Metastasis, in which
cancer cells migrate to other tissues and form new
tumors, is a major cause of both
cancer death and treatment failure. In a previous study,
benproperine (Benp) was identified as a
cancer cell migration inhibitor and an inhibitor of
actin-related protein 2/3 complex subunit 2 (ARPC2). However, Benp is a racemic mixture, and which stereoisomer is the active isomer remains unclear. In this study, we found that S-Benp is an active isomer and inhibits the migration and invasion of
cancer cells much more strongly than R-Benp, with no effect on normal cells. The
metastasis inhibitory effect of S-Benp was also verified in an animal model. Validating that inhibitors bind to their targets in cells and tissues has been a very challenging task in
drug discovery. The direct interactions between ARPC2 and S-Benp were verified by surface plasmon resonance analysis (SPR), a cellular thermal shift assay (CETSA), and
drug affinity responsive target stability (DARTS). In the mutant study with ARPC2F225A cells, S-Benp did not bind to ARPC2F225A according to CETSA and DARTS. Furthermore, we validated that S-Benp colocalized with ARPC2 in
cancer cells and directly bound to ARPC2 in
tumor tissues using Cy3-conjugated S-Benp according to CETSA. Finally, actin polymerization assays and immunocytochemistry showed that S-Benp suppressed actin remodeling such as lamellipodium formation. Taken together, our data suggest that S-Benp is an active stereoisomer of Benp and a potential
metastasis inhibitor via ARPC2 binding.