2010-09, Antonchick, Andrey P., Gerding-Reimers, Claas, Catarinella, Mario, Schürmann, Markus, Preut, Hans, Ziegler, Slava, Rauh, Daniel, Waldmann, Herbert

In biology-oriented synthesis the underlying scaffold classes of natural products selected in evolution are used to define biologically relevant starting points in chemical structure space for the synthesis of compound collections with focused structural diversity. Here we describe a highly enantioselective synthesis of natural-product-inspired 3,3'-pyrrolidinyl spirooxindoles--which contain an all-carbon quaternary centre and three tertiary stereocentres. This synthesis takes place by means of an asymmetric Lewis acid-catalysed 1,3-dipolar cycloaddition of an azomethine ylide to a substituted 3-methylene-2-oxindole using 1-3 mol% of a chiral catalyst formed from a N,P-ferrocenyl ligand and CuPF₆(CH₃CN)₄. Cellular evaluation has identified a molecule that arrests mitosis, induces multiple microtubule organizing centres and multipolar spindles, causes chromosome congression defects during mitosis and inhibits tubulin regrowth in cells. Our findings support the concept that compound collections based on natural-product-inspired scaffolds constructed with complex stereochemistry will be a rich source of compounds with diverse bioactivity.

2009-11-16, Catarinella, Mario, Grüner, Tamara, Strittmatter, Tobias, Marx, Andreas, Mayer, Thomas U.

Motor abgewürgt: Mithilfe eines auf Malachitgrün beruhenden Assays gelang die Identifizierung von BTB-1 (siehe Bild), dem ersten niedermolekularen Inhibitor von Kif18A. BTB-1 hemmt die ATPase-Aktivität der rekombinanten Motordomäne von Kif18A in vitro reversibel (rot=Mikrotubulus, blau-schwarze Struktur=Kif18A) und wird ein nützliches Hilfsmittel für die Analyse der mechanochemischen Eigenschaften von Kif18A sein.

2010-07-15, Svetlik, Jan, Veizerová, Lucia, Mayer, Thomas U., Catarinella, Mario

A synthesis of novel pyrazolopyridine, benzopyranopyrazolopyridine, and oxygen-bridged pyrazolo-, tetrazolo-, benzimidazo-, and thiazolopyrimidines via Hantzsch- and Biginelli-like condensations has been developed. The ability of these compounds to inhibit Eg5 activity has been examined. The results indicate that synthetic manipulations in the monastrol thiourea moiety are inefficient.

2009, Catarinella, Mario, Grüner, Tamara, Strittmatter, Tobias, Marx, Andreas, Mayer, Thomas U.

The survival and development of each organism relies on the equal partitioning of its genome during cell division. Errors in this process can lead to severe developmental defects and cancer in humans. Key for the accurate distribution of the genome is the mitotic spindle composed of dynamic microtubules (Mts).[1] The shape and function of the mitotic spindle depends on the concerted action of various kinesins, these are molecular motor proteins which as microtubule-stimulated ATPases convert chemical energy into mechanical force. Recently, we identified the kinesin-8 member Kif18A as a central component for the correct alignment of chromosomes at the spindle equator.[2] Furthermore, in vitro analyses revealed that Kif18A distinguishes itself from all other kinesins by its dual functionality: motility and depolymerase activity.[2 4] Owing to their fast and often reversible mode of action, small molecules are ideally suited to dissect the function and mechanism of proteins. Given the complex enzymatic characteristics of Kif18A, we set up a smallmolecule screen to identify inhibitors of Kif18A. Herein we report the discovery of BTB-1 (Figure 1a), the first smallmolecule inhibitor of Kif18A. We show that BTB-1 potently inhibits the ATPase activity of Kif18A (IC50=1.69 mm) but not of other tested key mitotic kinesins. BTB-1 blocks the motility of Kif18A in a reversible manner. Notably, BTB-1 inhibits Kif18A in an adenosine triphosphate(ATP)-competitive but microtubule-uncompetitive manner and slows down the progression of cells through mitosis.

2010, Catarinella, Mario

The accurate transmission of the genetic information from mother to daughter cells constitutes a crucial event in the life cycle of all eukaryotic cells (Morgan 2006). To fulfill this task cells rely on a specialized microtubule-based structure called the mitotic spindle (Wittmann et al. 2001). The assembly and the functions of the spindle apparatus are tightly regulated by the orchestrated interplay of dynamic microtubules and motor proteins (Wittmann et al. 2001). In addition to Cytoplasmic dynein, motor proteins of the kinesin superfamily are fundamental for the structure and function of the mitotic spindle (Goshima and Vale 2003; Zhu et al. 2005). Kif18A is a member of the kinesin-8 protein family characterized by its unique dual functionality, which couples a highly processive motor activity with the ability to destabilize microtubules by specifically depolymerizing them at their plus end (Mayr et al. 2007). Recent studies proposed Kif18A to be a key component in the process of chromosome alignment (Stumpff and Wordeman 2007). Notably, this kinesin is required to slow down the oscillatory movements of chromosomes happening in prometaphase after their binding to kinetochore-microtubules and to increase their switch rate across the spindle equatorial region, ultimately leading to the correct chromosome positioning at the metaphase plate (Stumpff et al. 2008). Other reports identified Kif18A as a regulatory partner of Cenp-E (Huang et al. 2009; Zhang and Matunis 2009), a member of the kinesin-7 family involved in chromosome alignment and checkpoint signaling pathways (Yao et al. 2000). Moreover, evidences derived from different works suggest a role of Kif18A in late mitotic events (Gatt et al. 2005; Mayr et al. 2007; Stumpff et al. 2008). The mechanisms by which Kif18A accomplishes these different functions are not fully understood, partially due to the lack of appropriate experimental tools. In this work we reported the functional characterization of the Kif18A small molecule inhibitor 4-chloro-2-nitrodiphenyl sulphone, named BTB-1. This compound originally identified in a reverse chemical genetics high-throughput screening (Grüner 2004), has been validated as a specific and potent inhibitor of the ATPase activity of Kif18A in vitro by three different unrelated approaches. BTB-1 has proven to be effective against Kif18A at low micromolar concentrations (IC₅₀ = 1.7 μM) through a reversible inhibitory mechanism. Moreover, detailed kinetics studies revealed its inhibitory activity to be ATP competitive and microtubule uncompetitive. When tested on human cells BTB-1 induced a mitotic specific phenotype, characterized by the presence of aberrant spindle structures, misaligned chromosomes, increased mitotic index and a prolonged time to complete mitosis. Collectively, these results led to the discovery of the first small molecule inhibitor of the mitotic kinesin Kif18A, which could be employed as a powerful tool to functionally dissect and characterize the different properties of this motor protein.

2007-07, Florian, Stefan, Hümmer, Stefan, Catarinella, Mario, Mayer, Thomas U.

To understand biological processes, biologists typically study how perturbations of protein functions affect the phenotype. Protein activity in living cells can be influenced in many different ways: by manipulation of the genomic information, by injecting inhibitory antibodies, or, more recently, by the use of ribonucleic acid-medicated interference (RNAi). All these methods have proven to be extremely helpful, as they possess a high degree of specificity. However, they are less suitable for experiments requiring precise timing and fast reversibility of the perturbation. The advantage of small molecules is that they specifically interact with their target on a fast time scale and often in a reversible manner. In the last 15 years, this approach, termed "chemical genetics," has received a lot of attention. The term genetics pays tribute to the analogy between chemical genetics and the classic genetic approach, where manipulations at the gene level are used to draw conclusions about the function of the corresponding protein. Chemical genetics has only recently been used as a systematic approach in biology. The term was coined in the 1990's, when combinatorial chemistry was developed as a fast method to synthesize large compound libraries [Mitchison (1994) "Towards a pharmacological genetics," Chem. Biol. 1, 3-6; Schreiber (1998) "Chemical genetics resulting from a passion for synthetic organic chemistry," Bioorg. Med. Chem. 6, 1127-1152].