PET Imaging for The Early Evaluation of Ocular Inflammation in Diabetic Rats by Using [18F]-DPA-714.

Ocular complications of diabetes mellitus (DM) are the leading cause of vision loss. Ocular inflammation often occurs in the early stage of DM; however, there are no proven quantitative methods to evaluate the inflammatory status of eyes in DM. The 18 kDa translocator protein (TSPO) is an evolutionarily conserved cholesterol binding protein localized in the outer mitochondrial membrane. It is a biomarker of activated microglia/macrophages; however, its role in ocular inflammation is unclear. In this study, fluorine-18-DPA-714 ([18F]-DPA-714) was evaluated as a specific TSPO probe by cell uptake, cell binding assays and micro positron emission tomography (microPET) imaging in both in vitro and in vivo models. Primary microglia/macrophages (PMs) extracted from the cornea, retina, choroid or sclera of neonatal rats with or without high glucose (50 mM) treatment were used as the in vitro model. Sprague-Dawley (SD) rats that received an intraperitoneal administration of streptozotocin (STZ, 60 mg/kg once) were used as the in vivo model. Increased cell uptake and high binding affinity of [18F]-DPA-714 were observed in primary PMs under hyperglycemic stress. These findings were consistent with cellular morphological changes, cell activation, and TSPO up-regulation. [18F]-DPA-714 PET imaging and biodistribution in the eyes of DM rats revealed that inflammation initiates in microglia/macrophages in the early stages (3 weeks and 6 weeks), corresponding with up-regulated TSPO levels. Thus, [18F]-DPA-714 microPET imaging may be an effective approach for the early evaluation of ocular inflammation in DM.

Synthesis and Characterization of a Novel PET Tracer for Noninvasive Evaluation of FGL1 Status in Tumors.

Fibrinogen-like protein 1 (FGL1) is a potential novel immune checkpoint target for malignant tumor diagnosis and therapy. Accurate detection of FGL1 levels in tumors via noninvasive PET imaging might be beneficial for managing the disease. To achieve this, multiple FGL1-targeting peptides (FGLP) were designed, and a promising candidate, 68Ga-NOTA-FGLP2, was identified through a high-throughput screening approach using microPET imaging of 68Ga-labeled peptides. Subsequent in vitro cell experiments showed that uptake values of 68Ga-NOTA-FGLP2 in FGL1 positive Huh7 tumor cells were significantly higher than those in FGL1 negative U87 MG tumor cells. Further microPET imaging showed that the Huh7 xenografts were clearly visualized with a favorable contrast. ROI analysis showed that the uptake values of the tracer in Huh7 xenografts were 2.63 ± 0.07% ID/g at 30 min p.i.. After treatment with an excess of unlabeled FGLP2, the tumor uptake significantly decreased to 0.54 ± 0.05% ID/g at 30 min p.i.. Moreover, the uptake in U87 MG xenografts was 0.44 ± 0.06% ID/g at the same time point. The tracer was excreted mainly through the renal system. 18F-FDG PET imaging was also performed in mice bearing Huh7 and U87 MG xenografts, respectively. However, there was no significant difference in the uptake between the tumors with different FGL1 expressions. Preclinical data indicated that 68Ga-NOTA-FGLP2 might be a suitable radiotracer for in vivo noninvasive visualization of tumors with abundant expression of FGL1. Further investigation of 68Ga-NOTA-FGLP2 for tumor diagnosis and therapy is undergoing.

PET imaging of retinal inflammation in mice exposed to blue light using [18F]-DPA-714.

Purpose: Positron emission tomography (PET) is widely used in high-precision imaging, which may provide a simple and noninvasive method for the detection of pathology and therapeutic effects. [18F]-DPA-714 is a second-generation translocator protein (TSPO) positron emission tomography radiotracer that shows great promise in a model of neuroinflammation. In this study, [18F]-DPA-714 micro-PET imaging was used to evaluate retinal inflammation in mice exposed to blue light, a well-established model of age-related macular degeneration (AMD) for molecular mechanism research and drug screening. Methods: C57BL/6J melanized mice were subjected to 10,000, 15,000, and 20,000 lux blue light for 5 days (8 h/day) to develop the retinal injury model, and the structure and function of the retina were assessed using hematoxylin-eosin (HE) staining, electroretinography (ERG), and terminal-deoxynucleotidyl transferase (TdT)-mediated nick-end labeling (TUNEL) immunostaining. Then, [18F]-DPA-714 was injected approximately 100 µCi through each tail vein, and static imaging was performed 1 h after injection. Finally, the mice eyeballs were collected for biodistribution and immune analysis. Results: The blue light exposure significantly destroyed the structure and function of the retina, and the uptake of [18F]-DPA-714 in the retinas of the mice exposed to blue light were the most significantly upregulated, which was consistent with the biodistribution data. In addition, the immunohistochemical, western blot, and immunofluorescence data showed an increase in microglial TSPO expression. Conclusions: [18F]-DPA-714 micro-PET imaging might be a good method for evaluating early inflammatory status during retinal pathology.

PET imaging of retinal inflammation in mice exposed to blue light using [18F]-DPA-714.

Purpose: Positron emission tomography (PET) is widely used in high-precision imaging, which may provide a simple and noninvasive method for the detection of pathology and therapeutic effects. [18F]-DPA-714 is a second-generation translocator protein (TSPO) positron emission tomography radiotracer that shows great promise in a model of neuroinflammation. In this study, [18F]-DPA-714 micro-PET imaging was used to evaluate retinal inflammation in mice exposed to blue light, a well-established model of age-related macular degeneration (AMD) for molecular mechanism research and drug screening. Methods: C57BL/6J melanized mice were subjected to 10,000, 15,000, and 20,000 lux blue light for 5 days (8 h/day) to develop the retinal injury model, and the structure and function of the retina were assessed using hematoxylin-eosin (HE) staining, electroretinography (ERG), and terminal-deoxynucleotidyl transferase (TdT)-mediated nick-end labeling (TUNEL) immunostaining. Then, [18F]-DPA-714 was injected approximately 100 µCi through each tail vein, and static imaging was performed 1 h after injection. Finally, the mice eyeballs were collected for biodistribution and immune analysis. Results: The blue light exposure significantly destroyed the structure and function of the retina, and the uptake of [18F]-DPA-714 in the retinas of the mice exposed to blue light were the most significantly upregulated, which was consistent with the biodistribution data. In addition, the immunohistochemical, western blot, and immunofluorescence data showed an increase in microglial TSPO expression. Conclusions: [18F]-DPA-714 micro-PET imaging might be a good method for evaluating early inflammatory status during retinal pathology.

Feasibility of in vivo CAR T cells tracking using streptavidin-biotin-paired positron emission tomography.

BACKGROUND: A novel reporter system, streptavidin (SA)- [68 Ga]Ga-labeled biotin ([68 Ga]Ga-DOTA-biotin), was constructed and its ability for PET imaging the behaviors of CAR T cells were also evaluated in this study. METHODS: In vitro activity and cytotoxicity of the SA transduced anti-CD19-CAR T (denoted as SA-CD19-CAR T) cells were determined. The feasibility of monitoring proliferation profiles of SA-CD19-CAR T cells using [68 Ga]Ga-DOTA-biotin was firstly investigated in a solid tumor model. Also, the pharmacodynamics and pharmacokinetics of the CAR T cells in whole-body hematologic neoplasms were evaluated by bioluminescence imaging and [68 Ga]Ga-DOTA-biotin PET imaging simultaneously. RESULTS: After transduction with SA, the activity and cytotoxicity of the modified CAR T cells were not affected. PET images revealed that the uptakes of [68 Ga]Ga-DOTA-biotin in CD19+ K562 solid tumors were 0.67 ± 0.32 ID%/g and 1.26 ± 0.13 ID%/g at 30 min and 96 h p.i. after administration of SA-CD19-CAR T cells respectively. It confirmed that the SA-CD19-CAR T cells could effectively inhibit the growth of Raji hematologic tumors. However, low radioactivity related to the proliferation of CD19-CAR T cells was detected in the Raji model. CONCLUSION: SA-CD19-CAR T cells were constructed successfully without disturbing the antitumor functions of the cells. The proliferation of the CAR T cells in solid tumors could be early detected by [68 Ga]Ga-DOTA-biotin PET imaging.

Immuno-PET Imaging of TNF-α in Colitis Using 89Zr-DFO-infliximab.

Tumor necrosis factor-alpha (TNF-α) neutralization has become increasingly important in the treatment of inflammatory bowel diseases (IBD). A series of monoclonal antibodies were approved in the clinic for anti-TNF-α therapy. However, a comprehensive assessment of TNF-α levels throughout the colon, which facilitates the diagnosis of IBD and predicts anti-TNF-α efficacy, remains challenging. Here, we radiolabeled infliximab with long-lived radionuclides 89Zr for immuno-positron emission tomography (PET) imaging of TNF-α in vivo. The increased TNF-α level was detected in the inflammatory colon of the dextran sodium sulfate-induced colitis mice. The immuno-PET imaging of 89Zr-desferrioxamine-infliximab reveals a high uptake (7.1 ± 0.3%ID/g) in the inflammatory colon, which is significantly higher than in the healthy control and blocked groups. The colon-to-muscle ratio reached more than 10 and was maintained at a high level for 10 h after injection. The ex vivo biodistribution study also verified the superior uptake in the inflammatory colon. This study provides an in vivo immune-PET approach to molecular imaging of the pro-inflammatory cytokine TNF-α. It is promising in diagnosing and predicting efficacy in both IBD and other autoimmune diseases.

Pharmacokinetic and pharmacodynamic studies of CD19 CAR T cell in human leukaemic xenograft models with dual-modality imaging.

In recent years, chimeric antigen receptor T (CAR T)-cell therapy has shown great potential in treating haematologic disease, but no breakthrough has been achieved in solid tumours. In order to clarify the antitumour mechanism of CAR T cell in solid tumours, the pharmacokinetic (PK) and pharmacodynamic (PD) investigations of CD19 CAR T cell were performed in human leukaemic xenograft mouse models. For PK investigation, we radiolabelled CD19 CAR T cell with 89 Zr and used PET imaging in the CD19-positive and the CD19-negative K562-luc animal models. For PD evaluation, optical imaging, tumour volume measurement and DNA copy-number detection were performed. Unfortunately, the qPCR results of the DNA copy number in the blood were below the detection limit. The tumour-specific uptake was higher in the CD19-positive model than in the CD19-negative model, and this was consistent with the PD results. The preliminary PK and PD studies of CD19 CAR T cell in solid tumours are instructive. Considering the less efficiency of CAR T-cell therapy of solid tumours with the limited number of CAR T cells entering the interior of solid tumours, this study is suggestive for the subsequent CAR T-cell design and evaluation of solid tumour therapy.

Tumor-selective blockade of CD47 signaling with a CD47/PD-L1 bispecific antibody for enhanced anti-tumor activity and limited toxicity.

CD47, an immune checkpoint receptor frequently unregulated in various blood and solid tumors, interacts with ligand SIPRα on innate immune cells, and conveys a "do not eat me" signal to inhibit macrophage-mediated tumor phagocytosis. This makes CD47 a valuable target for cancer immunotherapy. However, the therapeutic utility of CD47-SIRPα blockade monoclonal antibodies is largely compromised due to significant red blood cell (RBCs) toxicities and fast target-mediated clearance as a result of extensive expression of CD47 on normal cells. To overcome these limitations and further improve therapeutic efficacy, we designed IBI322, a CD47/PD-L1 bispecific antibody which attenuated CD47 activity in monovalent binding and blocked PD-L1 activity in bivalent binding. IBI322 selectively bound to CD47+PD-L1+ tumor cells, effectively inhibited CD47-SIRPα signal and triggered strong tumor cell phagocytosis in vitro, but only with minimal impact on CD47 single positive cells such as human RBCs. In addition, as a dual blocker of innate and adaptive immune checkpoints, IBI322 effectively accumulated in PD-L1-positive tumors and demonstrated synergistic activity in inducing complete tumor regression in vivo. Furthermore, IBI322 showed only marginal RBCs depletion and was well tolerated in non-human primates (NHP) after repeated weekly injections, suggesting a sufficient therapeutic window in future clinical development of IBI322 for cancer treatment.

68Ga-NOTA PET imaging for gastric emptying assessment in mice.

BACKGROUND: Positron emission tomography (PET) has the potential for visualization and quantification of gastric emptying (GE). The traditional Chinese medicine (TCM) has been recognized promising for constipation. This study aimed to establish a PET imaging method for noninvasive GE measurement and to evaluate the efficacy of a TCM on delayed GE caused by constipation using PET imaging. METHODS: [68Ga]Ga-NOTA was synthesized as the tracer and sesame paste with different viscosity were selected as test meals. The dynamic PET scans were performed after [68Ga]Ga-NOTA mixed with test meals were administered to normal mice. Two methods were utilized for the quantification of PET imaging. A constipation mouse model was treated with maren chengqi decoction (MCD), and the established PET imaging scans were performed after the treatment. RESULTS: [68Ga]Ga-NOTA was synthesized within 20 min, and its radiochemical purity was > 95%. PET images showed the dynamic process of GE. %ID/g, volume, and total activity correlated well with each other. Among which, the half of GE time derived from %ID/g for 4 test meals were 3.92 ± 0.87 min, 13.1 ± 1.25 min, 17.8 ± 1.31 min, and 59.7 ± 3.11 min, respectively. Constipation mice treated with MCD showed improved body weight and fecal conditions as well as ameliorated GE measured by [68Ga]Ga-NOTA PET. CONCLUSIONS: A PET imaging method for noninvasive GE measurement was established with stable radiotracer, high image quality, and reliable quantification methods. The efficacy of MCD on delayed GE was demonstrated using PET.

Feasibility study of 68Ga-labeled CAR T cells for in vivo tracking using micro-positron emission tomography imaging.

Clinical tracking of chimeric antigen receptor (CAR) T cells in vivo by positron emission tomography (PET) imaging is an area of intense interest. But the long-lived positron emitter-labeled CAR T cells stay in the liver and spleen for days or even weeks. Thus, the excessive absorbed effective dose becomes a major biosafety issue leading it difficult for clinical translation. In this study we used 68Ga, a commercially available short-lived positron emitter, to label CAR T cells for noninvasive cell tracking in vivo. CAR T cells could be tracked in vivo by 68Ga-PET imaging for at least 6 h. We showed a significant correlation between the distribution of 89Zr and 68Ga-labeled CAR T cells in the same tissues (lungs, liver, and spleen). The distribution and homing behavior of CAR T cells at the early period is highly correlated with the long-term fate of CAR T cells in vivo. And the effective absorbed dose of 68Ga-labeled CAR T cells is only one twenty-fourth of 89Zr-labeled CAR T cells, which was safe for clinical translation. We conclude the feasibility of 68Ga instead of 89Zr directly labeling CAR T cells for noninvasive tracking of the cells in vivo at an early stage based on PET imaging. This method provides a potential solution to the emerging need for safe and practical PET tracer for cell tracking clinically.

Self-Assembling Nonconjugated Poly(amide-imide) into Thermoresponsive Nanovesicles with Unexpected Red Fluorescence for Bioimaging.

Nonconjugated red fluorescent polymers have been increasingly studied to improve the biocompatibility and penetration depth over conventional fluorescent materials. However, the accessibility of such polymers remains challenging due to the scarcity of nonconjugated fluorophores and lacking relevant mechanism of red-shifted fluorescence. Herein, we discovered that the combination of hydrogen bonding and π-π stacking interactions provides nonconjugated poly(amide-imide) with a large bathochromic shift (>100 nm) from blue-green fluorescence to red emission. The amphiphilic PEGylated poly(amide-imide) derived from in situ PEGylation self-assembled into nanovesicles in water, which isolated the aminosuccinimide fluorophore from the solvents and suppressed the hydrogen bonds formation between aminosuccinimide fluorophores and water. Therefore, the fluorescence of PEGylated poly(amide-imide) in water was soundly retained. Furthermore, the strong hydrogen bonding and hydrophobic interactions with water provided PEGylated poly(amide-imide) with a reversible thermoresponsiveness and presented a concentration-dependent behavior. Finally, accompanied by the excellent biostability and photostability, PEGylated poly(amide-imide) exhibited as a good candidate for cell imaging.

Combinatory effects of vaccinia virus VG9 and the STAT3 inhibitor Stattic on cancer therapy.

The recombinant vaccinia virus VG9 and the STAT3 inhibitor Stattic were combined to kill cancer cells via both oncolytic activity and inhibition of STAT3 phosphorylation in cells. The combinatory anti-tumour activity of these compounds was superior to the activity of VG9 or Stattic alone in vivo. The inhibition of tumour growth occurred via increased apoptosis and autophagy pathways. Furthermore, the combinatory anti-tumour activity was more efficient than that of VG9 or Stattic alone on xenografts, especially in nude mice.

Evaluating the utility of human glucagon-like peptide 1 receptor gene as a novel radionuclide reporter gene: a promising molecular imaging tool.

Radiolabelled ligands of glucagon-like peptide 1 receptor (GLP-1R) have been used to image the GLP-1R-expressing tissues (e.g., islets and insulinoma). Here, we introduced human glucagon-like peptide 1 receptor (hglp-1r) gene as a novel radionuclide reporter gene to broaden its applications in molecular imaging in vivo. Transient and stable baculoviral vectors (BV) were re-constructed and used to transfer the hglp-1r gene or enhanced green fluorescent protein (egfp) reporter gene into the stem cells or tumor cells. Cell proliferation assay and flow cytometry analysis demonstrated that BV-mediated reporter gene transferring and expression was biosafe and highly efficient. The BV-mediated exogenous hGLP-1R in target cells showed same ligand-receptor binding characteristics compared with its counterpart in insulinoma cells. Furthermore, the ligand-receptor binding assay showed a high affinity (IC50 = 0.3708 nM) and robust correlation (R2 = 0.9264) between the fluorescein-tagged or radiolabeled ligand probes and exogenous hGLP-1R in target cells. The target cells transferred with BV-mediated hGLP-1R could be clearly visualized in nude mice by micro-PET, which was capable of the purposes of short-term tracking transplanted stem cells or long-term monitoring tumor formation. Then, the image-based analysis and bio-distribution analysis quantitatively confirmed high target-to-background ratio of hGLP-1R-expressing cells. This study also investigated the endogenous GLP-1R-expressing organs/tissues in nude mice in the hGLP-1R radionuclide reporter gene imaging. Summarily, we evaluated the utility of hglp-1r gene as a novel radionuclide reporter gene, and demonstrated that it was a favorable and promising candidate of molecular imaging tool, which would expand the spectrum of radionuclide reporter gene imaging systems.

A Method for Detecting Dynamic Mutation of Complex Systems Using Improved Information Entropy.

In this study, a nonlinear analysis method called improved information entropy (IIE) is proposed on the basis of constructing a special probability mass function for the normalized analysis of Shannon entropy for a time series. The definition is directly applied to several typical time series, and the characteristic of IIE is analyzed. This method can distinguish different kinds of signals and reflects the complexity of one-dimensional time series of high sensitivity to the changes in signal. Thus, the method is applied to the fault diagnosis of a rolling bearing. Experimental results show that the method can effectively extract the sensitive characteristics of the bearing running state and has fast operation time and minimal parameter requirements.

Microglial activation mediates chronic mild stress-induced depressive- and anxiety-like behavior in adult rats.

BACKGROUND: Depression is a heterogeneous disorder, with the exact neuronal mechanisms causing the disease yet to be discovered. Recent work suggests it is accompanied by neuro-inflammation, characterized, in particular, by microglial activation. However, microglial activation and its involvement in neuro-inflammation and stress-related depressive disorders are far from understood. METHODS: We utilized multiple detection methods to detect the neuro-inflammation in the hippocampus of rats after exposure to chronic mild stress (CMS). Male Sprague Dawley (SD) rats were subjected to chronic mild stressors for 12 weeks. Microglial activation and hippocampal neuro-inflammation were detected by using a combinatory approach of in vivo [18F] DPA-714 positron emission computed tomography (PET) imaging, ionized calcium-binding adapter molecule 1 and translocator protein (TSPO) immunohistochemistry, and detection of NOD-like receptor protein 3 (NLRP3) inflammasome and some inflammatory mediators. Then, the rats were treated with minocycline during the last 4 weeks to observe its effect on hippocampal neuro-inflammation and depressive-like behavior induced by chronic mild stress. RESULTS: The results show that 12 weeks of chronic mild stress induced remarkable depressive- and anxiety-like behavior, simultaneously causing hippocampal microglial activation detected by PET, immunofluorescence staining, and western blotting. Likewise, activation of NLRP3 inflammasome and upregulation of inflammatory mediators, such as interleukin-1ß (IL-1ß), IL-6, and IL-18, were also observed in the hippocampus after exposure to chronic stress. Interestingly, the anti-inflammatory mediators, such as IL-4 and IL-10, were also increased in the hippocampus following chronic mild stress, which may hint that chronic stress activates different types of microglia, which produce pro-inflammatory cytokines or anti-inflammatory cytokines. Furthermore, chronic minocycline treatment alleviated the depressive-like behavior induced by chronic stress and significantly inhibited microglial activation. Similarly, the activation of NLRP3 inflammasome and the increase of inflammatory mediators were not exhibited or significantly less marked in the minocycline treatment group. CONCLUSION: These results together indicate that microglial activation mediates the chronic mild stress-induced depressive- and anxiety-like behavior and hippocampal neuro-inflammation.

Retrofitting baculoviral vector with Sleeping Beauty transposon system: competent for long-term reporter gene imaging in vivo.

Reporter gene imaging is widely used for non-invasively detecting tumorigenesis, trafficking therapeutic cells, and monitoring treatment effect. Baculoviral vectors (BVs) have been utilized as transgenic vectors in the reporter gene imaging systems in recent years. However, BV-mediated report gene imaging can only provide short-term investigation due to its transient transgene expression, which is incompetent for the long-term applications. In the current study, we reconstructed a series of hybrid BVs with several elements, to investigate the feasibility of this hybrid BV-mediated long-term reporter gene imaging in vivo. We showed that with the indispensable assistance of a positive-selection process, hybrid BV containing Sleeping Beauty 100× (SB) transposon system (BV-SB) could significantly prolong the enhanced green fluorescent protein (eGFP) expression for at least 180 days in vitro at nearly 100% eGFP positive percentage and over 1011 arbitrary unit total fluorescence intensity, whereas other hybrid BV-mediated transgene expression gradually faded in only 20 days. Furthermore, BV-SB-mediated eGFP fluorescent reporter gene imaging monitored tumorigenesis in the nude mice for at least 35 days. In addition, we exploited the glucagon-like peptide 1 receptor (glp-1r) gene as a radionuclide reporter gene for in vivo micro-PET imaging. At 50th day post-tumor transplantation, the micro-PET imaging showed considerable radiotracer-receptor-binding in vivo, resulted by stable high level of BV-SB-mediated GLP-1R expression in tumor. In summary, we retrofitted BV with the SB transposon system to make it competent for the long-term reporter gene imaging in vivo, which might broaden the application scopes of BV in the long-term molecular imaging and other biomedicine research fields.

Non-invasive glucagon-like peptide-1 receptor imaging in pancreas with (18)F-Al labeled Cys(39)-exendin-4.

PURPOSE: Glucagon-like peptide-1 receptor (GLP-1R) is abundantly expressed on beta cells and may be an ideal target for the pancreas imaging. Monitoring the GLP-1R of pancreas could be benefit for understanding the pathophysiology of diabetes. In the present study, (18)F-Al labeled exendin-4 analog, (18)F-Al-NOTA-MAL-Cys(39)-exendin-4, was evaluated for PET imaging GLP-1R in the pancreas. METHODS: The targeting of (18)F-Al labeled exendin-4 analog was examined in healthy and streptozotocin induced diabetic rats. Rats were injected with (18)F-Al-NOTA-MAL-Cys(39)-exendin-4 and microPET imaging was performed at 1 h postinjection, followed by ex vivo biodistribution. GLP-1R expression in pancreas was determined through post mortern examinations. RESULTS: The pancreas of healthy rats was readily visualized after administration of (18)F-Al-NOTA-MAL-Cys(39)-exendin-4, whereas the pancreas of diabetic rats, as well as those from rats co-injected with excess of unlabeled peptides, was barely visible by microPET. At 60 min postinjection, the pancreatic uptakes were 1.02 ± 0.15%ID/g and 0.23 ± 0.05%ID/g in healthy and diabetic rats respectively. Under block, the pancreatic uptakes of non-diabetic rats reduced to 0.21 ± 0.07%ID/g at the same time point. Biodistribution data and IHC staining confirmed the findings of the microPET imaging. CONCLUSION: The favorable preclinical data indicated that (18)F-Al-NOTA-MAL-Cys(39)-exendin-4may be suitable for non-invasive monitoring functional pancreatic beta cells.

A new protocol for the synthesis of 4,7,12,15-tetrachloro[2.2]paracyclophane.

We report a green and convenient protocol to prepare 4,7,12,15-tetrachloro[2.2]paracyclophane, the precursor of parylene D, from 2,5-dichloro-p-xylene. In the first bromination step, with H2O2-HBr as a bromide source, this procedure becomes organic-waste-free and organic-solvent-free and can appropriately replace the existing bromination methods. The Winberg elimination-dimerization step, using aqueous sodium hydroxide solution instead of silver oxide for anion exchange, results in a significant improvement in product yield. Furthermore, four substituted [2.2]paracyclophanes were also prepared in this convenient way.

(18)F-Alfatide II PET/CT in healthy human volunteers and patients with brain metastases.

PURPOSE: We report the biodistribution and radiation dosimetry of an integrin αvß3 specific PET tracer (18)F-AlF-NOTA-E[PEG4-c(RGDfk)]2) (denoted as (18)F-Alfatide II). We also assessed the value of (18)F-Alfatide II in patients with brain metastases. METHODS: A series of torso (from the skull to the thigh) static images were acquired in five healthy volunteers (3 M, 2 F) at 5, 10, 15, 30, 45, and 60 min after injection of (18)F-Alfatide II (257 ± 48 MBq). Regions of interest (ROIs) were drawn manually, and the time-activity curves (TACs) were obtained for major organs. Nine patients with brain metastases were examined by static PET imaging with (18)F-FDG (5.55 MBq/kg) and (18)F-Alfatide II. RESULTS: Injection of (18)F-Alfatide II was well tolerated in all healthy volunteers, with no serious tracer-related adverse events found. (18)F-Alfatide II showed rapid clearance from the blood pool and kidneys. The total effective dose equivalent (EDE) and effective dose (ED) were 0.0277 ± 0.003 mSv/MBq and 0.0198 ± 0.002 mSv/MBq, respectively. The organs with the highest absorbed dose were the kidneys and the spleen. Nine patients with 20 brain metastatic lesions identified by MRI and/or CT were enrolled in this study. All 20 brain lesions were visualized by (18)F-Alfatide II PET, while only ten lesions were visualized by (18)F-FDG, and 13 by CT. CONCLUSION: F-Alfatide II is a safe PET tracer with a favorable dosimetry profile. The observed ED suggests that (18)F-Alfatide II is feasible for human studies. (18)F-Alfatide II has potential value in finding brain metastases of different cancers as a biomarker of angiogenesis.

A new (68)Ga-labeled BBN peptide with a hydrophilic linker for GRPR-targeted tumor imaging.

Bombesin (BBN) is a peptide exhibiting high affinity for the gastrin-releasing peptide receptor (GRPR), which is overexpressed on several types of cancers. Various GRPR antagonists and agonists have been labeled with radiometals for positron emission tomography (PET) imaging of GRPR-positive tumors. However, unfavorable hepatobiliary excretion such as high intestinal activity may prohibit their clinical utility for imaging abdominal cancer. In this study, the modified BBN peptide with a new hydrophilic linker was labeled with (68)Ga for PET imaging of GRPR-expressing PC-3 prostate cancer xenograft model. GRPR antagonists, MATBBN (Gly-Gly-Gly-Arg-Asp-Asn-D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-NHCH2CH3) and ATBBN (D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-NHCH2CH3), were conjugated with 1,4,7-triazacyclononanetriacetic acid (NOTA) and labeled with (68)Ga. Partition coefficient and in vitro stability were also determined. GRPR binding affinity of both tracers was investigated by competitive radioligand binding assay. The in vivo receptor targeting potential and pharmacokinetic of (68)Ga-NOTA-MATBBN were also evaluated in PC-3 prostate tumor model and compared with those of (68)Ga-NOTA-ATBBN. NOTA-conjugated BBN analogs were labeled with (68)Ga within 20 min with a decay-corrected yield ranging from 90 to 95 % and a radiochemical purity of more than 98 %. The specific activity of (68)Ga-NOTA-MATBBN and (68)Ga-NOTA-ATBBN was at least 16.5 and 11.9 GBq/µmol, respectively. The radiotracers were stable in phosphate-buffered saline and human serum. (68)Ga-NOTA-MATBBN was more hydrophilic than (68)Ga-NOTA-ATBBN, as indicated by their log P values (-2.73 ± 0.02 vs. -1.20 ± 0.03). The IC50 values of NOTA-ATBBN and NOTA-MATBBN were similar (102.7 ± 1.18 and 124.6 ± 1.21 nM). The accumulation of (68)Ga-labeled GRPR antagonists in the subcutaneous PC-3 tumors could be visualized via small animal PET. The tumors were clearly visible, and the tumor uptakes of (68)Ga-NOTA-MATBBN and (68)Ga-NOTA-ATBBN were determined to be 4.19 ± 0.32, 4.00 ± 0.41, 2.93 ± 0.35 and 4.70 ± 0.40, 4.10 ± 0.30, 3.14 ± 0.30 %ID/g at 30, 60, and 120 min, respectively. There was considerable accumulation and retention of (68)Ga-NOTA-ATBBN in the liver and intestines. In contrast, the abdominal area does not have much retention of (68)Ga-NOTA-MATBBN. Biodistribution data were in accordance with the PET results, showing that (68)Ga-NOTA-MATBBN had more favorable pharmacokinetics and higher tumor to background ratios than those of (68)Ga-NOTA-ATBBN. At 1 h postinjection, the tumor to liver and intestine of (68)Ga-NOTA-MATBBN were 8.05 ± 0.56 and 21.72 ± 3.47 and the corresponding values of unmodified counterpart were 0.85 ± 0.23 and 3.45 ± 0.43, respectively. GRPR binding specificity was demonstrated by reduced tumor uptake of radiolabeled tracers after coinjection of an excess of unlabeled BBN peptides. (68)Ga-NOTA-MATBBN exhibited GRPR-targeting properties both in vitro and in vivo. The favorable characterizations of (68)Ga-NOTA-MATBBN such as convenient synthesis, specific GRPR targeting, high tumor uptake, and satisfactory pharmacokinetics warrant its further investigation for clinical cancer imaging.