C5-Iminosugar modification of casein kinase 1δ lead 3-(4-fluorophenyl)-5-isopropyl-4-(pyridin-4-yl)isoxazole promotes enhanced inhibitor affinity and selectivity.

The quest for isoform-selective and specific ATP-competitive protein kinase inhibitors is of great interest, as inhibitors with these qualities will come with reduced toxicity and improved efficacy. However, creating such inhibitors is very challenging due to the high molecular similarity of kinases ATP active sites. To achieve selectivity for our casein kinase (CK) 1 inhibitor series, we elected to endow our previous CK1δ-hit, 3-(4-fluorophenyl)-5-isopropyl-4-(pyridin-4-yl)isoxazole (1), with chiral iminosugar scaffolds. These scaffolds were attached to C5 of the isoxazole ring, a position deemed favorable to facilitate binding interactions with the ribose pocket/solvent-open area of the ATP binding pocket of CK1δ. Here, we describe the synthesis of analogs of 1 ((-)-/(+)-34, (-)-/(+)-48), which were prepared in 13 steps from enantiomerically pure ethyl (3R,4S)- and ethyl (3S,4R)-1-benzyl-4-[(tert-butyldimethylsilyl)oxy]-5-oxopyrrolidine-3-carboxylate ((-)-11 and (+)-11), respectively. The synthesis involved the coupling of Weinreb amide-activated chiral pyrrolidine scaffolds with 4- and 2-fluoro-4-picoline and reaction of the resulting 4-picolyl ketone intermediates ((-)-/(+)-40 and (-)-/(+)-44) with 4-fluoro-N-hydroxybenzenecarboximidoyl chloride to form the desired isoxazole ring. The activity of the compounds against human CK1δ, -ε, and -α was assessed in recently optimized in vitro assays. Compound (-)-34 was the most active compound with IC50 values (CK1δ/ε) of 1/8 µM and displayed enhanced selectivity toward CK1δ.

Newly Developed CK1-Specific Inhibitors Show Specifically Stronger Effects on CK1 Mutants and Colon Cancer Cell Lines.

Protein kinases of the CK1 family can be involved in numerous physiological and pathophysiological processes. Dysregulated expression and/or activity as well as mutation of CK1 isoforms have previously been linked to tumorigenesis. Among all neoplastic diseases, colon and rectal cancer (CRC) represent the fourth leading cause of cancer related deaths. Since mutations in CK1δ previously found in CRC patients exhibited increased oncogenic features, inhibition of CK1δ is supposed to have promising therapeutic potential for tumors, which present overexpression or mutations of this CK1 isoform. Therefore, it is important to develop new small molecule inhibitors exhibiting higher affinity toward CK1δ mutants. In the present study, we first characterized the kinetic properties of CK1δ mutants, which were detected in different tumor entities. Subsequently, we characterized the ability of several newly developed IWP-based inhibitors to inhibit wild type and CK1δ mutants and we furthermore analyzed their effects on growth inhibition of various cultured colon cancer cell lines. Our results indicate, that these compounds represent a promising base for the development of novel CRC therapy concepts.

Alkyl and Aryl Thiol Addition to [1.1.1]Propellane: Scope and Limitations of a Fast Conjugation Reaction.

Herein the addition of different thiols to the strained carbon-carbon bond of [1.1.1]propellane (1) is reported. The reaction pathway was investigated, addition reactions with substituted thiols, hydrogen sulfide and protected cysteine were performed, and further modifications of the products were verified. The clean reaction proceeds by a radical chain process, which was confirmed by different deuterium labelling experiments. It shows high functional-group tolerance, since halo-, hydroxy-, methoxy-, carboxy-, amino- and nitro-substituted thiols could be added to 1 with few by-products in 16-90 % yield. Oxidation of the products allows tuning of the polarity and subsequent reactions of the products. The click-type reaction proceeds even faster with selenols, as was shown in a proof of concept. Thiol addition to 1 offers a facile tool for surface modification, conjugation and tuning of hydrophilicity in bio- and medicinal chemistry.

A biosemiotic interpretation of certain genital morphological structures in the spiders Dysdera erythrina and Dysdera crocata (Araneae: Dysderidae).

A biosemiotic approach to the interpretation of morphological data is apt to highlight morphological traits that have hitherto gone unnoticed for their crucial roles in intraspecific sign interpretation and communication processes. Examples of such traits include specific genital structures found in the haplogyne spiders Dysdera erythrina (Walckenaer 1802) and Dysdera crocata (Koch 1838). In both D. erythrina and D. crocata, the distal sclerite of the male bulb and the anterior diverticulum of the female endogyne exhibit a striking, previously unreported correspondence in size and shape, allowing for a precise match between these structures during copulation. In D. erythrina, the sclerite at the tip of the bulb and the anterior diverticulum are semi-circular in shape, whereas in D. crocata they are rectangular. From the perspective of biosemiotics, which studies the production and interpretation of signs and codes in living systems, these structures are considered the morphological zones of an intraspecific sign interpretation process. This process constitutes one of the necessary prerequisites for sperm transfer and the achievement of fertilization. Therefore, these morphological elements deserve particular attention as they hold higher taxonomic value compared to morphological traits of the bulb for which a relevant role in mating and fertilization has not been proven. Thus, an approach to species delimitation based on biosemiotics, with its specific evaluation of morphological structures, provides new insights for the multidisciplinary endeavour of modern integrative taxonomy.

Zebrafish as model system for the biological characterization of CK1 inhibitors.

Introduction: The CK1 family is involved in a variety of physiological processes by regulating different signaling pathways, including the Wnt/ß-catenin, the Hedgehog and the p53 signaling pathways. Mutations or dysregulation of kinases in general and of CK1 in particular are known to promote the development of cancer, neurodegenerative diseases and inflammation. There is increasing evidence that CK1 isoform specific small molecule inhibitors, including CK1δ- and CK1ε-specific inhibitors of Wnt production (IWP)-based small molecules with structural similarity to benzimidazole compounds, have promising therapeutic potential. Methods: In this study, we investigated the suitability of the zebrafish model system for the evaluation of such CK1 inhibitors. To this end, the kinetic parameters of human CK1 isoforms were compared with those of zebrafish orthologues. Furthermore, the effects of selective CK1δ inhibition during zebrafish embryonic development were analyzed in vivo. Results: The results revealed that zebrafish CK1δA and CK1δB were inhibited as effectively as human CK1δ by compounds G2-2 with IC50 values of 345 and 270 nM for CK1δA and CK1δB versus 503 nM for human CK1δ and G2-3 exhibiting IC50 values of 514 and 561 nM for zebrafish CK1δA and B, and 562 nM for human CK1δ. Furthermore, the effects of selective CK1δ inhibition on zebrafish embryonic development in vivo revealed phenotypic abnormalities indicative of downregulation of CK1δ. Treatment of zebrafish embryos with selected inhibitors resulted in marked phenotypic changes including blood stasis, heart failure, and tail malformations. Conclusion: The results suggest that the zebrafish is a suitable in vivo assay model system for initial studies of the biological relevance of CK1δ inhibition.

[Wilhelm Roux's concept of 'developmental mechanics']. / Wilhelm Roux (1850-1924) and seine konzeption der entwicklungsmechanik.

The aim of Roux's 'developmental mechanics' was to determine the causes of individual development. One of the central points of 'developmental mechanics' is the distinction between processes of 'self-differentiation' and processes of 'dependent differentiation'. In the case of self-differentiation the factors responsible for the differentiation of an egg or part of an egg lie in the part itself, whereas the case of dependent differentiation the specific development is driven essentially by external conditions. The principal type of dependent differentiation is 'functional adaptation', a concept by which Roux tried to explain in mechanistic, non-teleological way the adaptation of the inner structure of various organs (e.g. bones, cartilage, muscles) to their specific function. It is common to regard Roux's embryological experiments on problems of self-differentiation as more or less isolated from his research on functional adaptation. Such an approach is inadequate since 'developmental mechanics' rests on both research fields equally. This is also testified by the specific character of Roux's synthetic theory of ontogeny, which divides ontogeny into several phases according to their different shares of inherited and non-inherited, functional factors of differentiation.