte restricted antigen, asialoglycoprotein receptor (ASGPR). Our results show that the murine IFNα2 homolog (mIFNα2) fused to an ASGPR specific dAb, termed DOM26h-196-61, could be expressed in mammalian tissue culture systems and retains the desirable biophysical properties and activity of both fusion partners when measured in vitro. Furthermore a clear increase in in vivo targeting of the liver by mIFNα2-ASGPR dAb fusion protein, compared to that observed with either unfused mIFNα2 or mIFNα2 fused to an isotype control dAb VHD2 (which does not bind ASGPR) was demonstrated using microSPECT imaging. We suggest that these findings may be applicable in the development of a liver-targeted human IFN molecule with improved safety and patient compliance in comparison to the current standard of care, which could ultimately be used as a treatment for human hepatitis virus infections.

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Top-down control analysis was used to quantify how different mRNAs cause physiological changes in response to effectors. We analysed published DNA microarray data (Ideker et al., 2001 Science 292,929-934) that reported growth rates and significant changes in the concentrations of 997 mRNAs in wild type yeast and nine mutants in the presence and absence of galactose. Firstly, mRNAs were clustered into eight modules by similarity of expression profile. Multiplication of integrated response coefficients of mRNA modules to galactose by the elasticities of growth rate to each of these modules showed that more than 95% of the galactoseinduced change in doubling time was transmitted by one block, which contained three mRNAs involved in galactose metabolism. Secondly, the mRNAs were clustered into two blocks and the direct and indirect responses of each block of mRNAs to galactose were calculated. Most of the large change in block 2 was shown to be indirect, i.e. mediated by block 1. Top-down control analysis provides a novel and informative way to analyse and quantify gene expression data. The method is generally applicable to modern DNA microarray expression profiling to discover and quantify the relative importance of different groups of mRNA transcripts in mediating physiological responses.