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Capacity limits of asialoglycoprotein receptor-mediated liver targeting.

ABSTRACT: The abundant cell surface asialoglycoprotein receptor (ASGPR) is a highly selective receptor found on hepatocytes that potentially can be exploited as a selective shuttle for delivery. Various nucleic acid therapeutics that bind ASGPR are already in clinical development, but this receptor-mediated delivery mechanism can be saturated, which will likely result in reduced selectivity for the liver and therefore increase the likelihood for systemic adverse effects. Therefore, when aiming to utilize this mechanism, it is important to optimize both the administration protocol and the molecular properties. We here present a study using a novel ASGPR-targeted antibody to estimate ASGPR expression, turnover and internalization rates in vivo in mice. Using pharmacokinetic data (intravenous and subcutaneous dosing) and an in-silico target-mediated drug disposition (TMDD) model, we estimate an ASGPR expression level of 1.8 million molecules per hepatocyte. The half-life of the degradation of the receptor was found to be equal to 15 hours and the formed ligand-receptor complex is internalized with a half-life of 5 days. A biodistribution study was performed and confirmed the accuracy of the TMDD model predictions. The kinetics of the ASGPR shows that saturation of the shuttle at therapeutic concentrations is possible; however, simulation allows the dosing schedule to be optimized. The developed TMDD model can be used to support the development of therapies that use the ASGPR as a shuttle into hepatocytes.

SUBMITTER: Bon C

PROVIDER: S-EPMC5680813 | biostudies-literature | 2017 Nov/Dec

REPOSITORIES: biostudies-literature

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Capacity limits of asialoglycoprotein receptor-mediated liver targeting.

Bon Charlotte C Hofer Thomas T Bousquet-Mélou Alain A Davies Mark R MR Krippendorff Ben-Fillippo BF

mAbs 20170906 8

The abundant cell surface asialoglycoprotein receptor (ASGPR) is a highly selective receptor found on hepatocytes that potentially can be exploited as a selective shuttle for delivery. Various nucleic acid therapeutics that bind ASGPR are already in clinical development, but this receptor-mediated delivery mechanism can be saturated, which will likely result in reduced selectivity for the liver and therefore increase the likelihood for systemic adverse effects. Therefore, when aiming to utilize ...[more]

PMID: 28876162

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Project description:The hepatic carbohydrate-recognizing asialoglycoprotein receptor (ASGR1) mediates the endocytosis/lysosomal degradation of desialylated glycoproteins following binding to terminal galactose/N-acetylgalactosamine. Human heterozygote carriers of ASGR1 deletions exhibit ∼34% lower risk of coronary artery disease and ∼10% to 14% reduction of non-HDL cholesterol. Since the proprotein convertase PCSK9 is a major degrader of the low-density lipoprotein receptor (LDLR), we investigated the degradation and functionality of LDLR and/or PCSK9 by endogenous/overexpressed ASGR1 using Western blot and immunofluorescence in HepG2-naïve and HepG2-PCSK9-knockout cells. ASGR1, like PCSK9, targets LDLR, and both independently interact with/enhance the degradation of the receptor. This lack of cooperativity between PCSK9 and ASGR1 was confirmed in livers of wildtype (WT) and Pcsk9-/- mice. ASGR1 knockdown in HepG2-naïve cells significantly increased total (∼1.2-fold) and cell-surface (∼4-fold) LDLR protein. In HepG2-PCSK9-knockout cells, ASGR1 silencing led to ∼2-fold higher levels of LDLR protein and DiI (1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate)-LDL uptake associated with ∼9-fold increased cell-surface LDLR. Overexpression of WT-ASGR1/2 primarily reduced levels of immature non-O-glycosylated LDLR (∼110 kDa), whereas the triple Ala-mutant of Gln240/Trp244/Glu253 (characterized by loss of carbohydrate binding) reduced expression of the mature form of LDLR (∼150 kDa), suggesting that ASGR1 binds the LDLR in both a sugar-dependent and -independent fashion. The protease furin cleaves ASGR1 at the RKMK103↓ motif into a secreted form, likely resulting in a loss of function on LDLR. Altogether, we demonstrate that LDLR is the first example of a liver-receptor ligand of ASGR1. We conclude that silencing of ASGR1 and PCSK9 may lead to higher LDL uptake by hepatocytes, thereby providing a novel approach to further reduce LDL cholesterol levels.

Dataset Information

Capacity limits of asialoglycoprotein receptor-mediated liver targeting.

Publications

Capacity limits of asialoglycoprotein receptor-mediated liver targeting.

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