Experimentally guided computational antibody affinity maturation with de novo docking, modelling and rational design

Antibodies are an important class of therapeutics that have significant clinical impact for the treatment of severe diseases. Computational tools to support antibody drug discovery have been developing at an increasing rate over the last decade and typically rely upon a predetermined co-crystal structure of the antibody bound to the antigen for structural predictions. Here, we show an example of successful in silico affinity maturation of a hybridoma derived antibody, AB1, using just a homology model of the antibody fragment variable region and a protein-protein docking model of the AB1 antibody bound to the antigen, murine CCL20 (muCCL20). In silico affinity maturation, together with alanine scanning, has allowed us to fine-tune the protein-protein docking model to subsequently enable the identification of two single-point mutations that increase the affinity of AB1 for muCCL20. To our knowledge, this is one of the first examples of the use of homology modelling and protein docking for affinity maturation and represents an approach that can be widely deployed.

[1]  Á. Zaballos,et al.  NMR Solution Structure of Murine CCL20/MIP-3α, a Chemokine That Specifically Chemoattracts Immature Dendritic Cells and Lymphocytes through Its Highly Specific Interaction with the β-Chemokine Receptor CCR6* , 2001, The Journal of Biological Chemistry.

[2]  Ruhong Zhou,et al.  Replica exchange with solute tempering: efficiency in large scale systems. , 2007, The journal of physical chemistry. B.

[3]  C. Eigenbrot,et al.  High-affinity human antibodies from phage-displayed synthetic Fab libraries with a single framework scaffold. , 2004, Journal of molecular biology.

[4]  V. Dechavanne,et al.  Design of Experiment in CHO and HEK transient transfection condition optimization. , 2011, Protein expression and purification.

[5]  Brian D. Weitzner,et al.  Modeling and docking of antibody structures with Rosetta , 2017, Nature Protocols.

[6]  P. Kollman,et al.  Continuum Solvent Studies of the Stability of DNA, RNA, and Phosphoramidate−DNA Helices , 1998 .

[7]  Peter A. Kollman,et al.  Computational alanine scanning of the 1:1 human growth hormone–receptor complex , 2002, J. Comput. Chem..

[8]  Peter Timmerman,et al.  Affinity maturation of antibodies assisted by in silico modeling , 2008, Proceedings of the National Academy of Sciences.

[9]  B. Roux.  The calculation of the potential of mean force using computer simulations , 1995 .

[10]  Yoonjoo Choi,et al.  FREAD revisited: Accurate loop structure prediction using a database search algorithm , 2010, Proteins.

[11]  Woody Sherman,et al.  Protein and ligand preparation: parameters, protocols, and influence on virtual screening enrichments , 2013, Journal of Computer-Aided Molecular Design.

[12]  Simultaneous humanization and affinity optimization of monoclonal antibodies. , 2003, Methods in molecular biology.

[13]  Gerald M. Edelman,et al.  THE COVALENT STRUCTURE OF AN ENTIRE γG IMMUNOGLOBULIN MOLECULE , 1969 .

[14]  Tingjun Hou,et al.  Assessing the Performance of the MM/PBSA and MM/GBSA Methods. 1. The Accuracy of Binding Free Energy Calculations Based on Molecular Dynamics Simulations , 2011, J. Chem. Inf. Model..

[15]  Neelesh Soni,et al.  In silico methods for design of biological therapeutics. , 2017, Methods.

[16]  Alexander D. MacKerell,et al.  CHARMM general force field: A force field for drug‐like molecules compatible with the CHARMM all‐atom additive biological force fields , 2009, J. Comput. Chem..

[17]  E. Purisima,et al.  Assisted Design of Antibody and Protein Therapeutics (ADAPT) , 2017, PloS one.

[18]  Li Li,et al.  RDOCK: Refinement of rigid‐body protein docking predictions , 2003, Proteins.

[19]  Sebastian Kelm,et al.  SAbPred: a structure-based antibody prediction server , 2016, Nucleic Acids Res..

[20]  Chris Maloney,et al.  PubMed Central , 2017 .

[21]  P A Kollman,et al.  Free energy calculations on dimer stability of the HIV protease using molecular dynamics and a continuum solvent model. , 2000, Journal of molecular biology.

[22]  M. DeLisa,et al.  Computational affinity maturation of camelid single-domain intrabodies against the nonamyloid component of alpha-synuclein , 2018, Scientific Reports.

[23]  Jan H. Jensen,et al.  PROPKA3: Consistent Treatment of Internal and Surface Residues in Empirical pKa Predictions. , 2011, Journal of chemical theory and computation.

[24]  E. Purisima,et al.  Application of Assisted Design of Antibody and Protein Therapeutics (ADAPT) improves efficacy of a Clostridium difficile toxin A single-domain antibody , 2018, Scientific Reports.

[25]  Jennifer L. Knight,et al.  OPLS3: A Force Field Providing Broad Coverage of Drug-like Small Molecules and Proteins. , 2016, Journal of chemical theory and computation.

[26]  A. Stojadinovic,et al.  Affinity maturation of monoclonal antibodies by multi-site-directed mutagenesis. , 2014, Methods in molecular biology.

[27]  David L Mobley,et al.  Alchemical free energy methods for drug discovery: progress and challenges. , 2011, Current opinion in structural biology.

[28]  G. Edelman,et al.  The covalent structure of an entire gammaG immunoglobulin molecule. , 1969, Proceedings of the National Academy of Sciences of the United States of America.

[29]  Jérôme Lane,et al.  IMGT®, the international ImMunoGeneTics information system® , 2004, Nucleic Acids Res..

[30]  K. Takano.  ON SOLUTION OF , 1983 .

[31]  B. Nelson,et al.  Structure-Guided Combinatorial Engineering Facilitates Affinity and Specificity Optimization of Anti-CD81 Antibodies. , 2018, Journal of molecular biology.

[32]  E. Kabat,et al.  Sequences of proteins of immunological interest , 1991 .

[33]  Marc F Lensink,et al.  Docking, scoring, and affinity prediction in CAPRI , 2013, Proteins.

[34]  Youyong Li,et al.  Assessing the performance of MM/PBSA and MM/GBSA methods. 4. Accuracies of MM/PBSA and MM/GBSA methodologies evaluated by various simulation protocols using PDBbind data set. , 2014, Physical chemistry chemical physics : PCCP.

[35]  Fenglou Mao,et al.  Potential of mean force for protein–protein interaction studies , 2002, Proteins.

[36]  F. M. Ytreberg,et al.  Fast Calculation of Protein–Protein Binding Free Energies Using Umbrella Sampling with a Coarse-Grained Model , 2017, Journal of chemical theory and computation.

[37]  Jiye Shi,et al.  SAbDab: the structural antibody database , 2013, Nucleic Acids Res..

[38]  Robert Abel,et al.  Free Energy Perturbation Calculations of the Thermodynamics of Protein Side-Chain Mutations. , 2017, Journal of molecular biology.

[39]  Christophe Chipot,et al.  Calculating Free Energy Differences Using Perturbation Theory , 2007 .

[40]  P. Kollman,et al.  Calculating structures and free energies of complex molecules: combining molecular mechanics and continuum models. , 2000, Accounts of chemical research.

[41]  Jiye Shi,et al.  ABodyBuilder: Automated antibody structure prediction with data–driven accuracy estimation , 2016, mAbs.

[42]  Kouhei Tsumoto,et al.  Affinity Improvement of a Therapeutic Antibody by Structure-Based Computational Design: Generation of Electrostatic Interactions in the Transition State Stabilizes the Antibody-Antigen Complex , 2014, PloS one.

[43]  Bruce Tidor,et al.  Computational design of antibody-affinity improvement beyond in vivo maturation , 2007, Nature Biotechnology.

[44]  W. L. Jorgensen.  The Many Roles of Computation in Drug Discovery , 2004, Science.

[45]  Bruce Tidor,et al.  Progress in computational protein design. , 2007, Current opinion in biotechnology.

[46]  P. Chames,et al.  Improving the affinity and the fine specificity of an anti-cortisol antibody by parsimonious mutagenesis and phage display. , 1998, Journal of immunology.

[47]  Jan H. Jensen,et al.  Very fast empirical prediction and rationalization of protein pKa values , 2005, Proteins.

[48]  P. Kollman,et al.  Computational Alanine Scanning To Probe Protein−Protein Interactions: A Novel Approach To Evaluate Binding Free Energies , 1999 .

[49]  Christophe Chipot,et al.  Efficient determination of protein-protein standard binding free energies from first principles. , 2013, Journal of chemical theory and computation.

[50]  Adam Godzik,et al.  Flexible structure alignment by chaining aligned fragment pairs allowing twists , 2003, ECCB.

[51]  K D Wittrup,et al.  Directed evolution of antibody fragments with monovalent femtomolar antigen-binding affinity. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[52]  B. Berne,et al.  Replica exchange with solute scaling: a more efficient version of replica exchange with solute tempering (REST2). , 2011, The journal of physical chemistry. B.

[53]  Woody Sherman,et al.  Affinity enhancement of an in vivo matured therapeutic antibody using structure‐based computational design , 2006, Protein science : a publication of the Protein Society.

[54]  B. Berne,et al.  Replica exchange with solute tempering: a method for sampling biological systems in explicit water. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[55]  M. Karplus,et al.  Effective energy function for proteins in solution , 1999, Proteins.

[56]  J. Debreczeni,et al.  Structure and characterization of a high affinity C5a monoclonal antibody that blocks binding to C5aR1 and C5aR2 receptors , 2017, mAbs.