Development of Small Molecules that Specifically Inhibit the D-loop Activity of RAD51.

RAD51 is the central protein in homologous recombination (HR) DNA repair and represents a therapeutic target in oncology. Herein we report a novel class of RAD51 inhibitors that were identified by high throughput screening. In contrast to many previously reported RAD51 inhibitors, our lead compound 1 is capable of blocking RAD51-mediated D-loop formation (IC50 21.3 +/- 7.8 mu M) at concentrations that do not influence RAD51 binding to ssDNA. In human cells, 1 inhibits HR (IC50 13.1 +/- 1.6 mu M) without blocking RAD51's ability to assemble into subnuclear foci at sites of DNA damage. We determined that the active constituent of 1 is actually an oxidized derivative (termed RI(dI)-1 or 8) of the original screening compound. Our SAR campaign also yielded RI(dI)-2 (hereafter termed 9h), which effectively blocks RAD51's D-loop activity in biochemical systems (IC50 11.1 +/- 1.3 mu M) and inhibits HR activity in human cells (IC50 3.0 +/- 1.8 mu M).