Closantel is an Allosteric Inhibitor of Human Taspase1

Background: Taspase1 has been identified as an MLL proteolytical processing enzyme that has co-evolved in invertebrates and vertebrates with Trithorax/MLL/KMT2A protein family members. Taspase1 has some very unique features: firstly, a dimerization process triggers an intrinsic serine protease function that leads to a Asp233•Thr234 cleavage and an αββα heterotetrameric structure, while secondly, the N-terminal Threonine residue of the β-subunit is able to catalyze a site-specific hydrolysis at the consensus sequence Q-3X-2D-1•G1X2D3D4, present in MLL family members as well as in the transcription factor TFIIA. However, Taspase1 is not a classical enzyme as it cleaves its few target proteins in a stoichiometric 1:1 fashion. Thus, binding of a single substrate molecule and its hydrolysis occurs after the initial dimerization and auto-activation step. This property makes it nearly impossible to screen for potential inhibitors in a classical screening setup, and consequently entrance points for rational drug design are hard to obtain. Henceforth, a rather sophisticated approach is required to identify potential drugs that inhibit Taspase1. Methods: Here, we report the development of an HTRF reporter assay that allowed the identification of a first Taspase1 inhibitor, Closantel sodium. Findings: This drug has been identified in a library of FDA-approved drugs and binding occurs in a non-covalent fashion. In in vitro experiments the drug attenuated already activated as well as pre-mature Taspase1 with IC50 values in the low µM range. Our data indicate that Closantel sodium also interferes with the dimerization step and/or the intrinsic serine protease function necessary for auto-activation of Taspase1. Interpretation: Taspase1 is required to activate the AF4-MLL oncofusion proteins, and was also found to be overexpressed in several solid tumors. Therefore, this novel inhibitor might be useful for further validation of Taspase1 as a target for cancer therapy and for the design of more potent ligands for future clinical applications. Funding Information: This work has been conducted and performed within the framework of the DFG grants MA 1876/12-1 and MA 1876/13-1. Declaration of Interests: The authors declare that they have no competing interests.