Fluorescence lifetime-based tumor contrast enhancement using an EGFR antibody labeled near infrared fluorophore

Purpose: Imaging techniques for highly specific detection of cancer cells in-vivo can have applications ranging from preclinical drug discovery studies to clinical cancer diagnosis and surgical therapy. While fluorescence imaging using cancer targeted antibodies has shown promise, non-specific probe accumulation in tissue results in significant background fluorescence, reducing detection sensitivity using traditional intensity-based continuous wave (CW) fluorescence imaging. Here we demonstrate that fluorescence lifetime (FLT) imaging can provide significant tumor contrast enhancement over CW intensity in preclinical models of human breast cancer. Experimental Design: Mice bearing MDA-MB-231 tumors were injected with anti-epidermal growth factor receptor (EGFR) antibody conjugated to the fluorescent dye IRDye800CW (anti-EGFR-800). Time domain fluorescence imaging was performed in vivo and in situ up to 48 hours post dye injection. Results: Mice injected with anti-EGFR-800 showed a significantly longer FLT (0.7±0.03 ns) compared to the FLT of non-specific probe uptake in liver (0.63±0.05 ns), providing a dramatic improvement in sensitivity and specificity compared to CW intensity. IgG antibody conjugated IRDye800CW did not show an increased FLT compared to normal tissue, suggesting that the FLT increase of anti-EGFR-800 in tumors was due to receptor binding. Using serial surgery, we show that FLT allows the detection of smaller residual tumors in the surgical bed than possible using CW intensity. Conclusions: Our data suggest that FLT can significantly enhance tumor contrast using fluorescently labeled antibodies, thereby accelerating the efficient clinical application of these probes for margin assessment in image guided surgery and for highly specific detection of tumor receptors in-vivo.