PEGylation of a High-Affinity Anti-(+)Methamphetamine Single Chain Antibody Fragment Extends Functional Half-Life by Reducing Clearance

Abstract

Purpose Methamphetamine (METH) abuse is a worldwide drug problem, yet no FDA-approved pharmacological treatments are available for METH abuse. Therefore, we produced an anti- METH single chain antibody fragment (scFv7F9Cys) as a pharmacological treatment for METH abuse. ScFv’s have a short half-life due to their small size, limiting their clinical use. Thus, we examined the pharmacokinetic effects of conjugating poly(ethylene) glycol (-PEG) to scFv7F9Cys to extend its functional half-life. Methods The affinity of scFv7F9Cys and PEG conjugates to METH was determined in vitro via equilibrium dialysis saturation binding. Pharmacokinetic and parameters of scFv7F9Cys and scFv7F9Cys-PEG20K (30 mg/kg i.v. each) and their ability to bind METH in vivo were determined in male Sprague-Dawley rats receiving a subcutaneous infusion of METH (3.2 mg/kg/day). Results Of three PEGylated conjugates, scFv7F9Cys-PEG20K was determined the most viable therapeutic candidate. PEGylation of scFv7F9Cys did not alter METH binding functionality in vitro, and produced a 27-fold increase in the in vivo half-life of the antibody fragment. Furthermore, total METH serum concentrations increased following scFv7F9Cys or scFv7F9Cys-PEG20K administration, with scFv7F9Cys-PEG20K producing significantly longer changes in METH distribution than scFv7F9Cys. Conclusions PEGylation of scFv7F9Cys significantly increase the functional half-life of scFv7F9Cys, suggesting it may be a long-lasting pharmacological treatment option for METH abuse.

Repeated Transcranial Direct Current Stimulation (tDCS) on Methamphetamine Craving: A Randomized, Sham-controlled Study

“…Amphetaminetype stimulants are the most common abused substances after cannabis and compared to other substance use disorders, amphetamine use disorder constitutes a considerable piece of the global weight of disease [1]. The number of methamphetamine addicts and demand for the treatment of methamphetamine-related conditions is increasing in many parts of the world, including southwest Asia [1]; however, no proper pharmacological treatment has yet been approved by the Food and Drug Administration [2], and behavioral interventions, as the main rehabilitation for these patients [3], have been successful only to some extent [4].…”

The Development and Characterization of an scFv-Fc Fusion–Based Gene Therapy to Reduce the Psychostimulant Effects of Methamphetamine Abuse

“…The short half-life of scFvs, which is approximately 60 min due to high clearance rate, limits the circulating concentrations of anti-METH antibodies in mice. (Nanaware-Kharade et al, 2015;Reichard et al, 2016;Hay et al, 2018).…”

“…Wells were measured for light absorption at 480 nm on a Biotek HT Synergy plate reader. Concentrations of the anti-METH antibodies were interpolated from a standard curve of purified scFv7F9 protein (Reichard et al, 2016).…”

“…Serum pooled from vehicle mice on day 22 was used as a negative control to show mice not given the AAV-7F9-Fc vectors did not express 7F9-Fc protein. Purified scFv7F9 protein with a 6His tag was used as a positive control (Reichard et al, 2016). The Western blot membranes were stained with HRP anti-His tag antibody (Biolegend, #652504) diluted 1:2,000 in 0.5% (w/v) BSA.…”

The development and characterization of an scFv-Fc fusion based gene therapy to reduce the psychostimulant effects of methamphetamine abuse

“…In contrast, we have previously reported the design and testing of a singlechain variable fragment (scFv), which consists of only the variable regions of an IgG connected by a 50 amino acid linker (Hay et al, 2018). Because scFvs have a short half-life of roughly 60 min, limited circulating concentrations of anti-METH antibodies were observed in mice (Nanaware-Kharade et al, 2015;Reichard et al, 2016;Hay et al, 2018).…”