Dynamic fluorescence images were obtained from a subcutaneous human Kaposi's sarcoma tumor (KS1767) model immediately following the intravenous injection of an integrin-targeting cyanine dye conjugate, Cy5.5-c(KRGDf). The fluorescence images, acquired via an intensified charge-coupled device detection system, were used in conjunction with a pharmacokinetic (PK) model to determine kinetic properties of target binding in the presence and absence of a competitive ligand, free c(KRGDf). The results indicate that the conjugate dye behaves similarly in normal tissue to the free Cy5.5 dye while it possesses increased uptake in tumor tissue. The change in pharmacokinetic parameters obtained from dynamic imaging of Cy5.5-c(KRGDf) after administration of c(KRGDf) as a competitive ligand to the integrin receptor suggests that (i) the increased uptake of Cy5.5-c(KRGDf) is molecularly specific and that (ii) receptor turnover occurs within 24 h. In addition, PK analysis enables quantification of an in vivo c(KRGDf) binding constant attributable to integrin binding. In vivo pharmacokinetic analysis based on rapid and dynamic optical imaging may be potentially useful for evaluating the presence and turnover rate of disease markers that are potential targets of molecular medicine.
Frequency-domain photon migration measurements across the surface of a tissue-mimicking, semi-infinite phantom are acquired via an intensified charge-coupled device (ICCD) detection system and used in conjunction with the diffusion approximation to determine the optical properties. The absorption and reduced scattering coefficients are determined least accurately when relative measurements of average light intensity I are employed either alone or in a combination with relative modulation amplitude data I and/or relative phase shift data rel. The absorption and reduced scattering coefficients are found accurate to within 15 and 11%, respectively, of the values obtained from standard single-pixel measurements when rel measurements are employed alone or in combination with I data.
Molecular targeting with exogenous near-infrared excitable fluorescent agents using time-dependent imaging techniques may enable diagnostic imaging of breast cancer and prognostic imaging of sentinel lymph nodes within the breast. However, prior to the administration of unproven contrast agents, phantom studies on clinically relevant volumes are essential to assess the benefits of fluorescence-enhanced optical imaging in humans. Diagnostic 3-D fluorescence-enhanced optical tomography is demonstrated using 0.5 to 1 cm3 single and multiple targets differentiated from their surroundings by indocyanine green (micromolar) in a breast-shaped phantom (10-cm diameter). Fluorescence measurements of referenced ac intensity and phase shift were acquired in response to point illumination measurement geometry using a homodyned intensified charge-coupled device system modulated at 100 MHz. Bayesian reconstructions show artifact-free 3-D images (3857 unknowns) from 3-D boundary surface measurements (126 to 439). In a reflectance geometry appropriate for prognostic imaging of lymph node involvement, fluorescence measurements were likewise acquired from the surface of a semi-infinite phantom (8×8×8 cm3) in response to area illumination (12 cm2) by excitation light. Tomographic 3-D reconstructions (24,123 unknowns) were recovered from 2-D boundary surface measurements (3194) using the modified truncated Newton's method. These studies represent the first 3-D tomographic images from physiologically relevant geometries for breast imaging.
KEYWORDS: Tumors, Luminescence, In vivo imaging, Signal detection, Imaging systems, Cancer, Data acquisition, Charge-coupled devices, Animal model studies
The specificity of a novel EGF-Cy5.5 fluorescent optical probe was assessed using CW fluorescence imaging accomplished via an ICCD camera. Imaging was performed on mice with MDA-MB-468 cancer, known to overexpress EGFr, and contrasted against an analogous cell line, MDA-MB-435, that does not express EGFr. Fluorescence images on mice bearing s.c. inoculated tumors were obtained every 6 seconds for a period of 20 minutes following i.v. injection of ICG, Cy5.5, or EGF-Cy5.5 and every 24 hrs thereafter for up to 192 hrs. In addition, mice with MDA-MB-468 tumors were injected i.v. with anti-EGFr antibody C225 24 hrs prior to injection of EGF-Cy5.5. Monitoring the time-sensitive fluorescence intensity confirms that ICG and Cy5.5 show no favorable binding to tumor, regardless of EGFr expression level. In contrast, EGF-Cy5.5 exhibits selective accumulation only in the MDA-MB-468 tumor. Moreover, tumor uptake of EGF-Cy5.5 was blocked by pre-injection of C225 antibody, demonstrating specificity of the targeted contrast agent. Data further demonstrate that ICG and Cy5.5 fluorescence is completely absent from the tumor site, regardless of EGFr expression level, 24-hrs post injection. Similarly, little EGF-Cy5.5 fluorescence was detected in the EGFr-negative tumor after 24 hrs, however, for the MDA-MB-468 tumor, EGF-Cy5.5 fluorescence did not reach undetectable levels until 192 hrs.
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