Characterization of Organic Anion and Cation Transport in Three Human Renal Proximal Tubular Epithelial Models.

The polarised expression of specific transporters in proximal tubular epithelial cells is important for the renal clearance of many endogenous and exogenous compounds. Thus, ideally, the in vitro tools utilised for predictions would have a similar expression of apical and basolateral xenobiotic transporters as in vivo. Here, we assessed the functionality of organic cation and anion transporters in proximal tubular-like cells (PTL) differentiated from human induced pluripotent stem cells (iPSC), primary human proximal tubular epithelial cells (PTEC), and telomerase-immortalised human renal proximal tubular epithelial cells (RPTEC/TERT1).

Development of a membrane-disruption assay using phospholipid vesicles as a proxy for the detection of cellular membrane degradation.

Snakebite envenoming is a global health issue that affects millions of people worldwide, and that causes morbidity rates surpassing 450,000 individuals annually. Patients suffering from snakebite morbidities may experience permanent disabilities such as pain, blindness and amputations. The (local) tissue damage that causes these life-long morbidities is the result of cell- and tissue-damaging toxins present in the venoms.

Development of a high-throughput in vitro screening method for the assessment of cell-damaging activities of snake venoms.

Snakebite envenoming is a globally important public health issue that has devastating consequences on human health and well-being, with annual mortality rates between 81,000 and 138,000. Snake venoms may cause different pathological effects by altering normal physiological processes such as nervous transfer and blood coagulation. In addition, snake venoms can cause severe (local) tissue damage that may result in life-long morbidities, with current estimates pointing towards an additional 450,000 individuals that suffer from permanent disabilities such as amputations, contractions and blindness.

Structural and Molecular Determinants for Isoform Bias at Human Histamine H Receptor Isoforms.

The human histamine H receptor (hHR) is predominantly expressed in the CNS, where it regulates the synthesis and release of histamine and other neurotransmitters. Due to its neuromodulatory role, the hHR has been associated with various CNS disorders, including Alzheimer's and Parkinson's disease. Markedly, the hHR gene undergoes extensive splicing, resulting in 20 isoforms, of which 7TM isoforms exhibit variations in the intracellular loop 3 (IL3) and/or C-terminal tail.

Capturing time-dependent activation of genes and stress-response pathways using transcriptomics in iPSC-derived renal proximal tubule cells.

Transcriptomic analysis is a powerful method in the utilization of New Approach Methods (NAMs) for identifying mechanisms of toxicity and application to hazard characterization. With this regard, mapping toxicological events to time of exposure would be helpful to characterize early events. Here, we investigated time-dependent changes in gene expression levels in iPSC-derived renal proximal tubular-like cells (PTL) treated with five diverse compounds using TempO-Seq transcriptomics with the aims to evaluate the application of PTL for toxicity prediction and to report on temporal effects for the activation of cellular stress response pathways.

SAR exploration of the non-imidazole histamine H receptor ligand ZEL-H16 reveals potent inverse agonism.

Histamine H receptor (H R) agonists without an imidazole moiety remain very scarce. Of these, ZEL-H16 (1) has been reported previously as a high-affinity non-imidazole H R (partial) agonist. Our structure-activity relationship analysis using derivatives of 1 identified both basic moieties as key interaction motifs and the distance of these from the central core as a determinant for H R affinity.

Fusicoccin-A Targets Cancerous Inhibitor of Protein Phosphatase 2A by Stabilizing a C-Terminal Interaction with 14-3-3.

The cancerous inhibitor of protein phosphatase 2A (CIP2A) is an oncoprotein found overexpressed in many types of cancer. CIP2A has been shown to stabilize oncoproteins such as cMYC by shielding them from PP2A-mediated dephosphorylation. Here we report that the penultimate residue Ser904 in the C-terminus of CIP2A can be phosphorylated to create a binding site for the regulatory protein 14-3-3.

Fragment Screening Yields a Small-Molecule Stabilizer of 14-3-3 Dimers That Modulates Client Protein Interactions.

The development of protein-protein interaction (PPI) inhibitors has been a successful strategy in drug development. However, the identification of PPI stabilizers has proven much more challenging. Here we report a fragment-based drug screening approach using the regulatory hub-protein 14-3-3 as a platform for identifying PPI stabilizers.

Generation and characterization of iPSC-derived renal proximal tubule-like cells with extended stability.

The renal proximal tubule is responsible for re-absorption of the majority of the glomerular filtrate and its proper function is necessary for whole-body homeostasis. Aging, certain diseases and chemical-induced toxicity are factors that contribute to proximal tubule injury and chronic kidney disease progression. To better understand these processes, it would be advantageous to generate renal tissues from human induced pluripotent stem cells (iPSC).

4-(3-Aminoazetidin-1-yl)pyrimidin-2-amines as High-Affinity Non-imidazole Histamine H Receptor Agonists with in Vivo Central Nervous System Activity.

Despite the high diversity of histamine H receptor (HR) antagonist/inverse agonist structures, partial or full HR agonists have typically been imidazole derivatives. An in-house screening campaign intriguingly afforded the non-imidazole 4-(3-azetidin-1-yl)pyrimidin-2-amine as a partial HR agonist. Here, the design, synthesis, and structure-activity relationships of analogues are described.

A Photoswitchable Agonist for the Histamine H Receptor, a Prototypic Family A G-Protein-Coupled Receptor.

Spatiotemporal control over biochemical signaling processes involving G protein-coupled receptors (GPCRs) is highly desired for dissecting their complex intracellular signaling. We developed sixteen photoswitchable ligands for the human histamine H receptor (hH R). Upon illumination, key compound 65 decreases its affinity for the hH R by 8.

Synthesis and Characterization of a Bidirectional Photoswitchable Antagonist Toolbox for Real-Time GPCR Photopharmacology.

Noninvasive methods to modulate G protein-coupled receptors (GPCRs) with temporal and spatial precision are in great demand. Photopharmacology uses photons to control in situ the biological properties of photoswitchable small-molecule ligands, which bodes well for chemical biological precision approaches. Integrating the light-switchable configurational properties of an azobenzene into the ligand core, we developed a bidirectional antagonist toolbox for an archetypical family A GPCR, the histamine H receptor (HR).

A comprehensive analysis of the 14-3-3 interactome in barley leaves using a complementary proteomics and two-hybrid approach.

This study describes the identification of over 150 target proteins of the five 14-3-3 isoforms in 7-d-old barley (Hordeum vulgare) cv Himalaya seedlings using yeast two-hybrid screens complemented with 14-3-3 protein affinity purification and tandem mass spectrometry. Independent experiments for a subset of genes confirmed the yeast two-hybrid interactions, demonstrating a low false positive identification rate. These combined approaches resulted in the identification of more than 150 putative targets; 15% were previously reported to be 14-3-3 interactors, including, for example, Serpin, RF2A, WPK4 kinase, P-type proton-translocating adenosine triphosphatase, EF1A, glutamine synthetase, and invertases.