Summary The mitotic kinase AURORA-A is essential for cell cycle progression and is considered a priority cancer target. While the catalytic activity of AURORA-A is essential for its mitotic function, recent reports indicate an additional non-catalytic function, which is difficult to target by conventional small molecules. We therefore developed a series of chemical degraders (PROTACs) by connecting a clinical kinase inhibitor of AURORA-A to E3 ligase-binding molecules (e.g. thalidomide). One degrader induced rapid, durable and highly specific degradation of AURORA-A. In addition ,we found that the degrader complex was stabilized by cooperative binding between AURORA-A and CEREBLON. Degrader-mediated AURORA-A depletion caused an S-phase defect, which is not the cell cycle effect observed upon kinase inhibition, supporting an important non-catalytic function of AURORA-A during DNA replication. AURORA-A degradation induced rampant apoptosis in cancer cell lines, and thus represents a versatile starting point for developing new therapeutics to counter AURORA-A function in cancer.

In total, more than 700 proteins regulate chromatin function 18,[22][23] and they are often part of multi-domain protein complexes. Beside the catalytic subunit that controls chromatin accessibility, also subunits that recognize and interact with epigenetic modifications are crucial components of histone modifying complexes. 2 Despite the three classes of epigenetic readers, erasers, and writers, also epigenetic movers, shapers and insulators interact with chromatin structure. [24][25] Proteins that recognize post-translational modifications are classified as epigenetic readers. 26 Well-studied protein families for epigenetic readers are, e.g., bromodomains (BRDs), which recognize acetylated lysine residues. The BRDs have been extensively studied and successfully drugged in cancer treatment. 26 In Table 1, the bromodomain BRD4 of the bromodomain and extraterminal domain (BET) family is listed due to its prominent role in super-enhancers (SEs) organization and regulation of oncogene expression in cancer. 27 Targeting BRD4 by inhibiting the acetyl-lysine binding site with small molecules, e.g., the first BRD targeting inhibitor (JQ1), was shown to be an effective strategy for cancers like the aggressive NUT midline carcinoma (NMC). [28][29] Beside the outstanding role of BRD4, other BRDs are involved as epigenetic readers in various nucleosome remodeling complexes: in the ATP-dependent human complexes BAF (BRG1/BRMassociated factor) and PBAF (polybromo-associated BAF factor), two bromodomains, SMARCA2/ 4 (SWI/SNF-related, matrix-associated actin-dependent regulator of chromatin, subfamily A2/ 4), perturbate with the core subunits BRG1/BRM histone-DNA contacts. [30][31][32] Mutations in BAF components are one of the most frequently observed genetic alteration in cancer. [33][34] demonstrated how mutations and misregulations of histone lysine methyltransferases (KMTs), demethylases and methyl-lysine-binding proteins are connected to various diseases, thus making them effective therapeutic targets for cancer treatments. [46][47] The histone demethylation process is carried out by lysine demethylases like LSD1 48 and the JARID1 familiy 49 epigenetic erasers that are known to be perturbed in cancer, as previously listed in Table 1. Equally involved in cancer formation is the class of histone lysine methyltransferases (KMTs) which are categorized as epigenetic writers. KMTs comprise proteins like MLL1-3 and SET1D which are relevant drug targets, as shown in the non-exhaustive list in Table 1. Within histone lysine methylation, H3K4 methylation is an evolutionary conserved motif that marks active gene transcription 50-51 and is highly enriched at the promotor region and transcription start site. 51 The family of Histone lysine Methyltransferases and its adaptor proteins are described in the following chapter.while the pink colored c-Myc peptide MbIIIb binds to WDR5 on a shallow cleft on the surface, the so called WBM side (pdb entry: 3eg6 and 4y7r).WDR5 has emerged as a promising drug target for anti-cancer therapies as i...

Highlights d 25% of human E2s have a ubiquitination site (Lys +5 ) near the catalytic center d Flexibility of the active-site region enables Lys +5 autoubiquitination in UBE2S d Lys +5 -linked ubiquitin inhibits UBE2S by adopting a closed conformation d The extent of Lys +5 ubiquitination in UBE2S is regulated during the cell cycle

Histone H3K4 methylation serves as post-translational hallmark of actively transcribed genes and is introduced by histone methyltransferases (HMT) and its regulatory scaffolding proteins. One of these is the WD-repeat containing protein 5 (WDR5) that has also been associated with controlling long non-coding RNAs and transcription factors including MYC. The wide influence of dysfunctional HMTs complexes and the typically upregulated MYC levels in diverse tumor types suggested WDR5 as an attractive drug target. Indeed, protein-protein interface inhibitors for two protein interaction interfaces on WDR5 have been developed. While such compounds only inhibit a subset of WDR5 interactions, chemically induced proteasomal degradation of WDR5 might represent an elegant way to target all oncogenic function. This study presents the design, synthesis and evaluation of two diverse WDR5 degrader series based on two WIN site binding scaffolds and shows that linker nature and length strongly influence degradation efficacy.

Formation of the Aurora-A–MYCN complex increases levels of the oncogenic transcription factor MYCN in neuroblastoma cells by abrogating its degradation through the ubiquitin proteasome system. While some small-molecule inhibitors of Aurora-A were shown to destabilize MYCN, clinical trials have not been satisfactory to date. MYCN itself is considered to be `undruggable' due to its large intrinsically disordered regions. Targeting the Aurora-A–MYCN complex rather than Aurora-A or MYCN alone will open new possibilities for drug development and screening campaigns. To overcome the challenges that a ternary system composed of Aurora-A, MYCN and a small molecule entails, a covalently cross-linked construct of the Aurora-A–MYCN complex was designed, expressed and characterized, thus enabling screening and design campaigns to identify selective binders.