Mechanistic Studies of Extrinsic Apoptosis by Therapeutic Molecules in Cancer Cells

Abstract

Death receptor (DR) activation by ligands including TRAIL is the initiating step in extrinsic apoptosis. Despite considerable effort, the clinical benefits from DR agonists have been limited, and no such drug has demonstrated good efficacy in phase II clinical trials. This may be linked to observations that differential cell fate decisions can arise both between cell lines and among individual cells within the same cell line, contributing to resistance to TRAIL and DR therapeutic antibodies in general. Here, I show that DISC activity and caspase-8 dynamics play an important role in extrinsic apoptotic fate decisions for newly engineered ligands. Experiments in cells treated with TRAIL and two novel synthetic TRAIL analogues, T191 and T202, demonstrate that the maximum caspase-8 activation rate threshold is a critical barrier that has to be overcome. In HeLa, DU145, and HT29 cells, this is the major barrier of apoptosis. In HCT116 cells, other factors may also play critical roles in determining cell fate. I also explored potential sensitization strategies in this work in the hope of overcoming pro-survival barriers in TRAIL-resistant cells, and I found that glutamine depletion greatly increased receptor-mediated apoptosis in cells of various cancer types, and this sensitization effect may result from increased caspase-8 activity induced by changes in FLIP and/or TRAF2 levels. I have also determined that the combination of T202 with a proteasome inhibitor (bortezomib) or SMAC mimetic (LCL161) can produce a consistent synergistic effect in multiple breast cancer and colorectal cancer cell lines, pointing to a potentially promising strategy for T202-based treatment.