This invention is a means of identifying the mechanism of action of cell death-inducing small molecules. It may be used by drug development companies for the identification of drug targets and mechanisms of drug resistance for a particular small molecule.
Pharmaceutical companies most commonly identify anti-cancer drugs with large screens for agents that cause cell death. Target identification of these drugs occurs later in the process and significantly hampers drug development especially when mechanism of action is never defined. Current methods to identify mechanism of action, such as microarray approaches, are expensive, lack reproducibility, and cannot efficiently differentiate between compounds that engage common downstream signaling pathways. Better understanding of mechanism of action would lead to target identification as well as insights into drug resistance. Combination chemotherapy also relies on multi-drug administration for cancer treatment with the goal of evading spontaneous resistance to single agents. Chemotherapy may be improved with greater understanding of mechanism of action for increased combinatorial efficacy of drugs.
This invention uses RNA interference (RNAi) to knockdown genes known to be involved in mediating responses to a diverse set of chemotherapeutics, with short hairpin RNAs (shRNAs). Cell-lines were then exposed to twenty common chemotherapeutics and their response monitored. The distinct set of shRNAs which promoted resistance and sensitivity for each drug were clustered to produce a functional array. Since drugs with similar mechanisms of action cluster together, the mechanism of action of small molecules may be defined. This approach has already yielded the mechanism of action of two poorly characterized drugs, apigenin and NSC3852. Furthermore, it is highly-reproducible, simple, and flexible.
- Simple and reproducible so that it can be performed by any laboratory with the appropriate vectors and cells
be applied to a number of small molecules and combinations