Received for publication November 8, 2007.
Revised December 17, 2007.
Accepted for publication December 18, 2007.

A new mechanism of TAS-103 action discovered by target screening with drug-immobilized affinity beads

Abstract

TAS-103 is a quinoline derivative that displays anti-tumour activity in murine and human tumour models. TAS-103 has been reported to be a potent topoisomerase II poison. However, other studies have indicated that cellular susceptibility to TAS-103 is not correlated with topoisomerase II expression. Since the direct target of TAS-103 remained unclear, we searched for a TAS-103 binding protein using high-performance affinity latex beads. We obtained a component of the signal recognition particle (SRP) as a TAS-103 binding protein. This component is a 54-kDa subunit (SRP54) of SRP, which mediates the proper delivery of secretory proteins in cells. We fractioned 293T cell lysates using gel-filtration chromatography and performed a co-immunoprecipitation assay using 293T cells expressing FLAG-tagged SRP54. The results revealed that TAS-103 disrupts SRP complex formation and reduces the amount of SRP14 and SRP19. RNAi-mediated knockdown of SRP54 or SRP14 promoted accumulation of the exogenously expressed chimeric protein, IL-6-FLAG, inside cells. In conclusion, we identified Signal Recognition particle as a target of TAS-103 by using affinity latex beads. This provides new insights into the mechanism underlying the effects of chemotherapies comprising TAS-103 and also demonstrates the usefulness of the affinity beads.

Key words: Protein-binding, Structure/function/mechanism, Protein targets