P300 Modulates Endothelial Mechanotransduction of Fluid Shear Stress.

Purpose: P300 is a lysine acetyltransferase that plays a significant role in regulating transcription and the nuclear acetylome. While P300 has been shown to be required for the transcription of certain early flow responsive genes, relatively little is known about its role in the endothelial response to hemodynamic fluid stress. Here we sought to define the role of P300 in mechanotransduction of fluid shear stress in the vascular endothelium.

Dysfunctional mechanotransduction regulates the progression of PIK3CA-driven vascular malformations.

Somatic activating mutations in are common drivers of vascular and lymphatic malformations. Despite common biophysical signatures of tissues susceptible to lesion formation, including compliant extracellular matrix and low rates of perfusion, lesions vary in clinical presentation from localized cystic dilatation to diffuse and infiltrative vascular dysplasia. The mechanisms driving the differences in disease severity and variability in clinical presentation and the role of the biophysical microenvironment in potentiating progression are poorly understood.

Donor-Derived Engineered Microvessels for Cardiovascular Risk Stratification of Patients with Kidney Failure.

Cardiovascular disease is the cause of death in ≈50% of hemodialysis patients. Accumulation of uremic solutes in systemic circulation is thought to be a key driver of the endothelial dysfunction that underlies elevated cardiovascular events. A challenge in understanding the mechanisms relating chronic kidney disease to cardiovascular disease is the lack of in vitro models that allow screening of the effects of the uremic environment on the endothelium.

Identification and Optimization of RNA-Splicing Modulators as Huntingtin Protein-Lowering Agents for the Treatment of Huntington's Disease.

Huntington's disease (HD) is caused by an expanded CAG trinucleotide repeat in exon 1 of the huntingtin () gene. We report the design of a series of pre-mRNA splicing modulators that lower huntingtin (HTT) protein, including the toxic mutant huntingtin (mHTT), by promoting insertion of a pseudoexon containing a premature termination codon at the exon 49-50 junction. The resulting transcript undergoes nonsense-mediated decay, leading to a reduction of mRNA transcripts and protein levels.

Patient-derived extracellular matrix demonstrates role of COL3A1 in blood vessel mechanics.

Vascular Ehlers-Danlos Syndrome (vEDS) is a rare autosomal dominant disease caused by mutations in the COL3A1 gene, which renders patients susceptible to aneurysm and arterial dissection and rupture. To determine the role of COL3A1 variants in the biochemical and biophysical properties of human arterial ECM, we developed a method for synthesizing ECM directly from vEDS donor fibroblasts. We found that the protein content of the ECM generated from vEDS donor fibroblasts differed significantly from ECM from healthy donors, including upregulation of collagen subtypes and other proteins related to ECM structural integrity.

Microphysiological model of PIK3CA-driven vascular malformations reveals a role of dysregulated Rac1 and mTORC1/2 in lesion formation.

Somatic activating mutations of are associated with development of vascular malformations (VMs). Here, we describe a microfluidic model of -driven VMs consisting of human umbilical vein endothelial cells expressing activating mutations embedded in three-dimensional hydrogels. We observed enlarged, irregular vessel phenotypes and the formation of cyst-like structures consistent with clinical signatures and not previously observed in cell culture models.

Developing HDAC4-Selective Protein Degraders To Investigate the Role of HDAC4 in Huntington's Disease Pathology.

Huntington's disease (HD) is a lethal autosomal dominant neurodegenerative disorder resulting from a CAG repeat expansion in the huntingtin () gene. The product of translation of this gene is a highly aggregation-prone protein containing a polyglutamine tract >35 repeats (mHTT) that has been shown to colocalize with histone deacetylase 4 (HDAC4) in cytoplasmic inclusions in HD mouse models. Genetic reduction of HDAC4 in an HD mouse model resulted in delayed aggregation of mHTT, along with amelioration of neurological phenotypes and extended lifespan.

Microfluidic and Organ-on-a-Chip Approaches to Investigate Cellular and Microenvironmental Contributions to Cardiovascular Function and Pathology.

Over the past decade, advances in microfabrication and biomaterials have facilitated the development of microfluidic tissue and organ models to address challenges with conventional animal and cell culture systems. These systems have largely been developed for human disease modeling and preclinical drug development and have been increasingly used to understand cellular and molecular mechanisms, particularly in the cardiovascular system where the characteristic mechanics and architecture are difficult to recapitulate in traditional systems. Here, we review recent microfluidic approaches to model the cardiovascular system and novel insights provided by these systems.

Ultrasensitive quantitative measurement of huntingtin phosphorylation at residue S13.

Huntington's disease (HD) is a progressive neurodegenerative disorder caused by an expansion of a CAG triplet repeat (encoding for a polyglutamine tract) within the first exon of the huntingtin gene. Expression of the mutant huntingtin (mHTT) protein can result in the production of N-terminal fragments with a robust propensity to form oligomers and aggregates, which may be causally associated with HD pathology. Several lines of evidence indicate that N17 phosphorylation or pseudophosphorylation at any of the residues T3, S13 or S16, alone or in combination, modulates mHTT aggregation, subcellular localization and toxicity.

Structure-guided Discovery of Dual-recognition Chemibodies.

Small molecules and antibodies each have advantages and limitations as therapeutics. Here, we present for the first time to our knowledge, the structure-guided design of "chemibodies" as small molecule-antibody hybrids that offer dual recognition of a single target by both a small molecule and an antibody, using DPP-IV enzyme as a proof of concept study. Biochemical characterization demonstrates that the chemibodies present superior DPP-IV inhibition compared to either small molecule or antibody component alone.

What service users with psychotic disorders want in a mental health crisis or relapse: thematic analysis of joint crisis plans.

Purpose: Recent legislation and guidance in England emphasises the importance of service user choice in care planning. However, it is not obvious how best to facilitate choices in care planning, and some clinicians are concerned that service users may make 'unwise' decisions. This study aimed to examine mental health service users' preferences and priorities in the event of a future mental health crisis or relapse.

Disubstituted 1-aryl-4-aminopiperidine library synthesis using computational drug design and high-throughput batch and flow technologies.

A platform that incorporates computational library design, parallel solution-phase synthesis, continuous flow hydrogenation, and automated high throughput purification and reformatting technologies was applied to the production of a 120-member library of 1-aryl-4-aminopiperidine analogues for drug discovery screening. The application described herein demonstrates the advantages of computational library design coupled with a flexible, modular approach to library synthesis. The enabling technologies described can be readily adopted by the traditional medicinal chemist without extensive training and lengthy process development times.

N-substituted azaindoles as potent inhibitors of Cdc7 kinase.

Cdc7 kinase is responsible for the initiation and regulation of DNA replication and has been proposed as a target for cancer therapy. We have identified a class of Cdc7 inhibitors based on a substituted indole core. Synthesis of focused indole and azaindole analogs yielded potent and selective 5-azaindole Cdc7 inhibitors with improved intrinsic metabolic stability (ie 36).

2-Phenylamino-6-cyano-1H-benzimidazole-based isoform selective casein kinase 1 gamma (CK1γ) inhibitors.

Screening of the Amgen compound library led to the identification of 2-phenylamino-6-cyano-1H-benzimidazole 1a as a potent CK1 gamma inhibitor with excellent kinase selectivity and unprecedented CK1 isoform selectivity. Further structure-based optimization of this series resulted in the discovery of 1h which possessed good enzymatic and cellular potency, excellent CK1 isoform and kinase selectivity, and acceptable pharmacokinetic properties.

Admission hyperglycemia and acute myocardial infarction: outcomes and potential therapies for diabetics and nondiabetics.

Hyperglycemia, in both diabetic and nondiabetic patients, has a significant negative impact on the morbidity and mortality of patients presenting with an acute myocardial infarction (AMI). Contemporary evidence indicates that persistent hyperglycemia after initial hospital admission continues to exert negative effects on AMI patients. There have been a number of studies demonstrating the benefit of tight glucose control in patients presenting with AMI, but a lack of convincing clinical data has led to loose guidelines and poor implementation of glucose targets for this group of patients.

Rapid development of piperidine carboxamides as potent and selective anaplastic lymphoma kinase inhibitors.

Piperidine carboxamide 1 was identified as a novel inhibitor of anaplastic lymphoma kinase (ALK enzyme assay IC(50) = 0.174 μM) during high throughput screening, with selectivity over the related kinase insulin-like growth factor-1 (IGF1R). The X-ray cocrystal structure of 1 with the ALK kinase domain revealed an unusual DFG-shifted conformation, allowing access to an extended hydrophobic pocket.

Discovery of potent and highly selective thienopyridine Janus kinase 2 inhibitors.

Developing Janus kinase 2 (Jak2) inhibitors has become a significant focus for small molecule drug discovery programs in recent years due to the identification of a Jak2 gain-of-function mutation in the majority of patients with myeloproliferative disorders (MPD). Here, we describe the discovery of a thienopyridine series of Jak2 inhibitors that culminates with compounds showing 100- to >500-fold selectivity over the related Jak family kinases in enzyme assays. Selectivity for Jak2 was also observed in TEL-Jak cellular assays, as well as in cytokine-stimulated peripheral blood mononuclear cell (PBMC) and whole blood assays.

Evaluation of a commercial packed bed flow hydrogenator for reaction screening, optimization, and synthesis.

The performance of the ThalesNano H-Cube(®), a commercial packed bed flow hydrogenator, was evaluated in the context of small scale reaction screening and optimization. A model reaction, the reduction of styrene to ethylbenzene through a 10% Pd/C catalyst bed, was used to examine performance at various pressure settings, over sequential runs, and with commercial catalyst cartridges. In addition, the consistency of the hydrogen flow was indirectly measured by in-line UV spectroscopy.

Discovery of pyridazinopyridinones as potent and selective p38 mitogen-activated protein kinase inhibitors.

The p38 mitogen-activated protein kinase (MAPK) plays an important role in the production of proinflammatory cytokines, making it an attractive target for the treatment of various inflammatory diseases. A series of pyridazinopyridinone compounds were designed as novel p38 kinase inhibitors. A structure-activity investigation identified several compounds possessing excellent potency in both enzyme and human whole blood assays.

Structure-activity relationship (SAR) investigations of tetrahydroquinolines as BKCa agonists.

The membrane bound large-conductance, calcium-activated potassium channel (BKCa) is an important regulator of neuronal activity. Here we describe the identification and structure-activity relationship of a novel class of potent tetrahydroquinoline BKCa agonists. An example from this class of BKCa agonists was shown to depress the spontaneous neuronal discharges in an electrophysiological model of migraine.

Selective inhibitors of the mutant B-Raf pathway: discovery of a potent and orally bioavailable aminoisoquinoline.

The discovery and optimization of a novel series of aminoisoquinolines as potent, selective, and efficacious inhibitors of the mutant B-Raf pathway is presented. The N-linked pyridylpyrimidine benzamide 2 was identified as a potent, modestly selective inhibitor of the B-Raf enzyme. Replacement of the benzamide with an aminoisoquinoline core significantly improved kinase selectivity and imparted favorable pharmacokinetic properties, leading to the identification of 1 as a potent antitumor agent in xenograft models.

Design and synthesis of peripherally restricted transient receptor potential vanilloid 1 (TRPV1) antagonists.

Transient receptor potential vanilloid 1 (TRPV1) channel antagonists may have clinical utility for the treatment of chronic nociceptive and neuropathic pain. We recently advanced a TRPV1 antagonist, 3 (AMG 517), into clinical trials as a new therapy for the treatment of pain. However, in addition to the desired analgesic effects, this TRPV1 antagonist significantly increased body core temperature following oral administration in rodents.

4-Aminopyrimidine tetrahydronaphthols: a series of novel vanilloid receptor-1 antagonists with improved solubility properties.

8-(6-(4-(Trifluoromethyl)phenyl)pyrimidin-4-ylamino)-1,2,3,4-tetrahydronaphthalen-2-ol (4) and analogs (5-10) were shown to be potent inhibitors of human and rat TRPV1 in vitro with increased solubility over our previous series. Synthesis, SAR, and improvements in metabolic stability and absorption of these compounds are described herein.