Discovery of a novel series of selective macrocyclic PKCTheta inhibitors.

A series of macrocyclic PKCθ inhibitors based on a 1,3-dihydro-2H-imidazo[4,5-b]pyridin-2-one hinge binder has been studied. Different aromatic and heteroaromatic substituents have been explored in order to optimize potency, isoform selectivity as well as DMPK properties. The importance of the length of the macrocyclic linker has also been analyzed.

Identification of unknown impurities J, RRT 2.2, 2.4, 2.6 and 3.4 in tetralysal® capsules.

Tetralysal® is a Galderma oral drug product (DP) marketed for the treatment of acne. Tetralysal® is sold in capsules containing either 150 mg or 300 mg of the drug substance. In the British Pharmacopoeia monograph for Lymecycline Capsules, the impurities already specified in the drug substance (A-G), visible in the European Pharmacopoeia 〈1654〉, are also specified together with an unidentified impurity at RRT 1.

Structure-Based Optimization and Discovery of M3258, a Specific Inhibitor of the Immunoproteasome Subunit LMP7 (β5i).

Proteasomes are broadly expressed key components of the ubiquitin-dependent protein degradation pathway containing catalytically active subunits (β1, β2, and β5). LMP7 (β5i) is a subunit of the immunoproteasome, an inducible isoform that is predominantly expressed in hematopoietic cells. Clinically effective pan-proteasome inhibitors for the treatment of multiple myeloma (MM) nonselectively target LMP7 and other subunits of the constitutive proteasome and immunoproteasome with comparable potency, which can limit the therapeutic applicability of these drugs.

M3258 Is a Selective Inhibitor of the Immunoproteasome Subunit LMP7 (β5i) Delivering Efficacy in Multiple Myeloma Models.

Large multifunctional peptidase 7 (LMP7/β5i/PSMB8) is a proteolytic subunit of the immunoproteasome, which is predominantly expressed in normal and malignant hematolymphoid cells, including multiple myeloma, and contributes to the degradation of ubiquitinated proteins. Described herein for the first time is the preclinical profile of M3258; an orally bioavailable, potent, reversible and highly selective LMP7 inhibitor. M3258 demonstrated strong antitumor efficacy in multiple myeloma xenograft models, including a novel model of the human bone niche of multiple myeloma.

Establishing a systematic framework to characterise in vitro methods for human hepatic metabolic clearance.

Hepatic metabolic clearance is one of the most important factors driving the overall kinetics of chemicals including substances used in various product categories such as pesticides, biocides, pharmaceuticals, and cosmetics. A large number of in vitro systems from purified isozymes and subcellular organelles to hepatocytes in simple cultures and in complex scaffold setups are available for measuring hepatic metabolic clearance for different applications. However, there is currently no approach for systematically characterising and comparing these in vitro methods in terms of their design, applicability and performance.

Determination of low intrinsic clearance in vitro: the benefit of a novel internal standard in human hepatocyte incubations.

1. A novel method utilizing an internal standard in hepatocytes incubations has been developed and demonstrated to decrease the variability in the determination of intrinsic clearance (CL) in this system. The reduced variability was shown to allow differentiation of lower elimination rate constants from noise.

Comparison of methods for the prediction of human clearance from hepatocyte intrinsic clearance for a set of reference compounds and an external evaluation set.

1. We compared direct scaling, regression model equation and the so-called "Poulin et al." methods to scale clearance (CL) from in vitro intrinsic clearance (CL) measured in human hepatocytes using two sets of compounds.

Structure-activity relationship and properties optimization of a series of quinazoline-2,4-diones as inhibitors of the canonical Wnt pathway.

Wnt signaling pathway plays a critical role in numerous cellular processes, including tumor initiation, proliferation, invasion/infiltration, metastasis formation and resistance to chemotherapy. In a drug discovery project aimed at the identification of inhibitors of the canonical Wnt pathway, we selected a series of quinazoline 2,4-diones as starting point for the therapeutic treatment of glioblastoma multiforme. Despite of poor physico-chemical properties of hit compound 1, our medicinal chemistry effort allowed the discovery and characterization of lead compound 33 (SEN461), with improved ADME profile, good bioavailability and active in vitro and in vivo in glioblastoma, gastric and sarcoma tumors.

Expression of Tight Junction and Drug Efflux Transporter Proteins in an in vitro Model of Human Blood-Brain Barrier.

Interendothelial cell tight junctions (TJs) proteins contribute to maintain the structural and functional integrity of the blood-brain barrier (BBB) and several efflux transporters regulate transport of compounds across BBB. A unique double compartment-model of the BBB, consisting of cerebral endothelial cells isolated from cryopreserved human glial tumors, alone and in the presence of human astroglial cells derived from the same tissue preparation was established. Endothelial cell viability and transendothelial electrical resistance (TEER) were measured in this model and three representative TJ proteins - occludin (OCLN), zonula occludens-1 (ZO-1) and claudin-5 (CLN-5) - as well as several drug efflux transporters - P-glycoprotein (P-gp), multidrug resistance protein-1 and 2 (MRP-1 and MRP-2), organic anion-transporting polypeptide-1 and 3 (oatp1 and oatp3) were analyzed at both the protein and gene transcript level.

Evaluation of the novel in vitro systems for hepatic drug clearance and assessment of their predictive utility.

A valuable strategy for the assessment of in vitro systems is proposed which involves a tiered approach consisting of four levels valuable for both selection of probe compounds and designing experiments for evaluation. At level 1, a Preliminary Assessment is used to triage novel systems based on existing information generated using the methods employed in the development of the system. There is no special requirement for specific probes or experimental conditions.

Clearance-dependent underprediction of in vivo intrinsic clearance from human hepatocytes: comparison with permeabilities from artificial membrane (PAMPA) assay, in silico and caco-2 assay, for 65 drugs.

Underprediction of in vivo intrinsic clearance (CLint) from suspended human hepatocytes has recently been shown to be clearance-dependent although the mechanistic basis is currently unknown. One possible explanation is rate limiting transmembrane (passive) permeation into hepatocytes in vitro; evidence to support this has been minor to date, but there has not been a systematic exploration of the impact of passive permeability in vitro. To investigate the relationship between underprediction of in vivo CLint and potentially rate limiting permeability, permeability constants (Px, collated from published studies and determined experimentally in this study), using three alternative methodologies (parallel artificial membrane permeability assay (PAMPA), caco-2 permeability assay and calculated using an empirical model) were compared with CLint from suspended human hepatocytes for 65 drugs from a recently reported database of clearance predictions.

Comparison of cryopreserved HepaRG cells with cryopreserved human hepatocytes for prediction of clearance for 26 drugs.

Prediction of clearance in drug discovery currently relies on human primary hepatocytes, which can vary widely in drug-metabolizing enzyme activity. Potential alternative in vitro models include the HepaRG cell (from immortalized hepatoma cells), which in culture can express drug-metabolizing enzymes to an extent comparable to that of primary hepatocytes. Utility of the HepaRG cell will depend on robust performance, relative to that of primary hepatocytes, in routine high-throughput analysis.

Fruitful adrenergic α(2C)-agonism/α(2A)-antagonism combination to prevent and contrast morphine tolerance and dependence.

The functional in vitro study of the enantiomers of imidazolines 4-7 highlighted the role played by the nature of the ortho phenyl substituent in determining the preferred α(2C)-AR configuration. Indeed, the (S) enantiomers of 4-6 or (R) enantiomer of 7 behave as eutomers and activate this subtype as full agonists; the corresponding distomers are partial agonists. Because in clinical pain management with opioids α(2C)-AR agonists, devoid of the α(2A)-AR-mediated side effects, may represent an improvement over current therapies with clonidine like drugs, 4 and its enantiomers, showing α(2C)-agonism/α(2A)-antagonism, have been studied in vivo.

Novel alpha-7 nicotinic acetylcholine receptor agonists containing a urea moiety: identification and characterization of the potent, selective, and orally efficacious agonist 1-[6-(4-fluorophenyl)pyridin-3-yl]-3-(4-piperidin-1-ylbutyl) urea (SEN34625/WYE-103914).

Alpha-7 nicotinic acetylcholine receptor (alpha7 nAChR) agonists are promising therapeutic candidates for the treatment of cognitive impairment. We report a series of novel, potent small molecule agonists (4-18) of the alpha7 nAChR deriving from our continuing efforts in the areas of Alzheimer's disease and schizophrenia. One of the compounds of the series containing a urea moiety (16) was further shown to be a selective agonist of the alpha7 nAChR with excellent in vitro and in vivo profiles, brain penetration, and oral bioavailability and demonstrated in vivo efficacy in multiple behavioral cognition models.

Procognitive and neuroprotective activity of a novel alpha7 nicotinic acetylcholine receptor agonist for treatment of neurodegenerative and cognitive disorders.

The alpha7 nicotinic acetylcholine receptor (nAChR) is a promising target for treatment of cognitive dysfunction associated with Alzheimer's disease and schizophrenia. Here, we report the pharmacological properties of 5-morpholin-4-yl-pentanoic acid (4-pyridin-3-yl-phenyl)-amide [SEN12333 (WAY-317538)], a novel selective agonist of alpha7 nAChR. SEN12333 shows high affinity for the rat alpha7 receptor expressed in GH4C1 cells (K(i) = 260 nM) and acts as full agonist in functional Ca(2+) flux studies (EC(50) = 1.

Evaluation of the role of intestinal and liver metabolism in the conversion of two different ester prodrugs of sanfetrinem to the parent drug in vitro and in vivo using different rat tissues and a surgically prepared rat model.

To improve oral absorption of sanfetrinem, a broad-spectrum, beta-lactamase-stable antibiotic, two different ester prodrugs have been selected. Both prodrugs proved to be readily hydrolyzed after absorption before reaching the systemic circulation. The objective of this study was to evaluate the role of intestinal and liver metabolism in the conversion of the two prodrugs into the active compound.