Targets such as transcription factors are often challenging to study in recombinant or biochemical settings as they require native cellular localization, PTMs, and complexation to fold and function properly. Applying our proprietary chemoproteomics platform, Belharra has enabled ligand discovery campaigns in live cells which have uncovered tractable ligands for TFs important in disease pathology. Several examples of these ligand discovery and development efforts will be discussed.

Pioneer transcription factors (TFs) bind to and open closed chromatin, facilitating engagement by other regulatory factors involved in gene activation or repression. We present the chemical proteomic discovery of covalent small molecules that stereoselectively and site-specifically engage the pioneer TF, FOXA1. These compounds rapidly remodel FOXA1 interactions with chromatin in prostate cancer cell and create corresponding changes in chromatin accessibility through relaxing the DNA-binding preferences of FOXA1.

The Hippo pathway is a conserved signaling pathway that regulates organ size and cell fate by controlling cell proliferation and apoptosis. Here we report acrylamide small molecules that form a covalent bond with a conserved cysteine at the TEAD palmitate pocket. Binding studies showed profound stabilization of TEADs by the small molecules, and co-crystal structures reveal that the compounds mimic the binding mode of palmitate.   In mammalian cells, the compounds stabilize the TEAD•YAP1 interaction yet reduce TEAD and YAP1 protein levels and inhibit TEAD transcription factor activity. These compounds inhibit cell viability in several cancer cell lines.

Privileged chemical scaffolds for transcription factors (TFs) are not yet known. To enable discovery and optimization of targeted therapeutics for TFs, we have developed Strategian, a deep tensor factorization model trained on Talus Bio’s TF-Scan database. TF-Scan utilizes state-of-the-art DIA proteomics to observe compound-TF interactions across millions of data points in a native cellular environment. Strategian predicts the effects of untested compounds on TF activity, drawing from a drug-like universe of over 50B potential molecules. The model prioritizes candidates by potency and selectivity, followed by triage in TF-Scan. We identified novel scaffolds for STAT3, and direct inhibitors of IRF5.

Highly potent, exquisitely selective, and orally bioavailable inhibitor of SMARCA2 degrader identified using proprietary platform technology DNsD (Directed Neo-Substrate Degradation). Potential candidate has been identified for the treatment of SMARCA4 mutated cancers. In vitro and in vivo profile of potential candidate molecule will be presented.

Transcription factors are known to bind to cereblon in the presence of molecular glues and some reports implicate interactions with multiple zinc fingers. We present biophysical and structural assessments of the minimal binding domains of IKZF2 and other transcription factors, revealing that multiple zinc fingers interact with cereblon:glue complexes. In these examples, the binding modes are distinct and may have implications for the design of selective degraders.

BRM (SMARCA2) and BRG1 (SMARCA4) are mutually exclusive ATPase subunits of the mSWI/SNF (BAF) chromatin remodeling complex. BAF is an attractive therapeutic target because of its role in transcription, and mutations in the subunits of BAF are common in cancer and neurological disorders. Herein, we report the discovery of FHD-286, as a potent allosteric inhibitor of the dual ATPase subunits. FHD-286 is an orally bioavailable compound with antitumor activity in mouse xenograft models of uveal melanoma and acute myeloid leukemia and is being evaluated in Phase 1 clinical trials.

SBT-100 is approximately 15kD VHH derived nano-antibody which crosses the cell membrane and blood brain barrier (BBB) in less than 15 minutes in vivo. It binds to the transcription factor STAT3 (signal transducer activator of transcription 3) and inhibits its function. SBT-100 inhibits STAT3’s phosphorylation (i.e., activation), translocation to the nucleus, and binding to its DNA promoter.  It is effective at inhibiting human cancer growth in vitro and in vivo. Because it crosses the BBB, SBT-100 significantly suppresses the autoimmune response in multiple sclerosis and autoimmune uveitis, where is protects the retina and prevents blindness.