title: Identification of a drug target motif: an anti-tumor drug NK109 interacts with a PNxxxxP.
abstract: The synthetic compound NK109 shows anti-tumor effects against a number of human cancer cell lines. The mechanism of action is thought to involve the inhibition of DNA topoisomerase II. However, NK109 also exhibits potent anti-tumor activities against doxorubicin-, cisplatin- and etoposide-resistant human cell lines. This paper describes target validation of NK109 using biotinylated NK109 and a T7 phage library screening procedure. Phage particles displaying an affinity for NK109 were isolated and the DNA sequence determined. The amino acid sequences of selected peptides, and the results of mutation experiments by alanine scanning, confirmed that the binding target motif of NK109 is PNxxxxP. In silico analysis of the interaction between NK109 and the peptide, by docking simulation and molecular dynamics, supported this conclusion. The PNxxxxP motif exists in the C2 domain of protein kinase Calpha. NK109 was confirmed to bind the C2 domain from surface plasmon resonance analysis. Furthermore, NK109 moderately inhibited protein kinase C activity in vitro. Our results show that the anti-tumor activity of NK109 stems from interactions with multiple protein targets.
authors.affiliation: Genome and Drug Research Center, Tokyo University of Science, 2641 Yamazaki, Noda-shi, Chiba 278-8510, Japan.
authors.forename: Kengo
authors.initials: K
authors.lastname: Morohashi
authors.forename: Ayako
authors.initials: A
authors.lastname: Yoshino
authors.forename: Atsushi
authors.initials: A
authors.lastname: Yoshimori
authors.forename: Seiichi
authors.initials: S
authors.lastname: Saito
authors.forename: Seiichi
authors.initials: S
authors.lastname: Tanuma
authors.forename: Kengo
authors.initials: K
authors.lastname: Sakaguchi
authors.forename: Fumio
authors.initials: F
authors.lastname: Sugawara
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017123:Bacteriophage T7 / Q000235:genetics; D053119:Benzophenanthridines; D006801:Humans; D008969:Molecular Sequence Data; D010617:Phenanthridines / Q000494:pharmacology*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D051571:Protein Kinase C-alpha; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*
chemical_list: D000970:Antineoplastic Agents; D053119:Benzophenanthridines; C108284:NK 109; D010617:Phenanthridines; D047428:Protein Kinase Inhibitors; C496628:PRKCA protein, human; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha