In recent years numerous targeted therapies have been identified for therapeutic application in the treatment of cancer. While Chronic Lymphocytic Leukemia (CLL) is an obviously important clinical challenge, it is also is a disease well-suited for the development of novel agents, as readily available tumor cells facilitate validation of specific mechanisms of action in vivo. However, lacking from CLL therapeutic development have been CLL-specific targets and an appropriate in vivo model to test new therapies before transitioning to clinical investigation. Project 6 investigators in conjunction with others in the CLL Research Consortium (CRC) have actively developed multiple new therapies for CLL, including fludarabine, rituximab, and most recently, flavopiridol, as discussed in the previous application. We have further validated the Tcl-1 transgenic mouse model of CLL generated by Project 1 and advanced it into a tool that can be used for investigating new therapeutic agents for subsequent clinical development in CLL. This work and the interactions that have come forth through the CRC have resulted in 23 peer-reviewed publications. Here, we propose to continue active pre-clinical and translational development of new therapeutic agents in CLL. In Aim 1, we will expand our preliminary work with flavopiridol by performing detailed pharmacokinetic, pharmacodynamic, and pharmacogenomic studies as part of our planned Phase II clinical trial, including assessment of efficacy in patients with high risk genetic features such as del(17p13) and mutated p53. Relevant to the development of any effective agent is achieving an understanding of resistance mechanisms, which we will pursue using both CLL patient cells and the Tcl-1 transgenic mouse model of CLL. In Aim 2, we will continue pre-clinical development of the PDK1/Akt inhibitor OSU-03012, now approved for clinical development in CLL by the NCI RAID program, by a) examining the relationship of apoptosis induced by OSU-03012 to inhibition of PDK1/Akt in primary CLL cells; b) examining alternative signaling pathways inhibited by OSU-03012 and the mechanism(s) by which it induces caspase- and Bcl-2-independent apoptosis in CLL cells; and c) exploring synergy of OSU-03012 with other agents used for CLL treatment. In Aim 3, we will use the Tcl-1 transgenic mouse as a pre-clinical tool for developing CLL therapies by performing in vivo studies with flavopiridol, OSU-03012, and other novel therapies. These experiments will also incorporate limited pharmacokinetics and pharmacodynamics. Additionally, we will use the Tcl-1 mouse model to identify relevant mechanisms of drug resistance in vivo for therapies employed in the treatment of CLL. Each of these projects will be performed in collaboration with Projects 1-5 of the CRC, continuing our extensive interactions with these investigators. Overall, this project seeks to continue the comprehensive drug development effort by physician scientists, pharmacologists, and medicinal chemists at Ohio State University and other CRC institutions.