UBC Theses and Dissertations
A novel non-glucose based osmotic agent with liver uptake potential for peritoneal dialysis solutions Swaminathan, Meenakshi
Peritoneal Dialysis (PD), a treatment modality for end-stage renal disease (ESRD) uses high concentrations of glucose in its dialysis solutions. Since, 40% ESRD patients are diabetic, use of glucose-based PD solutions is non-rational because of the local and systemic adverse effects of glucose. There is a need for a biocompatible, non-glucose based osmotic agent that can replace glucose in peritoneal dialysis solutions. Recent studies on low molecular weight hyperbranched polyglycerol (HPG) as an osmotic agent show good ultrafiltration and reduction of peritoneal membrane injury but elimination through the kidneys. Our studies show that HPG is excreted via kidneys, and in ESRD patients, kidney function is impaired which could potentially result in poor excretion of this osmotic agent. N-acetylgalactosamine (GalNAc) carrying proteins and macromolecules has shown to be effectively up taken by the liver through asialoglycoprotein receptor (ASGPR). Thus, we hypothesize that HPG modified by GalNAc conjugation can be recognized by ASGPR in the liver, taken up and excreted through feces. GalNAc epoxide was synthesized by standard organic modifications and conjugated with HPG (3K) at two different densities (one or three GalNAc molecules per HPG) (denoted as HPG+1S or HPG+3S). The polymers were labeled with alexa647 and screened for effective internalization in a panel of hepatocyte cell lines with different levels of ASGPR expression using flow cytometry. Results of dose-dependent uptake of polymers and ASGPR staining of hepatocytes suggested receptor mediated internalization of HPG+3S in HepG2 and HuH7.5.1 cells. Kₘ values were assessed to determine the efficacious HPG conjugate followed by competitive inhibition with natural ligands of ASGPR in HepG2 cells. HEK293 cells ectopically expressing ASGPR1 and ASGPR1&2 genes were used to confirm HPG+3S uptake through ASGPR specifically. Mice tissue distribution studies of radiolabeled HPG and HPG+3S showed better uptake of HPG+3S by the liver than HPG. Percentage of HPG+3S excreted via feces was significantly more than HPG in mice with normal kidney function, which support the enhanced binding of HPG+3S to ASGPR in hepatocytes. In conclusion, both our in vitro and in vivo results substantiate our claim that GalNAc conjugated HPG can be rerouted to be excreted via feces.
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