carbocyanines and 1-4-7-10-tetraazacyclododecane--1-4-7-10-tetraacetic-acid

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.

Topics: Animals; Azides; Carbocyanines; Cell Line, Tumor; Doxorubicin; Female; Fluorescent Dyes; Heterocyclic Compounds, 1-Ring; Humans; Mice; Optical Imaging; Positron Emission Tomography Computed Tomography; Receptors, Immunologic; Receptors, Peptide; Tissue Distribution

Overexpression of neurotensin receptors (NTRs) has been suggested to play important roles in the growth and survival of a variety of tumor types. The aim of this study is to develop a dual-modality probe (64Cu -DOTA-NT-Cy5.5) for imaging NTR1 expression in vivo with both positron emission tomography (PET) and fluorescence. In this approach, the thiol group and N terminal amino group of neurotensin analogue (Cys-NT) were chemically modified with Cy5.5 dye and DOTA chelator, respectively. After radiolabeling with 64Cu, the resulting probe (64Cu-DOTA-NT-Cy5.5) was evaluated in NTR1 positive HT-29 tumor model. Small animal PET quantification analysis demonstrated that the tumor uptake was 1.91±0.22 and 1.79±0.16%ID/g at 1 and 4 h postinjection (p.i.), respectively. The tumor-to-muscle ratio was 17.44±3.25 at 4 h p.i. based on biodistribution. Receptor specificity was confirmed by the successful blocking experiment at 4 h p.i. (0.42±0.05%ID/g). In parallel with PET experiment, fluorescence imaging was also performed, which demonstrated prominent tumor uptake in HT-29 model. As a proof of concept, an imaging guided surgery was performed to the fluorescent moiety of this probe and could provide potential surgery guidance for NTR positive patients. In summary, our results clearly indicated that the dual-modality probe, 64Cu-DOTA-NT-Cy5.5, could serve as a promising agent to image NTR positive tumors in vivo.

Topics: Animals; Carbocyanines; Chelating Agents; Colonic Neoplasms; Copper Radioisotopes; Fluorescent Dyes; Heterocyclic Compounds, 1-Ring; Humans; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Positron-Emission Tomography; Radiopharmaceuticals; Receptors, Neurotensin; Tissue Distribution; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

In glioblastoma, EphB4 receptors, a member of the largest family of receptor tyrosine kinases, are overexpressed in both tumor cells and angiogenic blood vessels. The purpose of this study was to examine whether the EphB4-binding peptide TNYL-RAW labeled with both (64)Cu and near-infrared fluorescence dye Cy5.5 could be used as a molecular imaging agent for dual-modality positron emission tomography/computed tomography [PET/CT] and optical imaging of human glioblastoma in orthotopic brain tumor models.. TNYL-RAW was conjugated to Cy5.5 and the radiometal chelator 1,4,7,10-tetraazadodecane-N,N',N″,N‴-tetraacetic acid. The conjugate was then labeled with (64)Cu for in vitro binding and in vivo dual μPET/CT and optical imaging studies in nude mice implanted with EphB4-expressing U251 and EphB4-negative U87 human glioblastoma cells. Tumors and brains were removed at the end of the imaging sessions for immunohistochemical staining and fluorescence microscopic examinations.. μPET/CT and near-infrared optical imaging clearly showed specific uptake of the dual-labeled TNYL-RAW peptide in both U251 and U87 tumors in the brains of the nude mice after intravenous injection of the peptide. In U251 tumors, the Cy5.5-labeled peptide colocalized with both tumor blood vessels and tumor cells; in U87 tumors, the tracer colocalized only with tumor blood vessels, not with tumor cells.. Dual-labeled EphB4-specific peptide could be used as a noninvasive molecular imaging agent for PET/CT and optical imaging of glioblastoma owing to its ability to bind to both EphB4-expressing angiogenic blood vessels and EphB4-expressing tumor cells.

Topics: Animals; Autoradiography; Brain Neoplasms; Carbocyanines; Cell Line, Tumor; Fluorescence; Fluorescent Antibody Technique; Glioblastoma; Heterocyclic Compounds, 1-Ring; Humans; Infrared Rays; Kinetics; Luciferases; Male; Mice; Mice, Nude; Peptides; Positron-Emission Tomography; Receptor, EphB4; Surface Plasmon Resonance; Tissue Distribution; Tomography, X-Ray Computed; Xenograft Model Antitumor Assays

Development of a heptamethine cyanine based tumor-targeting PET imaging probe for noninvasive detection and diagnosis of breast cancer.. Tumor-specific heptamethine-cyanine DOTA conjugate complexed with Cu-64 (PC-1001) was synthesized for breast cancer imaging. In vitro cellular uptake studies were performed in the breast cancer MCF-7 and noncancerous breast epithelial MCF-10A cell lines to establish tumor specificity. In vivo time-dependent fluorescence and PET imaging of breast tumor xenografts in mice were performed. Blood clearance, biodistribution, and tumor-specific uptake and plasma binding of PC-1001 were quantified. Tumor histology (H&E staining) and fluorescence imaging were examined.. PC-1001 displayed similar fluorescence properties (ε=82,880cm(-1)M(-1), Ex/Em=750/820nm) to the parental dye. Time-dependent cellular accumulation indicated significantly higher probe uptake (>2-fold, 30min) in MCF-7 than MCF-10A cells and the uptake was observed to be mediated by organic anion transport peptides (OATPs) system. In vivo studies revealed that PC-1001 has desirable accumulation profile in tumor tissues, with tumor versus muscle uptake of about 4.3 fold at 24h and 5.8 fold at 48h post probe injections. Blood half-life of PC-1001 was observed to be 4.3±0.2h. Microscopic fluorescence imaging of harvested tumor indicated that the uptake of PC-1001 was restricted to viable rather than necrotic tumor cells.. A highly efficient tumor-targeting PET/fluorescence imaging probe PC-1001 is synthesized and validated in vitro in MCF-7 breast cancer cells and in vivo in mice breast cancer xenograft model.

Topics: Animals; Biological Transport; Breast Neoplasms; Carbocyanines; Cell Transformation, Neoplastic; Chemistry Techniques, Synthetic; Copper Radioisotopes; Female; Heterocyclic Compounds, 1-Ring; Humans; MCF-7 Cells; Mice; Positron-Emission Tomography; Spectrometry, Fluorescence; Tumor Burden

Engineered nanoparticles with theranostic functions have attracted a lot of attention for their potential role in the dawning era of personalized medicine. Iron oxide nanoparticles (IONPs), with their advantages of being non-toxic, biodegradable and inexpensive, are candidate platforms for the buildup of theranostic nanostructures; however, progress in using them has been limited largely due to inefficient drug loading and delivery. In the current study, we utilized dopamine to modify the surface of IONPs, yielding nanoconjugates that can be easily encapsulated into human serum albumin (HSA) matrices (clinically utilized drug carriers). This nanosystem is well-suited for dual encapsulation of IONPs and drug molecules, because the encapsulation is achieved in a way that is similar to common drug loading. To assess the biophysical characteristics of this novel nanosystem, the HSA coated IONPs (HSA-IONPs) were dually labeled with (64)Cu-DOTA and Cy5.5, and tested in a subcutaneous U87MG xenograft mouse model. In vivo positron emission tomography (PET)/near-infrared fluorescence (NIRF)/magnetic resonance imaging (MRI) tri-modality imaging, and ex vivo analyses and histological examinations were carefully conducted to investigate the in vivo behavior of the nanostructures. With the compact HSA coating, the HSA-IONPs manifested a prolonged circulation half-life; more impressively, they showed massive accumulation in lesions, high extravasation rate, and low uptake of the particles by macrophages at the tumor area.

Topics: Animals; Biocompatible Materials; Carbocyanines; Cell Line, Tumor; Chelating Agents; Copper Radioisotopes; Drug Carriers; Ferric Compounds; Heterocyclic Compounds, 1-Ring; Humans; Magnetic Resonance Imaging; Materials Testing; Metal Nanoparticles; Mice; Mice, Nude; Molecular Structure; Neoplasm Transplantation; Neoplasms; Positron-Emission Tomography; Spectroscopy, Near-Infrared; Transplantation, Heterologous

Previously, we used directed evolution to engineer mutants of the Ecballium elaterium trypsin inhibitor (EETI-II) knottin that bind to αvβ3 and αvβ5 integrin receptors with low nanomolar affinity, and showed that Cy5.5- or (64)Cu-DOTA-labeled knottin peptides could be used to image integrin expression in mouse tumor models using near-infrared fluorescence (NIRF) imaging or positron emission tomography (PET). Here, we report the development of a dual-labeled knottin peptide conjugated to both NIRF and PET imaging agents for multimodality imaging in living subjects. We created an orthogonally protected peptide-based linker for stoichiometric coupling of (64)Cu-DOTA and Cy5.5 onto the knottin N-terminus and confirmed that conjugation did not affect binding to αvβ3 and αvβ5 integrins. NIRF and PET imaging studies in tumor xenograft models showed that Cy5.5 conjugation significantly increased kidney uptake and retention compared to the knottin peptide labeled with (64)Cu-DOTA alone. In the tumor, the dual-labeled (64)Cu-DOTA/Cy5.5 knottin peptide showed decreased wash-out leading to significantly better retention (p < 0.05) compared to the (64)Cu-DOTA-labeled knottin peptide. Tumor uptake was significantly reduced (p < 0.05) when the dual-labeled knottin peptide was coinjected with an excess of unlabeled competitor and when tested in a tumor model with lower levels of integrin expression. Finally, plots of tumor-to-background tissue ratios for Cy5.5 versus (64)Cu uptake were well-correlated over several time points post injection, demonstrating pharmacokinetic cross validation of imaging labels. This dual-modality NIRF/PET imaging agent is promising for further development in clinical applications where high sensitivity and high resolution are desired, such as detection of tumors located deep within the body and image-guided surgical resection.

Topics: Animals; Carbocyanines; Cell Line, Tumor; Copper Radioisotopes; Cystine-Knot Miniproteins; Female; Heterocyclic Compounds, 1-Ring; Humans; Mice; Mice, Nude; Molecular Imaging; Molecular Structure; Neoplasms; Positron-Emission Tomography; Spectroscopy, Near-Infrared; Tissue Distribution

Combining two or more different imaging modalities in the same agent can be of considerable value in molecular imaging. We describe the use of streptavidin nanoparticle-based complexes as multimodality imaging agents to achieve tumor detection in a mouse model by both fluorescence and nuclear imaging. Up to four biotinylated functionalities can be readily attached to these streptavidin nanoparticles without apparent influence on their properties and with reasonable pharmacokinetics and therefore may be ideally suited for multimodality imaging. By binding a biotinylated anti-Her2 Herceptin antibody to provide tumor targeting, a biotinylated DOTA chelator labeled with (111)ln and a biotinylated Cy5.5 fluorophore to a streptavidin nanoparticle, we demonstrated multimodality imaging in SUM190 (Her2+) tumor bearing mice on both an IVIS fluorescence camera and a NanoSPECT/CT small animal nuclear camera. The imaging results show high tumor accumulation and strong tumor-to-normal tissue contrast by both fluorescence and nuclear imaging. The subsequent biodistribution study confirmed the specific tumor accumulation in that tumor accumulation of radioactivity at 40 h was 21 ID%/g and therefore much higher than all other tissues including liver, heart, kidney, spleen, and muscle that accumulated 8.7, 2.5, 6.9, 7.2, and 1.9 ID%/g, respectively. In conclusion, the streptavidin nanoparticle under development in this laboratory was used effectively for multimodality imaging of tumor in mice by fluorescence and nuclear detection. Presumably, other imaging modalities could also be considered.

Topics: Animals; Carbocyanines; Cell Line, Tumor; Cell Nucleus; Female; Fluorescence; Heterocyclic Compounds, 1-Ring; Humans; Mice; Mice, Nude; Nanoparticles; Neoplasms; Organometallic Compounds; Streptavidin; Xenograft Model Antitumor Assays

There is a critical need for molecular imaging agents to detect cell surface integrin receptors that are present in human cancers. Previously, we used directed evolution to engineer knottin peptides that bind with high affinity ( approximately 10 to 30 nmol/L) to integrin receptors that are overexpressed on the surface of tumor cells and the tumor neovasculature. To evaluate these peptides as molecular imaging agents, we site-specifically conjugated Cy5.5 or (64)Cu-1,4,7,10-tetra-azacyclododecane-N,N',N'',N'''-tetraacetic acid (DOTA) to their N termini, and used optical and positron emission tomography (PET) imaging to measure their uptake and biodistribution in U87MG glioblastoma murine xenograft models. NIR fluorescence and microPET imaging both showed that integrin binding affinity plays a strong role in the tumor uptake of knottin peptides. Tumor uptake at 1 hour postinjection for two high-affinity (IC(50), approximately 20 nmol/L) (64)Cu-DOTA-conjugated knottin peptides was 4.47% +/- 1.21% and 4.56% +/- 0.64% injected dose/gram (%ID/g), compared with a low-affinity knottin peptide (IC(50), approximately 0.4 mumol/L; 1.48 +/- 0.53%ID/g) and c(RGDyK) (IC(50), approximately 1 mumol/L; 2.32 +/- 0.55%ID/g), a low-affinity cyclic pentapeptide under clinical development. Furthermore, (64)Cu-DOTA-conjugated knottin peptides generated lower levels of nonspecific liver uptake ( approximately 2%ID/g) compared with c(RGDyK) ( approximately 4%ID/g) 1 hour postinjection. MicroPET imaging results were confirmed by in vivo biodistribution studies. (64)Cu-DOTA-conjugated knottin peptides were stable in mouse serum, and in vivo metabolite analysis showed minimal degradation in the blood or tumor upon injection. Thus, engineered integrin-binding knottin peptides show great potential as clinical diagnostics for a variety of cancers.

Topics: Amino Acid Sequence; Animals; Binding, Competitive; Brain Neoplasms; Carbocyanines; Cell Line, Tumor; Copper Radioisotopes; Glioblastoma; Heterocyclic Compounds, 1-Ring; Humans; Integrins; Mice; Mice, Nude; Molecular Sequence Data; Oligopeptides; Peptides; Positron-Emission Tomography; Spectroscopy, Near-Infrared; Substrate Specificity; Transplantation, Heterologous