727 E. Tyler St. Tempe
The challenge in drug development has been the implementation of novel active treatment regimens in cancer. Despite a deeper understanding of molecular oncogenesis many therapeutic strategies have failed in the clinic, raising the questions if currently developed drugs are not active enough against the targets or the “true” molecular targets – the synthetic lethal vulnerabilities in cancer genomes – have not been discovered yet.
Many therapeutic strategies were developed based on findings from structural molecular abnormalities such as mutations, amplifications or expression changes. However, functional information of interfering with the biological system (that is a cancer cell) and measuring the outcome after inhibiting individual genes is difficult to discern from these “static” research approaches. In order to obtain functional information of target gene interference that could be exploited for design of rational combinations in leukemias, our laboratory adopted small interfering RNA (siRNA) methodologies to suspension-leukemic cells in vitro. The challenge up to now has been the insufficient transfection of these leukemia cells for high-throughput siRNA (HT-siRNA). After successful establishment of a HT-siRNA platform for leukemia suspension cells, we performed the first cytotoxic drug-siRNA kinome sensitizer screen in leukemias. In siRNA kinome screens with the most commonly used leukemia drug Cytarabine, we identified a putative master sensitizer kinase in combination with Cytarabine. The target kinase and combination was validated in secondary siRNA screens and with a pharmacological inhibitor respectively. Based on these results a clinical trial has been proposed testing the identified rational combination.
A similar approach has been performed with 5-Azacytidine, a commonly used drug in myelodysplastic syndrome (MDS, often a pre-leukemia state). Of close to 900 genes individually silenced in combination with 5-Azacytidine, inhibition of only one genes potently sensitized to 5-Azacytidine. In validation experiments we confirmed the validity of this genes as a sensitizer to 5-Azacytidine by secondary siRNA screens. As well as showed strong sensitization of combining a novel small molecule inhibitor in clinical development targeting the identified gene, in combination with 5-Azacytidine, both in vitro and ex vivo. Clinical trials concepts have been developed for this novel combination as well.
Through the RNAi functional genomics approach we have identified new targetable concepts in myeloid cells alone or with commonly used leukemia drugs that yielded novel testable hypothesis to rapidly translate into clinical development and new treatment strategies in leukemias. These examples demonstrate the tremendous power and utility of functional genomics utilizing siRNA to identify, rational targets and combinations in cancer treatment.