In VivoClearance of Alpha-1 Acid Glycoprotein Is Influenced by the Extent of Its N-Linked Glycosylation and by Its Interaction with the Vessel Wall

Abstract

<jats:p>Alpha-1 acid glycoprotein (AGP) is a highly glycosylated plasma protein that exerts vasoprotective effects. We hypothesized that AGP’s N-linked glycans govern its rate of clearance from the circulation, and followed the disappearance of different forms of radiolabeled human AGP from the plasma of rabbits and mice. Enzymatic deglycosylation of human plasma-derived AGP (pdAGP) by Peptide: N-Glycosidase F yielded a mixture of differentially deglycosylated forms (PNGase-AGP), while the introduction of five Asn to Gln mutations in recombinant<jats:italic>Pichia pastoris</jats:italic>-derived AGP (rAGP-N(5)Q) eliminated N-linked glycosylation. PNGase-AGP was cleared from the rabbit circulation 9-fold, and rAGP-N(5)Q, 46-fold more rapidly than pdAGP, primarily via a renal route.<jats:italic>Pichia pastoris</jats:italic>-derived wild-type rAGP differed from pdAGP in expressing mannose-terminated glycans, and, like neuraminidase-treated pdAGP, was more rapidly removed from the rabbit circulation than rAGP-N(5)Q. Systemic hyaluronidase treatment of mice transiently decreased pdAGP clearance. AGP administration to mice reduced vascular binding of hyaluronic acid binding protein in the liver microcirculation and increased its plasma levels. Our results support a critical role of N-linked glycosylation of AGP in regulating its<jats:italic>in vivo</jats:italic>clearance and an influence of a hyaluronidase-sensitive component of the vessel wall on its transendothelial passage.</jats:p>

N-glycan PK Profiling Using a High Sensitivity nanoLCMS Work-Flow with Heavy Stable Isotope Labeled Internal Standard and Application to a Preclinical Study of an IgG1 Biopharmaceutical

“…The fast decrease in M5 can be explained by a second mechanism. M5 and in conclusion M5 containing glycoforms are probably removed by a specific clearance mechanism from circulation involving the mannose receptor which binds amongst others specifically to mannose containing glycoproteins and removes them from circulation with similar functions in humans and rabbits that were described previously ( 11 , 12 , 14 , 16 , 17 , 22 ). The incomplete removal of M5 from circulation observed in this study can be explained by the structural conformation of the Fc part.…”

Plasma protein N-glycan signatures of type 2 diabetes

“…Sialic acids influence plasma protein clearance and are directly involved in activation and control of the immune system. Moreover, their functions depend on the linkage type [22,37]. Our finding of increased sialylation in diabetes and with increasing BMI was specifically due to α2,6-sialylation, while α2,3-sialylation decreased.…”

Molecular Aspects of Human Alpha-1 Acid Glycoprotein — Structure and Function

“…In addition, the results from a mice study indicated that systemic hyaluronidase treatment decreased the initial clearance of AGP and that AGP administration reduced the binding of hyaluromic acid binding protein to the vessel wall of liver sinusoids. [99] These results suggest that AGP, including N-linked glycans, interact with hyaluronan or hyaluronidase-sensitive component of the vessel wall which influence the transendothelial passage of AGP. Based on these results, a new hepatic elimination pathway involving at least two types of receptors, namely an asialoglycoprotein receptor and another yet to be identified receptor, for AGP was proposed ( Figure 5).…”

“…[97] Pharmacoki-netic studies using mice and rabbits demonstrated that AGP was mainly distributed in the liver. [98,99] We also clarified that AGP was mainly incorporated into liver parenchymal cells via a receptor-mediated pathway, and the hemoglobin β-chain located on liver plasma membranes contributes to the intracellular uptake of AGP. [100,101] These data suggest that AGP is finally taken up by liver parenchymal cells via the hemoglobin β-chain and is then degraded or eliminated from the body.…”

“…In addition, McCurdy et al also obtained similar results using glycan-deficient AGP in rabbits. [99] An asialoglycoprotein receptor has been reported to be associated with the incorporation of AGP into liver tissue. [102] Regoeczi et al studied the pharmacokinetics of asialo-AGP in chickens and rabbits, and suggested the possibility of the presence of a naturally occurring terminal catabolic point of AGP that was related to hepatic uptake via a hepatic plasma membrane receptor (an asialoglycoprotein receptor).…”