9186 Phys. Chem. Chem. Phys., 2013, 15, 9186--9196 This journal is c the Owner Societies 2013 Cite this: Phys. Chem. Chem. Phys., 2013, 15, 9186 The conformational landscape of tartrate-based inhibitors of the TACE enzyme as revealed by Hamiltonian Replica Exchange simulation† Carlo Guardiani and Piero Procacci* The inhibitors of the Tumor Necrosis Factor-a Converting Enzyme represent promising tools for the treatment of Rheumatoid Arthritis, Multiple Sclerosis and other autoimmune diseases. In this work, using Hamiltonian Replica Exchange Molecular Dynamics simulations and atomistic force field we perform an accurate structural characterization of a group of tartrate-based inhibitors. The simulations highlight a correlation between the conformational landscape in bulk solvent and inhibition potency. Since the structures in bulk solvent are much more compact than the crystallographic bound state, we formulate the hypothesis of a two-step docking mechanism: (i) formation of an intermediate between the compact, hydroxyl exposing conformations in solution and the catalytic zinc ion; (ii) structural rearrangement in the active site of TACE of the zinc-tethered drug in the final binding conformation. Introduction Autoimmune diseases comprise more than 50 distinct diseases and syndromes and affect about 5% of the population in Europe and North America. 1 Examples of autoimmune disorders include Rheumatoid Arthritis (RA), Multiple Sclerosis (MS), juvenile diabetes and Crohn’s disease. Despite the broad array of clinical forms, a remarkable emerging feature of autoimmune diseases is the central role played by the cytokine Tumor Necrosis Factor alpha (TNFa). 2 TNFa is expressed as a homotrimeric 26 kDa membrane protein (mTNFa) that is cleaved by TNFa Converting Enzyme (TACE), a sheddase, belonging to the family of the matrix metalloproteinases (MMPs) that releases a 17 kDa soluble ectodomain of the cytokine (sTNFa). The current strategy for controlling the pro-inflammatory response in autoimmune diseases induced by an excess of the circulating sTNFa is that of selectively inhibiting the enzyme TACE responsible for the cleavage of the soluble ectodomain from the transmembrane precursor mTNFa. The catalytic domain of TACE is characterized by a long binding groove delimited by b-strand IV and helix C. 3 At the center of the groove there is a catalytic zinc ion coordinated by three histidines and by a water molecule. Effective inhibitors must displace the catalytic water molecule to bind the zinc ion and they must comprise a number of groups (P1, P2, P3, P1 0 , P2 0 , P3 0 ) capable of fitting into the binding pockets (S1, S2, S3, S1 0 , S2 0 , S3 0 ) located along the binding groove. As a conse- quence, TACE and MMP inhibitors are comprised of two main units: 4 a peptidomimetic backbone designed to recognize the primed pockets, and a zinc binding group (ZBG). Hydroxamate, as a powerful ZBG, is widely used in TACE synthetic inhibitors but its affinity for zinc is so high that it causes unspecific interactions with MMPs and other metal containing enzymes. This is why a number of ZBGs with affinity lower than that of hydroxamate are currently being tested. 5,6 A novel promising family of TACE inhibitors is represented by the bis-amides of L-tartaric acid first developed by Rosner et al. 7 These compounds feature a tartrate core linking a left hand side (LHS) and a right hand side (RHS) substituent through amide bonds. The tartrate core acts as a tridentate zinc chelator while the RHS (R)-1-(4-(1H-pyrazol-1-yl)phenyl)- ethanamine group fits into the S1 0 and S3 0 pockets. Thanks to the central position of the ZBG, these drugs can target both primed and non-primed pockets, the latter being often neglected by traditional hydroxamate inhibitors. Since the RHS phenyl-pyrazol unit exploits the peculiar tunnel that in TACE connects pockets S1 0 and S3 0 , this group was deemed to be optimal and subsequent research efforts were addressed to optimize the LHS substituent. 8 Department of Chemistry, University of Florence, Italy. E-mail: piero.procacci@unifi.it † Electronic supplementary information (ESI) available: Force field parameters, clustering analysis, effects of chirality on the conformational behavior of drug-6 and drug-39, tartrate inhibitors in DMSO, crystal-like conformations in DMSO, the docking mechanism of the tartrate-based TACE inhibitor. See DOI: 10.1039/ c3cp00108c Received 9th January 2013, Accepted 10th April 2013 DOI: 10.1039/c3cp00108c www.rsc.org/pccp PCCP PAPER Published on 15 April 2013. Downloaded by Universita Studi di Firenze on 11/07/2013 09:44:26. View Article Online View Journal | View Issue