Synthesis and Evaluation of Covalent 14-3-3σ Protein-Protein Interaction Stabilisers

The focus on protein-protein interactions (PPIs) for drug development has grown by targeting them with small molecules to treat diseases. Initially, inhibiting PPIs had greater success. Another approach involves stabilizing PPIs using small molecules which is often overlooked due to limited understanding of mechanisms. A compelling case study is 14-3-3 protein complexes, controlling diverse cellular functions. In humans, 7 isoforms (β, γ, σ, δ, η, ε, τ) exist. In interest is the σ isoform, with its exclusive cysteine residue (Cys38) in the binding interface, this unique feature makes it a prime focus for covalent protein modification 1.

Oestrogen receptor alpha (ERα) is a transcription factor driving 75% of breast cancers. ERα activation leads to breast cancer cell proliferation. Binding to 14-3-3 proteins prevent ERα dimerization2. Thus, stabilizing the 14-3-3/ERα PPI is a potential breast cancer treatment strategy. This project focused on designing, synthesizing, and evaluating novel covalent stabilizers targeting Cys38 on 14-3-3σ, based on WR-1065. The work extends a prior study on WR-1065's stabilizing effect via covalent modification of Cys38.

These molecules featured a shorter carbon linker, as depicted in Figure 1. Each molecule integrated a distinct electrophilic warhead to covalently target Cys38, aimed to enhance the interaction between 14-3-3/ERα and establish if they were isoform specific. The efficacy of these novel stabilizers was assessed using various analytical techniques.

In parallel, pyrrolidone1 was identified as a weak stabilizer of 14-3-3/ERα, primarily effective on 14-3-3/PMA2. Efforts remain to boost pyrrolidone1's effectiveness by characterizing derivatives, although their synthesis remains challenging.

Overall, various small molecules were found to be a greater success than others. This investigation has potentially uncovered a new mode of action that warrants further exploration.