Structure-based rational design of staurosporine-based fluorescent probe with broad-ranging kinase affinity for kinase panel application.

Selectivity profiling of compounds is important for kinase drug discovery. To this end, we aimed to develop a broad-range protein kinase assay by synthesizing a novel staurosporine-derived fluorescent probe based on staurosporine and kinase-binding related structural information. Upon structural analysis of staurosporine with kinases, a 4'-methylamine moiety of staurosporine was found to be located on the solvent side of the kinases, to which several linker units can be conjugated by either alkylation or acylation. However, such conjugation was suggested to reduce the binding affinities of the modified compound for several kinases, owing to the elimination of hydrogen bond donor moiety of NH-group from 4'-methylamine and/or steric hindrance by acyl moiety. Based on this structural information, we designed and synthesized a novel staurosporine-based probe without methyl group in order to retain the hydrogen bond donor, similar to unmodified staurosporine. The broad range of the kinase binding assay demonstrated that our novel fluorescent probe is an excellent tool for developing broad-ranging kinase binding assay.

Design, synthesis, and structure-activity relationship of TAK-418 and its derivatives as a novel series of LSD1 inhibitors with lowered risk of hematological side effects.

Lysine-specific demethylase 1 (LSD1) is an enzyme that demethylates methylated histone H3 lysine 4 (H3K4). Inhibition of LSD1 enzyme activity could increase H3K4 methylation levels and treat diseases associated with epigenetic dysregulation. However, known LSD1 inhibitors disrupt the interaction between LSD1 and cofactors such as GFI1B, causing the risk of hematological toxicity, including thrombocytopenia. Starting from a known LSD1 inhibitor (±)1 as a lead compound, a novel series of LSD1 inhibitors that do not induce the expression of GFI1 mRNA, an in vitro surrogate marker of LSD1-GFI1B dissociation, has been designed and synthesized. Initial structure-activity relationship (SAR) studies revealed the structural features key to avoiding GFI1 mRNA induction. Such SAR information enables optimization of LSD1 inhibitors with lowered risk of hematological side effects; TAK-418 ((1R,2R)-2n), the clinical candidate compound found through this optimization, has a hematological safety profile in rodents and humans. We further confirmed that oral administration of TAK-418 at 0.3 and 1 mg/kg for 2 weeks ameliorated memory deficits in mice with NMDA receptor hypofunction, suggesting potential of efficacy in neurodevelopmental disorders. TAK-418 warrants further investigation as a novel class of LSD1 inhibitors with a superior safety profile for the treatment of CNS disorders.

Strictly regulated agonist-dependent activation of AMPA-R is the key characteristic of TAK-653 for robust synaptic responses and cognitive improvement.

Agonistic profiles of AMPA receptor (AMPA-R) potentiators may be associated with seizure risk and bell-shaped dose-response effects. Here, we report the pharmacological characteristics of a novel AMPA-R potentiator, TAK-653, which exhibits minimal agonistic properties. TAK-653 bound to the ligand binding domain of recombinant AMPA-R in a glutamate-dependent manner. TAK-653 strictly potentiated a glutamate-induced Ca2+ influx in hGluA1i-expressing CHO cells through structural interference at Ser743 in GluA1. In primary neurons, TAK-653 augmented AMPA-induced Ca2+ influx and AMPA-elicited currents via physiological AMPA-R with little agonistic effects. Interestingly, TAK-653 enhanced electrically evoked AMPA-R-mediated EPSPs more potently than AMPA (agonist) or LY451646 (AMPA-R potentiator with a prominent agonistic effect) in brain slices. Moreover, TAK-653 improved cognition for both working memory and recognition memory, while LY451646 did so only for recognition memory, and AMPA did not improve either. These data suggest that the facilitation of phasic AMPA-R activation by physiologically-released glutamate is the key to enhancing synaptic and cognitive functions, and nonselective activation of resting AMPA-Rs may negatively affect this process. Importantly, TAK-653 had a wide safety margin against convulsion; TAK-653 showed a 419-fold (plasma Cmax) and 1017-fold (AUC plasma) margin in rats. These findings provide insight into a therapeutically important aspect of AMPA-R potentiation.

T-448, a specific inhibitor of LSD1 enzyme activity, improves learning function without causing thrombocytopenia in mice.

Dysregulation of histone H3 lysine 4 (H3K4) methylation has been implicated in the pathogenesis of several neurodevelopmental disorders. Targeting lysine-specific demethylase 1 (LSD1), an H3K4 demethylase, is therefore a promising approach to treat these disorders. However, LSD1 forms complexes with cofactors including growth factor independent 1B (GFI1B), a critical regulator of hematopoietic differentiation. Known tranylcypromine-based irreversible LSD1 inhibitors bind to coenzyme flavin adenine dinucleotide (FAD) and disrupt the LSD1-GFI1B complex, which is associated with hematotoxicity such as thrombocytopenia, representing a major hurdle in the development of LSD1 inhibitors as therapeutic agents. To discover LSD1 inhibitors with potent epigenetic modulation and lower risk of hematotoxicity, we screened small molecules that enhance H3K4 methylation by the inhibition of LSD1 enzyme activity in primary cultured rat neurons but have little impact on LSD1-GFI1B complex in human TF-1a erythroblasts. Here we report the discovery of a specific inhibitor of LSD1 enzyme activity, T-448 (3-((1S,2R)-2-(cyclobutylamino)cyclopropyl)-N-(5-methyl-1,3,4-thiadiazol-2-yl)benzamide fumarate). T-448 has minimal impact on the LSD1-GFI1B complex and a superior hematological safety profile in mice via the generation of a compact formyl-FAD adduct. T-448 increased brain H3K4 methylation and partially restored learning function in mice with NMDA receptor hypofunction. T-448-type LSD1 inhibitors with improved safety profiles may provide unique therapeutic approaches for central nervous system disorders associated with epigenetic dysregulation.

Platinum-catalyzed intramolecular vinylchalcogenation of alkynes with beta-phenylchalcogeno conjugated amides.

Platinum-catalyzed intramolecular vinylthiolation and -selenation of internal alkynes with vinylchalcogenides 1 having a carbamoyl group on cis-beta-position of vinyl moiety was developed. The conjugated six-membered lactam framework 2 was constructed in high yields. Density functional theory calculations for alkyne insertion processes suggest seven-membered platinacycle 3 is a kinetically favored intermediate of this catalytic system.

Palladium-catalyzed selenoacylation of allenes leading to the regioselective formation of functionalized allyl selenides.

Palladium-catalyzed regio- and stereoselective selenoacylation of allenes with selenol esters proceeded to produce functionalized allyl selenides with the acyl moiety at the inner carbon and the SePh group at the terminal carbon in high yields. A mechanism accounting for the observed regio- and stereoselectivities is proposed based on the results of DFT calculations.

Palladium-catalyzed intramolecular selenocarbamoylation of alkynes with carbamoselenoates: formation of alpha-alkylidene-beta-lactam framework.

Pd-catalyzed intramolecular selenocarbamoylation of alkynes leading to alpha-alkylidene-beta-lactams was developed. This reaction can be applied to thiocarbamoylation and to the synthesis of delta- and epsilon-lactams and a cyclobutanone.