Using Hapten Design to Discover Therapeutic Monoclonal Antibodies for Treating Methamphetamine Abuse

Abstract

When generating monoclonal antibodies (mAb) against small molecules, the chemical composition and molecular orientation of the drug-like hapten on the antigen is a crucial determinant. This is especially important when attempting to discover therapeutic mAb against the drugs of abuse (ϩ)-methamphetamine, and the related compound (ϩ)-3,4-methylenedioxymethamphetamine [(ϩ)-MDMA, the plus isomer in the racemic mixture known as MDMA or ecstasy]. The goal of these studies was to design and synthesize (ϩ)-METH-like haptens with structural attributes that could make them effective for generating monoclonal antibodies for treating medical problems associated with these stimulant drugs of abuse. Five prototype (ϩ)-METH-like haptens, which mimic structural aspects of these drugs, were synthesized and used to generate mAb. After screening for anti-(ϩ)-METH IgG antibodies in more than 25,000 potential mouse hybridoma cell lines, one prototype mAb from each of the five haptens was selected and studied in detail for molecular properties and preclinical efficacy. The amino acid sequences of the IgGvariable regions, structural models, affinity, and ligand specificity of each mAb were then used to help elucidate important therapeutic characteristics. Four of these antibodies exhibited high affinity and specificity to (ϩ)-METH and (ϩ)-MDMA; whereas one antibody (designated mAb4G9) exhibited high affinity and specificity to (ϩ)-METH, (ϩ)-MDMA, and (ϩ)-AMP, without significant cross-reactivity against other METH-like ligands, over-the-counter medications, or endogenous neurotransmitters. Considered together, discovery of mAb4G9 and the other antibodies in this report represent an important step in understanding the process for custom design of drug class-specific therapeutic antibodies for the treatment of drug addiction. (ϩ)-Methamphetamine [(ϩ)-METH] abuse has becomeAmerica's number one drug threat (NACo, 2005), and effective treatment strategies for abuse of (ϩ)-METH and related stimulants are greatly needed. Current pharmacotherapies for managing the acute cardiovascular system, central nervous system, and toxic effects are mostly supportive (Sato, 1992;Albertson et al., 1999;Richards et al., 1999); they do nothing to remove the drug from its sites of action in the brain. Also lacking are medications that can reduce or treat the medically crippling effects of (ϩ)-METH addiction. Monoclonal antibodies (mAb) provide an attractive potential medication that can target the drug instead of the site of action (Kosten and Owens, 2005). These high-affinity protein-based medications act as so-called pharmacokinetic antagonists, sequestering the drug in the bloodstream away from medically vulnerable tissues, such as the brain and heart. Unlike nicotine and cocaine where the effects are caused by a single, specific compound, drugs such as opiates (e.g., morphine), arylcyclohexylamines (e.g., phencyclidine) and amphetamines [e.g., (ϩ)-METH] are starting structures from which many pharmacologically similar compounds can be synthesize...

Crystal structures of a therapeutic single chain antibody in complex with two drugs of abuse—Methamphetamine and 3,4‐methylenedioxymethamphetamine

“…2(A)]. 8 Water molecules in the binding cavity Because METH and MDMA are predominantly hydrophobic, one would expect the binding cavities to be devoid of solvent molecules; however, in both complexes, there are two water molecules, Ow(5) and Ow (6), in the cavity separated by a distance of 2.8 Å indicating a strong hydrogen bonding interaction between them [ Fig. 5(A)].…”

“…Critical improvements have come through progressive knowledge-based design of haptens and the use of more effective antigenic carrier proteins for stimulating an immune response. 6 Generating high-specificity antibodies against METH is challenging due to the presence of pharmacologically active metabolites and the potential presence of some over the counter medications that could compete for binding and reduce efficacy. An additional challenge arises from the small size of the drug molecule.…”

“…11 We crystallized a scFv derived from one of our highest affinity antibodies, mAb6H4 (K D ¼ 10 nM). 6 This mAb was generated using a hapten called METH-P6 [ Fig. 2(A)].…”

Pharmacological effects of two anti-methamphetamine monoclonal antibodies

“…1,10 Large scale production of anti-METH mAb, purification and formulation are also described elsewhere. 10 The mAb4G9 is an IgG2b, k light chain mAb and the mAb7F9 is an IgG1, k light chain.…”

“…6,9,10 Furthermore, both mAbs have high affinity for METH and differing affinities for the related compounds (C)-amphetamine (AMP) and (C)-3,4-methylenedioxymethamphetamine (MDMA). MAb4G9 has a K D for METH of 16 nM and a K I for MDMA of 68 nM.…”