LAUSR

The existence of tightly integrated cross talk through multiple signaling and effector pathways has been appreciated in malignant cells. The realization of the plasticity of such networks is stimulating the development of combinational therapy to overcome the limitations of one-dimensional therapies. Synergistic pairs of siRNAs or siRNA and drug combinations are the new frontiers in identifying effective therapeutic combinations. To elucidate effective combinations, we developed a versatile protocol to screen siRNA libraries in triple-negative breast cancer cell models. This protocol outlines the steps to identify synergistic combinations of siRNA-siRNA or siRNA-drug combinations using siRNA libraries via a robotic screen. By focusing on smaller functional siRNA libraries, we present methodologies to identify synergistic siRNA pairings against cancerous cell growth and molecular targets to augment the activity of pro-apoptotic TRAIL protein. Here, we summarize the critical steps to undertake such combinational target identification, emphasizing critical factors that affect the outcome of the screens. Our experience suggests that siRNA library screening is an efficient protocol to identify complementary therapeutic pairs of new or already-existing drugs. This protocol is simple, robust and can be completed within a 1-week working period.