Insulin receptor antibody-iduronate 2-sulfatase fusion protein: pharmacokinetics, anti-drug antibody, and safety pharmacology in Rhesus monkeys

Abstract

Mucopolysaccharidosis (MPS) Type II is caused by mutations in the gene encoding the lysosomal enzyme, iduronate 2-sulfatase (IDS). The majority of MPSII cases affect the brain. However, enzyme replacement therapy with recombinant IDS does not treat the brain, because IDS is a large molecule drug that does not cross the blood-brain barrier (BBB). To enable BBB penetration, IDS has been re-engineered as an IgG-IDS fusion protein, where the IgG domain is a monoclonal antibody (MAb) against the human insulin receptor (HIR). The HIRMAb crosses the BBB via receptor-mediated transport on the endogenous BBB insulin receptor, and the HIRMAb domain of the fusion protein acts as a molecular Trojan horse to ferry the fused IDS into brain from blood. The present study reports on the first safety pharmacology and pharmacokinetics study of the HIRMAb-IDS fusion protein. Juvenile male Rhesus monkeys were infused intravenously (IV) weekly for 26 weeks with 0, 3, 10, or 30 mg/kg of the HIRMAb-IDS fusion protein. The plasma clearance of the fusion protein followed a linear pharmacokinetics profile, which was equivalent either with measurements of the plasma concentration of immunoreactive HIRMAb-IDS fusion protein, or with assays of plasma IDS enzyme activity. Anti-drug antibody (ADA) titers were monitored monthly, and the ADA response was primarily directed against the variable region of the HIRMAb domain of the fusion protein. No infusion related reactions or clinical signs of immune response were observed during the course of the study. A battery of safety pharmacology, clinical chemistry, and tissue histopathology showed no signs of adverse events, and demonstrate the safety profile of chronic treatment of primates with 3-30 mg/kg weekly IV infusion doses of the HIRMAb-IDS fusion protein.

Keywords: blood-brain barrier; drug delivery; lysosomal enzyme; monoclonal antibody.

Figure 1

Immunocytochemistry of Rhesus monkey brain stained with 15 ug/mL of either the biotinylated HIRMAb-IDS fusion protein (A) or the biotinylated human IgG1k isotype control antibody (B). The slides are counter-stained with hematoxylin. Both sections were developed under identical conditions. Magnification bar in panel B is 22 microns.

Figure 2

Plasma profile of immunoreactive HIRMAb-IDS fusion protein (open squares) and IDS enzyme activity (closed squares) at week 1 for 3 mg/kg (A), 10 mg/kg (B), and 30 mg/kg (C) infusion doses of the HIRMAb-IDS fusion protein. Mean ± SD (N=6–9).

Figure 3

Plasma area under the concentration curve (AUC) of immunoreactive HIRMAb-IDS fusion protein (open squares), in µg•min/mL, and AUC of plasma IDS enzyme activity (closed squares), in kilounits•min/mL, is plotted vs dose of HIRMAb-IDS fusion protein at week 1. Mean ± SD.

Figure 4

(A) Comparison of plasma area under the concentration curve (AUC) for the IDS enzyme activity vs the plasma AUC of immunoreactive HIRMAb-IDS fusion protein at week 1. Mean ± SD. (B) Comparison of the in vivo specific activity (closed bar) of the HIRMAb-IDS fusion protein in plasma in monkeys over 23 hours after infusion vs the in vitro IDS specific activity (open bar) of the infused HIRMAb-IDS fusion protein. Mean ± SD. The in vivo specific activity was determined from the slope of the plot in panel A.

Figure 5

Plasma profile of IDS enzyme activity at week 25 for 3 mg/kg (A), 10 mg/kg (B), and 30 mg/kg (C) infusion doses of the HIRMAb-IDS fusion protein. Mean ± SD (N=6–9).

Figure 6

Time course of ADA formation against the HIRMAb-IDS fusion protein. Data are mean ± SE (n=6–9 monkeys per time point). All plasma samples were diluted 1:50 in PBS. The dose of each group of monkeys is given in the inset of the figure.

Figure 7

Absorbance (A490) is plotted against dilution of pools of primate plasma from 1:50 to 1:10,000 in PBS for monkeys administered the HIRMAb-IDS fusion protein at weekly doses of 3 mg/kg (panel A), 10 mg/kg (panel B), and 30 mg/kg (panel C). The pool was produced from the week 24 blood samples. The capture reagent in the sandwich ELISA is the HIRMAb-IDS fusion protein (open circles), the HIRMAb alone (closed squares), or the human IgG1κ isotype control (closed triangles). Data are mean ± SD (N=3 replicates).