Targeting CD146 with a 64Cu-labeled antibody enables in vivo immunoPET imaging of high-grade gliomas.
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ABSTRACT:
SUBMITTER: Yang Y
PROVIDER: S-EPMC4664343 | biostudies-other | 2015 Nov
REPOSITORIES: biostudies-other
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Project description:UnlabelledThe critical challenge in abdominal aortic aneurysm (AAA) research is the accurate diagnosis and assessment of AAA progression. Angiogenesis is a pathologic hallmark of AAA, and CD105 is highly expressed on newly formed vessels. Our goal was to use (64)Cu-labeled anti-CD105 antibody Fab fragment for noninvasive assessment of angiogenesis in the aortic wall in a murine model of AAA.MethodsFab fragment of TRC105, a mAb that specifically binds to CD105, was generated by enzymatic papain digestion and conjugated to NOTA (1,4,7-triazacyclononane-1,4,7-triacetic acid) for (64)Cu labeling. The binding affinity/specificity of NOTA-TRC105-Fab was evaluated by flow cytometry and various ex vivo studies. BALB/c mice were anesthetized and treated with calcium phosphate to induce AAA and underwent weekly PET scans using (64)Cu-NOTA-TRC105-Fab. Biodistribution and autoradiography studies were also performed to confirm the accuracy of PET results.ResultsNOTA-TRC105-Fab exhibited high purity and specifically bound to CD105 in vitro. Uptake of (64)Cu-NOTA-TRC105-Fab increased from a control level of 3.4 ± 0.1 to 9.5 ± 0.4 percentage injected dose per gram (%ID/g) at 6 h after injection on day 5 and decreased to 7.2 ± 1.4 %ID/g on day 12, which correlated well with biodistribution and autoradiography studies (i.e., much higher tracer uptake in AAA than normal aorta). Of note, enhanced AAA contrast was achieved, due to the minimal background in the abdominal area of mice. Degradation of elastic fibers and highly expressed CD105 were observed in ex vivo studies.Conclusion(64)Cu-NOTA-TRC105-Fab cleared rapidly through the kidneys, which enabled noninvasive PET imaging of the aorta with enhanced contrast and showed increased angiogenesis (CD105 expression) during AAA. (64)Cu-NOTA-TRC105-Fab PET may potentially be used for future diagnosis and prognosis of AAA.
| S-EPMC4452422 | biostudies-literature
Project description:T cells are key players in inflammation, autoimmune diseases, and immunotherapy. Thus, holistic and noninvasive in vivo characterizations of the temporal distribution and homing dynamics of lymphocytes in mammals are of special interest. Herein, we show that PET-based T-cell labeling facilitates quantitative, highly sensitive, and holistic monitoring of T-cell homing patterns in vivo. We developed a new T-cell receptor (TCR)-specific labeling approach for the intracellular labeling of mouse T cells. We found that continuous TCR plasma membrane turnover and the endocytosis of the specific (64)Cu-monoclonal antibody (mAb)-TCR complex enables a stable labeling of T cells. The TCR-mAb complex was internalized within 24 h, whereas antigen recognition was not impaired. Harmful effects of the label on the viability, DNA-damage and apoptosis-necrosis induction, could be minimized while yielding a high contrast in in vivo PET images. We were able to follow and quantify the specific homing of systemically applied (64)Cu-labeled chicken ovalbumin (cOVA)-TCR transgenic T cells into the pulmonary and perithymic lymph nodes (LNs) of mice with cOVA-induced airway delayed-type hypersensitivity reaction (DTHR) but not into pulmonary and perithymic LNs of naïve control mice or mice diseased from turkey or pheasant OVA-induced DTHR. Our protocol provides consequent advancements in the detection of small accumulations of immune cells in single LNs and specific homing to the sites of inflammation by PET using the internalization of TCR-specific mAbs as a specific label of T cells. Thus, our labeling approach is applicable to other cells with constant membrane receptor turnover.
| S-EPMC4313850 | biostudies-other
Project description:Overexpression of the GRP78 receptor on cell surfaces has been linked with tumor growth, metastasis, and resistance to therapy. We developed a (64)Cu-labeled probe for PET imaging of tumor GRP78 expression based on a novel anti-GRP78 monoclonal antibody, MAb159.MAb159 was conjugated with the (64)Cu-chelator DOTA through lysines on the antibody. DOTA-human IgG was also prepared as a control that did not bind to GRP78. The resulting PET probes were evaluated in BXPC3 pancreatic cancer xenografts in athymic nude mice.The radiotracer was synthesized with a specific activity of 0.8 MBq/?g of antibody. In BXPC3 xenografts, (64)Cu-DOTA-MAb159 demonstrated prominent tumor accumulation (4.3 ± 1.2, 15.4 ± 2.6, and 18.3 ± 1.0 percentage injected dose per gram at 1, 17, and 48 after injection, respectively). In contrast, (64)Cu-DOTA-human IgG had low BXPC3 tumor accumulation (4.8 ± 0.5, 7.5 ± 0.7, and 4.6 ± 0.8 percentage injected dose per gram at 1, 17, and 48 h after injection, respectively).We demonstrated that GRP78 can serve as a valid target for pancreatic cancer imaging. The success of this approach will be valuable for evaluating disease course and therapeutic efficacy at the earliest stages of anti-GRP78 treatment. Moreover, these newly developed probes may have important applications in other types of cancer overexpressing GRP78.
| S-EPMC5448786 | biostudies-literature
Project description:Ewing sarcoma is a tumor of the bone and soft tissue characterized by diffuse cell membrane expression of CD99 (MIC2). Single-site, surgically resectable disease is associated with an excellent 5-year event-free survival; conversely, patients with distant metastases have a poor prognosis. Noninvasive imaging is the standard approach to identifying sites of metastatic disease. We sought to develop a CD99-targeted imaging agent for staging Ewing sarcoma and other CD99-expressing tumors.We identified a CD99 antibody with highly specific binding in vitro and labeled this antibody with (64)Cu. Mice with either subcutaneous Ewing sarcoma xenograft tumors or micrometastases were imaged with the (64)Cu-labeled anti-CD99 antibody and these results were compared with conventional MRI and 2[18F]fluoro-2-deoxy-D-glucose-positron emission tomography (FDG-PET) imaging.(64)Cu-labeled anti-CD99 antibody demonstrated high avidity for the CD99-positive subcutaneous tumors, with a high tumor-to-background ratio, greater than that demonstrated with FDG-PET. Micrometastases, measuring 1 to 2 mm on MRI, were not detected with FDG-PET but were readily visualized with the (64)Cu-labeled anti-CD99 antibody. Probe biodistribution studies demonstrated high specificity of the probe for CD99-positive tumors.(64)Cu-labeled anti-CD99 antibody can detect subcutaneous Ewing sarcoma tumors and metastatic sites with high sensitivity, outperforming FDG-PET in preclinical studies. This targeted radiotracer may have important implications for the diagnosis, surveillance, and treatment of Ewing sarcoma. Similarly, it may impact the management of other CD99 positive tumors.
| S-EPMC3946397 | biostudies-literature
Project description:Overexpression of CD146 has been correlated with aggressiveness, recurrence rate, and poor overall survival in hepatocellular carcinoma (HCC) patients. In this study, we set out to develop a CD146-targeting probe for high-contrast noninvasive in vivo positron emission tomography (PET) and near-infrared fluorescence (NIRF) imaging of HCCs. YY146, an anti-CD146 monoclonal antibody, was employed as a targeting molecule to which we conjugated the zwitterionic near-infrared fluorescence (NIRF) dye ZW800-1 and the chelator deferoxamine (Df). This enabled labeling of Df-YY146-ZW800 with (89)Zr and its subsequent detection using PET and NIRF imaging, all without compromising antibody binding properties. Two HCC cell lines expressing high (HepG2) and low (Huh7) levels of CD146 were employed to generate subcutaneous (s.c.) and orthotopic xenografts in athymic nude mice. Sequential PET and NIRF imaging performed after intravenous injection of (89)Zr-Df-YY146-ZW800 into tumor-bearing mice unveiled prominent and persistent uptake of the tracer in HepG2 tumors that peaked at 31.65 ± 7.15 percentage of injected dose per gram (%ID/g; n=4) 72 h post-injection. Owing to such marked accumulation, tumor delineation was successful by both PET and NIRF, which facilitated the fluorescence image-guided resection of orthotopic HepG2 tumors, despite the relatively high liver background. CD146-negative Huh7 and CD146-blocked HepG2 tumors exhibited significantly lower (89)Zr-Df-YY146-ZW800 accretion (6.1 ± 0.5 and 8.1 ± 1.0 %ID/g at 72 h p.i., respectively; n=4), demonstrating the CD146-specificity of the tracer in vivo. Ex vivo biodistribution and immunofluorescent staining corroborated the accuracy of the imaging data and correlated tracer uptake with in situ CD146 expression. Overall, (89)Zr-Df-YY146-ZW800 showed excellent properties as a PET/NIRF imaging agent, including high in vivo affinity and specificity for CD146-expressing HCC. CD146-targeted molecular imaging using dual-labeled YY146 has great potential for early detection, prognostication, and image-guided surgical resection of liver malignancies.
| S-EPMC4997246 | biostudies-literature
Project description:Complementary imaging modalities provide more information than either method alone can yield and we have developed a dual-mode imaging probe for combined magnetic resonance (MR) and positron emission tomography (PET) imaging. We have developed dual-mode PET/MRI active probes targeted to vascular inflammation and present synthesis of (1) an aliphatic amine polystyrene bead and (2) a novel superparamagnetic iron oxide nanoparticle targeted to macrophages that were both coupled to positron-emitting copper-64 isotopes. The amine groups of the polystyrene beads were directly conjugated with an amine-reactive form (isothiocyanate) of aza-macrocycle 1,4,7,10-tetraazacyclo-dodecane-1,4,7,10-tetraacetic acid (DOTA). Iron oxide nanoparticles are dextran sulfate coated, and the surface was modified to contain aldehyde groups to conjugate to an amine-activated DOTA. Incorporation of chelated Cu-64 to nanoparticles under these conditions, which is routinely used to couple DOTA to macromolecules, was unexpectedly difficult and illustrates that traditional conjugation methods do not always work in a nanoparticle environment. Therefore, we developed new methods to couple Cu-64 to nanoparticles and demonstrate successful labeling to a range of nanoparticle types. We obtained labeling yields of 24% for the amine polystyrene beads and 21% radiolabeling yield for the anionic dextran sulfate iron oxide nanoparticles. The new coupling chemistry can be generalized for attaching chelated metals to other nanoparticle platforms.
| S-EPMC2543939 | biostudies-literature
Project description:We report the use of multifunctional folic acid (FA)-modified dendrimers as a platform to radiolabel with 64Cu for PET imaging of folate receptor (FR)-expressing tumors. In this study, amine-terminated generation 5 (G5) poly(amidoamine) dendrimers were sequentially modified with fluorescein isothiocyanate (FI), FA, and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), followed by acetylation of the remaining dendrimer terminal amines. The as-formed multifunctional DOTA-FA-FI-G5·NHAc dendrimers were then radiolabeled with 64Cu via the DOTA chelation. We show that the FA modification renders the dendrimers with targeting specificity to cancer cells overexpressing FR in vitro. Importantly, the radiolabeled 64Cu-DOTA-FA-FI-G5·NHAc dendrimers can be used as a nanoprobe for specific targeting of FR-overexpressing cancer cells in vitro and targeted microPET imaging of the FR-expressing xenografted tumor model in vivo. The developed 64Cu-labeled multifunctional dendrimeric nanoprobe may hold great promise to be used for targeted PET imaging of different types of FR-expressing cancer.
| S-EPMC7473786 | biostudies-literature
Project description:Enhanced mitochondrial potential in carcinoma cells is an important characteristic of cancer. It is of great current interest to develop a radiotracer that is sensitive to mitochondrial potential changes at the early stage of tumor growth. In this report, we present the synthesis and evaluation of (64)Cu-labeled Lissamine rhodamine B (LRB), (64)Cu(DOTA-LRB) (DOTA-LRB = 2-(6-(diethylamino)-3-(diethyliminio)-3H-xanthen-9-yl)-5-(N-(2-(2-(4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclo-dodecan-1-yl)acetamido)ethyl)sulfamoyl)benzenesulfonate) as a new radiotracer for imaging tumors in athymic nude mice bearing U87MG human glioma xenografts by positron emission tomography (PET). We also explored its localization mechanism using Cu(DOTA-LRB) as the fluorescent probe in both the U87MG human glioma cell line and the cultured primary U87MG glioma cells. It was found that (64)Cu(DOTA-LRB) had the highest tumor uptake (6.54 ± 1.50, 6.91 ± 1.26, 5.68 ± 1.13, 7.58 ± 1.96, and 5.14 ± 1.50%ID/g at 0.5, 1, 2, 4, and 24 h postinjection, respectively) among many (64)Cu-labeled organic cations evaluated in the same animal model. The cellular staining study indicated that Cu(DOTA-LRB) was able to localize in mitochondria of U87MG glioma cells due to the enhanced negative mitochondrial potential. This statement is completely supported by the results from decoupling experiment with carbonylcyanide-m-chlorophenylhydrazone (CCCP). MicroPET data showed that the U87MG glioma tumors were clearly visualized as early as 30 min postinjection with (64)Cu(DOTA-LRB). (64)Cu(DOTA-LRB) remained stable during renal excretion, but underwent extensive degradation during hepatobiliary excretion. On the basis of the results from this study, it was concluded that (64)Cu(DOTA-LRB) represents a new class of promising PET radiotracers for noninvasive imaging of the MDR-negative tumors.
| S-EPMC3148415 | biostudies-literature
Project description:This report presents the synthesis and evaluation of (64)Cu(DO3A-xy-ACR) (DO3A-xy-ACR = 2,6-bis(dimethylamino)-10-(4-((4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecan-1-yl)methyl)benzyl)acridin-10-ium) as a radiotracer for imaging tumors in athymic nude mice bearing U87MG glioma xenografts by PET (positron emission tomography). The biodistribution data suggested that (64)Cu(DO3A-xy-ACR) was excreted mainly through the renal system with >65% of injected radioactivity being recovered from urine samples at 1 h postinjection (p.i.). The tumor uptake of (64)Cu(DO3A-xy-ACR) was 1.07 ± 0.23, 1.58 ± 0.55, 2.71 ± 0.66, 3.47 ± 1.19, and 3.52 ± 1.72%ID/g at 0.5, 1, 2, 4, and 24 h p.i., respectively. (64)Cu(DO3A-xy-ACR) had very high liver uptake (31.90 ± 3.98, 24.95 ± 5.64, 15.20 ± 4.29, 14.09 ± 6.82, and 8.18 ± 1.27%ID/g at 0.5, 1, 2, 4, and 24 h p.i., respectively) with low tumor/liver ratios. MicroPET studies showed that the tumors were clearly visualized as early as 30 min p.i. in the glioma-bearing mouse administered with (64)Cu(DO3A-xy-ACR). The high liver radioactivity accumulation was also seen. (64)Cu(DO3A-xy-ACR) had a relatively high metabolic stability during excretion via both renal and hepatobiliary routes, but it was completely decomposed in the liver homogenate. We explored the localization mechanism of Cu(DO3A-xy-ACR) using both U87MG human glioma and the cultured primary U87MG glioma cells. The results from the cellular staining assays showed that (64)Cu(DO3A-xy-ACR) is able to localize in the mitochondria of living U87MG glioma cells due to the enhanced negative mitochondrial potential as compared to normal cells. Although (64)Cu(DO3A-xy-ACR) is not an ideal PET radiotracer for tumor imaging due to its high liver uptake, the results from this study strongly suggest that (64)Cu-labeled acridinium cations are indeed able to localize in the energized mitochondria of tumor cells.
| S-EPMC3086098 | biostudies-literature
Project description:This purpose of this study is to determine the efficacy of a 45-amino acid Gp2 domain, engineered to bind to epidermal growth factor receptor (EGFR), as a positron emission tomography (PET) probe of EGFR in a xenograft mouse model. The EGFR-targeted Gp2 (Gp2-EGFR) and a nonbinding control were site-specifically labeled with 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelator. Binding affinity was tested toward human EGFR and mouse EGFR. Biological activity on downstream EGFR signaling was examined in cell culture. DOTA-Gp2 molecules were labeled with 64Cu and intravenously injected (0.6-2.3 MBq) into mice bearing EGFRhigh (n = 7) and EGFRlow (n = 4) xenografted tumors. PET/computed tomography (CT) images were acquired at 45 min, 2 h, and 24 h. Dynamic PET (25 min) was also acquired. Tomography results were verified with gamma counting of resected tissues. Two-tailed t tests with unequal variances provided statistical comparison. DOTA-Gp2-EGFR bound strongly to human (KD = 7 ± 5 nM) and murine (KD = 29 ± 6 nM) EGFR, and nontargeted Gp2 had no detectable binding. Gp2-EGFR did not agonize EGFR nor antagonize EGF-EGFR. 64Cu-Gp2-EGFR tracer effectively localized to EGFRhigh tumors at 45 min (3.2 ± 0.5%ID/g). High specificity was observed with significantly lower uptake in EGFRlow tumors (0.9 ± 0.3%ID/g, p < 0.001), high tumor-to-background ratios (11 ± 6 tumor/muscle, p < 0.001). Nontargeted Gp2 tracer had low uptake in EGFRhigh tumors (0.5 ± 0.3%ID/g, p < 0.001). Similar data was observed at 2 h, and tumor signal was retained at 24 h (2.9 ± 0.3%ID/g). An engineered Gp2 PET imaging probe exhibited low background and target-specific EGFRhigh tumor uptake at 45 min, with tumor signal retained at 24 h postinjection, and compared favorably with published EGFR PET probes for alternative protein scaffolds. These beneficial in vivo characteristics, combined with thermal stability, efficient evolution, and small size of the Gp2 domain validate its use as a future class of molecular imaging agents.
| S-EPMC5519506 | biostudies-literature