The Next Frontier of PROTACs: Development of Molecular Glues

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Targeted protein degradation (TPD) has emerged as a novel therapeutic approach for treating various diseases such as cancer, inflammation, neurodegenerative disorders, and infections. Many of these diseases are driven by aberrant expression of pathogenic proteins. TPD, with its broader scope of action, ability to target undruggable targets, and overcome drug resistance, has garnered significant attention.

Molecular glues and proteolysis targeting chimeras (PROTACs) are both protein degraders. A global surge in research on PROTAC technology has taken place. After PROTAC, molecular glue is expected to become another TPD research hotspot, which can regulate targets that are difficult to regulate with small molecules or antibodies.

Introduction to Molecular Glues

Molecular glues are small molecules with a M.W. less than 500 Daltons. They precisely control various biological processes, including signal transduction, transcription, chromatin regulation, protein folding, localization, and degradation over time. Molecular glues induce and enhance interactions between two proteins that lack intrinsic affinity. Molecular glues can promote the dimerization or co-localization of two proteins, resulting in distinct biological and pharmacological functions. When the target protein approaches the E3 ubiquitin ligase, it can undergo ubiquitination and degradation.

Fig. 1 Mechanism of action of molecular glues. (Dong, 2021)

The concept of "Molecular Glues" first appeared in the early 1990s derived from cyclosporin A and FK506. In 2000, rapamycin, approved by the FDA as an immunosuppressant, and its analogs were introduced. In 2014, researchers elucidated the mechanism of action of thalidomide analogs as degradation agents. Subsequently, molecular glues gradually gained attention and became a focus of research.

Fig. 2 Discovery of molecular glues. (Dong, 2021)

Molecular glues typically recruit target proteins for degradation by enhancing existing protein-protein interactions (PPI) or inducing the formation of new PPI complexes with substrate receptors for E3 ubiquitin ligases. Unlike traditional small molecule inhibitors or receptor antagonists, molecular glue degraders have several advantages. First, they drive the degradation of target proteins in a sub-stoichiometric and catalytic manner. Second, molecular glues can degrade previously inaccessible targets, as they do not require binding pockets on the target protein.

Molecular Glues and PROTACs

While both molecular glues and PROTACs are protein degraders, they differ in their mechanisms of action and molecular structural features. Molecular glues primarily induce or stabilize the PPI between the E3 ligase and substrate proteins, leading to protein degradation. On the other hand, PROTAC molecules consist of three parts: one end binds to the target protein, a linker connects to the E3 ligase ligand, and recruitment of the E3 ligase to the target protein leads to ubiquitination and subsequent degradation.

The mechanism of action of PROTACs is predictable and can be designed rationally based on the binding mode between the ligand and the target protein. In contrast, the discovery of molecular glues has a significant element of serendipity and cannot be obtained through large-scale screening of components like PROTACs. Currently, the discovery strategies for molecular glues include high-throughput screening, discovery from natural products, and chemical genomics screening.

Fig. 3 Molecular glues vs. PROTACs. (Sasso, 2023)

PROTAC molecules typically have a high molecular weight due to the presence of linkers and two ligand molecules, which results in poor cell permeability and pharmacokinetic characteristics. This limits their bioavailability. In contrast, molecular glues have lower molecular weights, higher cell permeability, and better oral absorption, meeting the criteria of drug-likeness, and showing greater clinical potential.

Types of Molecular Glues

1. Molecular Glues from Natural Products

The earliest drug discoveries involving the binding of two proteins to induce biological effects include cyclosporin A (CsA) and tacrolimus (FK506). These compounds, belonging to the cyclopeptide and macrolide structural types, respectively, bind to the receptor proteins cyclophilin and FK binding protein (FKBP). Despite differences in structure, both CsA and FK506 act as molecular glues by forming ternary complexes with calcineurin.

Another antibiotic with immunosuppressive properties, rapamycin, also binds to FKBP, forming a complex with the mammalian target of rapamycin (mTOR). mTOR is a crucial factor in cell growth and proliferation, and variations in its activity are closely related to tumors. Rapamycin induces the formation of FKBP-rapa-mTOR ternary complexes, acting as a molecular glue.

Fig. 4 Inducing PPI with molecular glues. (Che, 2018)

2. Molecular Glues from Imide Drugs

The first reported molecular glue was lenalidomide in 2014. Lenalidomide binds to the E3 ubiquitin ligase CUL4-CRBN protein and exhibits pleiotropic effects, leading to the ubiquitination of target proteins. CUL4, as an E3 ligase, is regulated by the CRBN protein. Lenalidomide, as well as analogs such as thalidomide and pomalidomide, can also bind to other proteins of this ligase, revealing the mechanism of action in treating chromosomal 5q deletion-associated myelodysplastic syndrome.

3. Molecular Glues from Arylsulfonamide Compounds

The arylsulfonamide derivative Indisulam was discovered through phenotypic anti-tumor screening and has been evaluated as an anti-tumor candidate in clinical trials. Although Indisulam exhibits selective anti-cancer activity, its mechanism of action and underlying targets have been studied for a considerable period. Indisulam acts as a molecular glue between E3 ubiquitin ligase CUL4-DCAF15 (DDB1 CUL4 associated factor 15) and splicing factor RNA binding motif protein 39 (RBM39). Other clinically tested sulfonamide drugs, such as tasisulam and CQS, also share the same mechanism of action.

Fig. 5 Chemical structures of arylsulfonamide degraders. (Frere, 2022)

Recent Advances in Molecular Glues

Currently, three molecular glue drugs have been approved and marketed globally: thalidomide, lenalidomide, and pomalidomide. These drugs, approved by the U.S. FDA, exhibit immunomodulatory, anti-inflammatory, and anti-tumor effects and are used to treat diseases such as multiple myeloma. Additionally, several molecular glue drugs are in various stages of clinical trials. Pharmaceutical companies such as Novartis, Bayer, BMS, Eisai, Nurix Therapeutics, and C4 Therapeutics are actively investing in the molecular glue field, indicating significant future market prospects for molecular glue drugs.

Table 1. Examples of molecular glues under clinical research.

• E7820 (Eisai, Inc., Phase II Clinical Trial)

Developed by Japan's Eisai, E7820 was initially designed as an angiogenesis inhibitor but later demonstrated anti-tumor properties in mouse experiments. Unlike acyl imide structures like lenalidomide, E7820 is a novel arylsulfonamide molecular glue that recruits DCAF15 to degrade RNA binding protein RBM39. Its DC50 is 17 nM, and it is currently in Phase II studies for relapsed or refractory AML, MDS, or CMML (NCT05024994).

• CFT7455 (C4 Therapeutics, Phase I/II Clinical Trial)

Selected and optimized from the C4 Therapeutics molecular glue compound library, CFT7455 contains a unique tricyclic imide structure. Similar to thalidomide and its derivatives, CFT7455 degrades IKZF1/3 by recruiting CRBN. Through structure-based rational design, the degradation efficiency of CFT7455 has significantly improved, with a DC50 reaching an impressive 170 pM. It is currently in Phase I/II clinical trials for the treatment of multiple myeloma and non-Hodgkin's lymphoma (NCT04756726).

• DKY709 (Novartis, Phase I/II Clinical Trial)

Optimized by Novartis based on the structure of pomalidomide, DKY709 selectively degrades the transcription factor IKZF2. It is currently being investigated for the treatment of various solid tumors (NCT03891953).

• CC-90009 (BMS/Celgene, Phase II Clinical Trial)

Developed by Celgene, CC-90009 is the first translational termination factor GSPT1 degrader to enter clinical trials. It was optimized after the identification of lead compounds through phenotype screening against AML cell lines. CC-90009 is currently in Phase II clinical trials for AML (NCT04336982) and MDS (NCT02848001).

Conclusion

While over 600 E3 ubiquitin ligases have been reported, only five have been used for molecular glue-mediated degradation: CRBN, DDB1, β-TrCP, DCAF15, and SIAH1. The E3 ubiquitin ligase library still holds untapped potential, and identifying new ligands for E3 ubiquitin ligases can expand the range of target proteins that can be degraded. Furthermore, the chemical space of molecular glue drugs warrants further exploration, as most reported molecular glues still exhibit high similarity to thalidomide and its derivatives. A deeper understanding of protein-protein interaction interfaces and structure-guided molecular glue design based on this understanding is needed to truly bring molecular glues into clinical applications.

References

1. Dong, G., et al., Molecular Glues for Targeted Protein Degradation: From Serendipity to Rational Discovery, J. Med. Chem., 2021, 64, 15, 10606-10620.
2. Sasso, J. M., et al., Molecular Glues: The Adhesive Connecting Targeted Protein Degradation to the Clinic, Biochemistry, 2023, 62, 3, 601-623.
3. Che, Y., et al., Inducing protein-protein interactions with molecular glues, Bioorganic & Medicinal Chemistry Letters, 2018, 28, 2585-2592.
4. Frere, G. A., et al., Chapter 1 - Emerging mechanisms of targeted protein degradation by molecular glues, Methods in Cell Biology, 2022, 169, 1-26.