In vitro activity: BMS-3 is a potent inhibitor of the LIM kinase (LIMK)with IC50 of 5nM and 6 nM for LIMK1 and LIMK2 respectively. Inhibition of LIMK1 by specific inhibitors (BMS-3) resulted in lower levels of actin polymerization during capacitation and a dramatic decrease in the percentage of sperm that undergo acrosomal exocytosis. Thus, we demonstrated for the first time that the master regulators of actin dynamics in somatic cells are present and active in mouse sperm. Combining the results of our present study with other results from the literature, we have proposed a working model regarding how LIMK1 and Cofilin control acrosomal exocytosis in mouse sperm.

Kinase Assay: BMS-3 is used to demonstrate the direct participation of LIMK1 in the phosphorylation of Cofilin. Inhibition of p-LIMK with 1-50 μM of BMS-3 results in a dose-dependent decrease of p-Cofilin after 10 min incubation in capacitating conditions. As a control, sperm are also incubated for 10 min under non-capacitating conditions which result in low levels of p-Cofilin. In the presence of 1 or 50 μM of BMS-3, actin polymerization levels are significantly lower compared to controls (DMSO).

Cell Assay: BMS-3 (Compound 2) causes a dose-dependent reduction in cell count and induces mitotic arrest by increases in total nuclear DNA intensity and histone H3 phosphorylation after 24 h treatment in A549 human lung cancer cells. BMS-3 inhibits A549 human lung cancer cells with EC50 value of 154 nM.