title: Improving the pharmacokinetic properties of biologics by fusion to an anti-HSA shark VNAR domain.
abstract: Advances in recombinant antibody technology and protein engineering have provided the opportunity to reduce antibodies to their smallest binding domain components and have concomitantly driven the requirement for devising strategies to increase serum half-life to optimise drug exposure, thereby increasing therapeutic efficacy. In this study, we adopted an immunization route to raise picomolar affinity shark immunoglobulin new antigen receptors (IgNARs) to target human serum albumin (HSA). From our model shark species, Squalus acanthias, a phage display library encompassing the variable binding domain of IgNAR (VNAR) was constructed, screened against target, and positive clones were characterized for affinity and specificity. N-terminal and C-terminal molecular fusions of our lead hit in complex with a naïve VNAR domain were expressed, purified and exhibited the retention of high affinity binding to HSA, but also cross-selectivity to mouse, rat and monkey serum albumin both in vitro and in vivo. Furthermore, the naïve VNAR had enhanced pharmacokinetic (PK) characteristics in both N- and C-terminal orientations and when tested as a three domain construct with naïve VNAR flanking the HSA binding domain at both the N and C termini. Molecules derived from this platform technology also demonstrated the potential for clinical utility by being available via the subcutaneous route of delivery. This study thus demonstrates the first in vivo functional efficacy of a VNAR binding domain with the ability to enhance PK properties and support delivery of multifunctional therapies.
authors.affiliation: Molecular Partners AG, Zürich-Schlieren, Switzerland.
authors.forename: Mischa R
authors.initials: MR
authors.lastname: Müller
authors.forename: Kenneth
authors.initials: K
authors.lastname: Saunders
authors.forename: Christopher
authors.initials: C
authors.lastname: Grace
authors.forename: Macy
authors.initials: M
authors.lastname: Jin
authors.forename: Nicole
authors.initials: N
authors.lastname: Piche-Nicholas
authors.forename: John
authors.initials: J
authors.lastname: Steven
authors.forename: Ronan
authors.initials: R
authors.lastname: O'Dwyer
authors.forename: Leeying
authors.initials: L
authors.lastname: Wu
authors.forename: Lam
authors.initials: L
authors.lastname: Khetemenee
authors.forename: Yulia
authors.initials: Y
authors.lastname: Vugmeyster
authors.forename: Timothy P
authors.initials: TP
authors.lastname: Hickling
authors.forename: Lioudmila
authors.initials: L
authors.lastname: Tchistiakova
authors.forename: Stephane
authors.initials: S
authors.lastname: Olland
authors.forename: Davinder
authors.initials: D
authors.lastname: Gill
authors.forename: Allan
authors.initials: A
authors.lastname: Jensen
authors.forename: Caroline J
authors.initials: CJ
authors.lastname: Barelle
mesh_terms: D000818:Animals; D000915:Antibody Affinity; D000918:Antibody Specificity; D001688:Biological Products / Q000493:pharmacokinetics*; D015195:Drug Design; D000882:Haplorhini; D006801:Humans; D051379:Mice; D015202:Protein Engineering / Q000379:methods; D051381:Rats; D011946:Receptors, Antigen / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000493:pharmacokinetics*; D012709:Serum Albumin / Q000276:immunology; D012754:Sharks; D061905:Single-Domain Antibodies / Q000235:genetics / Q000378:metabolism*
chemical_list: D001688:Biological Products; D011946:Receptors, Antigen; D011993:Recombinant Fusion Proteins; D012709:Serum Albumin; D061905:Single-Domain Antibodies
keywords: FcRn; IgNAR; albumin; biologic therapeutics; half-life; phage display; shark; single chain binding domain