RESUMEN
Human apurinic/apyrimidinic endonuclease (Ape1) plays an important role by processing the >10,000 highly toxic abasic sites generated in the genome of each cell every day. Ape1 has recently emerged as a target for inhibition, in that its overexpression in tumors has been linked with poor response to both radiation and chemotherapy and lower overall patient survival. Inhibition of Ape1 using siRNA or the expression of a dominant-negative form of the protein has been shown to sensitize cells to DNA-damaging agents, including various chemotherapeutic agents. However, potent small-molecule inhibitors of Ape1 remain to be found. To this end, we screened Ape1 against the NCI Diversity Set of small molecules and discovered aromatic nitroso, carboxylate, sulfonamide, and arylstibonic acid compounds with micromolar affinities for the protein. A further screen of a 37-compound arylstibonic acid sublibrary identified ligands with IC(50) values in the range of 4 to 300 nM. The negatively charged stibonic acids act by a partial-mixed mode and probably serve as DNA phosphate mimics. These compounds provide a useful scaffold for development of chemotherapeutic agents against Ape1.
Asunto(s)
Antimonio/farmacología , ADN-(Sitio Apurínico o Apirimidínico) Liasa/antagonistas & inhibidores , Endonucleasas/antagonistas & inhibidores , Antimonio/química , Antineoplásicos/farmacología , Línea Celular Tumoral , ADN/análisis , ADN/metabolismo , ADN-(Sitio Apurínico o Apirimidínico) Liasa/genética , ADN-(Sitio Apurínico o Apirimidínico) Liasa/aislamiento & purificación , Humanos , Concentración 50 Inhibidora , Cinética , Ligandos , Modelos Químicos , Osteosarcoma/tratamiento farmacológico , Osteosarcoma/enzimología , Osteosarcoma/genética , Osteosarcoma/patología , Proteínas Recombinantes/antagonistas & inhibidores , Proteínas Recombinantes/aislamiento & purificación , Electricidad Estática , Relación Estructura-Actividad , Especificidad por SustratoRESUMEN
The DNA repair enzyme uracil DNA glycosylase has been crystallized with a cationic 1-aza-2'-deoxyribose-containing DNA that mimics the ultimate transition state of the reaction in which the water nucleophile attacks the anomeric center of the oxacarbenium ion-uracil anion reaction intermediate. Comparison with substrate and product structures, and the previous structure of the intermediate determined by kinetic isotope effects, reveals an exquisite example of geometric strain, least atomic motion, and electrophile migration in biological catalysis. This structure provides a rare opportunity to reconstruct the detailed structural transformations that occur along an enzymatic reaction coordinate.