Ligand Design for BTK-targeting PROTAC^®

Currently, proteolysis targeting chimera (PROTAC^®) has emerged as a new modality with the possibility to promote drug discovery. This technique can be used to induce proteasomal degradation processes of specific proteins instead of inhibiting these proteins. Bruton’s tyrosine kinase (BTK) is a nonreceptor tyrosine kinase that has been expressed in hepatocellular carcinoma (Tec) family and is essential in normal B cell development. After collecting crystal structure of desired proteins, Creative Biolabs would like to offer cost-effective ligand design services for BTK kinase or other protein kinase targets and is pleased to introduce multiple ligand design strategies for clients’ challenging PROTAC^® projects.

• Introduction to BTK

BTK has an N-terminal region with a pleckstrin homology (PH) domain and a proline-rich Tec homology (TH) domain. The PH portion is the site of activation by G protein βγ subunits and phosphatidylinositol phosphates, and inhibition by protein kinase C. The remaining region of BTK encompasses Src homology (SH) domain SH3, followed by an SH2, and a C-terminal SH1 with kinase activity. The SH2 domain can mediate binding to tyrosine-phosphorylated motifs on other molecules, while the SH3 domain is responsible for binding to proline-rich peptide motifs. There are two regulatory tyrosine residues in BTK, Tyr-223, and Tyr-551, which participate in kinase activation. Distinct domains of BTK are essential for their physiological functions. The crystal structure of the PH domain has been described, which contributes to a better understanding of how point mutations in this domain inactivate BTK and cause disorders.

Fig.1 BTK kinase domain structure. (Wang, 2014)

BTK engages in transduction pathways evoked by the binding of a multitude of extracellular ligands to their cell surface receptors. Mutations in the human BTK gene result in XLA, which is a male immunodeficiency featured by a lack of mature B lymphocytes. To be noticed, aberrant regulation of BTK signaling has been associated with various types of B cell malignancies, such as mantle cell lymphoma, diffuse large B cell lymphoma, and chronic lymphocytic leukemia (CLL). Ibrutinib is a robust and selective irreversible inhibitor of BTK. This agent can covalently bind to BTK and has affected the treatment landscape for B cell carcinomas, especially CLL.

Fig.2 Regulation of BTK expression and activation. (Singh, 2017)

• Ligand Design Services for BTK at Creative Biolabs

Creative Biolabs has developed a stream of novel strategies to control protein degradation and one representative strategy is PROTAC^® technology. This tool uses a bifunctional-hybrid molecule that binds both E3 ubiquitin ligase and target protein, thereby causing the exposed lysine on the target protein being ubiquitinated by the E3 ligase complex, followed by the ubiquitin-proteasome system (UPS)-mediated protein degradation. Targeted degradation of BTK serves as a model system to understand the foundation of rapid targeted protein degradation. In the previous studies, members of an 11-compound PROTAC^® library are analyzed for their ability to form ternary complexes with BTK and Cereblon (CRBN, an E3 ligase element). Through in vitro and in vivo experiments, fast, efficient, selective, and prolonged BTK degradation has been demonstrated.

Fig.3 PROTAC^®s selectively degrade BTK. (Zorba, 2018)

PROTAC^®-based protein degradation is an emerging industry that holds a remarkable promise for targeting the undruggable proteome. Here, Creative Biolabs takes a series of cellular assays and biochemical techniques to investigate the efficient knockdown of BTK kinases. In particular, we devote to developing ideal ligands by unique PROTAC^® technology and providing custom ligand design services for BTK and other targets. Based on structural characteristics of diversified proteins, our teams are able to propose different formats of ligands to be studied, including small molecules, peptides, proteins, and recombinant antibodies.

• Key Benefits
  • Rational ligand design services for BTK-targeting PROTAC^®s with required degradation profiles
  • Advanced platforms and professional technicians ensure ligands with high potency and selectivity
  • Detailed reports involving design, protocols, models, and analysis

To date, PROTAC^®s can be used to target varieties of proteins, including protein kinases, transcription factors, skeleton proteins, and regulatory proteins. With extensive experience in molecular discovery fields, Creative Biolabs has been recognized as a long-term expert in successfully developing chimeric degraders for BTK and other kinase degradation. For more details, please don’t hesitate to contact us.

References

1. Wang, S.F. A Novel deletion mutation and structural abnormality in the Bruton's tyrosine kinase gene identified in a Chinese patient with X-linked agammaglobulinemia. Clin Lab. 2014, 60(5): 859-862.
2. Singh, S.P.; et al. Targeting Bruton’s tyrosine kinase expression levels through microRNAs in chronic lymphocytic leukemia treatment. Transl Cancer Res. 2017, 6(Suppl 3): S502-S507.
3. Zorba, A.; et al. Delineating the role of cooperativity in the design of potent PROTAC^®s for BTK. PNAS. 2018, 115(31): E7285-E7292.