Bioluminescence Tracks the in vivo effects of CCI-779 in a minimal residual disease model of HNSCC

Proc Amer Assoc Cancer Res, Volume 47, 2006 4470 Overexpression of eIF4E in “tumor-free” surgical margins of head and neck squamous cell carcinoma (HNSCC) patients is an independent predictor of recurrence and appears to be functionally active through activation of the Akt/mTOR pathway. mTOR inhibitors are cytostatic agents and hence best used in combination therapy or as single agents in minimal residual disease (MRD). We have previously shown using established and MRD models in nude mice that the rapamycin analog CCI-779 inhibits tumor growth. However to determine efficacy of this drug we have used traditional methods of tumor measurements and sacrifice of the mice. Bioluminescent imaging (BLI) is a sensitive, non-invasive method of detecting and quantifying minimal tumor cells. We hypothesize that BLI can provide us with a way of monitoring the in vivo effects of the mTOR inhibitor CCI-779 in a model of MRD thus offering an exciting preclinical strategy to assess tumor response. A HNSCC cell line FaDu was stably transfected with the reporter gene luciferase. 1× 106 FaDu9000 cells (clone with the highest luciferase activity) were pipetted into a surgical bed created in the dorsal flank of the animal. Ten animals were randomly assigned to one of two groups, control or drug (CCI-779 at 20mg/kg) to undergo BLI twice a week. After injecting luciferin, photons emitted from the luciferase expressing cells were quantified using the xenogen IVIS system which measures the photons/second in region of interest (ROI). The effects of the drug on the mTOR pathway were determined by analyzing peripheral blood mononuclear cells (PBMCs). The first scan was performed immediately after surgery and there was no statistically significant difference in photon counts/second between the control and treated group, indicating the number of viable cells implanted were similar in the two groups of mice. CCI-779 prevented tumor formation and the average tumor volume at three weeks in the treated mice (35+ 13mm3) compared to the control group (408+40mm3). Using BLI we were able to observe the effects of the drug on the longitudinal growth of tumor cells. There were significant differences in luciferase activity between the control (32×105 + 3.7×105) and treated group (3.2×105 + 1×105) of mice at three weeks. Analysis of PBMC’s for total 4E-BP1 showed a decrease in the phosphorylation of 4E-BP1 after treatment, indicating the antitumor effects were a result of mTOR inhibition. We have previously shown using a large sample size that CCI-779 prevents tumor formation and increases survival in MRD model of HNSCC. The methods of assaying tumor growth were by tumor volume using calipers. Using BLI we were able to both temporally and spatially measure tumor growth in our model of MRD and monitor response to therapy. This technology has great implications for accelerating the preclinical stages of drug development especially in drugs targeted at minimal residual disease.