Worsening of the Toxic Effects of (±)Cis-4,4′-DMAR Following Its Co-Administration with (±)Trans-4,4′-DMAR: Neuro-Behavioural, Physiological, Immunohistochemical and Metabolic Studies in Mice

Abstract

4,4’-Dimethylaminorex (4,4’-DMAR) is a new synthetic stimulant, and only a little information has been made available so far regarding its pharmaco-toxicological effects. The aim of this study was to investigate the effects of the systemic administration of both the single (±)cis (0.1–60 mg/kg) and (±)trans (30 and 60 mg/kg) stereoisomers and their co-administration (e.g., (±)cis at 1, 10 or 60 mg/kg + (±)trans at 30 mg/kg) in mice. Moreover, we investigated the effect of 4,4′-DMAR on the expression of markers of oxidative/nitrosative stress (8-OHdG, iNOS, NT and NOX2), apoptosis (Smac/DIABLO and NF-κB), and heat shock proteins (HSP27, HSP70, HSP90) in the cerebral cortex. Our study demonstrated that the (±)cis stereoisomer dose-dependently induced psychomotor agitation, sweating, salivation, hyperthermia, stimulated aggression, convulsions and death. Conversely, the (±)trans stereoisomer was ineffective whilst the stereoisomers’ co-administration resulted in a worsening of the toxic (±)cis stereoisomer effects. This trend of responses was confirmed by immunohistochemical analysis on the cortex. Finally, we investigated the potentially toxic effects of stereoisomer co-administration by studying urinary excretion. The excretion study showed that the (±)trans stereoisomer reduced the metabolism of the (±)cis form and increased its amount in the urine, possibly reflecting its increased plasma levels and, therefore, the worsening of its toxicity.

Genotoxicological Characterization of (±)cis-4,4′-DMAR and (±)trans-4,4′-DMAR and Their Association

“…So, the co-treatments demonstrated that the assumption of both racemates aggravates the genotoxic effect. This worsening of 4,4′-DMAR toxicity agrees with an in vivo study that assessed a range of physiological and neuro-behavioural parameters in mice [ 15 ] and highlights another aspect to consider, i.e., that when the drug’s chemical structure presents multiple combinations of enantiomers and diastereomers, it is important to evaluate whether a possible mixture, when consumed, behaves as a single molecule, given the unpredictability of illicit drugs’ composition [ 3 ].…”

“…Finally, for a more in-depth genotoxicological evaluation, the contribution of metabolites should be taken into account, also in consideration of the hypothesis formulated by Tirri et al: “the urinary excretion studies suggested that the worsening of physiological and neuro-behavioural parameters could be related to the inhibition of the metabolism of the (±)cis-4,4′-DMAR form by the (±)trans-4,4′-DMAR“ [ 15 ].…”

“…The (±)cis-4,4′-DMAR and (±)trans-4,4′-DMAR were synthetised and provided by the University of Ferrara Department of Organic Chemistry [ 15 ]. The test compounds were dissolved in absolute ethanol up to 80 mM stock solution and stored at –20 °C.…”

“…More recently, in a mouse in vivo study, the two DMAR stereoisomeric forms also demonstrated opposite toxicity: (±)trans-4,4′-DMAR did not show toxic effects; conversely, (±)cis-4,4′-DMAR exhibited severe levels of toxicity, including salivation, convulsions, hyperthermia and death, attributable to the serotonin syndrome. Critical data obtained following the co-administration of the two DMAR stereoisomers showed a worsening of the toxic effects caused by (±)cis-4,4′-DMAR, resulting in rapid and severe hyperthermia, convulsions and death [ 15 ].…”

Epigenetic Studies for Evaluation of NPS Toxicity: Focus on Synthetic Cannabinoids and Cathinones

“…This narrative review aims to find potential predictive markers of organ damage, considering existing knowledge of NPS toxicology and possibly related epigenetic changes, with a focus on synthetic cathinones and cannabinoids. Our previous study suggested that in vivo synthetic stimulants, such as 4,4′-Dimethylaminorex (4,4′-DMAR), involves physiological, neurobehavioral and neurotoxicological effects, as confirmed by immunohistochemical analysis [ 64 ]. In particular, we investigated the effect of 4,4′-DMAR on the expression of specific cerebral cortex markers (oxidative/nitrosative stress, apoptosis and heat shock proteins) commonly known to be indicative of brain damage [ 65 , 66 ].…”