Dynamics of ligand-protein interactions - impact on drug discovery

Introducing a new drug to market is a lengthy and expensive process (typically 10-15 years and $1.7 billion). Better understanding of how and why a drug molecule binds to a target and what changes in the atomistic structure and chemistry could improve the binding affinity and shorten the process. In addition to structure-based approaches, the role of thermodynamics and molecular motions in binding selectivity and efficiency have attracted increasing attention. Whilst calorimetric methods can quantify total free energy and entropy change, it is difficult to estimate contributions from the different components of entropy, one of the largest unknowns being the magnitude of the configurational entropy. Molecular dynamics (MD) simulations of the drug and target protein can provide more details of the different atomistic movements contributing to the total entropy change, thus potentially providing valuable clues for lead optimisation. In this study we use the well characterised N-terminal domain of the Hsp90 chaperone protein as a model system to study the changes in conformational flexibility (configurational entropy) upon binding of small molecule inhibitors using MD simulations, NMR and ITC. We show that the two inhibitors studied cause different changes in the protein dynamics. These effects were seen with NMR relaxation dispersion methods and with MD but the dynamic changes however are not reflected in the global ITC parameters. Here the water is assumed to have a dominating effect in the overall entropy change. However, as some Hsp90 clients have been shown to preferentially interact with only one conformation of the protein, we propose that the changes seen with NMR and MD could be of interest for drug design. Manipulating the dynamics by small molecules could favour interaction with a subset of client proteins, without affecting the interaction of others, all together providing specificity and potentially allowing to design an ‘ideal’ drug that only prevents the folding of ‘bad’ cancer related proteins without affecting Hsp90 functions in the normal cells. As the MD simulations also reflect these dynamic changes, we propose that simulations could be also used as a screening tool for selecting which inhibitors could be taken for further development in the lab

Guideline on the requirements of external quality assessment programs in molecular pathology

<p>Molecular pathology is an integral part of daily diagnostic pathology and used for classification of tumors, for prediction of prognosis and response to therapy, and to support treatment decisions. For these reasons, analyses in molecular pathology must be highly reliable and hence external quality assessment (EQA) programs are called for. Several EQA programs exist to which laboratories can subscribe, but they vary in scope, number of subscribers, and execution. The guideline presented in this paper has been developed with the purpose to harmonize EQA in molecular pathology. It presents recommendations on how an EQA program should be organized, provides criteria for a reference laboratory, proposes requirements for EQA test samples, and defines the number of samples needed for an EQA program. Furthermore, a system for scoring of the results is proposed as well as measures to be taken for poorly performing laboratories. Proposals are made regarding the content requirements of an EQA report and how its results should be communicated. Finally, the need for an EQA database and a participant manual are elaborated. It is the intention of this guideline to improve EQA for molecular pathology in order to provide more reliable molecular analyses as well as optimal information regarding patient selection for treatment.</p>

EMQN best practice guidelines for the molecular genetic diagnosis of hereditary hemochromatosis (HH)

International audienceMolecular genetic testing for hereditary hemochromatosis (HH) is recognized as a reference test to confirm the diagnosis of suspected HH or to predict its risk. The vast majority (typically >90%) of patients with clinically characterized HH are homozygous for the p.C282Y variant in the HFE gene, referred to as HFE-related HH. Since 1996, HFE genotyping was implemented in diagnostic algorithms for suspected HH, allowing its early diagnosis and prevention. However, the penetrance of disease in p.C282Y homozygotes is incomplete. Hence, homozygosity for p.C282Y is not sufficient to diagnose HH. Neither is p.C282Y homozygosity required for diagnosis as other rare forms of HH exist, generally referred to as non-HFE-related HH. These pose significant challenges when defining criteria for referral, testing protocols, interpretation of test results and reporting practices. We present best practice guidelines for the molecular genetic diagnosis of HH where recommendations are classified, as far as possible, according to the level and strength of evidence. For clarification, the guidelines' recommendations are preceded by a detailed description of the methodology and results obtained with a series of actions taken in order to achieve a wide expert consensus, namely: (i) a survey on the current practices followed by laboratories offering molecular diagnosis of HH; (ii) a systematic literature search focused on some identified controversial topics; (iii) an expert Best Practice Workshop convened to achieve consensus on the practical recommendations included in the guidelines