Fragment‐Based Lead Discovery: Screening and Optimizing Fragments for Thermolysin Inhibition

Fragment‐based drug discovery has gained a foothold in today's lead identification processes. We present the application of in silico fragment‐based screening for the discovery of novel lead compounds for the metalloendoproteinase thermolysin. We have chosen thermolysin to validate our screening approach as it is a well‐studied enzyme and serves as a model system for other proteases. A protein‐targeted virtual library was designed and screening was carried out using the program AutoDock. Two fragment hits could be identified. For one of them, the crystal structure in complex with thermolysin is presented. This compound was selected for structure‐based optimization of binding affinity and improvement of ligand efficiency, while concomitantly keeping the fragment‐like properties of the initial hit. Redesigning the zinc coordination group revealed a novel class of fragments possessing Ki values as low as 128 μM, thus they provide a good starting point for further hit evolution in a tailored lead design.

[1]  G. Klebe,et al.  More than a simple lipophilic contact: a detailed thermodynamic analysis of nonbasic residues in the s1 pocket of thrombin. , 2009, Journal of molecular biology.

[2]  Gerhard Klebe,et al.  SFCscore: Scoring functions for affinity prediction of protein–ligand complexes , 2008, Proteins.

[3]  Paul G Wyatt,et al.  Identification of N-(4-piperidinyl)-4-(2,6-dichlorobenzoylamino)-1H-pyrazole-3-carboxamide (AT7519), a novel cyclin dependent kinase inhibitor using fragment-based X-ray crystallography and structure based drug design. , 2008, Journal of medicinal chemistry.

[4]  Susanne Nyström,et al.  Identification of MMP-12 inhibitors by using biosensor-based screening of a fragment library. , 2008, Journal of medicinal chemistry.

[5]  Jan Schultz,et al.  Integration of fragment screening and library design. , 2007, Drug discovery today.

[6]  J. R. Pliego,et al.  On the mechanism of the reaction between aryl acetates and hydroxylamine , 2007 .

[7]  T Neumann,et al.  SPR-based fragment screening: advantages and applications. , 2007, Current topics in medicinal chemistry.

[8]  S. Homans,et al.  Water, water everywhere--except where it matters? , 2007, Drug discovery today.

[9]  P. Hajduk,et al.  A decade of fragment-based drug design: strategic advances and lessons learned , 2007, Nature Reviews Drug Discovery.

[10]  H. Fuchsbauer,et al.  A quenched fluorescent dipeptide for assaying dispase- and thermolysin-like proteases. , 2006, Analytical biochemistry.

[11]  Christian Rummey,et al.  In silico fragment-based discovery of DPP-IV S1 pocket binders. , 2006, Bioorganic & medicinal chemistry letters.

[12]  Gianni Chessari,et al.  Application of fragment screening and fragment linking to the discovery of novel thrombin inhibitors. , 2006, Journal of medicinal chemistry.

[13]  Simona Golic Grdadolnik,et al.  In silico fragment-based discovery of indolin-2-one analogues as potent DNA gyrase inhibitors. , 2005, Bioorganic & medicinal chemistry letters.

[14]  G. Klebe,et al.  DrugScore(CSD)-knowledge-based scoring function derived from small molecule crystal data with superior recognition rate of near-native ligand poses and better affinity prediction. , 2005, Journal of medicinal chemistry.

[15]  John J Irwin,et al.  Virtual screening against metalloenzymes for inhibitors and substrates. , 2005, Biochemistry.

[16]  Christopher W Murray,et al.  Fragment-based lead discovery: leads by design. , 2005, Drug discovery today.

[17]  Christopher W Murray,et al.  Fragment-based lead discovery using X-ray crystallography. , 2005, Journal of medicinal chemistry.

[18]  Kevin Cowtan,et al.  research papers Acta Crystallographica Section D Biological , 2005 .

[19]  Jeffrey W. Peng,et al.  Theory and applications of NMR-based screening in pharmaceutical research. , 2004, Chemical reviews.

[20]  A. Hopkins,et al.  Ligand efficiency: a useful metric for lead selection. , 2004, Drug discovery today.

[21]  M. McCoy,et al.  Non-peptidic small-molecule inhibitors of the single-chain hepatitis C virus NS3 protease/NS4A cofactor complex discovered by structure-based NMR screening. , 2004, Journal of medicinal chemistry.

[22]  Richard D. Taylor,et al.  Improved protein–ligand docking using GOLD , 2003, Proteins.

[23]  Neera Borkakoti,et al.  Discovery of novel low molecular weight inhibitors of IMPDH via virtual needle screening. , 2003, Bioorganic & medicinal chemistry letters.

[24]  Renxiao Wang,et al.  Comparative evaluation of 11 scoring functions for molecular docking. , 2003, Journal of medicinal chemistry.

[25]  Gerhard Klebe,et al.  Relibase: design and development of a database for comprehensive analysis of protein-ligand interactions. , 2003, Journal of molecular biology.

[26]  Gerhard Klebe,et al.  Utilising structural knowledge in drug design strategies: applications using Relibase. , 2003, Journal of molecular biology.

[27]  David S. Goodsell,et al.  Automated docking using a Lamarckian genetic algorithm and an empirical binding free energy function , 1998, J. Comput. Chem..

[28]  R J Read,et al.  Crystallography & NMR system: A new software suite for macromolecular structure determination. , 1998, Acta crystallographica. Section D, Biological crystallography.

[29]  Michael M. Hann,et al.  RECAP-Retrosynthetic Combinatorial Analysis Procedure: A Powerful New Technique for Identifying Privileged Molecular Fragments with Useful Applications in Combinatorial Chemistry , 1998, J. Chem. Inf. Comput. Sci..

[30]  P Willett,et al.  Development and validation of a genetic algorithm for flexible docking. , 1997, Journal of molecular biology.

[31]  Thomas Lengauer,et al.  A fast flexible docking method using an incremental construction algorithm. , 1996, Journal of molecular biology.

[32]  P. Reddy,et al.  Unexpected Formation of Cyclic Hydroxamic Acids from O-(2-Aminobenzoyl) Hydroxylamine , 1996 .

[33]  M. Browner,et al.  Matrilysin-inhibitor complexes: common themes among metalloproteases. , 1996, Biochemistry.

[34]  M. Browner,et al.  Crystal structures of matrilysin-inhibitor complexes , 1995 .

[35]  S. Sheriff.  Some Methods for Examining the Interactions between Two Molecules , 1993 .

[36]  Brian W. Matthews,et al.  Structural basis of the action of thermolysin and related zinc peptidases , 1988 .

[37]  K. Tanaka,et al.  Syntheses of cyclic hydroxamic acid derivatives, and their chelating abilities and biological activities. , 1988, Chemical & pharmaceutical bulletin.

[38]  J. L. Smith,et al.  Structure of myohemerythrin in the azidomet state at 1.7/1.3 A resolution. , 1987, Journal of molecular biology.

[39]  B. Matthews,et al.  Binding of N-carboxymethyl dipeptide inhibitors to thermolysin determined by X-ray crystallography: a novel class of transition-state analogues for zinc peptidases. , 1984, Biochemistry.

[40]  U. Singh,et al.  A NEW FORCE FIELD FOR MOLECULAR MECHANICAL SIMULATION OF NUCLEIC ACIDS AND PROTEINS , 1984 .

[41]  B. Matthews,et al.  Structure of thermolysin refined at 1.6 A resolution. , 1982, Journal of molecular biology.

[42]  B. Matthews,et al.  Binding of hydroxamic acid inhibitors to crystalline thermolysin suggests a pentacoordinate zinc intermediate in catalysis. , 1982, Biochemistry.

[43]  B. Matthews.  The γ Turn. Evidence for a New Folded Conformation in Proteins , 1972 .

[44]  H. Drucker,et al.  The role of calcium in thermolysin: effect on kinetic properties and autodigestion. , 1971, Archives of biochemistry and biophysics.

[45]  B. Vallee,et al.  Thermolysin: a zinc metalloenzyme. , 1969, Biochemical and biophysical research communications.

[46]  T. Creighton.  Methods in Enzymology , 1968, The Yale Journal of Biology and Medicine.

[47]  K. Morihara.  The specificities of various neutral and alkaline proteinases from microorganisms. , 1967, Biochemical and biophysical research communications.

[48]  L. M. Lima,et al.  Bioisosterism: a useful strategy for molecular modification and drug design. , 2005, Current medicinal chemistry.

[49]  Brian K. Shoichet,et al.  ZINC - A Free Database of Commercially Available Compounds for Virtual Screening , 2005, J. Chem. Inf. Model..

[50]  J. Gasteiger,et al.  Automatic generation of 3D-atomic coordinates for organic molecules , 1990 .

[51]  J. Gasteiger,et al.  ITERATIVE PARTIAL EQUALIZATION OF ORBITAL ELECTRONEGATIVITY – A RAPID ACCESS TO ATOMIC CHARGES , 1980 .

[52]  N. Heindel,et al.  A unique rearrangement of 3,4-dihydro-5H-1,3,4-benzotriazepin-5-ones to 3-methylamino-4(3H)-quinazolinones , 1977 .