In relation to the first work package, a detailed atomistic model of Cyclophilin A dynamics was produced via a novel combination of two molecular simulation methodologies, accelerated Molecular Dynamics (aMD) and Markov State Modelling (MSM). The results generated with this algorithm provided a clear rationale for millisecond loop motions in Cyclophilin A that link the major state with a transient state. Such transient state was predicted to be stabilised by a single amino-acid mutation. Thus a Cyclophilin A mutant was prepared to validate the computational prediction, and subjected to a battery of biophysical measurements (Including NMR, X-ray, ITC). Overall the findings validate the transient state computed by the molecular modelling protocol. Along these lines, a preliminary similar model for the related isoform CypD was generated, but delays in production of samples have prevented experimental validation over the timescale of the reporting period. Preliminary results have been presented at several public meetings, and a paper is being written up.
In relation to the second work package, work published since the proposal submission indicated that a number of literature compounds were in fact artefacts. Attention focussed on another class of literature Cyclophilin ligands, but co-workers were unable to establish that these compounds do in fact, bind to Cyclophilins. Thus there was relatively little scope to pursue structure-activity relationship studies, and efforts focussed on the third work package. Instead the fellow has pursued collaborative work with co-workers to validate computational protocols for predictions of binding affinities. This has led to a number of publications.
In relation to the third work package, a combination of computational modelling with experiments carried out by co-workers has led to the discovery of a structurally novel class of Cyclophilin ligands that engage with Cyclophilins in a novel way. These compounds are under evaluation for possible patent filing, and a publication will report our findings in due course.
