Thermodynamic and structural analysis of peptide- and allele-dependent properties of two HLA-B27 subtypes exhibiting differential disease association

Selected HLA-B27 subtypes are associated with spondyloarthropathies, but the underlying mechanism is not understood. To explain this association in molecular terms, a comparison of peptide-dependent dynamic and structural properties of the differentially disease-associated subtypes HLA-B*2705 and HLA-B*2709 was carried out. These molecules differ only by a single amino acid at the floor of the peptide binding groove. The thermostabilities of a series of HLA-B27 molecules complexed with nonameric and decameric peptides were determined and revealed substantial differences depending on the subtype as well as the residues at the termini of the peptides. In addition we present the crystal structure of the B*2709 subtype complexed with a decameric peptide. This structure provides an explanation for the preference of HLA-B27 for a peptide with an N-terminal arginine as secondary anchor and the lack of preference for tyrosine as peptide C terminus in B*2709. The data show that differences in thermodynamic properties between peptide-complexed HLA-B27 subtypes are correlated with a variety of structural properties

Design, Synthesis, and Pharmacological Characterization of a Neutral, Non Prodrug Thrombin Inhibitor with Good Oral Pharmacokinetics

Despite extensive research on small molecule thrombin inhibitors for oral application in the past decades, only a single double prodrug with very modest oral bioavailability has reached human therapy as a marketed drug. We have undertaken major efforts to identify neutral, non prodrug inhibitors. Using a holistic analysis of all available internal data, we were able to build computational models and apply these for the selection of a lead series with the highest possibility of achieving oral bioavailability. In our design, we relied on protein structure knowledge to address potency and identified a small window of favorable physicochemical properties to balance absorption and metabolic stability. Protein structure information on the pregnane X receptor helped in overcoming a persistent cytochrome P450 3A4 induction problem. The selected compound series was optimized to a highly potent, neutral, non prodrug thrombin inhibitor by designing, synthesizing, and testing derivatives. The resulting optimized compound, BAY1217224, has reached first clinical trials, which have confirmed the desired pharmacokinetic propertie

BAY 069, a Novel Trifluoromethyl pyrimidinedione Based BCAT1 2 Inhibitor and Chemical Probe

[Image: see text] The branched-chain amino acid transaminases (BCATs) are enzymes that catalyze the first reaction of catabolism of the essential branched-chain amino acids to branched-chain keto acids to form glutamate. They are known to play a key role in different cancer types. Here, we report a new structural class of BCAT1/2 inhibitors, (trifluoromethyl)pyrimidinediones, identified by a high-throughput screening campaign and subsequent optimization guided by a series of X-ray crystal structures. Our potent dual BCAT1/2 inhibitor BAY-069 displays high cellular activity and very good selectivity. Along with a negative control (BAY-771), BAY-069 was donated as a chemical probe to the Structural Genomics Consortium

Fab Mor03268 Triggers Absorption Shift of a Diagnostic Dye Via Packaging in a Solvent Shielded Fab Dimer Interface

Molecular interactions between near IR fluorescent probes and specific antibodies may be exploited to generate novel smart probes for diagnostic imaging. Using a new phage display technology, we developed such antibody Fab fragments with subnanomolar binding affinity for tetrasulfocyanine, a near IR in vivo imaging agent. Unexpectedly, some Fabs induced redshifts of the dye absorption peak of up to 44 nm. This is the largest shift reported for a biological system so far. Crystal structure determination and absorption spectroscopy in the crystal in combination with microcalorimetry and small angle X ray scattering in solution revealed that the redshift is triggered by formation of a Fab dimer, with tetrasulfocyanine being buried in a fully closed protein cavity within the dimer interface. The derived principle of shifting the absorption peak of a symmetric dye via packaging within a Fab dimer interface may be transferred to other diagnostic fluorophores, opening the way towards smart imaging probes that change their wavelength upon interaction with an antibod

Computationally Empowered Workflow Identifies Novel Covalent Allosteric Binders for KRASG12C

Due to its frequent mutations in multiple lethal cancers, KRAS is one of the most studied anticancer targets nowadays. Since the discovery of the druggable allosteric binding site containing a G12C mutation, KRASG12C has been the focus of attention in oncology research. We report here a computationally driven approach aimed at identifying novel and selective KRASG12C covalent inhibitors. The workflow involved initial enumeration of virtual molecules tailored for the KRAS allosteric binding site. Tools such as pharmacophore modeling, docking, and free energy perturbations were deployed to prioritize the compounds with the best profiles. The synthesized naphthyridinone scaffold showed the ability to react with G12C and inhibit KRASG12C. Analogues were prepared to establish structure activity relationships, while molecular dynamics simulations and crystallization of the inhibitor KRASG12C complex highlighted an unprecedented binding mod

Discovery and Characterization of the Potent and Selective P2X4 InhibitorN [4 3 Chlorophenoxy 3 sulfamoylphenyl] 2 phenylacetamide BAY 1797 and Structure Guided Amelioration of Its CYP3A4 Induction Profile

The P2X4 receptor is a ligand gated ion channel that is expressed on a variety of cell types, especially those involved in inflammatory and immune processes. High throughput screening led to a new class of P2X4 inhibitors with substantial CYP 3A4 induction in human hepatocytes. A structure guided optimization with respect to decreased pregnane X receptor PXR binding was started. It was found that the introduction of larger and more polar substituents on the ether linker led to less PXR binding while maintaining the P2X4 inhibitory potency. This translated into significantly reduced CYP 3A4 induction for compounds 71 and 73. Unfortunately, the in vivo pharmacokinetic PK profiles of these compounds were insufficient for the desired profile in humans. However, BAY 1797 10 was identified and characterized as a potent and selective P2X4 antagonist. This compound is suitable for in vivo studies in rodents, and the anti inflammatory and anti nociceptive effects of BAY 1797 were demonstrated in a mouse complete Freund s adjuvant CFA inflammatory pain mode

A Novel NAMPT Inhibitor Based Antibody Drug Conjugate Payload Class for Cancer Therapy

Inhibition of intracellular nicotinamide phosphoribosyltransferase NAMPT represents a new mode of action for cancer targeting antibody drug conjugates ADCs with activity also in slowly proliferating cells. To extend the repertoire of available effector chemistries, we have developed a novel structural class of NAMPT inhibitors as ADC payloads. A structure activity relationship driven approach supported by protein structural information was pursued to identify a suitable attachment point for the linker to connect the NAMPT inhibitor with the antibody. Optimization of scaffolds and linker structures led to highly potent effector chemistries which were conjugated to antibodies targeting C4.4a LYPD3 , HER2 c erbB2 , or B7H3 CD276 and tested on antigen positive and negative cancer cell lines. Pharmacokinetic studies, including metabolite profiling, were performed to optimize the stability and selectivity of the ADCs and to evaluate potential bystander effects. Optimized NAMPTi ADCs demonstrated potent in vivo antitumor efficacy in target antigen expressing xenograft mouse models. This led to the development of highly potent NAMPT inhibitor ADCs with a very good selectivity profile compared with the corresponding isotype control ADCs. Moreover, we demonstrate to our knowledge for the first time the generation of NAMPTi payload metabolites from the NAMPTi ADCs in vitro and in vivo. In conclusion, NAMPTi ADCs represent an attractive new payload class designed for use in ADCs for the treatment of solid and hematological cancer