In Vivo Fluorescence Lifetime Imaging Monitors Binding of Specific Probes to Cancer Biomarkers

Abstract

One of the most important factors in choosing a treatment strategy for cancer is characterization of biomarkers in cancer cells. Particularly, recent advances in Monoclonal Antibodies (MAB) as primary-specific drugs targeting tumor receptors show that their efficacy depends strongly on characterization of tumor biomarkers. Assessment of their status in individual patients would facilitate selection of an optimal treatment strategy, and the continuous monitoring of those biomarkers and their binding process to the therapy would provide a means for early evaluation of the efficacy of therapeutic intervention. In this study we have demonstrated for the first time in live animals that the fluorescence lifetime can be used to detect the binding of targeted optical probes to the extracellular receptors on tumor cells in vivo . The rationale was that fluorescence lifetime of a specific probe is sensitive to local environment and/or affinity to other molecules. We attached Near-InfraRed (NIR) fluorescent probes to Human Epidermal Growth Factor 2 (HER2/neu)-specific Affibody molecules and used our time-resolved optical system to compare the fluorescence lifetime of the optical probes that were bound and unbound to tumor cells in live mice. Our results show that the fluorescence lifetime changes in our model system delineate HER2 receptor bound from the unbound probe in vivo . Thus, this method is useful as a specific marker of the receptor binding process, which can open a new paradigm in the “image and treat” concept, especially for early evaluation of the efficacy of the therapy.

Evaluation of breast tissue with confocal strip-mosaicking microscopy: a test approach emulating pathology-like examination

“…[1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] Confocal microscopy is an emerging technology for rapid imaging of freshly excised tissue without the need for frozen-or fixed-section processing. Initial studies have described the major findings of invasive breast cancers using fluorescence confocal microscopy.…”

FLIM-FRET for Cancer Applications

“…A HER2 specific affibody molecule was linked to the NIR fluorescent dye to monitor the binding of optical probe to HER2+ cancer. A change in the fluorescence lifetime of HER2-DyLight750 conjugate NIR probe was observed when bound to the HER2 receptor-positive cancer cells (BT-474, high HER2 expressing human breast cancer cells), relative to unbound probe in a mouse xenograft model cells [102]. The fluorescence lifetime model was non-sensitive to the intensity of excitation light or the fluorophore concentration, giving the model major advantage over those applications utilizing fluorescence intensity measurements.…”

“…Conventional FLIM microscopy systems operate in visible range limiting the imaging capabilities due to autofluorescence, reduced SNR and light scattering and preventing a straightforward in vitro cell-based validation of NIR-FRET-FLIM systems for in vivo applications; thus most in vivo NIR-FLIM studies do not report the in vitro cell-based microscopic imaging validation of in vivo FLIM measurements [109]. Previous studies have used upgraded confocal microscopy systems with NIR laser and detectors or used two-photon excitations at NIR wavelengths [102, 110, 111]. For example, Ardeshirpour and group used a specially adapted Olympus FV1000 inverted laser scanning two-photon microscope for evaluating in vitro cell-based the lifetime of their NIR probe, which was used in in vivo FLIM imaging experiments [102].…”

Fluorescence-guided Tumor Visualization Using the Tumor Paint BLZ-100

“…In order to overcome these challenges, surgeons have employed various imaging techniques, including image-guided surgery based on preoperative MRI (magnetic resonance imaging) scans, intra-operative MRI (iMRI), and intra-operative ultrasound. There are several other newer techniques, which have demonstrated potential, including time resolved fluorescence spectroscopy [13][14][15][16][17][18] and Raman spectroscopy [19][20][21][22], but these techniques are very hard to deploy in the operating conditions and require complicated hardware.…”