AMG 319, a Novel Inhibitor of Phosphoinositide-3 Kinase Delta (PI3Kd), Demonstrates Activity in Lymphoma Pre-Clinical Models

Abstract 3718

The PI3k/AKT/mTOR signaling pathway has been found to be deregulated in patients with various subtypes of B-cell lymphoma including mantle cell lymphoma (MCL) and diffuse large B-cell lymphoma (DLBCL). Targeting this particular pathway has been evaluated in multiple pre-clinical models and clinical trials using pharmacological inhibitors with variable degrees of success. More selective and potent PI3K inhibitors, such as AMG 319 are being developed and need proper evaluation in currently relevant pre-clinical models (i.e. rituximab resistant models). To this end, we evaluated the activity of AMG 319 as a single agent or in combination with monoclonal antibodies, targeted agents and chemotherapy drugs in a panel of rituximab-sensitive (RSCL), rituximab-resistant cell lines (RRCL) and primary lymphoma cells isolated from patients with treatment naïve or relapsed/refractory (rel/ref) B- and T-cell lymphoma. Furthermore, we attempted to characterize the cell death pathways executed following in vitro exposure to AMG 319. RSCL, RRCL, T-cell lymphoma, Hodgkin's lymphoma (HL) and MCL cell lines were exposed to escalating doses of AMG 319 (0.1 to 100μM). Isolated primary lymphoma cells were exposed to lower doses of AMG 319 (1 and 10μM) as a single agent and in combination with bortezomib (BTZ) (5–10nM). Cell viability was determined utilizing presto blue and cell titer glo assays. ⁵¹Cr release studies were conducted to assess the effect of AMG 319 in rituximab or ofatumumab immunological activity. Lymphoma cell lines were exposed to AMG 319 (10μM) or DMSO (0.1%) for 48 hours. Subsequently cells were labeled with ⁵¹Cr and exposed to rituximab, ofatumumab or isotype control with human serum (25%) or effector cells isolated from healthy donors (effector/target ratio 40/1). Finally, to characterize the contribution of the intrinsic apoptotic pathway to AMG 319 activity, primary tumor cells isolated from lymphoma patients were exposed to AMG 319 with or without a pan-caspase inhibitor (Q-VD-Oph, 5μM) and changes in cell viability were detected. AMG 319 exhibited dose-dependant activity as a single agent against various cell lines including RSCL, RRCL, MCL and T-cell lymphoma cell lines. In addition, AMG 319 induced cell death in primary tumor cells (N=5) at lower doses than in cell lines. Preliminary experiments suggest a synergistic activity when AMG 319 is combined with BTZ in vitro, although further analysis is required with a larger number of primary tumor cell samples. Pre-exposure of one RRCL and one RSCL to AMG 319 enhanced the biological activity of rituximab and ofatumumab in terms of antibody dependent cellular cytotoxicity (ADCC) or complement mediated cytotoxicity (CMC), and further investigations with additional cell lines are ongoing. Finally, caspase inhibition diminished AMG 319 activity in primary tumor cells in vitro, suggesting that induction of apoptosis via the mitochondrial pathway plays a role in its anti-tumor activity. Our data suggests that AMG 319 as a single agent is active against NHL cells and potentiate the anti-tumor activity of BTZ and to a lesser degree monoclonal antibodies targeting CD20 antigen. Ongoing studies are aimed to evaluate AMG 319 in combination with chemotherapy agents in order to better optimize the activity of this novel PI3 kinase delta inhibitor in B-cell and T-cell lymphomas.