Engineering potent long-acting variants of the Wnt inhibitor DKK2.
Wnt signaling pathways are required for a wide variety of biological processes ranging from embryonic development to tissue repair and regeneration. Dickkopf-2 (DKK2) is classically defined as a canonical Wnt inhibitor, though it may play a role in activating non-canonical Wnt pathways in the context of endothelial network formation after acute injury. Here we report the discovery of a fusion partner for a DKK2 polypeptide that significantly improves the expression, biochemical properties and pharmacokinetics (PK) of the DKK2 polypeptide. Specifically, human serum albumin (HSA) was identified as a highly effective fusion partner. Substitution of selected amino acid residues in DKK2 designed to decrease heparan sulfate binding by HSA-DKK2 variants, further improved the PK properties of the molecule in rodents. The HSA-DKK2 variants were monomeric, as thermally stable as wild type, and active as measured by their ability to bind to and prevent phosphorylation of the Wnt coreceptor LRP6. Our engineering efforts resulted in potent long-lived variants of the canonical Wnt inhibitor DKK2, applicable for Wnt pathway manipulation either by systematic delivery or focused administration at sites of tissue injury.
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Richelle
R
Sopko
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Joshua W
JW
Mugford
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Andreas
A
Lehmann
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Renée I
RI
Shapiro
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Mia
M
Rushe
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Abhishek
A
Kulkarni
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Joseph
J
Worrall
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Joseph
J
Amatucci
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Dingyi
D
Wen
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Nels E
NE
Pederson
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Brenda K
BK
Minesinger
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Joseph W
JW
Arndt
Department of Cell and Protein Sciences, Biogen, Cambridge, MA 02142, USA.
Blake
B
Pepinsky
D000818:Animals; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D060488:Low Density Lipoprotein Receptor-Related Protein-6 / Q000037:antagonists & inhibitors*; D051379:Mice; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D012709:Serum Albumin / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D051153:Wnt Proteins / Q000037:antagonists & inhibitors*; D060449:Wnt Signaling Pathway / Q000187:drug effects*
C405060:Dkk2 protein, mouse; D036341:Intercellular Signaling Peptides and Proteins; D060488:Low Density Lipoprotein Receptor-Related Protein-6; C555476:Lrp6 protein, mouse; D011993:Recombinant Fusion Proteins; D012709:Serum Albumin; D051153:Wnt Proteins
Dickkopf-2; Wnt signaling; heparan sulfate; human serum albumin; tissue injury