Optimization of M3 Antagonist-PDE4 Inhibitor (MAPI) Dual Pharmacology Molecules for the Treatment of COPD.

Aiming at the inhaled treatment of pulmonary diseases, the optimization process of the previously reported MAPI compound 92a is herein described. The project was focused on overcoming the chemical stability issue and achieving a balanced bronchodilator/anti-inflammatory profile in rats in order to be confident in a clinical effect without having to overdose at one of the biological targets. The chemical strategy was based on fine-tuning of the substitution pattern in the muscarinic and PDE4 structural portions of the dual pharmacology compounds, also making use of the analysis of a proprietary crystal structure in the PDE4 catalytic site. Compound 10f was identified as a chemically stable, potent, and in vivo balanced MAPI lead compound, as assessed in bronchoconstriction and inflammation assays in rats after intratracheal administration. After the in-depth investigation of the pharmacological and solid-state profile, 10f proved to be safe and suitable for development.

Developing inhaled drugs for respiratory diseases: A medicinal chemistry perspective.

Despite the devastating impact of many lung diseases on human health, there is still a significant unmet medical need in respiratory diseases, for which inhaled delivery represents a crucial strategy. More guidance on how to design and carry out multidisciplinary inhaled projects is needed. When designing inhaled drugs, the medicinal chemist must carefully balance the physicochemical properties of the molecule to achieve optimal target engagement in the lung. Although the medicinal chemistry strategy is unique for each project, and will change depending on multiple factors, such as the disease, target, systemic risk, delivery device, and formulation, general guidelines aiding inhaled drug design can be applied and are summarised in this review.

Discovery of M3 Antagonist-PDE4 Inhibitor Dual Pharmacology Molecules for the Treatment of Chronic Obstructive Pulmonary Disease.

In this paper, we report the discovery of dual M3 antagonist-PDE4 inhibitor (MAPI) compounds for the inhaled treatment of pulmonary diseases. The identification of dual compounds was enabled by the intuition that the fusion of a PDE4 scaffold derived from our CHF-6001 series with a muscarinic scaffold through a common linking ring could generate compounds active versus both the transmembrane M3 receptor and the intracellular PDE4 enzyme. Two chemical series characterized by two different muscarinic scaffolds were investigated. SAR optimization was aimed at obtaining M3 nanomolar affinity coupled with nanomolar PDE4 inhibition, which translated into anti-bronchospastic efficacy ex vivo (inhibition of rat trachea contraction) and into anti-inflammatory efficacy in vitro (inhibition of TNFα release). Among the best compounds, compound 92a achieved the goal of demonstrating in vivo efficacy and duration of action in both the bronchoconstriction and inflammation assays in rat after intratracheal administration.

Leptospirosis from water sources.

Leptospirosis outbreaks have been associated with many common water events including water consumption, water sports, environmental disasters, and occupational exposure. The ability of leptospires to survive in moist environments makes them a high-risk agent for infection following contact with any contaminated water source. Water treatment processes reduce the likelihood of leptospirosis or other microbial agents causing infection provided that they do not malfunction and the distribution networks are maintained. Notably, there are many differences in water treatment systems around the world, particularly between developing and developed countries. Detection of leptospirosis in water samples is uncommonly performed by molecular methods.

Discovery of highly potent, selective, and brain-penetrant aminopyrazole leucine-rich repeat kinase 2 (LRRK2) small molecule inhibitors.

Leucine-rich repeat kinase 2 (LRRK2) has drawn significant interest in the neuroscience research community because it is one of the most compelling targets for a potential disease-modifying Parkinson's disease therapy. Herein, we disclose structurally diverse small molecule inhibitors suitable for assessing the implications of sustained in vivo LRRK2 inhibition. Using previously reported aminopyrazole 2 as a lead molecule, we were able to engineer structural modifications in the solvent-exposed region of the ATP-binding site that significantly improve human hepatocyte stability, rat free brain exposure, and CYP inhibition and induction liabilities. Disciplined application of established optimal CNS design parameters culminated in the rapid identification of GNE-0877 (11) and GNE-9605 (20) as highly potent and selective LRRK2 inhibitors. The demonstrated metabolic stability, brain penetration across multiple species, and selectivity of these inhibitors support their use in preclinical efficacy and safety studies.

Haemoglobin and red cell counts in leptospirosis patients infected with different serovars.

INTRODUCTION: The aim of the study was to compare haemoglobin and red cell counts between patients known to be infected with a range of leptospiral serovars. METHODS: The study retrospectively compared the haemoglobin and red cell count results from the first blood samples taken from 207 patients at presentation to a Queensland Health hospital. RESULTS: Significant differences were observed in haemoglobin and red cell counts in those infected with Leptospira interrogans serovars Szwajizak and Canicola when compared with most of the other serovars. CONCLUSIONS: These findings suggest that haemoglobin and red cell counts may be useful in differentiating leptospiral serovars in leptospirosis patients.

Haemoglobin and red cell counts in leptospirosis patients infected with different serovars

Introduction The aim of the study was to compare haemoglobin and red cell counts between patients known to be infected with a range of leptospiral serovars. Methods The study retrospectively compared the haemoglobin and red cell count results from the first blood samples taken from 207 patients at presentation to a Queensland Health hospital. Results Significant differences were observed in haemoglobin and red cell counts in those infected with Leptospira interrogans serovars Szwajizak and Canicola when compared with most of the other serovars. Conclusions These findings suggest that haemoglobin and red cell counts may be useful in differentiating leptospiral serovars in leptospirosis patients. .

Discovery of a Highly Selective, Brain-Penetrant Aminopyrazole LRRK2 Inhibitor.

The modulation of LRRK2 kinase activity by a selective small molecule inhibitor has been proposed as a potentially viable treatment for Parkinson's disease. By using aminopyrazoles as aniline bioisosteres, we discovered a novel series of LRRK2 inhibitors. Herein, we describe our optimization effort that resulted in the identification of a highly potent, brain-penetrant aminopyrazole LRRK2 inhibitor (18) that addressed the liabilities (e.g., poor solubility and metabolic soft spots) of our previously disclosed anilino-aminopyrimidine inhibitors. In in vivo rodent PKPD studies, 18 demonstrated good brain exposure and engendered significant reduction in brain pLRRK2 levels post-ip administration. The strategies of bioisosteric substitution of aminopyrazoles for anilines and attenuation of CYP1A2 inhibition described herein have potential applications to other drug discovery programs.

Discovery of highly potent, selective, and brain-penetrable leucine-rich repeat kinase 2 (LRRK2) small molecule inhibitors.

There is a high demand for potent, selective, and brain-penetrant small molecule inhibitors of leucine-rich repeat kinase 2 (LRRK2) to test whether inhibition of LRRK2 kinase activity is a potentially viable treatment option for Parkinson's disease patients. Herein we disclose the use of property and structure-based drug design for the optimization of highly ligand efficient aminopyrimidine lead compounds. High throughput in vivo rodent cassette pharmacokinetic studies enabled rapid validation of in vitro-in vivo correlations. Guided by this data, optimal design parameters were established. Effective incorporation of these guidelines into our molecular design process resulted in the discovery of small molecule inhibitors such as GNE-7915 (18) and 19, which possess an ideal balance of LRRK2 cellular potency, broad kinase selectivity, metabolic stability, and brain penetration across multiple species. Advancement of GNE-7915 into rodent and higher species toxicity studies enabled risk assessment for early development.

Salvage robot-assisted partial nephrectomy for the management of renal cell carcinoma following failed stereotactic radiotherapy.

Nephron-sparing surgery is currently the standard of care for the management of small renal masses. While both neoadjuvant and adjuvant conventional external beam radiotherapy have failed to demonstrate an oncologic benefit for the treatment of renal cell carcinoma, more recent work aims to explore the utility of stereotactic radiotherapy. We present the case of a 70-year-old woman who failed primary treatment of a small renal mass with the CyberKnife radiotherapy system and describe her successful salvage treatment with robot-assisted partial nephrectomy. This case demonstrates the safety of robotic surgery for the management of renal tumors following failed stereotactic radiotherapy.

Mapping the structural requirements of inducers and substrates for decarboxylation of weak acid preservatives by the food spoilage mould Aspergillus niger.

Moulds are able to cause spoilage in preserved foods through degradation of the preservatives using the Pad-decarboxylation system. This causes, for example, decarboxylation of the preservative sorbic acid to 1,3-pentadiene, a volatile compound with a kerosene-like odour. Neither the natural role of this system nor the range of potential substrates has yet been reported. The Pad-decarboxylation system, encoded by a gene cluster in germinating spores of the mould Aspergillus niger, involves activity by two decarboxylases, PadA1 and OhbA1, and a regulator, SdrA, acting pleiotropically on sorbic acid and cinnamic acid. The structural features of compounds important for the induction of Pad-decarboxylation at both transcriptional and functionality levels were investigated by rtPCR and GCMS. Sorbic and cinnamic acids served as transcriptional inducers but ferulic, coumaric and hexanoic acids did not. 2,3,4,5,6-Pentafluorocinnamic acid was a substrate for the enzyme but had no inducer function; it was used to distinguish induction and competence for decarboxylation in combination with the analogue chemicals. The structural requirements for the substrates of the Pad-decarboxylation system were probed using a variety of sorbic and cinnamic acid analogues. High decarboxylation activity, ~100% conversion of 1mM substrates, required a mono-carboxylic acid with an alkenyl double bond in the trans (E)-configuration at position C2, further unsaturation at C4, and an overall molecular length between 6.5Å and 9Å. Polar groups on the phenyl ring of cinnamic acid abolished activity (no conversion). Furthermore, several compounds were shown to block Pad-decarboxylation. These compounds, primarily aldehyde analogues of active substrates, may serve to reduce food spoilage by moulds such as A. niger. The possible ecological role of Pad-decarboxylation of spore self-inhibitors is unlikely and the most probable role for Pad-decarboxylation is to remove cinnamic acid-type inhibitors from plant material and allow uninhibited germination and growth of mould spores.

Total synthesis of (-)-ulapualide A, a novel tris-oxazole macrolide from marine nudibranchs, based on some biosynthesis speculation.

A new, second generation, total synthesis of ulapualide A (1), whose stereochemistry was recently determined from X-ray analysis of its complex with the protein actin, is described. The synthesis is designed and based on some speculation of the biosynthetic origin of the contiguous tris-oxazole unit in ulapualide A, alongside that of the related co-metabolites that contain only two oxazole rings, e.g. 6 and 7. The mono-oxazole carboxylic acid 67b and the mono-oxazole secondary 55b alcohol which, together, contain all of the 10 asymmetric centres in the natural metabolite, were first elaborated using a combination of contemporary asymmetric synthesis protocols. Esterification of 67b with 55b under Yamaguchi conditions gave the ester 77 which was then converted into the omega-amino acid 18a following simultaneous deprotection of the t-butyl ester and the N-Boc protecting groups. Macrolactamisation of 18a, using HATU, now gave the key intermediate macrolactam 17, containing two of the three oxazole rings in ulapualide A (1). A number of procedures were used to introduce the third oxazole ring in ulapualide A from 17, including: a) cyclodehydration to the oxazoline 78a followed by oxidation using nickel peroxide leading to 76; b) dehydration to the enamide 79, followed by conversion into the methoxyoxazoline 78b, via 80, and elimination of methanol from 78b using camphorsulfonic acid. The tris-oxazole macrolide 76 was next converted into the aldehyde 82b in four straightforward steps, which was then reacted with N-methylformamide, leading to the E-alkenylformamide 83. Removal of the TBDPS protection at C3 in 83 finally gave (-)-ulapualide A, whose 1H and 13C NMR spectroscopic data were indistinguishable from those obtained for naturally derived material. It is likely that the tris-oxazole unit in ulapualide A (1) is derived in nature from a cascade of cyclodehydrations from an acylated tris-serine precursor, e.g.9, followed by oxidation of the resulting tris-oxazoline intermediate, i.e.10. It is also plausible to speculate that the biosynthesis of metabolites related to ulapualide A, e.g. the bis-oxazole 6 and the imide 7, involve cyclisations of just two of the serine units in 9. These speculations were given some credence by carrying out pertinent interconversions involving the bis-oxazole amide 24, the enamide 25, the imide 26, the oxazoline 27 and the tris-oxazole 30 as model compounds. An alternative strategy to the tris-oxazole macrolide intermediate 76 was also examined, involving preliminary synthesis of the aldehyde 73, containing a shortened (C25-C34) side chain from 67b and 47b. A Wadsworth-Emmons olefination reaction between 73 and the phosphonate ester 74 led smoothly to the E-alkene 75, but we were not able to reduce selectively the conjugated enone group in 75 to 76 without simultaneous reduction of the oxazole alkene bond, using a variety of reagents and reaction conditions.

The structural determinants of macrolide-actin binding: in silico insights.

By the use of x-ray structures and flexible docking, we have developed the first in silico ligand-based view of the structural determinants of the binding of small molecule mimics of gelsolin, natural products bound to actin. Our technique highlights those residues on the actin binding site forming important hydrophobic and hydrogen-bonding interactions with the ligands. Significantly, through the flexible docking of toxin fragments, we have also identified potential residues on the actin binding site that have yet to be exploited. Guided by these observations, we have demonstrated that kabiramide C can be modified to produce a structure with a predicted binding energy increased by 20% while the molecular mass is reduced by 20%, clearly indicating the potential for future elaboration of structures targeting this important component of the cytoskeleton.

Sarcopenia in the Caenorhabditis elegans pharynx correlates with muscle contraction rate over lifespan.

In muscles, sarcopenia, the loss of muscle mass, is the major cause of aging-related functional decline and frailty. Several factors are correlated with sarcopenia during aging, including contraction-related cellular injury, oxidative stress, endocrine changes and reduced regenerative potential. However the involvement of these factors has not been experimentally investigated. Here, we report that contraction-related injury may significantly promote the progression of sarcopenia in the pharynx of the nematode, Caenorhabditis elegans, a model of aging in non-regenerative tissues. Both functional and structural declines in the pharynx during aging were significantly delayed in mutants with reduced muscle contraction rates. We also examined the role of bacteria in pharynx muscle decline during aging, as previous studies reported that antimicrobial treatments could extend C. elegans lifespan. Although microbial infection may have enhanced functional decline in the pharynx during aging, it was not the sole cause of decreased pumping rates in old animals. This study identifies contraction-related injury as a factor affecting the initiation and progression of sarcopenia during aging. Further, characterization of the specific types of damage induced by muscle contraction will be helpful for understanding the underlying causes of sarcopenia.

A catalytic asymmetric bioorganic route to enantioenriched tetrahydro- and dihydropyranones.

A conceptually novel approach to hetero Diels-Alder adducts of carbonyl compounds is described using as the key steps an antibody-mediated kinetic resolution of hydroxyenones followed by a ring-closure process. Various beta-hydroxyenones proved to be very good substrates for antibodies 84G3- and 93F3-catalyzed retro-aldol reactions, allowing the preparation of highly enantiomerically enriched (up to 99% ee) precursors of pyranones. An attractive feature of this methodology is the possibility to convert these acyclic-enantioenriched beta-hydroxyenones into tetrahydropyranones by a conventional Michael-type addition procedure or into the corresponding dihydropyranones using an alternative palladium-catalyzed oxidative ring closure. For the palladium-mediated cyclization, a biphasic system has been implemented that allows the direct preparation of enantiopure dihydropyranones from the corresponding racemic aldol precursors using a sequential antibody-resolution/palladium-cyclization strategy, without isolation of the intermediate enantioenriched hydroxyenones. This bioorganic route is best applied to the preparation of hetero Diels-Alder adducts otherwise derived from less nucleophilic dienes and unactivated dienophiles.

Behavioral deficits during early stages of aging in Caenorhabditis elegans result from locomotory deficits possibly linked to muscle frailty.

Many behavioral responses require the coordination of sensory inputs with motor outputs. Aging is associated with progressive declines in both motor function and muscle structure. However, the consequences of age-related motor deficits on behavior have not been clearly defined. Here, we examined the effects of aging on behavior in the nematode, Caenorhabditis elegans. As animals aged, mild locomotory deficits appeared that were sufficient to impair behavioral responses to sensory cues. In contrast, sensory ability appeared well maintained during aging. Age-related behavioral declines were delayed in animals with mutations in the daf-2/insulin-like pathway governing longevity. A decline in muscle tissue integrity was correlated with the onset of age-related behavioral deficits, although significant muscle deterioration was not. Treatment with a muscarinic agonist significantly improved locomotory behavior in aged animals, indicating that improved neuromuscular signaling may be one strategy for reducing the severity of age-related behavioral impairments.