Combination of Farnesyl Transferase Inhibitor (Tipifarnib) with Perifosine Induces Apoptosis through phos-PDK1 in Human Lymphoma and Leukemia Cell Lines.

Abstract

Introduction: Novel agents as anti-cancer therapy are used in the setting of specific molecular abnormalities that provide a survival advantage for malignant cells. One such agent, tipifarnib, is theoretically targeted at Ras mutations which are present in a number of different human cancers. Our previous experience with the FTIs (David et al, in press Blood) has demonstrated that they are ideal agents to combine with other targeted agents. We have investigated the combination of the AKT inhibitor perifosine with tipifarnib in human leukemia and lymphoma cell lines with the hypothesis that the combination of 2 targeted agents will disrupt separate survival pathways and ultimately result in synergistic tumor cell death.Methods: In this study we used the human leukemia cell lines HL-60, Jurkat, and the lymphoma cell line HT. Western blot analysis was used to assess for the effect of either single agent perifosine, tipifarnib, or the combination on AKT, p-AKT, PDK-1, and caspase cleavage. Flow cytometry was utilized to assess for Annexin V staining following combination therapy.Results:Dose escalation studies demonstrated that doses of tipifarnib up to 5μm demonstrated a significant cell death in HL-60 and HT cells. Perifosine doses of 1–5uM also induced cell death in both HL-60 and HT cells. When apoptosis was assessed using western blot analysis of caspase 3 activity and cleavage, the combination of perifosine and tipifarnib demonstrated significant apoptosis using low doses of both agents. The apoptosis was associated with downregulation of phos-PDK1, with a resultant downregulation in p-AKT. The level of phos-PDK1 was completely inhibited in less than 24 hrs in both the HL-60 and HT cell lines in combination than when either agent was given alone.Conclusion: The combination of perifosine, and AKT targeted agent, with tipifarnib, a Ras targeted agent, appear to induce significant cell death in lymphoma and leukemia cell lines with rapid downregulation of p-AKT via the PDK-1 pathway. This apoptosis occurs in vitro using concentrations well below those that have been achieved in current clinical trials using these agents. Additional studies are being carried out to further delineate the mechanism of synergy as well as to further explore the impact of sequence of administration using this combination. Further studies are also planned to xplore the impact of the combination on primary human leukemia and lymphoma cells from the blood and bone marrow.