Biology subjects

Heering, J.

Publications and source records attributed to Heering, J..

5 recordsLinked to original sources

bioRxiv2025

Optimization and evaluation of complementary degrader discovery assays for application in screening

Targeted protein degradation (TPD) mediated by molecular glues is an innovative pharmaceutical paradigm. By binding to and modulating the surface of an E3-ligase component, molecular glue degraders can facilitate the recruitment of a specific target protein (or vice versa) and, ultimately, invoke target degradation. This mode of action results in specific challenges for the development of rational discovery strategies, and complex hit validation workflows may be required to reliably eliminate compounds that elicit non-specific effects. With the aim to guide screening efforts, we optimized two orthogonal cell-based, target-centric assays for degrader discovery: (1) a time-resolved FRET assay directly quantifying levels of target protein and its degradation (signal inhibition), and (2) an assay coupling TPD to cell growth (signal rescue). To enable a deeper understanding of the individual assays strengths and limitations, we compared their statistical performance as well as respective hit populations by screening a specifically designed collection of about 1000 compounds containing well annotated reference compounds and known frequent hitters. We found that the signal rescue format reliably and specifically captured active target degraders while it efficiently filtered out interfering or frequent hitter compounds. Importantly, this format achieved to retrieve lower potency hits, which might be desirable in order to confidently include as many diverse chemical starting points as possible at the start of a drug discovery project.

pharmacology and toxicology↗

bioRxiv2024

STRUCTURE-BASED DESIGN OF PROTACS FOR THE DEGRADATION OF SOLUBLE EPOXIDE HYDROLASE

Soluble epoxide hydrolase (sEH) represents a promising target for inflammation-related diseases as it hydrolyzes highly anti-inflammatory epoxy-fatty acids (EpFAs) to the less active corresponding diols.1 sEH harbours two distinct catalytic domains, the C-terminal hydrolase domain and the N-terminal phosphatase domain which are connected by a proline-rich linker.2 Although potent inhibitors of enzymatic activity are available for both domains, sEH-PROTACs offer the unique ability to simultaneously degrade both domains, mimicking the sEH knockout phenotype associated with beneficial effects as reducing inflammation, attenuating neuroinflammation, and delaying the progression of Alzheimers disease. Herein, we report the structure-based development of a potent sEH-PROTAC as a useful tool compound for the investigation of sEH. In order to facilitate a rapid testing of the synthesized compounds a cell-based sEH degradation assay was developed based on the HiBiT-technology. A structure-activity-relationship (SAR) investigation was performed, based on the crystal structure of previously published sEH inhibitor FL217 where we identified two possible exit vectors. We designed and synthesized a set of 24 PROTACs with varying linkers in a combinatorial manner. Furthermore, co-crystallization of sEH with two selected PROTACs allowed us to explore the binding mode and rationalize the appropriate linker length. After biological and physicochemical investigation, the most suitable PROTAC 23 was identified and applied to degrade sEH in primary human macrophages, marking the successful translation and applicability to non-artificial systems.

biochemistry↗

bioRxiv2023

Discovery of a new activator of Slack potassium channels with robust efficacy in models of histamine-independent and chronic itch

Various disorders are accompanied by histamine-independent itching, which is often resistant to the currently available therapies. In this study, we hypothesized that pharmacological activation of Slack (Kcnt1, KNa1.1), a potassium channel highly expressed in itch-sensitive sensory neurons, has therapeutic potential for the treatment of itching. Based on the Slack-activating antipsychotic drug, loxapine, we designed a series of new derivatives with improved pharmacodynamic and pharmacokinetic profiles that enabled us to validate Slack as a pharmacological target in vivo. One of these new Slack activators, compound 6, exhibited negligible dopamine D2 and D3 receptor binding, unlike loxapine. We found that compound 6 displayed potent on-target antipruritic activity in multiple mouse models of acute histamine-independent and chronic itch without motor side effects. These properties make compound 6 a lead molecule for the development of new antipruritic therapies targeting Slack.

pharmacology and toxicology↗

bioRxiv2023

Targeting LC3/GABARAP for degrader development and autophagy modulation

Recent successes in developing small-molecule degraders that act through the ubiquitin system have spurred efforts to extend this technology to other mechanisms, including the autophagosomal-lysosomal pathway. Therefore, reports of autophagosome tethering compounds (ATTECs) have received considerable attention from the drug development community. ATTECs are based on the target recruitment to LC3/GABARAP, a family of membrane-bound proteins that tether autophagy receptors to the autophagosome. In order to validate the existing ligands, we rigorously tested target engagement of reported ATTEC ligands and handles. Surprisingly, using various biophysical methods, most available ligands did not interact with their designated target LC3. Intrigued by the idea of developing ATTECs, we evaluated the druggability of LC3/GABARAP by in silico docking and large scale crystallographic fragment screening. The data revealed that most fragments bound to the HP2, but not the HP1 pocket of the LC3-interacting region (LIR) docking site, suggesting favorable druggability of this binding pocket. Here, we present diverse comprehensively validated ligands for future ATTEC development.

biochemistry↗

bioRxiv2021

Nurr1 modulation mediates neuroprotective effects of statins

The ligand-sensing transcription factor Nurr1 emerges as a promising therapeutic target for neurodegenerative pathologies but Nurr1 ligands for functional studies and therapeutic validation are lacking. Here we report pronounced Nurr1 modulation by statins for which clinically relevant neuroprotective effects have been demonstrated. Several statins directly affected Nurr1 activity in cellular and cell-free settings with low micromolar to sub-micromolar potencies. Simvastatin exhibited anti-inflammatory effects in astrocytes which were abrogated by Nurr1 knockdown. Differential gene expression analysis in native and Nurr1 silenced cells revealed strong proinflammatory effects of Nurr1 knockdown while simvastatin treatment induced several neuroprotective mechanisms via Nurr1, for example, in energy utilization and reduced apoptosis. These findings suggest Nurr1 involvement in the well-documented but mechanistically elusive neuroprotection by statins.

neuroscience↗