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Investigating freezing-induced acidity changes in citrate buffers.
No full textCitrate buffers are commonly utilized in the field of biomolecule stabilization. We investigate their applicability in the frozen state within a range of initial pHs (2.5 to 8.0) and concentrations (0.02 to 0.60 M). Citrate buffer solutions subjected to various cooling and heating temperatures are examined in terms of the freezing-induced acidity changes, revealing that citrate buffers acidify upon cooling. The acidity is assessed with sulfonephthalein molecular probes frozen in the samples. Optical cryomicroscopy combined with differential scanning calorimetry was employed to investigate the causes of the observed acidity changes. The buffers partly crystallize and partly vitrify in the ice matrix; these processes influence the resulting pH and allow designing the optimal storage temperatures in the frozen state. The freezing-induced acidification apparently depends on the buffer concentration; at each pH, we suggest pertinent concentration, at which freezing causes minimal acidification
Unaltered hepatic wound healing response in male rats with ancestral liver injury.
No full textThe possibility that ancestral environmental exposure could result in adaptive inherited effects in mammals has been long debated. Numerous rodent models of transgenerational responses to various environmental factors have been published but due to technical, operational and resource burden, most still await independent confirmation. A previous study reported multigenerational epigenetic adaptation of the hepatic wound healing response upon exposure to the hepatotoxicant carbon tetrachloride (CCl4) in male rats. Here, we comprehensively investigate the transgenerational effects by repeating the original CCl4 multigenerational study with increased power, pedigree tracing, F2 dose-response and suitable randomization schemes. Detailed pathology evaluations do not support adaptive phenotypic suppression of the hepatic wound healing response or a greater fitness of F2 animals with ancestral liver injury exposure. However, transcriptomic analyses identified genes whose expression correlates with ancestral liver injury, although the biological relevance of this apparent transgenerational transmission at the molecular level remains to be determined. This work overall highlights the need for independent evaluation of transgenerational epigenetic inheritance paradigms in mammals
Rational Chemical Design of Molecular Glue Degraders.
No full textTargeted protein degradation with molecular glue degraders has arisen as a powerful therapeutic modality for eliminating classically undruggable disease-causing proteins through proteasome-mediated degradation. However, we currently lack rational chemical design principles for converting protein-targeting ligands into molecular glue degraders. To overcome this challenge, we sought to identify a transposable chemical handle that would convert protein-targeting ligands into molecular degraders of their corresponding targets. Using the CDK4/6 inhibitor ribociclib as a prototype, we identified a covalent handle that, when appended to the exit vector of ribociclib, induced the proteasome-mediated degradation of CDK4 in cancer cells. Further modification of our initial covalent scaffold led to an improved CDK4 degrader with the development of a but-2-ene-1,4-dione ("fumarate") handle that showed improved interactions with RNF126. Subsequent chemoproteomic profiling revealed interactions of the CDK4 degrader and the optimized fumarate handle with RNF126 as well as additional RING-family E3 ligases. We then transplanted this covalent handle onto a diverse set of protein-targeting ligands to induce the degradation of BRD4, BCR-ABL and c-ABL, PDE5, AR and AR-V7, BTK, LRRK2, HDAC1/3, and SMARCA2/4. Our study undercovers a design strategy for converting protein-targeting ligands into covalent molecular glue degraders
Molecular Insights into the Binding Interaction of the First Class of Potent Small Molecules Disrupting the YAP-TEAD Protein-Protein Interaction
No full textDisruption of the YAP-TEAD protein-protein interaction is an attractive therapeutic strategy for oncology to suppress tumour progression and cancer metastasis. YAP binds to TEAD at a large flat binding interface (~3500 Å2) devoid of a well-defined druggable pocket, so it has been difficult to design low molecular weight compounds to abrogate this protein-protein interaction directly. Recently, work by Furet and coworkers (ChemMedChem 2022, DOI: 10.1002/cmdc.202200303) reported the discovery of the first class of small molecules able to efficiently disrupt the transcriptional activity of TEAD by binding to a specific interaction site of the YAP-TEAD binding interface. Using high-throughput in-silico docking and structure-based drug design, they identified a virtual screening hit from a hot-spot derived from their previously rationally designed peptidic inhibitor. Given the advances in rapid high-throughput screening and rational approaches to peptidic ligand discovery for challenging targets, we analyzed the pharmacophore features involved in transferring from their peptidic to small molecule inhibitor that could enable small molecule discovery for such targets. Here, we show that pharmacophore analysis augmented by solvation analysis of molecular dynamics trajectories can guide the designs while binding free energy calculations provide greater insight into the binding conformation and energetics accompanying the association event. The computed binding free energy estimates agree with experimental findings and offer useful insight into structural determinants that influence ligand binding to the TEAD interaction surface, even for such a shallow binding site. In general, our results provide rationale for the structure-based design efforts in compound optimization that led to a significant gain in potency among the first class of potent small molecule YAP:TEAD protein-protein inhibitors
A novel molecule promotes readthrough at premature stop codons by inducing co-translational degradation of eRF1 involving GCN1, RNF14 and RNF25
No full textDrugs that promote translational readthrough at premature termination codons (PTC) are a promising for treating a wide variety of severe genetic diseases caused by nonsense mutations. Here, we present two novel and potent readthrough promoters – NVS1.1 and NVS2.1 – that restore substantial levels of functional full-length CFTR and IDUA proteins in disease models for cystic fibrosis and Hurler syndrome, respectively. In contrast to other readthrough promoters that affect decoding of the stop codon, the NVS compounds stimulate PTC suppression by triggering rapid proteasomal degradation of the translation termination factor eRF1. Interestingly, this appears to occur by trapping eRF1 in the terminating ribosome, causing ribosome stalls and subsequent ribosome collisions, which activate a novel branch of the ribosome quality control (RQC) network that involves the translational stress sensor GCN1 and the catalytic activity of the E3 ubiquitin ligases RNF14 and RNF2
Micellar Catalysis as a Tool for Suzuki-Miyaura Cross-Couplings in Water
No full textA scalable and environmentally benign protocol for Suzuki-Miyaura cross-coupling in water using the vitamin E derived surfactant TPGS-750-M is reported. The protocol is the simplest and most standard for such transformations under micellar catalysis that has been amply utilized within our own research and development portfolio. The impact of the synthetic procedure is reenforced by the green Process Mass Intensity and Total Carbon Release
SIMPD: an algorithm for generating simulated time splits for validating machine learning approaches.
No full textTime-split cross-validation is broadly recognized as the gold standard for validating predictive models intended for use in medicinal chemistry projects. Unfortunately this type of data is not broadly available outside of large pharmaceutical research organizations. Here we introduce the SIMPD (simulated medicinal chemistry project data) algorithm to split public data sets into training and test sets that mimic the differences observed in real-world medicinal chemistry project data sets. SIMPD uses a multi-objective genetic algorithm with objectives derived from an extensive analysis of the differences between early and late compounds in more than 130 lead-optimization projects run within the Novartis Institutes for BioMedical Research. Applying SIMPD to the real-world data sets produced training/test splits which more accurately reflect the differences in properties and machine-learning performance observed for temporal splits than other standard approaches like random or neighbor splits. We applied the SIMPD algorithm to bioactivity data extracted from ChEMBL and created 99 public data sets which can be used for validating machine-learning models intended for use in the setting of a medicinal chemistry project. The SIMPD code and simulated data sets are available under open-source/open-data licenses at github.com/rinikerlab/molecular_time_series
Secukinumab versus guselkumab in the complete resolution of ustekinumab-resistant psoriatic plaques: The ARROW study
No full textInterleukin (IL)-23–independent IL-17A production has been suggested to be involved in persistent manifestations of psoriatic disease, including anti–IL-12/23–refractory psoriatic plaques; this study aimed to test this hypothesis by investigating the clinical and molecular effects of direct IL-17A (with secukinumab) versus selective IL-23 inhibition (with guselkumab) in patients with anti–IL-12/23 (ustekinumab)–refractory psoriatic plaques. A 16-week, randomized, open-label, parallel-group, Phase IIa study (ARROW, NCT03553823) was conducted in patients with ≥1 active psoriatic plaque (total clinical score [TCS] ≥6) at screening despite treatment with ustekinumab, and a Psoriasis Area and Severity Index (PASI) score 1–10. Patients were randomized 1:1 to receive secukinumab 300 mg (n = 20) or guselkumab 100 mg (n = 20). Biopsies from one refractory (‘target plaque’) were obtained at baseline and Week 16. The primary endpoint was the proportion of patients whose ustekinumab-refractory target plaque achieved clear/almost clear status (TCS 0–2) at Week 16. Transcriptomic and histological analyses were conducted on target plaques to determine the molecular effects of direct IL-17A versus selective IL-23 inhibition. At Week 16, target plaque clear/almost clear status was achieved in 60.0% of patients treated with secukinumab versus 40.0% of patients treated with guselkumab (p = 0.1715). Molecular analyses identified that secukinumab modulated a greater proportion of psoriasis disease transcriptome genes (72.1% vs. 48.0%) and resulted in more histological responders (72.2% vs. 53.3%) compared with guselkumab. Secukinumab demonstrated a greater clinical and molecular effect on ustekinumab-refractory psoriatic plaques versus guselkumab. These results are consistent with the hypothesis that IL-23–independent IL-17 mechanisms may be relevant to the inflammation driving refractory manifestations of psoriasis
"Where chemocatalysis meets biocatalysis ... in water. Nature–ally!"
No full textChemoenzymatic catalysis, by definition, involves the merging of sequential reactions using both chemocatalysis and biocatalysis, oftentimes in a single reaction vessel. Several creative approaches and solutions that enable such combinations to be realized and applied to synthetic problems are presented herein, reflecting the state-of-the-art in this blossoming field. Coverage has been sectioned into three parts: (1) the initial discussion addresses historical developments that led to this area of research; (2) the second portion describes key developments post initial discoveries that have advanced this area, and (3) the third section presents reports covering the literature from roughly 2018 to the present, highlighting the latest achievements that provide attractive solutions to the main question of compatibility between biocatalysis (used predominantly in aqueous media), and chemocatalysis (that remains mainly performed in organic solvents). The discussion concludes with a brief overview of our follow up “review” in this 2-part series, where the realities of chemoenzymatic catalysis, both today and in the near future, as seen by several company representatives, are offered as insight as to where the field stands, and the prognosis for its future in organic synthesis