Protein-Ligand Useful Simulation for Fragment-Based Drug-Design – PLUS

Docking is one method to obtain structural information about protein-small molecule complexes using the 3D structure of the free protein. Nevertheless, in the context of FBDD, docking results are problematic and ranking of the proposed 3D structures by the docking programme largely differs between the programs. Therefore, we have made a program that is able to rank the docking results according to NMR experimental data. The NMR data correspond to chemical shift perturbations observed on the protein NMR spectrum upon fragment binding. The experimental data are compared to theoretical data (back-calculated data) that are calculated for each docking result. This back-calculation is based on empirical equations. The best docking solutions correspond to the structures that display the best agreement between the experimental and theoretical values. The program can be used as a filter and is compatible with any docking program.

The CSPsim program we have developed has been tested and compared with another structural method, X-Ray crystallography, to demonstrate its efficiency. The program can also evidence protein conformational change events, which occur upon fragment binding. The program developed here is a robust and efficient tool to facilitate the Fragment-Based Drug Design process. A project with PULSALYS, SATT of Lyon St-Étienne, is currently financed to add a graphical interface and add an option to filter docking results according to 1D NMR data.

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The projet PLUS (One partner) resulted in 7 publications including 1 Book chapter about Fragment-Based Drug Design, as well as 10 international Communications and more than 15 national communications with two poster prizes. The program CSPsim is now at the “Agence de Protection des Programmes”.

The growing popularity of Fragment-Based Drug Design (FBDD) shows that this methodology is more and more recognized as a tangible alternative to high throughput screening, and a successful method of hit identification and lead conception. The methodology is also reported as a novel powerful method for Chemical Biology. The FBDD approach consists of identifying low-molecular weight compounds (fragments) that weakly bind to the target protein. These very simple molecules with few chemical functional groups usually display low affinity for the target (high uM to low mM), compared to the bigger, more complex molecules used in classical HTS campaigns. Optimization of the fragment hits, usually by addition of new chemical functions or by linking of two fragment hits binding in adjacent pockets, is strongly driven by structural information. X-Ray crystallography is the key technique for this structure-based drug design process, but its applicability is strongly case dependent, due to the low affinity of the fragments. Docking calculations can also be done but they still suffer from the fact that the scoring functions used to rank the docked ligand positions are not optimized enough for interactions involving small and weak ligands such as fragments. Therefore, new methods including experimental data are required to allow a fast evaluation of the binding site and binding mode of the fragments once they have been detected as ligands in screening experiments. Nuclear Magnetic Resonance (NMR) is one of the most powerful techniques to screen and identity hit fragments. The method also enables the fragment binding site identification, thanks to qualitative interpretation of the Chemical Shift Perturbation (CSP) observed in the protein NMR spectra upon addition of the fragment. Here, we plan to use quantitative CSP to further characterize the binding properties of the fragments and investigate their binding site(s) and their binding mode(s) when bound to the target protein. The originality of the proposal is to analyze the binding properties of very weak affinity ligands (10 mM > KD > 10 uM). The final deliverable will be a software dedicated to the assessment of the fragment positions when bound to the protein target. This proposal aims at providing new important insights into the complexity and versatility of low-affinity protein-ligand interactions, and will have a strong impact both in the field of Chemical Biology and Drug Design.
The coordinator of the proposal has introduced and developed the FBDD approach in the LSA Laboratory. A 250-fragment library was designed and NMR methods are currently used to (1) apply the FBDD techniques to design new active molecules in collaboration with chemists groups and (2) study the fragment-protein interactions in terms of affinity, specificity and binding properties, with 3 articles published in 2010.