Discovery of RXFP2 genetic association in resistant hypertensive men and RXFP2 antagonists for the treatment of resistant hypertension.

Hypertension remains a leading cause of cardiovascular and kidney diseases. Failure to control blood pressure with ≥ 3 medications or control requiring ≥ 4 medications is classified as resistant hypertension (rHTN) and new therapies are needed to reduce the resulting increased risk of morbidity and mortality. Here, we report genetic evidence that relaxin family peptide receptor 2 (RXFP2) is associated with rHTN in men, but not in women. This study shows that adrenal gland gene expression of RXFP2 is increased in men with hypertension and the RXFP2 natural ligand, INSL3, increases adrenal steroidogenesis and corticosteroid secretion in human adrenal cells. To address the hypothesis that RXFP2 activation is an important mechanism in rHTN, we discovered and characterized small molecule and monoclonal antibody (mAb) blockers of RXFP2. The novel chemical entities and mAbs show potent, selective inhibition of RXFP2 and reduce aldosterone and cortisol synthesis and release. The RXFP2 mAbs have suitable rat pharmacokinetic profiles to evaluate the role of RXFP2 in the development and maintenance of rHTN. Overall, we identified RXFP2 activity as a potential new mechanism in rHTN and discovered RXFP2 antagonists for the future interrogation of RXFP2 in cardiovascular and renal diseases.

Modulation of macrophage inflammatory function through selective inhibition of the epigenetic reader protein SP140.

BACKGROUND: SP140 is a bromodomain-containing protein expressed predominantly in immune cells. Genetic polymorphisms and epigenetic modifications in the SP140 locus have been linked to Crohn's disease (CD), suggesting a role in inflammation. RESULTS: We report the development of the first small molecule SP140 inhibitor (GSK761) and utilize this to elucidate SP140 function in macrophages. We show that SP140 is highly expressed in CD mucosal macrophages and in in vitro-generated inflammatory macrophages. SP140 inhibition through GSK761 reduced monocyte-to-inflammatory macrophage differentiation and lipopolysaccharide (LPS)-induced inflammatory activation, while inducing the generation of CD206+ regulatory macrophages that were shown to associate with a therapeutic response to anti-TNF in CD patients. SP140 preferentially occupies transcriptional start sites in inflammatory macrophages, with enrichment at gene loci encoding pro-inflammatory cytokines/chemokines and inflammatory pathways. GSK761 specifically reduces SP140 chromatin binding and thereby expression of SP140-regulated genes. GSK761 inhibits the expression of cytokines, including TNF, by CD14+ macrophages isolated from CD intestinal mucosa. CONCLUSIONS: This study identifies SP140 as a druggable epigenetic therapeutic target for CD.

Investigation of Janus Kinase (JAK) Inhibitors for Lung Delivery and the Importance of Aldehyde Oxidase Metabolism.

The Janus family of tyrosine kinases (JAK1, JAK2, JAK3, and TYK2) play an essential role in the receptor signaling of cytokines that have been implicated in the pathogenesis of severe asthma, and there is emerging interest in the development of small-molecule-inhaled JAK inhibitors as treatments. Here, we describe the optimization of a quinazoline series of JAK inhibitors and the results of mouse lung pharmacokinetic (PK) studies where only low concentrations of parent compound were observed. Subsequent investigations revealed that the low exposure was due to metabolism by aldehyde oxidase (AO), so we sought to identify quinazolines that were not metabolized by AO. We found that specific substituents at the quinazoline 2-position prevented AO metabolism and this was rationalized through computational docking studies in the AO binding site, but they compromised kinome selectivity. Results presented here highlight that AO metabolism is a potential issue in the lung.

The Discovery of in Vivo Active Mitochondrial Branched-Chain Aminotransferase (BCATm) Inhibitors by Hybridizing Fragment and HTS Hits.

The hybridization of hits, identified by complementary fragment and high throughput screens, enabled the discovery of the first series of potent inhibitors of mitochondrial branched-chain aminotransferase (BCATm) based on a 2-benzylamino-pyrazolo[1,5-a]pyrimidinone-3-carbonitrile template. Structure-guided growth enabled rapid optimization of potency with maintenance of ligand efficiency, while the focus on physicochemical properties delivered compounds with excellent pharmacokinetic exposure that enabled a proof of concept experiment in mice. Oral administration of 2-((4-chloro-2,6-difluorobenzyl)amino)-7-oxo-5-propyl-4,7-dihydropyrazolo[1,5-a]pyrimidine-3-carbonitrile 61 significantly raised the circulating levels of the branched-chain amino acids leucine, isoleucine, and valine in this acute study.

A phenotypic screening approach in cord blood-derived mast cells to identify anti-inflammatory compounds.

Mast cells are unique hematopoietic cells that are richly distributed in the skin and mucosal surfaces of the respiratory and gastrointestinal tract. They play a key role in allergic inflammation by releasing a cocktail of granular constituents, including histamine, serine proteases, and various eicosanoids and cytokines. As such, a number of drugs target either inhibition of mast cell degranulation or the products of degranulation. To identify potential novel drugs and mechanisms in mast cell biology, assays were developed to identify inhibitors of mast cell degranulation and activation in a phenotypic screen. Due to the challenges associated with obtaining primary mast cells, cord blood-derived mononuclear cells were reproducibly differentiated to mast cells and assays developed to monitor tryptase release and prostaglandin D2 generation. The tryptase assay was particularly sensitive, requiring only 500 cells per data point, which permitted a set of approximately 12,000 compounds to be screened robustly and cost-effectively. Active compounds were tested for concomitant inhibition of prostaglandin D2 generation. This study demonstrates the robustness and effectiveness of this approach in the identification of potential novel compounds and mechanisms targeting mast cell-driven inflammation, to enable innovative drug discovery efforts to be prosecuted.

Small-molecule modulation of cellular chaperones to treat protein misfolding disorders.

The correct folding of proteins is a fundamental process in the normal physiological functioning of cells, and is mediated by cellular chaperones including members of the Hsp70 family. Many diseases are caused by a failure of cellular chaperones to adequately maintain correct protein folding, and has led to the development of a therapeutic strategy to upregulate the activity of cellular chaperones in order to ameliorate intrinsic folding deficits. A large range of pharmacological agents that can induce cellular chaperones and correct deficits associated with misfolded proteins are known. This review surveys the mechanisms and compounds that have been used to modulate cellular chaperones, and discusses the continuing challenges in translating this approach into clinical improvements in the treatment of protein misfolding disorders.

Lanthanide reagents in solid phase synthesis.

From solid-supported ytterbium(III) catalysts to linkers cleaved by electron transfer from samarium(II) species, lanthanide reagents are beginning to find widespread application in solid phase organic synthesis. This tutorial review introduces the use of lanthanide(III) Lewis acids and lanthanide(IV) oxidants in solid phase chemistry before concentrating on the growing use of lanthanide(II) reagents in the area.

Modeled regional climate change and California endemic oak ranges.

In the coming century, anthropogenic climate change will threaten the persistence of restricted endemic species, complicating conservation planning. Although most efforts to quantify potential shifts in species' ranges use global climate model (GCM) output, regional climate model (RCM) output may be better suited to predicting shifts by restricted species, particularly in regions with complex topography or other regionally important climate-forcing factors. Using a RCM-based future climate scenario, we found that potential ranges of two California endemic oaks, Quercus douglasii and Quercus lobata, shrink considerably (to 59% and 54% of modern potential range sizes, respectively) and shift northward. This result is markedly different from that obtained by using a comparable GCM-based scenario, under which these species retain 81% and 73% of their modern potential range sizes, respectively. The difference between RCM- and GCM-based scenarios is due to greater warming and larger precipitation decreases during the growing season predicted by the RCM in these species' potential ranges. Based on the modeled regional climate change, <50% of protected land area currently containing these species is expected to contain them under a future midrange "business-as-usual" path of greenhouse gas emissions.

Could CO(2)-induced land-cover feedbacks alter near-shore upwelling regimes?

The response of marine and terrestrial environments to global changes in atmospheric carbon dioxide (CO(2)) concentrations will likely be governed by both responses to direct environmental forcing and responses to Earth-system feedbacks induced by that forcing. It has been proposed that anthropogenic greenhouse forcing will intensify coastal upwelling in eastern boundary current regions [Bakun, A. (1990) Science 247, 198-201]. Focusing on the California Current, we show that biophysical land-cover-atmosphere feedbacks induced by CO(2) radiative forcing enhance the radiative effects of CO(2) on land-sea thermal contrast, resulting in changes in eastern boundary current total seasonal upwelling and upwelling seasonality. Specifically, relative to CO(2) radiative forcing, land-cover-atmosphere feedbacks lead to a stronger increase in peak- and late-season near-shore upwelling in the northern limb of the California Current and a stronger decrease in peak- and late-season near-shore upwelling in the southern limb. Such changes will impact both marine and terrestrial communities [Bakun, A. (1990) Science 247, 198-201; Soto, C. G. (2001) Rev. Fish Biol. Fish. 11, 181-195; and Agostini, V. N. & Bakun, A. (2002) Fish. Oceanogr. 11, 129-142], and these and other Earth-system feedbacks should be expected to play a substantial role in shaping the response of eastern boundary current regions to CO(2) radiative forcing.

Chicxulub and climate: radiative perturbations of impact-produced S-bearing gases.

We use one-dimensional (1D) atmospheric models coupled to a sulfate aerosol model to investigate climate forcing and short-term response to stratospheric sulfate aerosols produced by the reaction of S-bearing gases and water vapor released in the Chicxulub impact event. A 1D radiation model is used to assess the climate forcing due to the impact-related loading of S-bearing gases. The model suggests that a climate forcing 100 times larger than that from the Pinatubo volcanic eruption is associated with the Chicxulub impact event for at least 2 years after the impact. In particular, we find a saturation effect in the forcing, that is, there is no significant difference in the maximum forcing between the highest (approximately 300 Gt) and lowest (approximately 30 Gt) estimated stratospheric S-loading from the Chicxulub impact. However, higher S-loads increase the overall duration of the forcing by several months. We use a single column model for a preliminary investigation of the short-term climate response to the impact-related production of sulfate aerosols (the lack of horizontal feedbacks limits the usefulness of the single column model to the first few days after the impact). Compared with the present steady-state climate, the introduction of large amounts of sulfate aerosols in the stratosphere results in a significant cooling of the Earth's surface. A long-term climate response can only be investigated with the use of a three-dimensional atmospheric model, which allows for the atmospheric circulation to adjust to the perturbation. Overall, although the climate perturbation to the forcing appears to be relatively large, the geologic record shows no sign of a significant long-term climatic shift across the K/T boundary, which is indicative of a fast post-impact climatic recovery.