Abstract
Novel Psychoactive Substances (NPS) include several classes of substances such as synthetic cannabinoids (SCBs), an emerging alternative to marijuana, easily purchasable on internet. SCBs are more dangerous than Δ9-Tetrahydrocannabinol as a consequence of their stronger affinities for the CB1 and CB2 receptors, which may result in longer duration of distinct effects, greater potency, and toxicity. The information on SCBs cytotoxicity, genotoxicity, mutagenicity, and long-term effects is scarce. This fact suggests the urgent need to increase available data and to investigate if some SCBs have an impact on the stability of genetic material. Therefore, the aim of the present study was the evaluation of the mutagenic effect of different SCBs belonging to indole- and indazole-structures. The analyzes were conducted in vitro on human TK6 cells and mutagenicity were measured as micronucleus fold increase by flow cytometry. Our results have highlighted, for the first time, the mutagenic capacity of four SCBs, in particular in terms of chromosomal damage induction. We underline the serious potential toxicity of SCBs that suggests the need to proceed with the studies of other different synthetic compounds. Moreover, we identified a method that allows a rapid but effective screening of NPS placed on the market increasingly faster.
Highlights
The production and use of novel psychoactive substances (NPS) are constantly increasing worldwide and, in Europe, the control of psychoactive substances has become a primary concern of governance and citizens
It was necessary to select a suitable mutagenesis test and a proper assay system. Among those validated by the Organization of Economic Cooperation and Development (OECD), we identified the “In Vitro Mammalian Cell Micronucleus Test” as the micronucleus (MN) represents a sensitive biomarker of mutagenicity, in terms of chromosomal damage [33]
For the first time, to highlight the mutagenic capacity of STS-135, APINAC, JWH-018-Cl, and BB-22 and confirmed what was previously demonstrated by Koller et al, first for JWH-018 for 5F-AKB-48
Summary
The production and use of novel psychoactive substances (NPS) are constantly increasing worldwide and, in Europe, the control of psychoactive substances has become a primary concern of governance and citizens. The Guava ViaCount assay showed that all the substances under study do not induce cytotoxicity and cytostasis at all concentrations tested, except for APINAC at 75 μM concentration These results agree with those previously published on tumor cell lines by Koller et al [30,32] and Couceiro et al [39], and the absence of cytotoxicity could be considered a positive result, but in reality, it is not at all from the point of view of genotoxicity. STS-135 stimulated apoptosis of TK6 cells, at all tested concentrations equal to or greater than 25 μM, 5F-AKB-48, APINAC, and JWH-018 only starting from 50 μM concentration, while JWH-018-Cl and BB-22 did not increase the percentage of apoptotic cells, with significant consequences in terms of mutagenicity, because it highlights the cell’s inability to counteract, through this mechanism of selective death, the transmission of genetic damage immediately to the daughter cells. It guarantees a much more objective result, that is not affected by the subjectivity of operator interpretation, as it happens in microscopy, with a considerable reduction in analysis times [34], which allows for a much faster screening of the effects induced by the NPS, increasingly faster placed on the market
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