Development and testing of AAV-delivered single-chain variable fragments for the treatment of methamphetamine abuse

Abstract

Methamphetamine (METH) substance abuse disorders have major impact on society, yet no medications have proven successful at preventing METH relapse or cravings. Anti-METH monoclonal antibodies can reduce METH brain concentrations; however, this therapy has limitations, including the need for repeated dosing throughout the course of addiction recovery. An adeno-associated viral (AAV)-delivered DNA sequence for a single-chain variable fragment could offer long-term, continuous expression of anti-METH antibody fragments. For these studies, we injected mice via tail vein with 1 x 1012 vector genomes of two AAV8 scFv constructs and measured long-term expression of the antibody fragments. Mice expressed each scFv for at least 212 days, achieving micromolar scFv concentrations in serum. In separate experiments 21 days and 50 days after injecting mice with AAV-scFvs mice were challenged with METH in vivo. The circulating scFvs were capable of decreasing brain METH concentrations by up to 60% and sequestering METH in serum for 2 to 3 hrs. These results suggest that AAV-delivered scFv could be a promising therapy to treat methamphetamine abuse.

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

“…In both this study (data not shown) and as previously reported with the same concentration of AAV, no lasting adverse effects were observed as a result of the high AAV dose (Hay et al, 2018).…”

“…One pharmacological approach is to use anti-METH monoclonal antibodies (mAbs) as pharmacokinetic antagonists to block or blunt METH effects. Unlike a small molecule medication, which would selectively block multiple receptors for extended periods of time, mAbs sequester METH in the circulatory system Stevens, Tawney, et al, 2014;Hay et al, 2018) and reduce METH entry into critical tissues like the central nervous system (CNS) and the cardiovascular system. Pharmacokinetic assays demonstrated that an anti-METH mAb therapy, administered as a bolus i.v.…”

“…Adeno-associated viruses (AAV) is commonly used as gene therapy vectors, because it is a non-pathogenic virus that can deliver up to 4.5 kilobases of DNA to host tissues (Zolotukhin, 2005). Previous studies have shown that a single dose of AAV, encoding an entire mAb sequence, reaches maximal expression of mAb in 3-4 weeks and expression persists for at least 8 months Rosenberg et al, 2012Rosenberg et al, , 2013Hay et al, 2018). Hicks et al (2012) showed that an anti-nicotine AAV based mAb therapy elicited over 4-fold greater circulating concentrations of high affinity mAbs with one injection compared to an anti-nicotine vaccine, which generated variable amounts of polyclonal antibodies and required one to two additional booster doses.…”

“…A full IgG antibody sequence can fit into an AAV capsid, but requires post-translational autocleavage of the heavy and light chains of the IgG antibody before dimerizing back together through disulfide bonds (Fang et al, 2005;Ho et al, 2013). 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).…”

“…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).…”