Anders Johansson, Research and Early Development, Cardiovascular, Renal and Metabolism, Biopharmaceuticals R&D, AstraZeneca, Gothenburg, Sweden

Oxetane-containing ring systems have shown great utility in medicinal chemistry to modulate physicochemical properties. There is an increasing number of examples of how incorporation of oxetanes can have a beneficial impact on the pharmacokinetic (PK) properties of compounds. A less explored aspect is how oxetanes affect metabolic pathways. An exaggerated dependence on one specific isoenzyme increases the risk of drug-drug interactions with co-administered drugs. It is not unusual to observe strong dependence of one isoform of cytochrome P450, most notably CYP3A4. This talk will illustrate that mEH-catalyzed (microsomal epoxide hydrolase) hydrolysis is an important metabolic pathway for structurally diverse oxetanes. To aid rationalize substrate binding, a homology model based on the Bombyx mori EH crystal structure was used. The introduction of an oxetane into a lead series could potentially decrease the dependence on cytochromes P450. However, human mEH biology is less studied and prediction of human PK properties could be less reliable if a large proportion of the metabolism of a clinical candidate proceeds via mEH. An early assessment of mEH metabolism to understand the complete metabolic fate of oxetane-containing compounds will be of great benefit to the design process.

In lead optimization programs, drug designers are often faced with questions that have conformational equilibria at their core: Is the ligand bound in, or close to, its minimum-energy solution conformation? Can binding be improved by conformational restriction, preorganizing the compound in a suitable conformation? Can the compound adopt a different set of conformations in the plasma membrane, shielding polar moieties and facilitating permeability?

To answer these questions in a routine fashion, we have developed the ReSCoSS workflow (Relevant Solution Conformer Sampling and Selection). ReSCoSS is a complete workstream, from drawing a molecule in 2D, 3D conversion, exhaustive conformational sampling, clustering and selection to ab initio minimization. It is designed to deliver a selection of the most relevant minimum-energy conformers in a range of polar and nonpolar solvents, which can then be of use in drug design applications as well as for the prediction of physicochemical properties such as logP and pKa. I will introduce the design principle of ReSCoSS and discuss how we have assessed the performance of the workflow using experimental data.

GPCRs play a pivotal role in transmitting signals at the cellular level and structural insights can be exploited to support structure-based drug discovery endeavors. Despite advances in GPCR crystallography, it remains challenging to obtain crystal structures particularly of active states. Molecular dynamics (MD) simulations have been used to explore the conformational landscape of GPCRs. Nevertheless, the search for physiologically relevant conformations, allosteric pockets and ligand binding events using classical MD simulations is still impractical.

The work I will present shows how by applying adaptive MD it is possible to computationally access, in a rigorous and timely manner, key functional states of a GPCR without a priori structural information of the active state [1]. Moreover, I will show that this cutting-edge MD methodology can help elucidating GPCRs regulatory mechanisms and opening avenues for the study of ligands binding to elusive yet pharmacologically important GPCR states, providing key insights into ligands mode of action [2].

[1] S. Lovera, A. Cuzzolin, S. Kelm, G. De Fabritiis, Z. A. Sands. Reconstruction of apo A2A receptor activation pathways reveal ligand-competent intermediates and state-dependent cholesterol hotspots. Sci Rep 9, 14199 (2019)

[2] S. Lovera, E.J.B. Landin, G. De Fabritiis, S. Kelm, J. Mercier, D. McMillan, R.B. Sessions, R.J. Taylor, Z. A. Sands, L. Joedicke, M.P. Crump. The Aminotriazole Antagonist Cmpd-1 Stabilises a Distinct Inactive State of the Adenosine 2A Receptor. Angewandte Chemie Int. Ed. 58, 1-6 (2019)

Proteins present particular challenges for property calculations due to their conformational flexibility in solution and the sensitivity of the structure to environmental parameters such as buffer strength and pH. We present a novel method for calculating thermodynamically averaged properties using a conformational ensemble which correctly takes into account the variability of both the structure and the charge state (protonation) of the protein.

The validity of this approach will be demonstrated using various benchmark calculations with experimental reference data, and additional applications will be shown with an emphasis toward modeling developability criteria for therapeutic antibodies.

First-generation gene therapies for Haemophilia A exhibited large patient-to-patient variations in FVIII levels and a drop of expression over time that may at least in part be caused by the large size of the FVIII expression cassette, resulting in vector genome sizes exceeding the wild type AAV genome.

Here we present the design and preliminary characterization of FLT210, a next generation AAV-FVIII cassette, which we believe possesses the attributes required to become the best-in-class vector to treat Haemophilia A patients. It includes the optimisation to the ‘FVIII-SQ’ protein and coding sequence by in-silico design and the enhancement of the level of FVIII expression and secretion by de-novo design of the regulatory regions.

Recombinant expression of botulinum neurotoxins enables protein engineering to exploit the modularity of these proteins, with potential for an increased range of indications. Re-targeting of these molecules by domain replacement yields Targeted Secretion Inhibitors (TSIs), with differentiated cell targeting, further broadening therapeutic potential.

Molecular modelling approaches are being used to support and enhance early stage discovery processes. Detailed analysis of protein interactions, linker options and general architecture allow prioritisation of molecules moving into experimental production. Coupled with increasing throughput of molecules for screening, these strategies extend the early exploration of design space and support selection of candidates with optimal function and improved developability.