Björn Riefke

Hydrophilic Cyanine Dyes as Contrast Agents for Near-infrared Tumor Imaging: Synthesis, Photophysical Properties and Spectroscopic In vivo Characterization¶

Abstract We have synthesized a group of glucamine and gluosamine-substituted cyanine dyes structurally related to indocyanine green (ICG) and have characterized these compounds with regard to their potential as contrast agents for biomedical optical imaging. The compounds reported herein exhibit increased hydrophilicity and less plasma protein binding (<50%), and are thus expected to have different pharmacokinetic properties compared with ICG. Furthermore, we measured enhanced fluorescence quantum yields (7–15%) in a physiological environment with respect to ICG. For the derivative with the highest hydrophilicity (5a) the efflux from tumor and normal tissue was monitored by intensity-modulated diffuse optical spectroscopy after intravenous injection into tumor-bearing rats. In comparison with ICG, 5a exhibited a considerably enhanced tissue-efflux half-life (73 min versus less than 10 min for ICG in tumor tissue), a two-fold higher initial tissue absorption coefficient compared to ICG, and finally, it generated an elevated tumor-to-tissue concentration gradient up to 1 h after injection. In conclusion, compounds such as 5a are promising contrast agents for optical imaging, and could facilitate highly sensitive and specific detection of breast cancer or other malignancies by utilizing mechanisms similar to contrast-enhanced magnetic resonance imaging or computerized tomography.

Macromolecular Contrast Agents for Optical Imaging of Tumors: Comparison of Indotricarbocyanine-labeled Human Serum Albumin and Transferrin¶

Abstract Macromolecules accumulate in solid tumors and can thus be used as carriers for the delivery of attached contrast agents to tumors. We report the synthesis and use of serum protein–dye conjugates consisting of transferrin (Tf) or human serum albumin (HSA) and an indotricarbocyanine (ITCC) derivative as contrast agents for the optical imaging of tumors. The compounds were characterized with respect to their photophysical properties and tested in vitro for their ability to bind to tumor cells and in vivo for their potential to delineate experimental tumors. In contrast to HAS-ITTC, Tf-ITCC showed receptor-mediated uptake by HT29 human colon cancer cells in vitro. After intravenous injection into HT29 tumor-bearing nude mice both compounds induced increased fluorescence contrast of tumors in vivo. After 24 h the contrast between tumor and normal tissue was significantly higher for Tf-ITCC than for HAS-ITCC. Dye-induced fluorescence was found to be predominantly located in perinecrotic areas of the tumor. Furthermore, Tf-ITCC produced fluorescence of viable tumor cells, whereas HAS-ITCC fluorescence was recorded along connective tissue. We conclude that ITCC-labeled Tf and HSA can serve as macromolecular contrast agents for the optical imaging of tumors, with Tf-ITCC showing higher efficiency.

Cyanine dyes as contrast agents in biomedical optical imaging.

As light exhibits highest penetration into living tissue in the near-infrared (NIR) region between 700 and 900 nm, tissue probing with NIR light provides the opportunity to detect tumors or other abnormalities up to several centimeters below the tissue surface (1). To gain a deeper understanding of physiological and molecular processes in vivo, an important role is increasingly attributed to the acquisition of optical signals generated by exogenously administered fluorescent probes (2–4). Particularly, NIRabsorbing cyanine dyes are potentially suited as fluorescent contrast agents, e.g. for fluorescence-guided endoscopy or optical mammography (1). The fact, that fluorescent dyes can be detected at low concentrations and nonionizing, harmless radiation can be applied repeatedly to the patient renders this technology particularly attractive. In the area of organic dyes, the class of cyanine dyes has proven to be most promising for biomedical applications (5). Thus, the objective of our work is the synthesis of novel cyanine dyes and their characterization as fluorescent contrast agents. Following different concepts, a hydrophilic indotricarbocyanine, which is supposed to act as extracellular contrast agent similar to MR agents, was studied on the one hand. In a different approach we demonstrate that receptor-mediated intracellular tumor targeting using cyanine dye-labeled peptides, which are structurally derived from natural ligands of heptahelical receptors, is a feasible way to achieve specific accumulation and high contrasts in animal tumors. One particular approach was adapted from the radiodiagnostic agent OctreoScan® and involved the use of the peptide octreotate which is known to effectively bind to the somatostatin receptor (SSTR).