Non-invasive imaging of caspase-3 activation by TRAIL/FasL adenoviral gene therapy in human glioma cancer model

Objectives We tried to monitor caspase-3-dependent apoptosis by adenoviral gene therapy using a caspase-3 sensor in human glioma cancer model.

Methods Previously, we had reported sensitive caspase-3 biosensor based on a split luciferase technology. Human glioma cancer cell line (D54) expressing caspase-3 sensor and renilla luciferase (Rluc) was established (referred to as a D54-CR). For in vitro experiment, Ad-TRAIL (tumor necrosis factor-related apoptosis inducing ligand)/or -FasL at 5, 25, and 50 MOI (multiplicity of infection) was transduced to D54-CR cells, followed by serial measurement of luciferase activity at designated time points. Inhibition study with Z-VAD (caspase inhibitor) was performed in D54-CR cells transduced with Ad-TRAIL/or -FasL to examine the selectivity of the caspase-3 sensor. FACS analysis was done to determine the percentage of 1) cleaved caspase-3/ or PARP and 2) Annexin-V positive cells in transduced D54-CR cells. After establishment of subcutaneous D54-CR tumor model, in vivo bioluminescence imaging (BLI) was acquired to determine the baseline of bioluminescence activity prior to Ad-FasL treatment. Ad-null/or -FasL (1×108 pfu/mouse) was intratumorally administered and BLI imaging was acquired at 24 h and 48 h post-injection.

Results BLI signals in D54-CR were increased in a time- and virus dose-dependent manner after transduction (P<0.001). Treatment of Z-VAD completely blocked the BLI activity in transduced cells (P<0.005). At 24 h and 48 h after intratumoral injection of Ad-FasL, in vivo BLI imaging showed ~4.1 and ~6.2 fold increase of BLI activity compared with control (P<0.01), which is corresponding to degree of apoptosis.

Conclusions To this end, we have shown the utility of a new surrogate biomarker for imaging caspase-3 mediated cell death by Ad-FasL/-TRAIL therapy in human glioma cancer model using the caspase-3 sensor.

Research Support This work was supported by the Nuclear Research and Development Program of National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (MEST), the Grant of the Korean Ministry of Education, Science and Technology (the Regional Core Research Program/Anti-aging and Well-being Research Center), and the Brain Korea 21 Project in 2011.