Fusion to a highly stable consensus albumin binding domain allows for tunable pharmacokinetics.
A number of classes of proteins have been engineered for high stability using consensus sequence design methods. Here we describe the engineering of a novel albumin binding domain (ABD) three-helix bundle protein. The resulting engineered ABD molecule, called ABDCon, is expressed at high levels in the soluble fraction of Escherichia coli and is highly stable, with a melting temperature of 81.5°C. ABDCon binds human, monkey and mouse serum albumins with affinity as high as 61 pM. The solution structure of ABDCon is consistent with the three-helix bundle design and epitope mapping studies enabled a precise definition of the albumin binding interface. Fusion of a 10 kDa scaffold protein to ABDCon results in a long terminal half-life of 60 h in mice and 182 h in cynomolgus monkeys. To explore the link between albumin affinity and in vivo exposure, mutations were designed at the albumin binding interface of ABDCon yielding variants that span an 11 000-fold range in affinity. The PK properties of five such variants were determined in mice in order to demonstrate the tunable nature of serum half-life, exposure and clearance with variations in albumin binding affinity.
Janssen Research & Development, L.L.C., 1400 Welsh & McKean Rd., Spring House, Pennsylvania, PA 19454, USA sjacobs9@its.jnj.com.
Steven A
SA
Jacobs
Janssen Research & Development, L.L.C., 1400 Welsh & McKean Rd., Spring House, Pennsylvania, PA 19454, USA.
Alan C
AC
Gibbs
Janssen Research & Development, L.L.C., 1400 Welsh & McKean Rd., Spring House, Pennsylvania, PA 19454, USA.
Michelle
M
Conk
Janssen Research & Development, L.L.C., 1400 Welsh & McKean Rd., Spring House, Pennsylvania, PA 19454, USA.
Fang
F
Yi
Janssen Research & Development, L.L.C., 1400 Welsh & McKean Rd., Spring House, Pennsylvania, PA 19454, USA.
Diane
D
Maguire
Janssen Research & Development, L.L.C., 1400 Welsh & McKean Rd., Spring House, Pennsylvania, PA 19454, USA.
Colleen
C
Kane
Janssen Research & Development, L.L.C., 1400 Welsh & McKean Rd., Spring House, Pennsylvania, PA 19454, USA.
Karyn T
KT
O'Neil
D000418:Albumins / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D016384:Consensus Sequence; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015202:Protein Engineering / Q000379:methods*; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism / Q000493:pharmacokinetics*
D000418:Albumins; D011993:Recombinant Fusion Proteins
albumin; half-life extension; phamacokinetics; scaffold