Multivalent Poly(ethylene glycol)-Containing Conjugates for In Vivo Antibody Suppression

Abstract

Poly(ethylene glycol) (PEG) was incorporated into multivalent conjugates of the N-terminal domain of beta(2)GPI (domain 1). PEG was incorporated to reduce the rate of elimination of the conjugates from plasma and to putatively improve their efficacy as toleragens for the suppression of anti-beta(2)GPI antibodies and the treatment of antiphospholipid syndrome (APS). Three structurally distinct types of multivalent platforms were constructed by incorporating PEG into the platform structures in different ways. The amount of PEG incorporated ranged from about 5000 g per mole to about 30000 g per mole. The platforms were functionalized with either four or eight aminooxy groups. The conjugates were prepared by forming oxime linkages between the aminooxy groups and N-terminally glyoxylated domain 1 polypeptide. The plasma half-life of each conjugate, labeled with (125)I, was measured in both mice and rats. The half-lives of the conjugates ranged from less than 10 min to about 1 h in mice, and from less than 3 h to about 19 h in rats. The ability of five tetravalent conjugates to suppress anti-domain 1 antibodies in immunized rats was also measured. Incorporation of PEG in the conjugates significantly reduced the doses required for suppression, and the amount of reduction correlated with the amount of PEG incorporated.

Multivalent Compounds for Antigen-Specific B Cell Tolerance and Treatment of Autoimmune Diseases

“…incorporation of PEG into the conjugates reduced their rates of clearance from circulation, and the amount of reduction depended on the amount of PEG incorporated [93]. Antibody suppression studies with immunized rats also showed increases in efficacy that correlate with the amount of PEG [93].…”

“…One of these conjugates, LJP 1027, was found to clear much more slowly from rats than from mice [92], and it was effective at suppressing anti-β 2 GPI antibodies in vivo in domain 1-immunized rats at the highest dose used (88 mmol/kg) [93]. Those results made it apparent that keeping the drug in longer circulation could improve its efficacy, so toleragens were developed with PEG incorporated into their structure [93]. Attachment of PEG to peptides, proteins, and small molecule drugs is a popular strategy to decrease their rates of clearance from circulation [94][95][96].…”

“…Antibody suppression studies with immunized rats also showed increases in efficacy that correlate with the amount of PEG [93]. It was somewhat surprising that the structural differences in the way the PEG was incorporated did not translate into significant differences in toleragenic activity.…”

Effect of the nonreducing end of Shigella dysenteriae type 1 O-specific oligosaccharides on their immunogenicity as conjugates in mice

“…Covalent attachment of the keto-derivatized oligosaccharides 1C-10C to BSA was achieved by oxime formation (26,27,(30)(31)(32)(33) between the saccharides' keto group and aminooxypropyl groups bound to the protein through a thioether linkage (26,27,(34)(35)(36)(37). For this purpose, BSA was reacted with succinimidyl 3-(bromoacetamido)propionate (36) 30, as described (26,27,38) (Fig.…”

Site-specific protein immobilization through N-terminal oxime linkages

“…12 Reaction of a N-terminally glyoxylated b 2 -glycoprotein (domain 1) with an aminooxy-terminated poly(ethylene glycol) (PEG)-based polymer has been reported. 13,14 After modification with PLP, myoglobin, green fluorescent protein, RNase A, and thioredoxin have also been conjugated to a linear PEG alkoxyamine in solution. 11 On surfaces, carbohydrates have been immobilized on an aminooxy-terminated polymer film via an oxime bond at the reducing terminus.…”