Photoactivatable Fluorescent Tags for Dual-Modality Positron Emission Tomography Optical Imaging

Abstract

Fluorescent protein conjugates are vital tools in a wide range of scientific disciplines from basic biochemical research to applications in clinical pathology and intraoperative surgery. We report the synthesis and characterization of photoactivatable fluorophores (PhotoTags) based on the functionalization of coumarin, fluorescein, BODIPY, rhodamine B, and cyanine dyes with a photochemically active aryl azide group. Photochemical labeling experiments using human serum albumin produced fluorescent proteins in high yields under irradiation with ultraviolet light for <15 min. We also synthesized DFO-RhodB-PEG 3 -ArN 3  a photoactivatable compound that can be radiolabeled with 89 Zr for applications in optical imaging and positron emission tomography. One-pot 89 Zr-radiolabeling and light-induced protein conjugation produced [ 89 Zr]ZrDFO-RhodB-PEG 3 -azepin-trastuzumab. Proofof-concept studies in vitro and in vivo confirmed that [ 89 Zr]ZrDFO-RhodB-PEG 3 -azepin-trastuzumab is a potential dual-modality agent for detecting human epidermal growth factor receptor 2 (HER2/neu) expression. Overall, the PhotoTag technology represents a rapid, synthetically versatile, and user-friendly approach for generating novel protein conjugates.

Multifunctional Photo-Cross-Linking Probes: From Target Protein Searching to Imaging Applications

“…Guillou et al designed a series of aryl azide probes containing metal chelating groups and fluorophores, including coumarin, BODIPY, fluorescein, cyanine, and rhodamine B to conjugate the fluorescent chelator with antibody drugs. 49 After optimization, the photoactivatable imaging agent [ 89 Zr]ZrDFO-RhodB-PEG 3azepin-trastuzumab successfully targeted HER2/neu receptors in vivo (Figure 7B). In this example, fluorogenic photo-crosslinkers conjugate with therapeutic antibodies that selectively bind to the malignant tumor site.…”

“…In vivo imaging is another fascinating application of fluorogenic photo-cross-linkers. Guillou et al designed a series of aryl azide probes containing metal chelating groups and fluorophores, including coumarin, BODIPY, fluorescein, cyanine, and rhodamine B to conjugate the fluorescent chelator with antibody drugs . After optimization, the photoactivatable imaging agent [ 89 Zr]­ZrDFO-RhodB-PEG 3 -azepin-trastuzumab successfully targeted HER2/neu receptors in vivo (Figure B).…”

Heptadentate chelates for 89Zr-radiolabelling of monoclonal antibodies

“…[24][25][26][27][28] These new PhotoTags bear a photochemically active aryl azide (ArN 3 ) and can be used to modify mAbs with radiometal ion complexes for PET, and with fluorophores for optical imaging in vitro or in vivo. 29 Upon irradiation, the ArN 3 absorbs UV light at wavelengths where most proteins do not absorb (365-450 nm), hence avoiding photodegradation of the protein. Photon absorption forms a highly reactive electrophile (ketenimine intermediate) which further reacts with a nucleophile, such as the primary ε-NH 2 group of Lys residues present on the surface of antibodies, to form a new a covalent bond.…”

Diagnostic and Therapeutic Radiopharmaceuticals

“…Other strategies to produce high radioactivity concentrations in tumor tissues have been reported, such as pre-targeting approaches , and the design of radiopharmaceuticals possessing differential metabolism in tumors versus organs. , Three papers have analyzed PET/SPECT images of animal cancer models to study the change of accumulation of the antibody trastuzumab with lowering hyaluronan concentration, the stereo­isomerism of 18 F-radiolabeled non-natural cyclic amino acids, , and the therapeutic effect of nanocapsules sealed with 153 Sm Development of multimodal probes combining PET (and SPECT) imaging with fluorescence − and photodynamic treatment. First-in-human studies of PET radiopharmaceuticals.…”

“…Development of multimodal probes combining PET (and SPECT) imaging with fluorescence − and photodynamic treatment. …”