Dataset Information

Cerenkov-Activated Sticky Tag for In Vivo Fluorescence Imaging.

ABSTRACT: A big challenge in the clinical use of Cerenkov luminescence (CL) imaging is its low signal intensity, which is several orders of magnitude below ambient light. Consequently, highly sensitive cameras, sufficient shielding from background light, and long acquisition times are required. To alleviate this problem, we hypothesized a strategy to convert the weak CL signal into a stronger fluorescence signal by using CL-activated formation of nitrenes from azides to locally fix a fluorescent probe in tissue by the formation of a covalent bond. CL-activated drug delivery was also evaluated using the same azide chemistry. The specific delivery of the CL-activated drug to cancer cells could reduce systemic toxicity, which is a limitation in chemotherapy. Methods: A cyanine-class near-infrared fluorescent dye, Cy7, and doxorubicin were synthetically attached to polyfluorinated aryl azide to form Cy7 azide and DOX azide, respectively. Fibrosarcoma cells were incubated with ¹⁸F-FDG and exposed to Cy7 azide with subsequent fluorescence imaging. For CL-activated tagging in vivo, tumor-bearing mice were injected first with ⁹⁰Y-DOTA-RGD, targeting ?_v?₃ integrins, and then with the Cy7 azide. Fluorescence signal was imaged over time. Breast cancer cells were incubated with DOX azide and ⁶⁸Ga, after which cell viability was quantified using an assay. Results: CL photoactivation of Cy7 azide in vitro showed significantly higher fluorescence signal from ¹⁸F-FDG-treated than untreated cells. In vivo, CL photoactivation could be shown by using the tumor-specific, integrin-targeting ⁹⁰Y-DOTA-RGD and the localized activation of Cy7 azide. Here, localized CL-induced fluorescence was detected in the tumors and remained significantly higher over several days than in tumors without CL. We also established as a next step CL-activated drug delivery of DOX azide by showing significantly decreasing cell viability of breast cancer cells in a CL dose-dependent manner in vitro using CL photoactivation of DOX azide. Conclusion: We were able to develop a CL-activated "sticky tag" that converts the low CL signal into a stable and long-lasting, highly intense fluorescence signal. This fluorescent footprint of the radioactive signal might be clinically used for intraoperative surgery. The CL-targeted drug delivery strategy may potentially be used for dual-step targeted therapy.

SUBMITTER: Das S

PROVIDER: S-EPMC5750525 | biostudies-literature | 2018 Jan

REPOSITORIES: biostudies-literature

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Cerenkov-Activated Sticky Tag for In Vivo Fluorescence Imaging.

Das Sudeep S Haedicke Katja K Grimm Jan J

Journal of nuclear medicine : official publication, Society of Nuclear Medicine 20170914 1

A big challenge in the clinical use of Cerenkov luminescence (CL) imaging is its low signal intensity, which is several orders of magnitude below ambient light. Consequently, highly sensitive cameras, sufficient shielding from background light, and long acquisition times are required. To alleviate this problem, we hypothesized a strategy to convert the weak CL signal into a stronger fluorescence signal by using CL-activated formation of nitrenes from azides to locally fix a fluorescent probe in ...[more]

PMID: 28912146

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Project description:The development of novel multimodality imaging agents and techniques represents the current frontier of research in the field of medical imaging science. However, the combination of nuclear tomography with optical techniques has yet to be established. Here, we report the use of the inherent optical emissions from the decay of radiopharmaceuticals for Cerenkov luminescence imaging (CLI) of tumors in vivo and correlate the results with those obtained from concordant immuno-PET studies.In vitro phantom studies were used to validate the visible light emission observed from a range of radionuclides including the positron emitters (18)F, (64)Cu, (89)Zr, and (124)I; beta-emitter (131)I; and alpha-particle emitter (225)Ac for potential use in CLI. The novel radiolabeled monoclonal antibody (89)Zr-desferrioxamine B [DFO]-J591 for immuno-PET of prostate-specific membrane antigen (PSMA) expression was used to coregister and correlate the CLI signal observed with the immuno-PET images and biodistribution studies.Phantom studies confirmed that Cerenkov radiation can be observed from a range of positron-, beta-, and alpha-emitting radionuclides using standard optical imaging devices. The change in light emission intensity versus time was concordant with radionuclide decay and was also found to correlate linearly with both the activity concentration and the measured PET signal (percentage injected dose per gram). In vivo studies conducted in male severe combined immune deficient mice bearing PSMA-positive, subcutaneous LNCaP tumors demonstrated that tumor-specific uptake of (89)Zr-DFO-J591 could be visualized by both immuno-PET and CLI. Optical and immuno-PET signal intensities were found to increase over time from 24 to 96 h, and biodistribution studies were found to correlate well with both imaging modalities.These studies represent the first, to our knowledge, quantitative assessment of CLI for measuring radiotracer uptake in vivo. Many radionuclides common to both nuclear tomographic imaging and radiotherapy have the potential to be used in CLI. The value of CLI lies in its ability to image radionuclides that do not emit either positrons or gamma-rays and are, thus, unsuitable for use with current nuclear imaging modalities. Optical imaging of Cerenkov radiation emission shows excellent promise as a potential new imaging modality for the rapid, high-throughput screening of radiopharmaceuticals.

Project description:Live-cell fluorescence imaging of methanogenic archaea has been limited due to the strictly anoxic conditions required for growth and issues with autofluorescence associated with electron carriers in central metabolism. Here, we show that the fluorescence-activating and absorption-shifting tag (FAST) complexed with the fluorogenic ligand 4-hydroxy-3-methylbenzylidene-rhodanine (HMBR) overcomes these issues and displays robust fluorescence in Methanococcus maripaludis. We also describe a mechanism to visualize cells under anoxic conditions using a fluorescence microscope. Derivatives of FAST were successfully applied for protein abundance analysis, subcellular localization analysis, and determination of protein-protein interactions. FAST fusions to both formate dehydrogenase (Fdh) and F420-reducing hydrogenase (Fru) displayed increased fluorescence in cells grown on formate-containing medium, consistent with previous studies suggesting the increased abundance of these proteins in the absence of H2. Additionally, FAST fusions to both Fru and the ATPase associated with the archaellum (FlaI) showed a membrane localization in single cells observed using anoxic fluorescence microscopy. Finally, a split reporter translationally fused to the alpha and beta subunits of Fdh reconstituted a functionally fluorescent molecule in vivo via bimolecular fluorescence complementation. Together, these observations demonstrate the utility of FAST as a tool for studying members of the methanogenic archaea. IMPORTANCE Methanogenic archaea are important members of anaerobic microbial communities where they catalyze essential reactions in the degradation of organic matter. Developing additional tools for studying the cell biology of these organisms is essential to understanding them at a mechanistic level. Here, we show that FAST, in combination with the fluorogenic ligand HMBR, can be used to monitor protein dynamics in live cells of M. maripaludis. The application of FAST holds promise for future studies focused on the metabolism and physiology of methanogenic archaea.

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Cerenkov-Activated Sticky Tag for In Vivo Fluorescence Imaging.

Publications

Cerenkov-Activated Sticky Tag for In Vivo Fluorescence Imaging.

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