Derivatization of Microcystins Can Increase Target Inhibition while Reducing Cellular Uptake

Microcystins, a large family of nonribosomal cyclic heptapeptides known for their hepatotoxicity, are among the best-studied cyanobacterial toxins. Recently, they have been discussed as leads for the development of anticancer drug substances. Their main mode-of-action is inhibition of the eukaryotic serine/threonine protein phosphatases 1 and 2A. Unlike many cytotoxins that can cross cell membranes by passive diffusion, microcystins depend on active uptake via organic anion transporting polypeptides 1B1 or 1B3. Both phosphatase inhibition and transportability strongly depend on the structure of the individual microcystin. Here, we present how chemical modification of positions 2 and 4 of the microcystin core structure can alter these two properties. Aiming to reduce transportability and increase phosphatase inhibition, we used pharmacophore modeling to investigate the phosphatase inhibition potential of microcystins derivatized with small molecules containing a variety of functional groups. The respective derivatives were synthesized using click chemistry. We discovered that some derivatized microcystins can address a yet undescribed subpocket of the protein phosphatase 1. The derivatized microcystins were tested for phosphatase 1 inhibition and cytotoxicity on transporter-expressing cell lines, revealing that target inhibition and transportability of microcystins can independently be influenced by the physicochemical properties, especially of the residue located in position 2 of the microcystin. Derivatization with small acids or amino acids resulted in microcystins with a favorable ratio of inhibition to transportability, making these derivatives potentially suitable for drug development.