Kenichiro Tsuda

In silico discovery of small-molecule Ras inhibitors that display antitumor activity by blocking the Ras–effector interaction

Mutational activation of the Ras oncogene products (H-Ras, K-Ras, and N-Ras) is frequently observed in human cancers, making them promising anticancer drug targets. Nonetheless, no effective strategy has been available for the development of Ras inhibitors, partly owing to the absence of well-defined surface pockets suitable for drug binding. Only recently, such pockets have been found in the crystal structures of a unique conformation of Ras⋅GTP. Here we report the successful development of small-molecule Ras inhibitors by an in silico screen targeting a pocket found in the crystal structure of M-Ras⋅GTP carrying an H-Ras–type substitution P40D. The selected compound Kobe0065 and its analog Kobe2602 exhibit inhibitory activity toward H-Ras⋅GTP-c-Raf-1 binding both in vivo and in vitro. They effectively inhibit both anchorage-dependent and -independent growth and induce apoptosis of H-rasG12V–transformed NIH 3T3 cells, which is accompanied by down-regulation of downstream molecules such as MEK/ERK, Akt, and RalA as well as an upstream molecule, Son of sevenless. Moreover, they exhibit antitumor activity on a xenograft of human colon carcinoma SW480 cells carrying the K-rasG12V gene by oral administration. The NMR structure of a complex of the compound with H-Ras⋅GTPT35S, exclusively adopting the unique conformation, confirms its insertion into one of the surface pockets and provides a molecular basis for binding inhibition toward multiple Ras⋅GTP-interacting molecules. This study proves the effectiveness of our strategy for structure-based drug design to target Ras⋅GTP, and the resulting Kobe0065-family compounds may serve as a scaffold for the development of Ras inhibitors with higher potency and specificity.

Ab initio MO studies of interaction mechanisms of Protein Kinase C with cell membranes

Protein Kinase C (PKC) is a family of regulatory enzymes. It is considered that binding with phorbol ester which are PKC activators, increases affinity of PKC for the membranes and consequently induces its conformation change. Electrostatic interactions between PKC and the membrane is assumed to be important, and performed ab initio MO calculations of one domain of PKC consisting of 50 amino acids and its complex with the ester is performed to investigate how the electrostatic potential of PKC changes through docking with the substrate. From the calculation, it is shown that the electrostatic potential of PKC near the binding site is dramatically affected through the binding, suggesting attractive interactions with the cell membrane.

Synthesis and binding selectivity of 7- and 15-decylbenzolactone-V8 for the C1 domains of protein kinase C isozymes.

Benzolactone-V8 (4) is a lactone analogue of the artificial tumor promoter benzolactam-V8 (1). To investigate the effect of hydrophobic substituents at positions 7 and 15 of 4 on binding selectivity for protein kinase C (PKC) isozymes, 7- and 15-decylbenzolactone-V8 (7, 8) were synthesized and their binding affinities for synthetic PKC isozyme C1 peptides were examined. Compound 8 showed moderate selectivity for novel PKC isozymes similar to 9-decylbenzolactone-V8 (5), while 7 was less selective. Compounds 7 and 8 showed no significant selectivity among novel PKC isozymes unlike 8-decylbenzolactone-V8 (6). These results indicate that the introduction of a hydrophobic substituent at position 8 of 4 is most effective in the development of PKC epsilon- and PKCeta-selective binders.

Wearable and low-stress ambulatory blood pressure monitoring technology for hypertension diagnosis

We propose a highly wearable, upper-arm type, oscillometric-based blood pressure monitoring technology with low-stress. The low-stress is realized by new developments in the hardware and software design. In the hardware design, conventional armband; cuff, is almost halved in volume thanks to a flexible plastic core and a liquid bag which enhances the fitness and pressure uniformity over the arm. Reduced air bag volume enables smaller motor pump size and battery leading to a thinner, more compact and more wearable unified device. In the software design, a new prediction algorithm enabled to apply less stress (and less pain) on arm of the patient. Proof-of-concept experiments on volunteers show a high accuracy on both technologies. This paper mainly introduces hardware developments. The system is promising for less-painful and less-stressful 24-hour blood pressure monitoring in hypertension managements and related healthcare solutions.

Detection of Drug Administration Behavior with Swallowing Sounds

In recent years, chronic diseases have become the main causes of death around the world, and medication non-adherence among patients with chronic diseases is a common problem. A system for detecting drug administration behavior in daily life is strongly required. Currently, there is not a system for detecting this behavior by using wearable sensors. In this paper, we propose a wearable sensing method for detecting drug administration behavior in daily life by using swallowing sound, which is available and suitable for daily monitoring. To recognize the behavior from swallowing activities, a classification methodology using wavelet based features as feature vectors and artificial neural network as classifier is proposed. A high classification accuracy of 85.4% was achieved in classifying two swallowing activities of drinking water and taking a capsule with water. Furthermore, we also propose a compensation method for time-dependent change based on the frequency characteristics of swallowing sound.