Yue Wu

Bio: Yue Wu is an academic researcher from Tsinghua University. The author has contributed to research in topics: Ligand (biochemistry) & Prodrug. The author has an hindex of 9, co-authored 13 publications receiving 442 citations.

PROTACs: great opportunities for academia and industry.

Abstract: Although many kinds of therapies are applied in the clinic, drug-resistance is a major and unavoidable problem. Another disturbing statistic is the limited number of drug targets, which are presently only 20–25% of all protein targets that are currently being studied. Moreover, the focus of current explorations of targets are their enzymatic functions, which ignores the functions from their scaffold moiety. As a promising and appealing technology, PROteolysis TArgeting Chimeras (PROTACs) have attracted great attention both from academia and industry for finding available approaches to solve the above problems. PROTACs regulate protein function by degrading target proteins instead of inhibiting them, providing more sensitivity to drug-resistant targets and a greater chance to affect the nonenzymatic functions. PROTACs have been proven to show better selectivity compared to classic inhibitors. PROTACs can be described as a chemical knockdown approach with rapidity and reversibility, which presents new and different biology compared to other gene editing tools by avoiding misinterpretations that arise from potential genetic compensation and/or spontaneous mutations. PRTOACs have been widely explored throughout the world and have outperformed not only in cancer diseases, but also in immune disorders, viral infections and neurodegenerative diseases. Although PROTACs present a very promising and powerful approach for crossing the hurdles of present drug discovery and tool development in biology, more efforts are needed to gain to get deeper insight into the efficacy and safety of PROTACs in the clinic. More target binders and more E3 ligases applicable for developing PROTACs are waiting for exploration.

PROTAC-induced BTK degradation as a novel therapy for mutated BTK C481S induced ibrutinib-resistant B-cell malignancies

Abstract: Dear Editor, Non-Hodgkin’s lymphoma (NHL) is a type of cancer that mainly develops from B-cell malignancies, causing 231,400 deaths in 2015 globally. According to the American Cancer Society, around 66,000 new cases of NHL are diagnosed each year in the United States. Bruton’s tyrosine kinase (BTK) is an enzyme encoded by the BTK gene in human. BTK is expressed in all cell lineages of the hematopoietic system except for T cells. As a cytoplasmic tyrosine kinase of the TEC family, BTK plays a crucial role in B cell development, differentiation, and signaling. BTK is closely associated with chronic B-cell receptor (BCR) activation, and is critical for the survival of B-cell neoplasms. Inhibition of BTK kinase activity has been proven to be an important and practical way of treating NHL. Ibrutinib is a first-inclass, covalent BTK inhibitor that is able to bind C481 (Cysteine481) of BTK with an ideal IC50 of 0.5 nM. 4,5 Ibrutinib was approved by the FDA in 2013 to 2015 for the treatment of several types of NHL, specifically relapsed/refractory mantle cell lymphoma (MCL), chronic lymphocytic leukemia (CLL), and Waldenström macroglobulinaemia (WM). Currently, there is an ongoing clinical trial on ibrutinib for its efficacy in DLBCL treatment. However, resistance to ibrutinib has been reported in various lymphomas, including CLL and MCL, due to a C481S (cysteine to serine mutation at position 481) BTK mutation. Because ibrutinib cannot form a covalent bond with the hydroxyl group of serine, C481S mutation increases the IC50 against BTK-C481S phosphorylation from 2.2 nM to 1 μM. Thus, there is an urgent need to develop a new strategy against C481S mutation-induced resistance. In addition, ibrutinib has been reported to induce a variety of side effects, including arthralgias, myalgias, atrial fibrillation, ecchymosis, and major hemorrhage. As ibrutinib shows inhibition of EGFR, ITK and TEC family kinases, with IC50s of around 10–100 nM, the pathogeny could be correlated to these known off-target effects of ibrutinib. Proteolysis-targeting chimera (PROTAC) has emerged as a novel chemical approach for the selective degradation of cellular proteins, known as chemical knockdown of a protein of interest. PROTAC molecules (PROTACs) are small molecules capable of bringing a target protein into the proximity of an E3 ligase of interest, causing consequent degradation of the target protein (Fig. 1a). These heterobifunctional molecules consist of three components: a target protein-binding moiety (targeting arm, TA), a degradation machinery-recruiting unit (degradation arm, DA), and a linker that couples these two functionalities. Typically, the utilized degradation machinery is the ubiquitin–proteasome system (UPS) that recruits an E3 ubiquitin ligase followed by ubiquitination of the target protein and its subsequent degradation by the proteasome. In 2015, Crews and Bradner independently reported that BRD4, which has been implicated in several different cancers, could be efficiently degraded through the PROTAC technique. This presented the PROTAC technique as a promising alternative approach against cancer. Unlike traditional drugs, PROTACs aim to eliminate proteins with aberrant functions, rather than inhibiting their activity. Therefore, the acquired resistance caused by the C481S BTK mutant could, in principle, be overcome by using PROTAC molecules. In this study, for the first time, we report the development of BTK-targeting degraders using the PROTAC strategy. These PROTACs could efficiently degrade ibrutinib-sensitive BTK-WT (wild type). More importantly, our newly designed PROTACs also significantly induced the degradation of ibrutinib-resistant BTK-C481S (50% degradation efficiency at 30 nM). Furthermore, our PROTAC molecules efficiently inhibited cell proliferation and colony formation, while exhibited no obvious inhibition (>1000 nM) of ITK, EGFR, and TEC, which are major off-targets of ibrutinib. These data demonstrate the strong potential for developing PROTAC-based therapeutic molecules. To develop BTK-targeting PROTAC degraders, a BTK-targeting arm was conjugated to a BTK-degradation arm by linkers with variable lengths (Fig. 1a, b, for details, please see Supplementary Information). As a result, the PROTAC molecules should bind both BTK and E3 ligase through the corresponding targeting arm and degradation arm, respectively. The drugs ibrutinib and spebrutinib were selected as the BTK-binding ligands, and pomalidomide (CRBN ligand) and RG-7112 (MDM2 ligand) were employed as corresponding E3 ligase binding partners. Based on our design principles (for details, please see Supplementary Information), a variety of BTK-targeting PROTAC molecules were prepared and evaluated. We found that CRBN-recruiting PROTACs were generally more effective than MDM2-recruiting ones. Among these CRBN-recruiting PROTACs, P13I with the conjugation of ibrutinib and pomalidomide demonstrated the best degrading ability (Fig. 1b). P13I induced 73% degradation of BTK at 10 nM and 89% at 100 nM in human Burkitt’s lymphoma, RAMOS cells. As a further validation, we examined the kinetics of P13I-induced BTK degradation in human ABC-DLBCL, HBL-1 cells. Western blot analysis showed that BTK degradation began at roughly 4 h, and was completed by around 24 h (Fig. 1c). With P13I treatment, halflives of BTK and C481S BTK were 4 h and 3 h, respectively. The results demonstrated that P13I could accelerate BTK degradation (for details, please see SI). Moreover, P13I could also efficiently degrade BTK in other NHL cell lines including MCL (Mino cells) and MM cell lines with a DC50 (50% protein degradation concentration) of 9.2 nM and 11.4 nM, respectively (Fig. 1d). Control experiments clearly demonstrated that ibrutinib, pomalidomide or unconjugated PROTAC arms (TA and DA) could not induce degradation of BTK (Fig. 1e). In contrast, ibrutinib or pomalidomide could competitively inhibit the degradation effect of P13I. Additionally, MG-132, a proteasome inhibitor, could completely disable the PROTAC effect. The same competitive

Discovery of a first-in-class CDK2 selective degrader for AML differentiation therapy.

Abstract: The discovery of effective therapeutic treatments for cancer via cell differentiation instead of antiproliferation remains a great challenge. Cyclin-dependent kinase 2 (CDK2) inactivation, which overcomes the differentiation arrest of acute myeloid leukemia (AML) cells, may be a promising method for AML treatment. However, there is no available selective CDK2 inhibitor. More importantly, the inhibition of only the enzymatic function of CDK2 would be insufficient to promote notable AML differentiation. To further validate the role and druggability of CDK2 involved in AML differentiation, a suitable chemical tool is needed. Therefore, we developed first-in-class CDK2-targeted proteolysis-targeting chimeras (PROTACs), which promoted rapid and potent CDK2 degradation in different cell lines without comparable degradation of other targets, and induced remarkable differentiation of AML cell lines and primary patient cells. These data clearly demonstrated the practicality and importance of PROTACs as alternative tools for verifying CDK2 protein functions.

Target Elucidation by Cocrystal Structures of NADH-Ubiquinone Oxidoreductase of Plasmodium falciparum (PfNDH2) with Small Molecule To Eliminate Drug-Resistant Malaria

Abstract: Drug-resistant malarial strains have been continuously emerging recently, which posts a great challenge for the global health. Therefore, new antimalarial drugs with novel targeting mechanisms are urgently needed for fighting drug-resistant malaria. NADH-ubiquinone oxidoreductase of Plasmodium falciparum (PfNDH2) represents a viable target for antimalarial drug development. However, the absence of structural information on PfNDH2 limited rational drug design and further development. Herein, we report high resolution crystal structures of the PfNDH2 protein for the first time in Apo-, NADH-, and RYL-552 (a new inhibitor)-bound states. The PfNDH2 inhibitor exhibits excellent potency against both drug-resistant strains in vitro and parasite-infected mice in vivo via a potential allosteric mechanism. Furthermore, it was found that the inhibitor can be used in combination with dihydroartemisinin (DHA) synergistically. These findings not only are important for malarial PfNDH2 protein-based drug development but ...

Design, Synthesis, and Evaluation of Highly Potent FAK-Targeting PROTACs

Abstract: Focal adhesion kinase (FAK), a cytoplasmic protein tyrosine kinase, exerts kinase-dependent enzymatic functions and kinase-independent scaffolding functions, both of which are crucial in cancer development, early embryonic development, and reproduction. However, previous efforts for FAK blocking mainly focus on kinase inhibitors. Proteolysis targeting chimeras (PROTACs) are heterobifunctional molecules that allow direct post-translational knockdown of proteins via ubiquitination of a target protein by E3 ubiquitin ligase and subsequent proteasomal degradation. Here, we designed and synthesized a FAK PROTAC library with FAK inhibitor (PF562271 or VS6063) and CRBN E3 ligand. A novel FAK-targeting PROTAC, FC-11, showed a rapid and reversible FAK degradation with a picomolar of DC50 in various cell lines in vitro, which imply that FAK-PROTACs could be useful as expand tools for studying functions of FAK in biological system and as potential therapeutic agents.

PROTAC targeted protein degraders: the past is prologue

Abstract: Targeted protein degradation (TPD) is an emerging therapeutic modality with the potential to tackle disease-causing proteins that have historically been highly challenging to target with conventional small molecules. In the 20 years since the concept of a proteolysis-targeting chimera (PROTAC) molecule harnessing the ubiquitin–proteasome system to degrade a target protein was reported, TPD has moved from academia to industry, where numerous companies have disclosed programmes in preclinical and early clinical development. With clinical proof-of-concept for PROTAC molecules against two well-established cancer targets provided in 2020, the field is poised to pursue targets that were previously considered ‘undruggable’. In this Review, we summarize the first two decades of PROTAC discovery and assess the current landscape, with a focus on industry activity. We then discuss key areas for the future of TPD, including establishing the target classes for which TPD is most suitable, expanding the use of ubiquitin ligases to enable precision medicine and extending the modality beyond oncology. Targeted protein degradation with proteolysis-targeting chimeras (PROTACs) has the potential to tackle disease-causing proteins that have historically been highly challenging to target with conventional small molecules. This article summarizes the first two decades of PROTAC discovery and discusses key areas for the future of this therapeutic modality, including establishing the target classes for which it is most suitable and extending its application beyond oncology.

PROTACs: great opportunities for academia and industry.

Abstract: Although many kinds of therapies are applied in the clinic, drug-resistance is a major and unavoidable problem. Another disturbing statistic is the limited number of drug targets, which are presently only 20–25% of all protein targets that are currently being studied. Moreover, the focus of current explorations of targets are their enzymatic functions, which ignores the functions from their scaffold moiety. As a promising and appealing technology, PROteolysis TArgeting Chimeras (PROTACs) have attracted great attention both from academia and industry for finding available approaches to solve the above problems. PROTACs regulate protein function by degrading target proteins instead of inhibiting them, providing more sensitivity to drug-resistant targets and a greater chance to affect the nonenzymatic functions. PROTACs have been proven to show better selectivity compared to classic inhibitors. PROTACs can be described as a chemical knockdown approach with rapidity and reversibility, which presents new and different biology compared to other gene editing tools by avoiding misinterpretations that arise from potential genetic compensation and/or spontaneous mutations. PRTOACs have been widely explored throughout the world and have outperformed not only in cancer diseases, but also in immune disorders, viral infections and neurodegenerative diseases. Although PROTACs present a very promising and powerful approach for crossing the hurdles of present drug discovery and tool development in biology, more efforts are needed to gain to get deeper insight into the efficacy and safety of PROTACs in the clinic. More target binders and more E3 ligases applicable for developing PROTACs are waiting for exploration.

Trends in kinase drug discovery: targets, indications and inhibitor design.

Abstract: The FDA approval of imatinib in 2001 was a breakthrough in molecularly targeted cancer therapy and heralded the emergence of kinase inhibitors as a key drug class in the oncology area and beyond. Twenty years on, this article analyses the landscape of approved and investigational therapies that target kinases and trends within it, including the most popular targets of kinase inhibitors and their expanding range of indications. There are currently 71 small-molecule kinase inhibitors (SMKIs) approved by the FDA and an additional 16 SMKIs approved by other regulatory agencies. Although oncology is still the predominant area for their application, there have been important approvals for indications such as rheumatoid arthritis, and one-third of the SMKIs in clinical development address disorders beyond oncology. Information on clinical trials of SMKIs reveals that approximately 110 novel kinases are currently being explored as targets, which together with the approximately 45 targets of approved kinase inhibitors represent only about 30% of the human kinome, indicating that there are still substantial unexplored opportunities for this drug class. We also discuss trends in kinase inhibitor design, including the development of allosteric and covalent inhibitors, bifunctional inhibitors and chemical degraders. The FDA approval of imatinib in 2001 heralded the emergence of kinase inhibitors as a key drug class in the oncology area and beyond. This article analyses the landscape of approved and investigational therapies that target kinases and trends within it, including the most popular targets of kinase inhibitors, their expanding range of indications and strategies for kinase inhibitor design.