Identification of a highly neurotoxic α-synuclein species inducing mitochondrial damage and mitophagy in Parkinson's disease.

Exposure of cultured primary neurons to preformed α-synuclein fibrils (PFFs) leads to the recruitment of endogenous α-synuclein and its templated conversion into fibrillar phosphorylated α-synuclein (pα-synF) aggregates resembling those involved in Parkinson's disease (PD) pathogenesis. Pα-synF was described previously as inclusions morphologically similar to Lewy bodies and Lewy neurites in PD patients. We discovered the existence of a conformationally distinct, nonfibrillar, phosphorylated α-syn species that we named "pα-syn*.

Pharmacological Inhibition of c-Jun N-terminal Kinase Reduces Food Intake and Sensitizes Leptin's Anorectic Signaling Actions.

The role for c-Jun N-terminal Kinase (JNK) in the control of feeding and energy balance is not well understood. Here, by use of novel and highly selective JNK inhibitors, we investigated the actions of JNK in the control of feeding and body weight homeostasis. In lean mice, intraperitoneal (i.

Rho Kinase (ROCK) Inhibitors and Their Therapeutic Potential.

Rho kinases (ROCKs) belong to the serine-threonine family, the inhibition of which affects the function of many downstream substrates. As such, ROCK inhibitors have potential therapeutic applicability in a wide variety of pathological conditions including asthma, cancer, erectile dysfunction, glaucoma, insulin resistance, kidney failure, neuronal degeneration, and osteoporosis. To date, two ROCK inhibitors have been approved for clinical use in Japan (fasudil and ripasudil) and one in China (fasudil).

Pyridopyrimidinone Derivatives as Potent and Selective c-Jun N-Terminal Kinase (JNK) Inhibitors.

A novel series of 2-aminopyridopyrimidinone based JNK (c-jun N-terminal kinase) inhibitors were discovered and developed. Structure-activity relationships (SARs) were systematically developed utilizing biochemical and cell based assays and in vitro and in vivo drug metabolism and pharmacokinetic (DMPK) studies. Through the optimization of lead compound 1, several potent and selective JNK inhibitors with high oral bioavailability were developed.

Serum- and Glucocorticoid-Inducible Kinase 1 Confers Protection in Cell-Based and in In Vivo Neurotoxin Models via the c-Jun N-Terminal Kinase Signaling Pathway.

Serum glucocorticoid kinase 1 (SGK1) has been shown to be protective in models of Parkinson's disease, but the details by which it confers benefit is unknown. The current study was designed to investigate the details by which SGK1 confers neuroprotection. To do this we employed a cellular neurodegeneration model to investigate c-Jun N-terminal kinase (JNK) signaling and endoplasmic reticulum (ER) stress induced by 6-hydroxydopamine.

Structural basis and biological consequences for JNK2/3 isoform selective aminopyrazoles.

Three JNK isoforms, JNK1, JNK2, and JNK3 have been reported and unique biological function has been ascribed to each. It is unknown if selective inhibition of these isoforms would confer therapeutic or safety benefit. To probe JNK isoform function we designed JNK2/3 inhibitors that have >30-fold selectivity over JNK1.

Bis-aryl urea derivatives as potent and selective LIM kinase (Limk) inhibitors.

The discovery/optimization of bis-aryl ureas as Limk inhibitors to obtain high potency and selectivity and appropriate pharmacokinetic properties through systematic SAR studies is reported. Docking studies supported the observed SAR. Optimized Limk inhibitors had high biochemical potency (IC50 < 25 nM), excellent selectivity against ROCK and JNK kinases (>400-fold), potent inhibition of cofilin phosphorylation in A7r5, PC-3, and CEM-SS T cells (IC50 < 1 μM), and good in vitro and in vivo pharmacokinetic properties.

Design and synthesis of highly potent and isoform selective JNK3 inhibitors: SAR studies on aminopyrazole derivatives.

The c-jun N-terminal kinase 3 (JNK3) is expressed primarily in the brain. Numerous reports have shown that inhibition of JNK3 is a promising strategy for treatment of neurodegeneration. The optimization of aminopyrazole-based JNK3 inhibitors with improved potency, isoform selectivity, and pharmacological properties by structure-activity relationship (SAR) studies utilizing biochemical and cell-based assays, and structure-based drug design is reported.

Synthesis and SAR of novel isoxazoles as potent c-jun N-terminal kinase (JNK) inhibitors.

The design and synthesis of isoxazole 3 is described, a potent JNK inhibitor with two fold selectivity over p38. Optimization of this scaffold led to compounds 27 and 28 which showed greatly improved selectivity over p38 by maintaining the JNK3 potency of compound 3. Extensive SAR studies will be described as well as preliminary in vivo data of the two lead compounds.

Rho kinase inhibitors: a patent review (2012 - 2013).

Introduction: The Rho kinase/ROCK is critical in vital signal transduction pathways central to many essential cellular activities. Since ROCK possess multiple substrates, modulation of ROCK activity is useful for treatment of many diseases.

Areas Covered: Significant progress has been made in the development of ROCK inhibitors over the past two years (Jan 2012 to Aug 2013).

A small molecule bidentate-binding dual inhibitor probe of the LRRK2 and JNK kinases.

Both JNK and LRRK2 are associated with Parkinson's disease (PD). Here we report a reasonably selective and potent kinase inhibitor (compound 6) that bound to both JNK and LRRK2 (a dual inhibitor). A bidentate-binding strategy that simultaneously utilized the ATP hinge binding and a unique protein surface site outside of the ATP pocket was applied to the design and identification of this kind of inhibitor.

Synthesis and biological evaluation of urea derivatives as highly potent and selective rho kinase inhibitors.

RhoA and its downstream effector ROCK mediate stress fiber formation and cell contraction through their effects on the phosphorylation of myosin light chain (MLC). Inhibition of the RhoA/ROCK pathway has proven to be a promising strategy for several indications such as cardiovascular disease, glaucoma, and inflammatory disease. In 2010, our group reported urea-based ROCK inhibitors as potential antiglaucoma agents.

Amino acid derived quinazolines as Rock/PKA inhibitors.

SAR and lead optimization studies for Rock inhibitors based on amino acid-derived quinazolines are described. Studies demonstrated that these amino acid derived quinazolinones were mainly pan-Rock (I & II) inhibitors. While selectivity against other kinases could be achieved, selectivity for most of these compounds against PKA was not achieved.

Inhibition of JNK mitochondrial localization and signaling is protective against ischemia/reperfusion injury in rats.

To build upon recent findings that mitochondrial JNK signaling is inhibited by selectively blocking the interaction between JNK and Sab, we utilized a cell-permeable peptide to demonstrate that ischemia/reperfusion (I/R) injury could be protected in vivo and that JNK mitochondrial signaling was the mechanism by which reactive oxygen species (ROS) generation, mitochondrial dysfunction, and cardiomyocyte cell death occur. We also demonstrated that 5 mg/kg SR-3306 (a selective JNK inhibitor) was able to protect against I/R injury, reducing infarct volume by 34% (p < 0.05) while also decreasing I/R-induced increases in the activity of creatine phosphokinase and creatine kinase-MB.

Blocking c-Jun N-terminal kinase (JNK) translocation to the mitochondria prevents 6-hydroxydopamine-induced toxicity in vitro and in vivo.

Because oxidative stress and mitochondrial dysfunction are well known contributors to Parkinson disease (PD), we set out to investigate the role mitochondrial JNK plays in the etiology of 6-hydroxydopamine-induced (6-OHDA) oxidative stress, mitochondrial dysfunction, and neurotoxicity in SHSY5Y cells and neuroprotection and motor behavioral protection in vivo. To do this, we utilized a cell-permeable peptide of the outer mitochondrial membrane protein, Sab (SH3BP5), as an inhibitor of JNK mitochondrial translocation. In vitro studies showed that 6-OHDA induced JNK translocation to the mitochondria and that inhibition of mitochondrial JNK signaling by Tat-Sab(KIM1) protected against 6-OHDA-induced oxidative stress, mitochondrial dysfunction, and neurotoxicity.

Structural mechanisms of allostery and autoinhibition in JNK family kinases.

c-Jun N-terminal (JNK) family kinases have a common peptide-docking site used by upstream activating kinases, substrates, scaffold proteins, and phosphatases, where the ensemble of bound proteins determines signaling output. Although there are many JNK structures, little is known about mechanisms of allosteric regulation between the catalytic and peptide-binding sites, and the activation loop, whose phosphorylation is required for catalytic activity. Here, we compare three structures of unliganded JNK3 bound to different peptides.

Atmospheric oxygen inhibits growth and differentiation of marrow-derived mouse mesenchymal stem cells via a p53-dependent mechanism: implications for long-term culture expansion.

Large scale expansion of human mesenchymal stem cells (MSCs) is routinely performed for clinical therapy. In contrast, developing protocols for large scale expansion of primary mouse MSCs has been more difficult due to unique aspects of rodent biology. Currently, established methods to isolate mouse MSCs select for rapidly dividing subpopulations that emerge from bone marrow cultures following long-term (months) expansion in atmospheric oxygen.

Enzyme kinetics and interaction studies for human JNK1β1 and substrates activating transcription factor 2 (ATF2) and c-Jun N-terminal kinase (c-Jun).

c-Jun N-terminal kinase (JNK) is a stress signal transducer linked to cell death, and survival. JNK1 has been implicated in obesity, glucose intolerance, and insulin resistance. In this study we report the kinetic mechanism for JNK1β1 with transcription factors ATF2 and c-Jun along with interaction kinetics for these substrates.

Discovery of potent and selective covalent inhibitors of JNK.

The mitogen-activated kinases JNK1/2/3 are key enzymes in signaling modules that transduce and integrate extracellular stimuli into coordinated cellular response. Here, we report the discovery of irreversible inhibitors of JNK1/2/3. We describe two JNK3 cocrystal structures at 2.

Discovery and optimization of indole and 7-azaindoles as Rho kinase (ROCK) inhibitors (part-II).

Therapeutic interventions with Rho kinase (ROCK) inhibitors may effectively treat several disorders such as hypertension, stroke, cancer, and glaucoma. Herein we disclose the optimization and biological evaluation of potent novel ROCK inhibitors based on substituted indole and 7-azaindole core scaffolds. Substitutions on the indole C3 position and on the indole NH and/or amide NH positions all yielded potent and selective ROCK inhibitors (25, 42, and 50).

Discovery and optimization of indoles and 7-azaindoles as Rho kinase (ROCK) inhibitors (part-I).

Rho kinase (ROCK) inhibitors are potential therapeutic agents to treat disorders such as hypertension, multiple sclerosis, cancers, and glaucoma. Here, we disclose the synthesis, optimization, biological evaluation of potent indole and 7-azaindole based ROCK inhibitors that have high potency on ROCK (IC(50)=1 nM) with 740-fold selectivity over PKA (47). Moreover, 47 showed very good DMPK properties making it a good candidate for further development.

Convergent synthesis and discovery of a natural product-inspired paralog-selective Hsp90 inhibitor.

A convergent synthesis of benzoquinone ansamycin analogs is described that proceeds by a sequence of metallacycle-mediated alkyne-alkyne coupling, followed by site- and stereoselective dihydroxylation and global carbamate formation. These studies have led to (1) validation of alkyne-alkyne coupling to produce geldanamycin analogs that lack the problematic quinone, (2) the discovery that C6-C7 bis-carbamate functionality is compatible with Hsp90 inhibition, and (3) the identification of 1 as a nonquinone geldanamycin-inspired paralog-selective Hsp90 inhibitor.

Small Molecule c-jun-N-terminal Kinase (JNK) Inhibitors Protect Dopaminergic Neurons in a Model of Parkinson's Disease.

There are currently no drugs to treat neurodegeneration in Parkinson's disease (PD) and all existing medications only treat symptoms, lose efficacy over time, and produce untoward side effects. In the current work, we report the first highly selective, orally bioavailable, c-jun-N-terminal kinase (JNK) inhibitor for protection of dopaminergic neurons in vitro and in vivo. At 300 nM this compound showed statistically significant protection of primary dopaminergic neurons exposed to 1-methyl-4-phenylpyridinium (MPP(+)), had pharmacokinetic properties in rodents consistent with twice daily (b.