Chimia最新文献

Serine hydrolases (SHs) represent a functionally defined enzyme class with significant roles in human physiology, yet many of the approximately 240 human SH's lack selective chemical inhibitors required for functional validation. To address this, we developed a medium throughput screening platform based on multiplexed gel-based activity-based protein profiling (ABPP). Using mouse brain proteome as a source, we performed a target-agnostic screen of 1,664 covalent compounds, leading to the identification of three micromolar-potent fatty acid amide hydrolase (FAAH) inhibitors featuring urea warheads and one epoxide-based inhibitor targeting an unannotated protein. An α/β-hydrolase domain-containing protein 2 (ABHD2) interaction was identified via chemical proteomics, which was validated with overexpression studies in U2OS cells. While the current approach is biased toward metabolic SHs and constrained by gel resolution, it provides a scalable workflow to facilitate the discovery of selective hits and mechanism-of-action studies for underexplored proteins.

AKT is a critical mediator of the phosphoinositide 3-kinase (PI3K) signalling cascade, playing a key role in regulating essential cellular processes. The identification of AKT as one of the most dysregulated pathways in cancer led to the development of multiple classes of inhibitors. Despite numerous inhibitors entering clinical investigation and leading to the FDA approval of Capivasertib, an ATP-competitive AKT inhibitor, in November 2023, AKT modulation through inhibition was characterised by toxicity and poor clinical efficacy. Targeted Protein Degradation (TPD) spearheaded by PROTACs boasted a paradigmatic shift in drug discovery and was demonstrated to be a valid therapeutic alternative to modulate AKT. To date, numerous AKT-targeting PROTACs have been disclosed. The majority of them outperformed the inhibitors in suppressing AKT activity, nurturing higher potency and improved selectivity. Notably, enhanced antiproliferative effects, sustained by more robust and prolonged inactivation of the AKT downstream signalling was observed. This review highlights AKT as a central therapeutic target in oncology and focuses on AKT modulation through a targeted protein degradation approach mainly using PROTACs. The review aims at illustrating all the AKT-targeting PROTACs disclosed in literature to date, a powerful new pharmacological tool that might remarkably expand the scope of AKT-targeted therapies and further elucidate the role of AKT in both normal and cancer-related phenotypes.

The endocannabinoid system is a key homeostatic regulator that influences multiple physiological processes across nervous, immune, and metabolic systems in humans. Despite decades of pharmaceutical research, only a few clinically relevant outcomes have been achieved. This limited success is partly attributed to the exceptional complexity and signaling promiscuity of endogenous cannabinoids acting through the cannabinoid receptor types 1 and 2. Here, we review the development, application, and potential of labeled small molecules as tool compounds, with a specific focus on cannabinoid receptor type 1 research, the most abundant G-proteincoupled receptor in the mammalian brain. Technical and scientific advancements in spectroscopy have enabled the application of radionuclide and fluorescent probes with improved spatial and temporal resolution. In parallel, interdisciplinary collaboration, cross-validation, and rigorous pharmacological characterization established highquality standards for the development of labeled probes. Together, these developments open up new avenues for probe-based investigations of cannabinoid receptor type 1 biology and molecular pharmacology.

IRE1α is an important ER stress sensor located on the ER membrane with dual kinase and ribonuclease activity. It plays a crucial role in restoring ER proteostasis and is associated with various human diseases. Targeting IRE1α has become a promising therapeutic approach. Many IRE1α modulators have been identified in recent years, and some of these have demonstrated excellent pre-clinical efficacy. The modulation of IRE1α RNase activity by small molecules can be achieved through two main mechanisms: directly binding to the RNase domain to block RNA splicing, or allosteric modulation of its activity through binding to the kinase domain. Apart from monovalent inhibitors and activators, proteolysis targeting chimeras have been reported to degrade IRE1α and block its downstream signalling by recruiting the E3 ligase-ubiquitin system. In this review we summarize the recent advances of targeting IRE1α with small molecules, including inhibitors, activators, and bifunctional molecules, providing an insight into future development of chemical modalities targeting IRE1α.

Oligonucleotides, both RNA and DNA, are fundamental to life despite being composed of a limited set of simple molecular building blocks. Chemists have long strived to add additional components, especially orthogonal, unnatural base pairs (UBPs). These increase the informational content of nucleic acids and provide site-specific anchors for labelling, enabling applications in aptamer enhancement, RNA structure elucidation, pathway tracing, sequencing, and the construction of semi-synthetic organisms. For this, suitable enzymes and techniques are required to incorporate and later analyse expanded alphabet genetic material. In this review we aim to outline some challenges, achievements, and possibilities that this field encompasses.

The fluorogen-activating aptamer, Spinach, was a milestone in cellular RNA imaging and conceptually similar to the green fluorescent protein (GFP). Since then, more than ten FLAP systems have been developed across a broad spectral range. All of these systems differ fundamentally from GFP in that FLAPs rely on noncovalent fluorogen-RNA interactions, whereas GFP integrates its fluorophore covalently. In this article, we discuss recently developed FLAPs, including the first covalent FLAP, coPepper, as a promising new strategy to overcome current limitations in FLAP-based RNA imaging. Moreover, the bioconjugation chemistry developed for coFLAPs has immediate impact on covalent RNA labeling, RNA drug targeting, and in general, on covalent drug design.

Protein arginylation is a conserved post-translational modification in eukaryotes, involving the conjugation of arginine residues to proteins by the enzyme arginyl-tRNA transferase. Historically associated with targeted degradation, recent studies have expanded this view by uncovering its broader regulatory influence across diverse cellular functions. This review first examines the established roles of arginylation in protein degradation through the Ubiquitin-Proteasome System and Autophagy-Lysosome System. It then highlights its non-degradative functions, including the modulation of protein-protein interactions, complex assembly, protein stability, and crosstalk with other post-translational modifications. Emerging evidence supports the notion that arginylation functions in a context dependent manner, simultaneously affecting both the stability and functional behaviour of proteins. Together, these works reveal arginylation as a dynamic and versatile mechanism that extends well beyond proteolysis, positioning it as a key global regulator of cellular functioning.

Jumonji C histone lysine demethylases (JmjC-KDMs) are key chromatin regulators best known for catalysing histone lysine demethylation. There is growing evidence that JmjC-KDMs have a broader catalytic scope. This review summarises recent advances on JmjC-KDM activities beyond histone lysine demethylation, including arginine demethylation and arginine hydroxylation. We discuss how emerging insights into sequence-reactivity and inter-domain relationships, combinatorial post-translational modifications (PTMs), and cellular context shape substrate selectivity and enzymatic outcomes. These findings highlight substantial mechanistic flexibility within the JmjC-KDM family and may help prompt reconsideration of how their biochemistry is connected to physiological roles. We discuss implications for JmjC-KDM inhibitor development and outline outstanding questions, guiding future research concerning their roles in epigenetic regulation.

The bacterial pore-forming toxin α-hemolysin has dimensions appropriate for capture and translocation of DNA strands in a single-molecule electrophoresis experiment. We used the nanopore's properties to study G-quadruplex unfolding in the human telomere repeat sequence with and without the presence of DNA lesions introduced into either the GGG tracks of a potential G-quadruplex (via oxidation) or the TTA loops (via photodimerization). Different topological folds of the G-quadruplex could be distinguished by either their current-time signatures or by their unfolding rates when a dA25 tail was added. Another more compact four-stranded structure, the i-motif, was also studied and found to be exceptionally well folded and stable inside the nanopore cavity. Comparisons are made between early studies with a home-built nanopore device vs. the currently available instrument from Oxford Nanopore Technologies (ONT), showing that we approach, but do not yet achieve, single-molecule sequencing of DNA damage sites in human telomere repeats. These studies aid in our understanding of the structure and dynamics of non-canonically folded DNA, its behaviour in crowded environments that mimic intracellular conditions, and the ability to use nanopore sequencing to identify DNA damage sites in this oxidation-prone segment of the genome.