OXavidin for Tissue Targeting Biotinylated Therapeutics

Abstract

Avidin is a glycoprotein from hen egg white that binds biotin with very high affinity. Here we describe OXavidin, a product containing aldehyde groups, obtained by ligand-assisted sugar oxidation of avidin by sodium periodate. OXavidin chemically reacts with cellular and tissue proteins through Schiff's base formation thus residing in tissues for weeks while preserving the biotin binding capacity. The long tissue residence of OXavidin as well as that of OXavidin/biotinylated agent complex occurs in normal and neoplastic tissues and immunohistochemistry shows a strong and homogenous stromal localization. Once localized in tissue/tumor, OXavidin becomes an “artificial receptor” for intravenous injected biotin allowing tumor targeting with biotinylated therapeutics like radioisotopes or toxins. Moreover, present data also suggest that OXavidin might be useful for the homing of biotinylated cells. Overall, OXavidin exhibits a remarkable potential for many different therapeutic applications.

Biochemical and Biological Characterization of a New Oxidized Avidin with Enhanced Tissue Binding Properties

“…Therefore, to evaluate their diffusion kinetics from a treated tissue, either 125 I-radiolabeled avidin or streptavidin was injected in the muscle of one hind limb of each mouse, simulating the recently described approach of cancer pretargeted radiotherapy named IART (37)(38)(39), and at different time points the mice were sacrificed, and the treated and controlateral limb (muscle), blood, liver, and kidney were collected, and radioactivity was measured by a ␥ counter. Previously published data from our group showed that the residence time of 125 I-labeled avidin and streptavidin in the treated tissue is very short, with ϳ8% of the injected dose/100 mg observed after 1 h and less than 1% ID/100 mg after 24 h. In agreement to previous pharmacokinetics data on avidin and streptavidin intravenously injected, the avidin diffused from the injected tissue, exhibited a faster clearance from the blood than streptavidin, and accumulated in the liver and kidney, whereas streptavidin preferentially accumulated in the kidney (35).…”

“…6 indicate that the half-life of OXavidin HABA in the treated limb is ϳ2 weeks as opposed to ϳ2 h previously reported for avidin (35). Both avidins were almost undetectable in blood, liver, and kidney 2 days after the intramuscular injection (data not shown).…”

“…After purification by size exclusion chromatography, 125 I-PEG-avidin was injected in the muscle of mice as described previously, and its tissue residence was compared with the one of radiolabeled avidin and oxidized avidin. The results indicated that 24 h after injection, less than 2% ID/100 mg of PEG-avidin and avidin was present in the treated limb versus ϳ16% of oxidized avidin (35). These findings, besides ruling out the possibility of improving the tissue permanence of avidin simply by increasing its size as for intravenously injected PEG-avidin, provide further evidence of the relationship between oxidized avidin tissue permanence and aldehyde groups.…”

“…Equal amounts of 125 I-labeled oxidized avidin or avidin were then intramuscularly injected in one hind limb of each mouse, and the tissue permanence was evaluated at different time points. One hour after injection, the amount of oxidized avidin in the treated limb was twice that of avidin, and the difference increased with time; avidin was almost undetectable after 24 or 48 h, whereas oxidized avidin was always higher than 10% ID/100 mg of tissue (35). These findings suggested that oxidation allows the avidin to react with tissue proteins in vivo by the formation of Schiff's bases.…”

“…In the perspective of improving avidin pharmacokinetic properties for tissue engineering applications, an oxidized avidin has been recently described showing that, once injected in tissues, might be useful in taking up intravenously injected radioactive biotin (35). Here we describe in detail the procedure for obtaining such an avidin variant by sodium periodate oxidation of the sugar pyranosydic rings after saturation with the low affinity ligand 4-hydroxyazobenzene-2Ј-carboxylic acid (HABA) 2 (36).…”