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Proteolysis-targeting chimeras (PROTAC) offers a promising potential alternative to protein inhibition for therapeutic intervention. Current PROTAC molecules require significant linker optimization and possess high molecular weight, which can limit their applications. Recently, an advanced PROTAC technology was developed, namely CLIPTAC (in-cell click-formed proteolysis-targeting chimeras). The main potential advantage of CLIPTAC is a significant reduction of the molecular weight and polar surface area of the separate reaction partners compared to the pre-assembled PROTAC molecule. Creative Biolabs is proud to provide the featured CLIPTAC platform that is designated for the development of such PROTACs.
In the CLIPTAC approach, the binding domain for the E3 ubiquitin ligase and the binding domain for the target molecule are initially present on two different molecules. These individual precursors have a lower molecular weight than regular PROTACs and therefore show improved cell permeability. Upon sequential administration of the two precursors, a functional PROTAC complex is formed in the cell via click chemistry. Both CRBN-based and VHL-based CLIPTACs have been designed to induce ubiquitylation and degradation of the target proteins.
Among the reported click chemistry reactions, the inverse electron demand Diels-Alder (IEDDA) cycloaddition between tetrazine and trans-cyclo-octene (TCO) was selected for the synthesis of CRBN-based CLIPTAC. This reaction is fast and high yielding, does not require the presence of a catalyst, and has found numerous biological applications, especially in the optical imaging field. The scientists designed and synthesized a tetrazine tagged thalidomide derivative (Tz-thalidomide) that can self-assemble with a TCO-tagged inhibitor of the protein of interest: the resulting CLIPTAC recruits the E3 ligase CRBN to the protein of interest resulting in its ubiquitination and then degradation. The CRBN-based CLIPTAC can simultaneously recruit target proteins and the E3 ligase CRBN and promote the proteasomal degradation of the target protein.
Fig.1 Schematic diagram of CRBN-based CLIPTACs.
To facilitate the discovery process for new PROTACs, another click chemistry, namely the Huisgen 1,3-dipolar cycloaddition reaction, was selected for the synthesis of VHL-based CLIPTAC. This reaction is typically high yielding, requiring stoichiometric quantities of each component and boasts excellent functional group compatibility under mild reaction conditions. In this approach, ligands for VHL were modified with terminal alkynes with linkers containing varying ethylene glycol units. A ligand for the target protein would have to be modified with an azide at a suitable position on the molecule so as to minimally influence the affinity of the ligand for its target protein and serve as the azide input for the “click reaction”. The VHL-based CLIPTAC can simultaneously recruit target proteins and the E3 ligase VHL and promote the proteasomal degradation of the target protein.
Fig.2 Schematic diagram of VHL-based CLIPTACs.
The CLIPTAC platform provides an advanced strategy to design new PROTACs, which can be extended to the destruction of any protein of interest via in-cell click-based formation of the PROTAC from a tagged ligand of these proteins and a tagged E3 ligase recruiter.
Based on the CLPTAC platform, Creative Biolabs systematically deploys a series of services to fully explore the potential of PROTAC as a new generation of small molecule drugs. If you are interested in our services, please directly contact us and consult our technical supports for more details.