Kinetics differences among microcystins variants : any relationship with toxicity?

Microcystins (MC) toxicity is congener-specific; however, the in vitro inhibition of PP1/PP2A (the key molecular event of MC toxicity) by single MC variants is comparable. Consequently, MC toxicokinetics, including both transporter-mediated uptake and biotransformation reaction, seems to be the critical point to explain the variant-specific toxic potential. Glutathione conjugation, either occurring spontaneously or catalyzed by GST, is the accepted main step in MC detoxification. We characterized the in vitro human conjugation of different variants (MC-LR, MC-RR, MC-YR, MC-LW and MC-LF), characterized by different structures and lipophilicity, the latter experimentally determined in the same conditions using the OECD guideline n° 117. The ranking from the most to the least lipophilic is : MC-LF>MC-LW>MC-LR>MC-YR>MCRR. Using single human hepatic recombinant isoforms (GSTA1, A2, A4, M1, T1 T2, P1, and O1) and human liver cytosol (HLC, pool of 200 donors) the kinetic parameters Vmax, Km and Cli were calculated for MC-LR, MCRR, MC-YR, and MC-LW. All tested recombinant GSTs were active in conjugating MCs, with comparable catalytic efficiency although the ranking was different, as well as the kinetic behavior. The spontaneous reaction was generally predominant when compared to enzymatic reaction. The reaction between MC-LF and GSH (both spontaneous and enzymatic) was very limited and no kinetic parameter could be calculated. This variant is therefore poorly detoxified in the human liver and further investigations are recommended on its toxicity. The variant which is most efficiently detoxified (Cli values about 2-3 folds higher) is MC-RR, which is the least acutely toxic. In addition, with MC-RR and only to a lesser extent for the other in the presence of GSH depletion, such as the one occurring being co-exposed to paracetamol or other GSH-depleting xenobiotics, the enzymatic reaction was by far predominant, giving relevance to the possible interindividual differences due to the activity of polymorphic GST isoforms. Acknowledgements: This study was partially supported by: the European Food Safety Authority (EFSA) under the grant agreement no.GA/EFSA/SCER/2015/01.