Design, Synthesis, and Evaluation of WD-Repeat-Containing Protein 5 (WDR5) Degraders

Abstract

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...

WIN site inhibition disrupts a subset of WDR5 function

“…We reasoned that comparison of the impact of WDR5 loss versus WIN site inhibition could resolve whether WIN site inhibitors block some or all of the function of WDR5, and would be timely given that the arsenal of WDR5 inhibitors has recently moved beyond those targeting the WIN site to those that trigger WDR5 degradation 26 , 27 . We also reasoned that monitoring the impact of WDR5 loss on H3K4me3 and transcriptional patterns would allow us to test the idea that WDR5 regulates transcription via modulation of H3K4 methylation.…”

PROTACs: great opportunities for academia and industry (an update from 2020 to 2021)

“…In 2021, Knapp group reported different WDR5 degraders based on two diverse WIN site binding scaffolds ( OICR-9429 modified scaffold and pyrroloimidazole modified scaffold) and different E3 ligase ligands. 277 In MV4-11 cells, OICR-9429 modified scaffold derived degrader 158 (Fig. 40 ) induced the degradation of WDR5 of 58% with a DC 50 value of 53 nM, and pyrroloimidazole modified scaffold derived degrader 159 (Fig.…”

PROTAC degraders as chemical probes for studying target biology and target validation

“…A variety of biophysical assay techniques including surface plasmon resonance (SPR), isothermal titration calorimetry (ITC) and differential scanning calorimetry (DSF), microscale thermophoresis (MST), fluorescence polarization (FP) has been employed to measure binary complex formation. [41][42][43][44] DSF has gained popularity due to the ease of use of this screening technique and its cost effectiveness. 43 Two detection methods measuring protein denaturation as a readout for ligand binding are commonly used.…”

“…Comparison of NanoBRET affinities measured in lysed as well as intact cells provides also a tool studying cell penetration of PROTACs as mentioned above. 42 Kinetic of PROTAC dissociation from the POI and E3 ligase can be determined using washout experiments. 31,47 Determining the affinity of a PROTAC to the E3 as well as the POI represents an important parameter for PROTAC optimization, but if binary complex formation is a limiting factor, if a certain affinity is required for degradation, or if high affinity may even hinder ternary complex formation between the E3, the PROTAC and the POI due to the onset of an early hook effect, is currently a matter of debate.…”