Vaccine-driven pharmacodynamic dissection and mitigation of fenethylline psychoactivity.

Fenethylline, also known by the trade name Captagon, is a synthetic psychoactive stimulant that has recently been linked to a substance-use disorder and 'pharmacoterrorism' in the Middle East. Although fenethylline shares a common phenethylamine core with other amphetamine-type stimulants, it also incorporates a covalently linked xanthine moiety into its parent structure. These independently active pharmacophores are liberated during metabolism, resulting in the release of a structurally diverse chemical mixture into the central nervous system. Although the psychoactive properties of fenethylline have been reported to differ from those of other synthetic stimulants, the in vivo chemical complexity it manifests upon ingestion has impeded efforts to unambiguously identify the specific species responsible for these effects. Here we develop a 'dissection through vaccination' approach, called DISSECTIV, to mitigate the psychoactive effects of fenethylline and show that its rapid-onset and distinct psychoactive properties are facilitated by functional synergy between theophylline and amphetamine. Our results demonstrate that incremental vaccination against a single chemical species within a multi-component mixture can be used to uncover emergent properties arising from polypharmacological activity. We anticipate that DISSECTIV will be used to expose unidentified active chemical species and resolve pharmacodynamic interactions within other chemically complex systems, such as those found in counterfeit or illegal drug preparations, post-metabolic tissue samples and natural product extracts.

An Anticaffeine Antibody-Oligonucleotide Conjugate for DNA-Directed Immobilization in Environmental Immunoarrays.

The development of fast and cheap high-throughput platforms for the detection of environmental contaminants is of particular importance to understand the human-related impact on the environment. The application of DNA-directed immobilization (DDI) of IgG molecules is currently limited to the clinical diagnostics scenario, possibly because of the high costs of production of such addressable platforms. We here describe the efficient and specific hybridization of an antibody-oligonucleotide conjugate to a short 12-mer capture probe. The specific antibody used is a monoclonal antibody against caffeine, a stimulant and important anthropogenic marker. With this work, we hope to contribute to broadening the application potential of DDI to environmental markers in order to develop cheaper and more stable high-throughput screening platforms for standard routine analysis of pollutants in a variety of complex matrices.

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

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.

Caffeine ingestion and antigen-stimulated human lymphocyte activation after prolonged cycling.

This study investigated the effect of caffeine ingestion on antigen-stimulated T- (CD4(+) and CD8(+) ) and natural killer (NK)- (CD3(-) CD56(+) ) cell activation after prolonged, strenuous cycling. In a randomized cross-over design, nine male endurance cyclists (age: 22 ± 3 years, VO(2peak) : 62 ± 4 mL/kg/min, mean ± SD) cycled for 90 min at 70% VO(2peak) 60 min after ingesting 6 mg/kg body mass of caffeine (CAF) or placebo (PLA). Venous blood samples were obtained before supplementation, pre-exercise, immediately post-exercise and 1 h post-exercise. Whole blood was stimulated with Pediacel (five in one) vaccine. At 1 h post-exercise the number of antigen-stimulated CD4(+) cells expressing CD69 decreased on CAF compared with PLA [15 (17) × 10(6) vs 23 (22) × 10(6) cells/L, P<0.05]. In addition, the geometric mean fluorescence intensity (GMFI) of CD69 expression on antigen-stimulated CD8(+) cells decreased on CAF compared with PLA 1 h post-exercise [78 (10)% vs 102 (24)%, P<0.05]. At the same time-point GMFI of CD69 expression on antigen-stimulated CD3(-) CD56(+) cells was increased on CAF compared with PLA [103 (9)% vs 87 (8)%, P<0.05]. These findings suggest that caffeine reduces antigen-stimulated CD69 expression on T cells while at the same time increases NK-cell activation 1 h after intensive cycling.

Impact of distinct chemical structures for the development of a methamphetamine vaccine.

(+)-Methamphetamine (METH) use and addiction has grown at alarming rates over the past two decades, while no approved pharmacotherapy exists for its treatment. Immunopharmacotherapy has the potential to offer relief through producing highly specific antibodies that prevent drug penetration across the blood-brain barrier thus decreasing reinforcement of the behavior. Current immunotherapy efforts against methamphetamine have focused on a single hapten structure, namely linker attachment at the aromatic ring of the METH molecule. Hapten design is largely responsible for immune recognition, as it affects presentation of the target antigen and thus the quality of the response. In the current paper we report the systematic generation of a series of haptens designed to target the most stable conformations of methamphetamine as determined by molecular modeling. On the basis of our previous studies with nicotine, we show that introduction of strategic molecular constraint is able to maximize immune recognition of the target structure as evidenced by higher antibody affinity. Vaccination of GIX(+) mice with six unique METH immunoconjugates resulted in high antibody titers for three particularly promising formulations (45-108 µg/mL, after the second immunization) and high affinity (82, 130, and 169 nM for MH2, MH6, and MH7 hapten-based vaccines, respectively). These findings represent a unique approach to the design of new vaccines against methamphetamine abuse.

Cerebral microglia mediate sleep/wake and neuroinflammatory effects of methamphetamine.

Methamphetamine and modafinil exert their wake-promoting effects by elevating monoaminergic tone. The severity of hypersomnolence that occurs subsequent to induced wakefulness differs between these two agents. Microglia detects and modulates CNS reactions to agents such as D-methamphetamine that induce cellular stress. We therefore hypothesized that changes in the sleep/wake cycle that occur subsequent to administration of D-methamphetamine are modulated by cerebral microglia. In CD11b-herpes thymidine kinase transgenic mice (CD11b-TK(mt-30)), activation of the inducible transgene by intracerebroventricular (icv) ganciclovir results in toxicity to CD11b-positive cells (i.e. microglia), thereby reducing cerebral microglial cell counts. CD11b-TK(mt-30)and wild type mice were subjected to chronic icv ganciclovir or vehicle administration with subcutaneous mini-osmotic pumps. D-methamphetamine (1 and 2 mg/kg), modafinil (30 and 100 mg/kg) and vehicle were administered intraperitoneally to these animals. In CD11b-TK(mt-30) mice, but not wild type, icv infusion of ganciclovir reduced the duration of wake produced by D-methamphetamine at 2 mg/kg by nearly 1h. Nitric oxide synthase (NOS) activity, studied ex vivo, and NOS expression were elevated in CD11b-positive cerebral microglia from wild type mice acutely exposed to d-methamphetamine. Additionally, CD11b-positive microglia, but not other cerebral cell populations, exhibited changes in sleep-regulatory cytokine expression in response to d-METH. Finally, CD11b-positive microglia exposed to d-methamphetamine in vitro exhibited increased NOS activity relative to pharmacologically-naïve cells. CD11b-positive microglia from the brains of neuronal NOS (nNOS)-knockout mice failed to exhibit this effect. We propose that the effects of D-METH on sleep/wake cycles are mediated in part by actions on microglia, including possibly nNOS activity and cytokine synthesis.

Decrease of lymphoproliferative response by amphetamine is mediated by dopamine from the nucleus accumbens: influence on splenic met-enkephalin levels.

Despite the mesocorticolimbic dopaminergic pathway being one of the main substrates underlying stimulating and reinforcing effects induced by psychostimulant drugs, there is little information regarding its role in their effects at the immune level. We have previously demonstrated that acute exposure to amphetamine (5 mg/kg, i.p.) induced an inhibitory effect on the splenic T-cell proliferative response, along with an increase in the methionine(met)-enkephalin content at limbic and immune levels, 4 days after drug administration. In this study, we investigated if a possible dopamine mechanism underlies these amphetamine-induced effects by administering D1 and D2 dopaminergic antagonists or a dopaminergic terminal neurotoxin before the drug. Pre-treatment with either SCH-23390 (0.1 mg/kg, i.p.) or raclopride (0.1 mg/kg, i.p.), a D1 or D2 dopaminergic receptor antagonist, respectively, abrogated the effects of amphetamine on the lymphoproliferative response and on met-enkephalin levels of the spleen. The amphetamine-induced increase in limbic met-enkephalin content was suppressed by SCH-23390 but not by raclopride pre-treatment. Finally, an intra-accumbens 6-hydroxy-dopamine injection administered 2 weeks previously prevented amphetamine-induced effects on the lymphoproliferative response and on met-enkephalin levels in the prefrontal cortex and spleen. These findings strongly suggest that D1 and D2 dopaminergic receptors are involved in amphetamine-induced effects at immune level as regards the lymphoproliferative response and the changes in spleen met-enkephalin content, whereas limbic met-enkephalin levels were modulated only by the D1 dopaminergic receptors. In addition, this study showed that a mesolimbic component modulated amphetamine-induced effects on the immune response, as previously shown at a behavioral level.

Influencing Antibody-Mediated Attenuation of Methamphetamine CNS Distribution through Vaccine Linker Design.

Active vaccination examining a single hapten engendered with a series of peptidic linkers has resulted in the production of antimethamphetamine antibodies. Given the limited chemical complexity of methamphetamine, the structure of the linker species embedded within the hapten could have a substantial effect on the ultimate efficacy of the resulting vaccines. Herein, we investigate linker effects by generating a series of methamphetamine haptens that harbor a linker with varying amino acid identity, peptide length, and associated carrier protein. Independent changes in each of these parameters were found to result in alterations in both the quantity and quality of the antibodies induced by vaccination. Although it was found that the consequence of the linker design was also dependent on the identity of the carrier protein, we demonstrate overall that the inclusion of a short, structurally simple, amino acid linker benefits the efficacy of a methamphetamine vaccine in limiting brain penetration of the free drug.

Active vaccination attenuates the psychostimulant effects of α-PVP and MDPV in rats.

Recreational use of substituted cathinones continues to be an emerging public health problem in the United States; cathinone derivatives α-pyrrolidinopentiophenone (α-PVP) and 3,4-methylenedioxypyrovalerone (MDPV), which have been linked to human fatalities and show high potential for abuse liability in animal models, are of particular concern. The objective of this study was to develop an immunotherapeutic strategy for attenuating the effects of α-PVP and MDPV in rats, using drug-conjugate vaccines created to generate antibodies with neutralizing capacity. Immunoconjugates (α-PVP-KLH and MDPV-KLH) or the control carrier protein, keyhole limpet hemocyanin (KLH), were administered to groups (N = 12) of male Sprague-Dawley rats on Weeks 0, 2 and 4. Groups were administered α-PVP or MDPV (0.0, 0.25, 0.5, 1.0, 5.0 mg/kg, i.p.) in acute drug challenges and tested for changes in wheel activity. Increased wheel activity produced by α-PVP or MDPV in the controls was attenuated in the α-PVP-KLH and MDPV-KLH vaccinated groups, respectively. Rectal temperature decreases produced by MDPV in the controls were reduced in duration in the MDPV-KLH vaccine group. A separate group (N = 19) was trained to intravenously self-administer α-PVP (0.05, 0.1 mg/kg/inf) and vaccinated with KLH or α-PVP-KLH, post-acquisition. Self-administration in α-PVP-KLH rats was initially higher than in the KLH rats but then significantly decreased following a final vaccine booster, unlike the stable intake of KLH rats. The data demonstrate that active vaccination provides functional protection against the effects of α-PVP and MDPV, in vivo, and recommend additional development of vaccines as potential therapeutics for mitigating the effects of designer cathinone derivatives.

An enzyme-linked immunosorbent assay (ELISA) for methylphenidate (Ritalin ) in urine.

A direct enyzme-linked immunosorbent assay (ELISA) for urinary immunoreactive methylphenidate (Ritalin), in which a standard 96-well microtiter plate is used, is described. For this ELISA, a methylphenidate-thyroglobulin conjugate is immobilized to the microtiter plate and competes with methylphenidate in the standard or urine sample for antibody-binding sites. After washing, the sheep methylphenidate antibody bound to immobilized methylphenidate is detected with peroxidase-labelled goat antisheep IgG. Following a further wash, tetramethylbenzidine is added, color is developed, and the plate is read at 450 nm on an ELISA plate reader. This method is unaffected by drugs of abuse and is suitable for routine use in the toxicology laboratory.

Cross-reactivity studies and predictive modeling of "Bath Salts" and other amphetamine-type stimulants with amphetamine screening immunoassays.

INTRODUCTION: The increasing abuse of amphetamine-like compounds presents a challenge for clinicians and clinical laboratories. Although these compounds may be identified by mass spectrometry-based assays, most clinical laboratories use amphetamine immunoassays that have unknown cross-reactivity with novel amphetamine-like drugs. To date, there has been a little systematic study of amphetamine immunoassay cross-reactivity with structurally diverse amphetamine-like drugs or of computational tools to predict cross-reactivity. METHODS: Cross-reactivities of 42 amphetamines and amphetamine-like drugs with three amphetamines screening immunoassays (AxSYM(®) Amphetamine/Methamphetamine II, CEDIA(®) amphetamine/Ecstasy, and EMIT(®) II Plus Amphetamines) were determined. Two- and three-dimensional molecular similarity and modeling approaches were evaluated for the ability to predict cross-reactivity using receiver-operator characteristic curve analysis. RESULTS: Overall, 34%-46% of the drugs tested positive on the immunoassay screens using a concentration of 20,000 ng/mL. The three immunoassays showed differential detection of the various classes of amphetamine-like drugs. Only the CEDIA assay detected piperazines well, while only the EMIT assay cross-reacted with the 2C class. All three immunoassays detected 4-substituted amphetamines. For the AxSYM and EMIT assays, two-dimensional molecular similarity methods that combined similarity to amphetamine/methamphetamine and 3,4-methylenedioxymethampetamine most accurately predicted cross-reactivity. For the CEDIA assay, three-dimensional pharmacophore methods performed best in predicting cross-reactivity. Using the best performing models, cross-reactivities of an additional 261 amphetamine-like compounds were predicted. CONCLUSIONS: Existing amphetamines immunoassays unevenly detect amphetamine-like drugs, particularly in the 2C, piperazine, and ß-keto classes. Computational similarity methods perform well in predicting cross-reactivity and can help prioritize testing of additional compounds in the future.

A methamphetamine vaccine attenuates methamphetamine-induced disruptions in thermoregulation and activity in rats.

BACKGROUND: There are no approved pharmacotherapies for d-methamphetamine (METH) addiction and existing therapies have limited efficacy. Advances in using immunotherapeutic approaches for cocaine and nicotine addiction have stimulated interest in creating a similar approach for METH addiction. This study investigated whether active vaccination against METH could potentially attenuate responses to METH in vivo. METHODS: Male Sprague Dawley rats (n = 32) received a four-boost series with one of three candidate anti-METH vaccines (MH2[R], MH6, and MH7) or a control keyhole limpet hemocyanin conjugate vaccine. Effects of METH on rectal temperature and wheel activity at 27°C ambient temperature were determined. The most efficacious vaccine, MH6, was then contrasted with keyhole limpet hemocyanin conjugate vaccine in a subsequent experiment (n = 16), wherein radiotelemetry determined home cage locomotor activity and body temperature at 23°C ambient temperature. RESULTS: The MH6 vaccine produced high antibody titers with nanomolar affinity for METH and sequestered METH in the periphery of rats. In experiment 1, the thermoregulatory and psychomotor responses produced by METH at 27°C were blocked in the MH6 group. In experiment 2, METH-induced decreases in body temperature and locomotor activity at 23°C were also attenuated in the MH6 group. A pharmacokinetic study in experiment 2 showed that MH6-vaccinated rats had higher METH serum concentrations, yet lower brain METH concentrations, than control rats, and METH concentrations correlated with individual antibody titer. CONCLUSIONS: These data demonstrate that active immunopharmacotherapy provides functional protection against physiological and behavioral disruptions induced by METH.

A vaccine against methamphetamine attenuates its behavioral effects in mice.

BACKGROUND: Vaccines have treatment potential for methamphetamine (MA) addiction. We tested whether a conjugate vaccine against MA (succinyl-methamphetamine-keyhole limpet hemocyanin carrier protein; SMA-KLH) would generate MA antibodies and alter MA-induced behaviors. METHODS: Mice were injected with SMA-KLH and received booster administrations 3 and 20 weeks later. Serum antibody titers reached peak levels by 4-6 weeks, remained at a modest level through 18 weeks, peaked again at 22 weeks after the second boost, and were still elevated at 35 weeks. At 7 weeks, groups of vaccinated and non-vaccinated mice were administered one of three MA doses (1, 2 or 3 mg/kg) to assess locomotor activity. RESULTS: Non-vaccinated mice showed dose-dependent effects of MA with hypolocomotion at the lowest dose and elevated activity levels at the highest dose. Both dose effects were reduced in SMA-KLH groups, particularly low dose-induced hypolocomotion at later times post MA administration. Separate groups of vaccinated and non-vaccinated mice were trained in MA place conditioning at 30 weeks with either 0 (vehicle) or 0.5mg/kg MA. Although times spent in the MA-paired side did not differ between groups on test vs. baseline sessions, SMA-KLH mice conditioned with MA showed reduced conditioned approach behaviors and decreased conditioned activity levels compared to control groups. CONCLUSION: These data suggest SMA-KLH attenuates the ability of MA to support place conditioning and reduces or delays its locomotor effects. Overall, results support SMA-KLH as a candidate MA vaccine.

Cross-reactivities of various phenethylamine-type designer drugs to immunoassays for amphetamines, with special attention to the evaluation of the one-step urine drug test Instant-View™, and the Emit® assays for use in drug enforcement.

Cross-reactivities of 76 kinds of phenethylamine-type designer drugs and related compounds to the urine drug tests Instant-View ™ (IV) (the Methamphetamine (MA) test, the Amphetamine 300 test, and the MDMA test) have been investigated. An on-site urine test kit consisting of these three IV tests has been evaluated for the on-site screening of MA users, and the kit has been found to have satisfactory specificity for drug enforcement purposes by separately detecting both MA and its metabolite amphetamine. The cross-reactivity profiles of Emit(®) II Plus Amphetamines Assay, Emit(®) II Plus Ecstasy assay, and Emit(®) d.a.u.(®) Amphetamine Class assay have also been investigated and discussed.

Insights into scFv:drug binding using the molecular dynamics simulation and free energy calculation.

Molecular dynamics simulations and free energy calculation have been performed to study how the single-chain variable fragment (scFv) binds methamphetamine (METH) and amphetamine (AMP). The structures of the scFv:METH and the scFv:AMP complexes are analyzed by examining the time-dependence of their RMSDs, by analyzing the distance between some key atoms of the selected residues, and by comparing the averaged structures with their corresponding crystallographic structures. It is observed that binding an AMP to the scFv does not cause significant changes to the binding pocket of the scFv:ligand complex. The binding free energy of scFv:AMP without introducing an extra water into the binding pocket is much stronger than scFv:METH. This is against the first of the two scenarios postulated in the experimental work of Celikel et al. (Protein Science 18, 2336 (2009)). However, adding a water to the AMP (at the position of the methyl group of METH), the binding free energy of the scFv:AMP-H2O complex, is found to be significantly weaker than scFv:METH. This is consistent with the second of the two scenarios given by Celikel et al. Decomposition of the binding energy into ligand-residue pair interactions shows that two residues (Tyr175 and Tyr177) have nearly-zero interactions with AMP in the scFv:AMP-H2O complex, whereas their interactions with METH in the scFv:METH complex are as large as -0.8 and -0.74 kcal mol(-1). The insights gained from this study may be helpful in designing more potent antibodies in treating METH abuse.

Effects of amphetamine on NK-related cytotoxicity in rats differing in locomotor reactivity and social position.

The effect of i.p. administration of 1mg/kg of amphetamine (AMPH) on natural killer cells cytotoxicity (NKCC) and number of large granular lymphocytes (LGL-NK) together with plasma corticosterone (CORT) level and WBC was evaluated in male Wistar rats differing in two behavioral features: locomotor reactivity to novelty (high, HR and low, LR responders) and social position (dominants, D and subordinates, S). In the majority of animals AMPH evoked (30 min after administration) an increase in NKCC and LGL (NK) number accompanied by lymphopenia, neutrocytosis, monocytosis, and an increase in CORT level. Changes in NKCC (LU20) showed substantial individual variability: in HR group approximately 513Delta%, p <0.01 (relative to the control); LR group approximately 56Delta%, p >.05; D group approximately 441Delta%, p >0.001; S group approximately 216Delta%, p >0.05; HR/D group approximately 643Delta%, p <.001; HR/S group approximately 414Delta%, p <.001; LR/D group approximately 191Delta%, p >.05; and LR/S group approximately -19Delta%, p .05. The increase in CORT level, lymphopenia, and neutrocytosis indicated a stress-like reaction to AMPH. No significant correlation between NKCC and CORT level was found. The results obtained indicate that AMPH can evoke an increase in NK-related cytotoxic activity quantitatively related to high behavioral reactivity to novelty and social dominance, however NKCC is not related to the AMPH-induced CORT changes.

Comparison of capillary electrophoresis-based immunoassay with fluorescence polarization immunoassay for the immunodetermination of methamphetamine using various methamphetamine antibodies.

An accurate and simple immunoassay using capillary electrophoresis (CE) with laser-induced fluorescence (LIF) was performed for the detection of methamphetamine (MA) in urine. The CE-LIF was conducted with an untreated fused-silica column using antiserum and a tracer of fluorescein isothiocyanate (FITC)-labeled MA. This CE-LIF system was compared with fluorescence polarization immunoassay (FPIA) in a TDx analyzer in the photo-check mode using the same FITC-labeled tracer and the same antiserum. Various antibodies, not only those prepared by our own immunogens but also those from commercial sources, were screened and characterized in both assay systems with regard to sensitivity, precision, and cross-reactivity. Both systems satisfied analytical precision and gave similar cross-reactivity patterns. However, the CE-LIF-based immunoassay was approximately one order superior to FPIA in sensitivity, requiring less volume of sample, antiserum, and tracer for the assay. Considering that the FPIA system is well known to be a useful tool for screening antibodies and detecting drugs, the CE-LIF-based immunoassay system, which is seemingly more advantageous than the FPIA system, appears to have great power for the characterization of antibodies and for the detection of MA in urine.

Production and characterization of monoclonal antibody that simultaneously recognizes methamphetamine and its major metabolite.

A series of monoclonal antibodies (mAbs) that react with methamphetamine-bovine serum albumin (MA-BSA) were established by intrasplenic immunization method. Among established 36 clones, two typical mAbs, designated NK-1 and NK-2, were described. The inhibition assay of enzyme-linked immunosorbent assay (ELISA) analysis using methamphetamine analogs indicated that NK-1 showed considerable reactivity not only MA-BSA but also methamphetamine and its major metabolite, para-hydroxymethamphetamine (p-hydroxymethamphetamine). The cross-reactivity between NK-1 and the methamphetamine analogs with modified alkyl side chain, indicates that methyl groups of R5 and R7 in the methamphetamine molecules are important for the maximum affinity. The length of alkyl side chain on methamphetamine significantly affected the binding affinity of NK-1. The results may suggest that NK-1 will recognize not only methamphetamine but also the bridge part of the methamphetamine that binds the methamphetamine molecules to a carrier protein.

Pharmacokinetic antagonism of (+)-methamphetamine discrimination by a low-affinity monoclonal anti-methamphetamine antibody.

Animals were trained to discriminate 5 or 10 mg/kg cocaine (rats), or 3 mg/kg (+)-amphetamine (pigeons) from saline, after which dose-response curves were determined for (+)-methamphetamine and other drugs before and after administration of a (+)-methamphetamine-specific monoclonal antibody (K(D) =250 nM). In rats trained to discriminate 10 mg/kg cocaine from saline, intravenous (+)-methamphetamine was about three times more potent as a discriminative stimulus than intraperitoneal (+)-methamphetamine. Also in these rats, intraperitoneal (+)-methamphetamine and (+)-amphetamine were about equipotent as discriminative stimuli, and were about three times more potent than intraperitoneal cocaine. In pigeons trained to discriminate 3 mg/kg intramuscular (i.m.) (+)-amphetamine from saline, (+)-methamphetamine and (+)-amphetamine were nearly equipotent, while cocaine was slightly less potent. In rats trained to discriminate 5 or 10 mg/kg cocaine from saline, intravenous administration of 1 g/kg of the antibody shifted both intravenous and intraperitoneal dose-response curves for (+)-methamphetamine discrimination approximately threefold to the right at 1 or 4 days after administration of the antibody. In pigeons trained to discriminate 3 mg/kg intramuscular (+)-amphetamine from saline, a similar shift of the (+)-methamphetamine dose-response curve to the right also lasted for 4-7 days. However, the antibody did not affect the (+)-amphetamine dose-response curve (pigeons), or the cocaine (rats) dose-response curve. The data show that a low affinity anti-(+)-methamphetamine-specific antibody can produce a specific antagonism of an effect of (+)-methamphetamine that is closely associated with its abuse.

Immunological consequences of methamphetamine protein glycation.

The drug of abuse methamphetamine has been found to participate in the aberrant glycation of proteins. The importance of this chemical process has been shown wherein mouse albumin was readily modified with methamphetamine, and injection of this protein into mice yields a significant immune response, even in the absence of adjuvants. Competition experiments revealed that although methamphetamine binds weakly to the elicited antibodies, the primary epitope is composed of both the methamphetamine moiety and glucose-derived cross-linking region. Implications of this phenonomenon in the context of drug addiction are discussed.