Multivalent Albumin-Neonatal Fc Receptor Interactions Mediate a Prominent Extension of the Serum Half-Life of a Therapeutic Protein.
Mol Pharm
; 18(6): 2397-2405, 2021 06 07.
Article
in En
| MEDLINE
| ID: mdl-33983743
ABSTRACT
Human serum albumin (HSA) has been used to extend the serum half-life of therapeutic proteins owing to its exceptionally long serum half-life via the neonatal Fc receptor (FcRn)-mediated recycling mechanism. In most cases, only one HSA molecule was conjugated to a therapeutic protein, leading to a limited extension of the serum half-life. In this study, we hypothesized that conjugation of multiple HSA molecules to a therapeutic protein significantly further extends the serum half-life via multivalent HSA-FcRn interactions. We chose urate oxidase (Uox), a tetrameric therapeutic protein used for the treatment of gout, as a model. In previous studies, only one HSA molecule was site-specifically conjugated to one Uox because of poor conjugation yield of the relatively slow bio-orthogonal chemistry, strain-promoted azide-alkyne cycloaddition (SPAAC). To increase the number of HSA molecules conjugated to one Uox, we employed the faster bio-orthogonal chemistry, inverse electron demand Diels-Alder reaction (IEDDA). We site-specifically introduced the phenylalanine analog with a fast-reacting tetrazine group (frTet) into position 174 of each subunit of Uox. We then achieved site-specific HSA conjugation to each subunit of Uox via IEDDA, generating Uox conjugated to four HSA molecules (Uox-HSA4), with a small portion of Uox conjugated to three HSA molecules (Uox-HSA3). We characterized Uox-HSA4 as well as Uox variants conjugated to one or two HSA molecules prepared via SPAAC (Uox-HSA1 or Uox-HSA2). The enzyme activity of all three Uox-HSA conjugates was comparable to that of unmodified Uox. We found out that an increase in HSA molecules conjugated to Uox (multiple albumin-conjugated therapeutic protein) enhanced FcRn binding and consequently prolonged the serum half-life in vivo. In particular, the conjugation of four HSA molecules to Uox led to a prominent extension of the serum half-life (over 21 h), which is about 16-fold longer than that of Uox-WT.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Urate Oxidase
/
Receptors, Fc
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Histocompatibility Antigens Class I
/
Excipients
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Serum Albumin, Human
Limits:
Animals
Language:
En
Journal:
Mol Pharm
Journal subject:
BIOLOGIA MOLECULAR
/
FARMACIA
/
FARMACOLOGIA
Year:
2021
Document type:
Article