In vitro activity: MBQ-167 is a dual inhibitor of Rac/Cdc42 (Ras-related C3 botulinum toxin substrate and cell division control protein 42 homolog), with IC50 values of 103 nM for Rac 1/2/3 and 78 nM for Cdc42 in metastatic MDA-MB-231 cells, respectively. Consequently, MBQ-167 significantly decreases Rac and Cdc42 downstream effector p21-activated kinase (PAK) signaling and the activity of STAT3, without affecting Rho, MAPK, or Akt activities. MBQ-167 also inhibits breast cancer cell migration, viability, and mammosphere formation. Moreover, MBQ-167 affects cancer cells that have undergone epithelial-to-mesenchymal transition by a loss of cell polarity and inhibition of cell surface actin-based extensions to ultimately result in detachment from the substratum. Prolonged incubation (120 hours) in MBQ-167 decreases metastatic cancer cell viability with a GI50 of approximately 130 nmol/L, without affecting noncancer mammary epithelial cells. The loss in cancer cell viability is due to MBQ-167-mediated G2-M cell-cycle arrest and subsequent apoptosis, especially of the detached cells. In vivo, MBQ-167 inhibits mammary tumor growth and metastasis in immunocompromised mice by approximately 90%. In conclusion, MBQ-167 is 10× more potent than other currently available Rac/Cdc42 inhibitors and has the potential to be developed as an anticancer drug, as well as a dual inhibitory probe for the study of Rac and Cdc42.

Kinase Assay: MBQ-167 is a dual inhibitor of Rac/Cdc42 (Ras-related C3 botulinum toxin substrate and cell division control protein 42 homolog), with IC50 values of 103 nM for Rac 1/2/3 and 78 nM for Cdc42 in metastatic MDA-MB-231 cells, respectively.

Cell Assay: MDA-MB-231 cells are treated with vehicle or MBQ-167 at 250 or 500 nM for 24 h. Cells are fixed, permeabilized, and stained with Rhodamine phalloidin to visualize F-actin, and with p-tyrosine or vinculin to visualize focal adhesions.