Strategies that target multiple components are usually required for treatment of diseases originating from complex biological systems. The multicomponent system consisting of the DR4 major histocompatibility complex type II molecule, the glycopeptide CI1259-273 from type II collagen, and a T-cell receptor is associated with development of rheumatoid arthritis (RA). We introduced non-native amino acids and amide bond isosteres into CI1259-273 and investigated the effect on binding to DR4 and the subsequent T-cell response. Molecular dynamics simulations revealed that complexes between DR4 and derivatives of CI1259-273 were highly dynamic. Signaling in the overall multicomponent system was found to depend on formation of an appropriate number of dynamic intramolecular hydrogen bonds between DR4 and CI1259-273, together with the positioning of the galactose moiety of CI1259-273 in the DR4 binding groove. Interestingly, the system tolerated modifications at several positions in CI1259-273, indicating opportunities to use analogues to increase our understanding of how rheumatoid arthritis develops and for evaluation as vaccines to treat RA.

Resistance development in insects significantly threatens the important benefits obtained by insecticide usage in vector control of disease-transmitting insects. Discovery of new chemical entities with insecticidal activity is highly desired in order to develop new insecticide candidates. Here, we present the design, synthesis, and biological evaluation of phenoxyacetamide-based inhibitors of the essential enzyme acetylcholinesterase 1 (AChE1). AChE1 is a validated insecticide target to control mosquito vectors of, e.g., malaria, dengue, and Zika virus infections. The inhibitors combine a mosquito versus human AChE selectivity with a high potency also for the resistance-conferring mutation G122S; two properties that have proven challenging to combine in a single compound. Structure activity relationship analyses and molecular dynamics simulations of inhibitor protein complexes have provided insights that elucidate the molecular basis for these properties. We also show that the inhibitors demonstrate in vivo insecticidal activity on disease-transmitting mosquitoes. Our findings support the concept of noncovalent, selective, and resistance-breaking inhibitors of AChE1 as a promising approach for future insecticide development.

The field of medicinal chemistry is constantly evolving. Aided by advances within techniques as well as knowledge of biological systems, increasingly complex targets and drugs can be considered. This thesis includes two projects focusing on the design of drugs targeting multicomponent systems, referring to systems for which multiple components must be considered during the drug design process.

In the first project, the long-term goal is to develop a vaccine against the autoimmune disease rheumatoid arthritis (RA). The cause of RA is unknown, but it is genetically linked to expression of class II MHC proteins that present antigens to T-cell receptors (TCRs), responsible for initiating an immune response. A glycopeptide fragment, CII259–273, from type II collagen has shown promising results as a vaccine against arthritis resembling RA in mice. CII259–273 binds to the class II MHC protein followed by presentation to the TCR, forming a multicomponent system.

We have used molecular dynamics (MD) simulations to study the effect that modifications of CII259–273 have on the multicomponent system. Non-native amino acids and amide bond isosteres have been introduced. This has demonstrated the importance of retaining the backbone conformation of CII259–273, as well as the hydrogen bonds formed to the backbone. The ability to introduce such modifications would be of value to affect the potency towards the MHC protein, and prevent degradation of the glycopeptide. The studies have revealed a multicomponent system that is highly sensitive to even small modifications that can affect the dynamics of the entire complex.

In the second project, the long-term goal is to develop a broad-spectrum antidote against nerve agents. Nerve agents are extremely toxic compounds that act by covalently inhibiting the enzyme acetylcholinesterase (AChE), which is essential for termination of nerve signalling. A major limitation of current antidotes is that their efficiency is dependent on the type of nerve agent. A broad-spectrum antidote must be able to bind to the multicomponent system consisting of AChE covalently inhibited by different nerve agents. It will then act by performing a nucleophilic attack on the nerve agent adduct, thus breaking the covalent bond to AChE.

We have used statistical molecular design (SMD) and quantitative structure-activity relationship (QSAR) modelling to identify a fragment with a potency for AChE inhibited by different nerve agents. A nucleophilic component able to restore the enzyme to the active form was thereafter introduced. This resulted in a functional reactivator, efficient for multiple nerve agents. Furthermore, the mechanism of reactivation has been investigated through structural studies, enabled by a combination of X-ray crystallography and molecular modelling. A high flexibility of the reactivator, as well as the ability to bind to AChE in multiple conformations, are defined as important properties for a broad-spectrum antidote.

Scientific disciplines such as medicinal- and environmental chemistry, pharmacology, and toxicology deal with the questions related to the effects small organic compounds exhort on biological targets and the compounds' physicochemical properties responsible for these effects. A common strategy in this endeavor is to establish structure-activity relationships (SARs). The aim of this work was to illustrate benefits of performing a statistical molecular design (SMD) and proper statistical analysis of the molecules' properties before SAR and quantitative structure-activity relationship (QSAR) analysis. Our SMD followed by synthesis yielded a set of inhibitors of the enzyme acetylcholinesterase (AChE) that had very few inherent dependencies between the substructures in the molecules. If such dependencies exist, they cause severe errors in SAR interpretation and predictions by QSAR-models, and leave a set of molecules less suitable for future decision-making. In our study, SAR- and QSAR models could show which molecular sub-structures and physicochemical features that were advantageous for the AChE inhibition. Finally, the QSAR model was used for the prediction of the inhibition of AChE by an external prediction set of molecules. The accuracy of these predictions was asserted by statistical significance tests and by comparisons to simple but relevant reference models.

Class II major histocompatibility complex (MHC) proteins are involved in initiation of immune responses to foreign antigens via presentation of peptides to receptors of CD4(+) T-cells. An analogous presentation of self-peptides may lead to autoimmune diseases, such as rheumatoid arthritis (RA). The glycopeptide fragment CII259-273, derived from type II collagen, is presented by A(q) MHCII molecules in the mouse and has a key role in development of collagen induced arthritis (CIA), a validated model for RA. We have introduced hydroxyethylene amide bond isosteres at the Ala(261)-Gly(262) position of CII259-273. Biological evaluation showed that A(q) binding and T cell recognition were dramatically reduced for the modified glycopeptides, although static models predicted similar binding modes as the native type II collagen fragment. Molecular dynamics (MD) simulations demonstrated that introduction of the hydroxyethylene isosteres disturbed the entire hydrogen bond network between the glycopeptides and A(q). As a consequence the hydroxyethylene isosteric glycopeptides were prone to dissociation from A(q) and unfolding of the beta(1)-helix. Thus, the isostere induced adjustment of the hydrogen bond network altered the structure and dynamics of A(q)/glycopeptide complexes leading to the loss of A(q) affinity and subsequent T cell response.