Harnessing PROTAC technology to combat stress hormone receptor activation

Counteracting the overactivation of glucocorticoid receptors (GR) is an important therapeutic goal in stress-related psychiatry and beyond. The only clinically approved GR antagonist lacks selectivity and induces unwanted side effects. To complement existing tools of small-molecule-based inhibitors, we present a highly potent, novel catalytically-driven GR degrader, KH-103, based on proteolysis-targeting chimera technology. This selective degrader enables immediate and reversible GR depletion that is independent of genetic manipulation and circumvents transcriptional adaptations to inhibition. KH-103 achieves passive inhibition, preventing agonistic induction of gene expression, and significantly averts the GR’s genomic effects compared to two currently available inhibitors. Application in primary-neuron cultures revealed the dependency of a glucocorticoid-induced increase in spontaneous calcium activity on GR. Finally, we present a proof of concept for application in-vivo. KH-103 opens opportunities for a more lucid interpretation of GR functions with translational potential. ### Competing Interest Statement The authors have declared no competing interest. * AAV : Adeno-associated virus ALS : Amyotrophic Lateral Sclerosis BBB : Blood-brain-barrier Bp : Base pair ChIP-seq : Chromatin immunoprecipitation sequencing CPM : Counts per million CRBN : Cereblon Cyps : Cytochromes P450 DEG : Differentially expressed gene DEX : Dexamethasone DMSO : Dimethyl sulfoxide dTAG : Degradation TAG Dusp1 : Dual-specificity phosphatase 1 EGFP : Enhanced green fluorescent protein FDR : False discovery rate FKBP12 : FK506-binding protein 12 FKBP5 : FK506-binding Protein 51 GC : Glucocorticoid GR : Glucocorticoid receptor H : Hour(s) HEK293 : Human embryonic kidney 293 Histone 3 : H3 HPA : Hypothalamus-Pituitary-Adrenal Hz : Hertz IL-6 : Interleukin-6 KD : Dissociation constant Kda : Kilodalton LBD : Ligand binding domain logFC : Log fold change MAP2 : Microtubule-associated protein 2 MIF : Mifepristone Min : Minute(s) MPro : SARS-CoV-2 main protease MR : Mineralocorticoid receptor N2a : Neuro 2a PDB : Protein data bank PEG : Poly(ethylene glycol) Per1 : Period circadian regulator 1 PROTAC : Proteolysis targeting chimeras PXR : Pregnane X receptor RT-qPCR : Reverse transcription-quantitative polymerase chain reaction Sgk1 : Serum/glucocorticoid regulated kinase 1 TAU : Tubulin-associated unit TSS : Transcription start site UPS : Ubiquitin-proteasome system VDAC : Voltage-dependent anion channel * AAV : Adeno-associated virus ALS : Amyotrophic Lateral Sclerosis BBB : Blood-brain-barrier Bp : Base pair ChIP-seq : Chromatin immunoprecipitation sequencing CPM : Counts per million CRBN : Cereblon Cyps : Cytochromes P450 DEG : Differentially expressed gene DEX : Dexamethasone DMSO : Dimethyl sulfoxide dTAG : Degradation TAG Dusp1 : Dual-specificity phosphatase 1 EGFP : Enhanced green fluorescent protein FDR : False discovery rate FKBP12 : FK506-binding protein 12 FKBP5 : FK506-binding Protein 51 GC : Glucocorticoid GR : Glucocorticoid receptor H : Hour(s) HEK293 : Human embryonic kidney 293 Histone 3 : H3 HPA : Hypothalamus-Pituitary-Adrenal Hz : Hertz IL-6 : Interleukin-6 KD : Dissociation constant Kda : Kilodalton LBD : Ligand binding domain logFC : Log fold change MAP2 : Microtubule-associated protein 2 MIF : Mifepristone Min : Minute(s) MPro : SARS-CoV-2 main protease MR : Mineralocorticoid receptor N2a : Neuro 2a PDB : Protein data bank PEG : Poly(ethylene glycol) Per1 : Period circadian regulator 1 PROTAC : Proteolysis targeting chimeras PXR : Pregnane X receptor RT-qPCR : Reverse transcription-quantitative polymerase chain reaction Sgk1 : Serum/glucocorticoid regulated kinase 1 TAU : Tubulin-associated unit TSS : Transcription start site UPS : Ubiquitin-proteasome system VDAC : Voltage-dependent anion channel