Linking ATP and allosteric sites to achieve superadditive binding with bivalent EGFR kinase inhibitors.

Bivalent molecules consisting of groups connected through bridging linkers often exhibit strong target binding and unique biological effects. However, developing bivalent inhibitors with the desired activity is challenging due to the dual motif architecture of these molecules and the variability that can be introduced through differing linker structures and geometries. We report a set of alternatively linked bivalent EGFR inhibitors that simultaneously occupy the ATP substrate and allosteric pockets. Crystal structures show that initial and redesigned linkers bridging a trisubstituted imidazole ATP-site inhibitor and dibenzodiazepinone allosteric-site inhibitor proved successful in spanning these sites. The re-engineered linker yielded a compound that exhibited significantly higher potency (~60 pM) against the drug-resistant EGFR L858R/T790M and L858R/T790M/C797S, which was superadditive as compared with the parent molecules. The enhanced potency is attributed to factors stemming from the linker connection to the allosteric-site group and informs strategies to engineer linkers in bivalent agent design.

Pitfalls and Considerations in Determining the Potency and Mutant Selectivity of Covalent Epidermal Growth Factor Receptor Inhibitors.

Enzyme inhibitors that form covalent bonds with their targets are being increasingly pursued in drug development. Assessing their biochemical activity relies on time-dependent assays, which are distinct and more complex compared with methods commonly employed for reversible-binding inhibitors. To provide general guidance to the covalent inhibitor development community, we explored methods and reported kinetic values and experimental factors in determining the biochemical activity of various covalent epidermal growth factor receptor (EGFR) inhibitors. We showcase how liquid handling and assay reagents impact kinetic parameters and potency interpretations, which are critical for structure-kinetic relationships and covalent drug design. Additionally, we include benchmark kinetic values with reference inhibitors, which are imperative, as covalent EGFR inhibitor kinetic values are infrequently consistent in the literature. This overview seeks to inform best practices for developing new covalent inhibitors and highlight appropriate steps to address gaps in knowledge presently limiting assay reliability and reproducibility.

Linking ATP and allosteric sites to achieve superadditive binding with bivalent EGFR kinase inhibitors.

Bivalent molecules consisting of groups connected through bridging linkers often exhibit strong target binding and unique biological effects. However, developing bivalent inhibitors with the desired activity is challenging due to the dual motif architecture of these molecules and the variability that can be introduced through differing linker structures and geometries. We report a set of alternatively linked bivalent EGFR inhibitors that simultaneously occupy the ATP substrate and allosteric pockets. Crystal structures show that initial and redesigned linkers bridging a trisubstituted imidazole ATP-site inhibitor and dibenzodiazepinone allosteric-site inhibitor proved successful in spanning these sites. The reengineered linker yielded a compound that exhibited significantly higher potency (~60 pM) against the drug-resistant EGFR L858R/T790M and L858R/T790M/C797S, which was superadditive as compared with the parent molecules. The enhanced potency is attributed to factors stemming from the linker connection to the allosteric-site group and informs strategies to engineer linkers in bivalent agent design.

Synthesis, Structure, Hirshfeld Surface Analysis, Non-Covalent Interaction, and In Silico Studies of 4-Hydroxy-1-[(4-Nitrophenyl)Sulfonyl]Pyrrolidine-2-Carboxyllic Acid.

The new compound 4-hydroxy-1-[(4-nitrophenyl)sulfonyl]pyrrolidine-2-carboxyllic acid was obtained by the reaction of 4-hydroxyproline with 4-nitrobenzenesulfonyl chloride. The compound was characterized using single crystal X-ray diffraction studies. Spectroscopic methods including NMR, FTIR, ES-MS, and UV were employed for further structural analysis of the synthesized compound. The title compound was found to have crystallized in an orthorhombic crystal system with space group P212121. The S1-N1 bond length of 1.628 (2) Å was a strong indication of the formation of the title compound. The absence of characteristic downfield 1H NMR peak of pyrrolidine ring and the presence of S-N stretching vibration at 857.82 cm-1 on the FTIR are strong indications for the formation of the sulfonamide. The experimental study was complemented with computations at the B3LYP/6-311G + + (d,p) level of theory to gain more understanding of interactions in the compound at the molecular level. Noncovalent interaction, Hirsfeld surface analysis and interaction energy calculations were employed in the analysis of the supramolecular architecture of the compound. Predicted ADMET parameters, awarded suitable bioavailability credentials, while the molecular docking study indicated that the compound enchants promising inhibition prospects against dihydropteroate synthase, DNA topoisomerase, and SARS-CoV-2 spike. Graphical Abstract: Herein we present the solid state structure, noncovalent interaction and spectroscopic analysis of a prospective bioactive compound 4-hydroxy-1-[(4-nitrophenyl)sulphonyl]pyrrolidine-2-carboxyllic acid. Supplementary Information: The online version contains supplementary material available at 10.1007/s10870-023-00978-0.

Structural Basis for Inhibition of Mutant EGFR with Lazertinib (YH25448).

Lazertinib (YH25448) is a novel third-generation tyrosine kinase inhibitor (TKI) developed as a treatment for EGFR mutant non-small cell lung cancer. To better understand the nature of lazertinib inhibition, we determined crystal structures of lazertinib in complex with both WT and mutant EGFR and compared its binding mode to that of structurally related EGFR TKIs. We observe that lazertinib binds EGFR with a distinctive pyrazole moiety enabling hydrogen bonds and van der Waals interactions facilitated through hydrophilic amine and hydrophobic phenyl groups, respectively. Biochemical assays and cell studies confirm that lazertinib effectively targets EGFR(L858R/T790M) and to a lesser extent HER2. The molecular basis for lazertinib inhibition of EGFR reported here highlights previously unexplored binding interactions leading to improved medicinal chemistry properties compared to clinically approved osimertinib (AZD9291) and offers novel strategies for structure-guided design of tyrosine kinase inhibitors.

Structure and Computational Studies of New Sulfonamide Compound: {(4-nitrophenyl)sulfonyl}tryptophan.

Synthesis of sulfonamide through an indirect method that avoids contamination of the product with no need for purification has been carried out using the indirect process. Here, we report the synthesis of a novel sulfonamide compound, ({4-nitrophenyl}sulfonyl)tryptophan (DNSPA) from 4-nitrobenzenesulphonylchloride and L-tryptophan precursors. The slow evaporation method was used to form single crystals of the named compound from methanolic solution. The compound was characterized by X-ray crystallographic analysis and spectroscopic methods (NMR, IR, mass spectrometry, and UV-vis). The sulfonamide N-H NMR signal at 8.07-8.09 ppm and S-N stretching vibration at 931 cm-1 indicate the formation of the target compound. The compound crystallized in the monoclinic crystal system and P21 space group with four molecules of the compound in the asymmetric unit. Molecular aggregation in the crystal structure revealed a 12-molecule aggregate synthon sustained by O-H⋯O hydrogen bonds and stabilised by N-H⋯O intermolecular contacts. Experimental studies were complemented by DFT calculations at the B3LYP/6-311++G(d,p) level of theory. The computed structural and spectroscopic data are in good agreement with those obtained experimentally. The energies of interactions between the units making up the molecule were calculated. Molecular docking studies showed that DNSPA has a binding energy of -6.37 kcal/mol for E. coli DNA gyrase (5MMN) and -6.35 kcal/mol for COVID-19 main protease (6LU7).

Architecture of the NADPH oxidase family of enzymes.

The NADPH Oxidases (NOX) catalyze the deliberate production of reactive oxygen species (ROS) and are established regulators of redox-dependent processes across diverse biological settings. Proper management of their activity is controlled through a conserved electron transfer (ET) cascade from cytosolic NADPH substrate through the plasma membrane to extracellular O2. After decades-long investigations of their biological functions, including potential as drug targets, only very recently has atomic-resolution information of NOX enzymes been made available. In this graphical review, we summarize the present structural biology understanding of the NOX enzymes afforded by X-ray crystallography and cryo-electron microscopy. Combined molecular-level insights predominantly informed by DUOX1 full-length Cryo-EM structures suggest a general structural basis for the control of their catalytic activity by intracellular domain-domain stabilization.

Polypharmacology of some medicinal plant metabolites against SARS-CoV-2 and host targets: Molecular dynamics evaluation of NSP9 RNA binding protein.

Medicinal plants as rich sources of bioactive compounds are now being explored for drug development against COVID-19. 19 medicinal plants known to exhibit antiviral and anti-inflammatory effects were manually curated, procuring a library of 521 metabolites; this was virtually screened against NSP9, including some other viral and host targets and were evaluated for polypharmacological indications. Leads were identified via rigorous scoring thresholds and ADMET filtering. MM-GBSA calculation was deployed to select NSP9-Lead complexes and the complexes were evaluated for their stability and protein-ligand communication via MD simulation. We identified 5 phytochemical leads for NSP9, 23 for Furin, 18 for ORF3a, and 19 for IL-6. Ochnaflavone and Licoflavone B, obtained from Lonicera japonica (Japanese Honeysuckle) and Glycyrrhiza glabra (Licorice), respectively, were identified to have the highest potential polypharmacological properties for the aforementioned targets and may act on multiple pathways simultaneously to inhibit viral entry, replication, and disease progression. Additionally, MD simulation supports the robust stability of Ochnaflavone and Licoflavone B against NSP9 at the active sites via hydrophobic interactions, H-bonding, and H-bonding facilitated by water. This study promotes the initiation of further experimental analysis of natural product-based anti-COVID-19 therapeutics.