Site-specific albumination of a therapeutic protein with multi-subunit to prolong activity in vivo

JOURNAL OF CONTROLLED RELEASE

卷:207

Article

Lim, Sung In¹  Hahn, Young S.²  Kwon, Inchan^1,3 

[1] Univ Virginia, Dept Chem Engn, Charlottesville, VA 22904 USA

[2] Univ Virginia, Dept Microbiol Immunol & Canc Biol, Charlottesville, VA 22908 USA

[3] Gwangju Inst Sci & Technol, Sch Mat Sci & Engn, Gwangju 500712, South Korea

关键词: Non-natural amino acid;    Site-specific albumination;    Therapeutic protein;    Urate oxidase;    Strain-promoted azide-alkyne cycloaddition;   

DOI  :  10.1016/j.jconrel.2015.04.004

来源: Elsevier

【 摘 要 】

Albumin fusion/conjugation (albumination) has been an effective method to prolong in vivo half-life of therapeutic proteins. However, its broader application to proteins with complex folding pathway or multi-subunit is restricted by incorrect folding, poor expression, heterogeneity, and loss of native activity of the proteins linked to albumin. We hypothesized that the site-specific conjugation of albumin to a permissive site of a target protein will expand the utilities of albumin as a therapeutic activity extender to proteins with a complex structure. We show here the genetic incorporation of a non-natural amino acid (NNAA) followed by chemoselective albumin conjugation to prolong therapeutic activity in vivo. Urate oxidase (Uox), a therapeutic enzyme for treatment of hyperuricemia, is a homotetramer with multiple surface lysines, limiting conventional approaches for albumination. Incorporation of p-azido-L-phenylalanine into two predetermined positions of Uox allowed site-specific linkage of dibenzocyclooctyne-derivatized human serum albumin (HSA) through strain-promoted azide-alkyne cycloaddition (SPAAC). The bio-orthogonality of SPAAC resulted in the production of a chemically well-defined conjugate, Uox-HSA, with a retained enzymatic activity. Uox-HSA had a half-life of 8.8 h in mice, while wild-type Uox had a half-life of 1.3 h. The AUC increased 5.5-fold (1657 vs. 303 mU/mL x h). These results clearly demonstrated that site-specific albumination led to the prolonged enzymatic activity of Uox in vivo. Site-specific albumination enabled by NNAA incorporation and orthogonal chemistry demonstrates its promise for the development of long-acting protein therapeutics with high potency and safety. (C) 2015 Elsevier B.V. All rights reserved.

【 授权许可】

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