Cerenkov radiation allows in vivo optical imaging of positron emitting radiotracers.

Phys Med Biol. 2010 Jan 21;55(2):483-95

Authors: Spinelli AE, D'Ambrosio D, Calderan L, Marengo M, Sbarbati A, Boschi F

In this paper, we showed that Cerenkov radiation (CR) escaping from the surface of small living animals injected with (18)F-FDG can be detected with optical imaging techniques. (18)F decays by emitting positrons with a maximum energy of 0.635 MeV; such positrons, when travelling into tissues faster than the speed of light in the same medium, are responsible of CR emission. A detailed model of the CR spectrum considering the positron energy spectrum was developed in order to quantify the amount of light emission. The results presented in this work were obtained using a commercial optical imager equipped with charged coupled detectors (CCD). Our data open the door to optical imaging (OI) in vivo of the glucose metabolism, at least in pre-clinical research. We found that the heart and bladder can be clearly identified in the animal body reflecting the accumulation of the (18)F-FDG. Moreover, we describe two different methods based on the spectral analysis of the CR that can be used to estimate the depth of the source inside the animal. We conclude that (18)F-FDG can be employed as it is as a bimodal tracer for positron emission tomography (PET) and OI techniques. Our results are encouraging, suggesting that it could be possible to apply the proposed approach not only to beta(+) but also to pure beta(-) emitters.

PMID: 20023328 [PubMed - indexed for MEDLINE]

Design and optimization of coin-shaped microreactor chips for PET radiopharmaceutical synthesis.

J Nucl Med. 2010 Feb;51(2):282-7

Authors: Elizarov AM, van Dam RM, Shin YS, Kolb HC, Padgett HC, Stout D, Shu J, Huang J, Daridon A, Heath JR

An integrated elastomeric microfluidic device, with a footprint the size of a postage stamp, has been designed and optimized for multistep radiosynthesis of PET tracers. METHODS: The unique architecture of the device is centered around a 5-microL coin-shaped reactor, which yields reaction efficiency and speed from a combination of high reagent concentration, pressurized reactions, and rapid heat and mass transfer. Its novel features facilitate mixing, solvent exchange, and product collection. New mixing mechanisms assisted by vacuum, pressure, and chemical reactions are exploited. RESULTS: The architecture of the reported reactor is the first that has allowed batch-mode microfluidic devices to produce radiopharmaceuticals of sufficient quality and quantity to be validated by in vivo imaging. CONCLUSION: The reactor has the potential to produce multiple human doses of (18)F-FDG; the most impact, however, is expected in the synthesis of PET radiopharmaceuticals that can be made only with low yields by currently available equipment.

PMID: 20124050 [PubMed - indexed for MEDLINE]

Molecular neuroimaging in rodents: assessing receptor expression and function.

Eur J Neurosci. 2009 Nov;30(10):1860-9

Authors: Mueggler T, Baltes C, Rudin M

Multimodal non-invasive neuroimaging in rodents constitutes an attractive tool for studying neurobiological processes in vivo. At present, imaging studies of brain anatomy and function as well as the investigation of structure-function relationships belong to the standard repertoire of neuroscientists. Molecular imaging adds a new perspective. The mapping of the receptor distribution and receptor occupancy can nowadays be complemented by specific readouts of receptor function either by visualizing the activity of signaling pathways or mapping the physiological consequences of receptor stimulation. Molecular information is obtained through the use of imaging probes that combine a target-specific ligand with a reporter moiety that generates a signal that can be detected from outside the body. For imaging probes targeting the central nervous system, penetration of the intact blood-brain barrier constitutes a major hurdle. Molecular imaging generates specific information and therefore has a large potential for disease phenotyping (diagnostics), therapy development and monitoring of treatment response. Molecular imaging is still in its infancy and major developments in imaging technology, probe design and data analysis are required in order to make an impact. Rodent molecular neuroimaging will play an important role in the development of these tools.

PMID: 19912336 [PubMed - indexed for MEDLINE]

Biodistribution and Imaging of 1-(2-Deoxy-beta-D-Ribofuranosyl)-2,4-Difluoro-5-[(123/125)I]Iodobenzene (dRF[(123/125)I]IB), A Nonpolar Thymidine-Mimetic Nucleoside, in Rats and Tumor-Bearing Mice.

Nucleosides Nucleotides Nucleic Acids. 2009 May;28(5):379-93

Authors: Stahlschmidt A, Khalili P, Sun W, Machulla HJ, Knaus EE, Wiebe LI

1-(2-Deoxy-beta-D-ribofuranosyl)-2,4-difluoro-5-iodobenzene (dRFIB) is a putative bioisostere of iododeoxyuridine (IUdR). The advantages of dRFIB over IUdR for in vivo studies include resistance to both phosphorolytic cleavage of the nucleoside bond and de-iodination. dRFIB was radioiodinated (dRF(123/125)IB) by copper-catalyzed exchange using commercial sodium [(123/125)I]iodide. The in vivo biodistribution of dRF[(125)I]IB in BALBc mice and imaging of dRF[(123)I]IB in Sprague-Dawley rats are reported. In vivo data for rats show rapid clearance of radioactivity from blood (>95%ID in 15 minutes), extensive excretion in urine (56%ID/24 hours), concentration in the hepatobiliary-small intestine system and very little fecal excretion ( approximately 3%ID/24 hours). Pharmacokinetic data for dRF[(125)I]IB (i.v. 48.7 ug/kg) in rats (t(1/2)[h] = 0.51 +/- 0.14, AUC(inf)[mug.min/mL] = 3.7 +/- 0.4, Cl[L/kg/h] = 0.75 +/- 0.12, Vss[L/kg] = 0.96 +/- 0.18) confirm previously reported dose-dependent pharmacokinetics. Scintigraphic images of rats dosed with dRF[(123)I]I were compatible with rapid soft-tissue clearance and extensive accumulation of radioactivity in bladder/urine and liver/small intestine. In tumor-bearing mice, thyroid and stomach radioactivity was indicative of moderate deiodination. An unidentified polar radioactive metabolite was detected in serum.

PMID: 20183590 [PubMed - in process]

Evaluation of Bifunctional Chelates for the Development of Gallium-Based Radiopharmaceuticals.

Bioconjug Chem. 2010 Feb 22;

Authors: Ferreira CL, Lamsa E, Woods M, Duan Y, Fernando P, Bensimon C, Kordos M, Guenther K, Jurek P, Kiefer GE

Ga radioisotopes, including the generator-produced positron-emitting isotope (68)Ga (t(1/2) = 68 min), are of increasing interest for the development of new radiopharmaceuticals. Bifunctional chelates (BFCs) that can be efficiently radiolabeled with Ga to yield complexes with good in vivo stability are needed. To this end, we undertook a systematic comparison of four BFCs containing different chelating moieties: two novel BFCs, p-NO(2)-Bn-Oxo (1-oxa-4,7,10-triazacyclododecane-4,7,10-triacetic acid) and p-NO(2)-Bn-PCTA (3,6,9,15-tetraazabicyclo [9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid), and two more commonly used BFCs, p-NO(2)-Bn-DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) and p-NO(2)-Bn-NOTA (1,4,7-triazacyclononane-1,4,7-triacetic acid). Each BFC was compared with respect to radiolabeling conditions, radiochemical yield, stability, and in vivo clearance properties. p-NO(2)-Bn-PCTA, p-NO(2)-Bn-Oxo, and p-NO(2)-Bn-NOTA were all more efficiently radiolabeled with Ga compared to p-NO(2)-Bn-DOTA. p-NO(2)-Bn-DOTA required longer reaction time, higher concentrations of BFC, or heating to obtain equivalent radiochemical yields. Better stability was observed for p-NO(2)-Bn-NOTA and p-NO(2)-Bn-PCTA compared to p-NO(2)-Bn-DOTA and p-NO(2)-Bn-Oxo, especially with respect to transmetalation to transferrin. Ga-radiolabled p-NO(2)-Bn-Oxo was found to be kinetically labile and therefore unstable in vivo. Ga-radiolabeled p-NO(2)-Bn-NOTA and p-NO(2)-Bn-PCTA were relatively inert, while Ga-radiolabeled p-NO(2)-Bn-DOTA had intermediate stability, losing >20% of Ga in less than one hour when incubated with apo-transferrin. Similar stability differences were seen when incubating at pH 2. In vivo PET imaging and biodistribution studies in mice showed that (68)Ga-radiolabeled p-NO(2)-Bn-PCTA, p-NO(2)-Bn-NOTA, and p-NO(2)-Bn-DOTA all cleared through the kidneys. While there was no statistical difference in the biodistribution results of (68)Ga-radiolabeled p-NO(2)-Bn-PCTA and p-NO(2)-Bn-DOTA, (68)Ga-radiolabeled p-NO(2)-Bn-NOTA cleared more rapidly from blood and muscle tissue but retained at up to 5 times higher activity in the kidneys.

PMID: 20175523 [PubMed - as supplied by publisher]

Impact of hydrogel nanoparticle size and functionalization on in vivo behavior for lung imaging and therapeutics.

Mol Pharm. 2009 Nov-Dec;6(6):1891-902

Authors: Liu Y, Ibricevic A, Cohen JA, Cohen JL, Gunsten SP, Fréchet JM, Walter MJ, Welch MJ, Brody SL

Polymer chemistry offers the possibility of synthesizing multifunctional nanoparticles which incorporate moieties that enhance diagnostic and therapeutic targeting of cargo delivery to the lung. However, since rules for predicting particle behavior following modification are not well-defined, it is essential that probes for tracking fate in vivo are also included. Accordingly, we designed polyacrylamide-based hydrogel particles of differing sizes, functionalized with a nona-arginine cell-penetrating peptide (Arg(9)), and labeled with imaging components to assess lung retention and cellular uptake after intratracheal administration. Radiolabeled microparticles (1-5 microm diameter) and nanoparticles (20-40 nm diameter) without and with Arg(9) showed diffuse airspace distribution by positron emission tomography imaging. Biodistribution studies revealed that particle clearance and extrapulmonary distribution was, in part, size dependent. Microparticles were rapidly cleared by mucociliary routes but, unexpectedly, also through the circulation. In contrast, nanoparticles had prolonged lung retention enhanced by Arg(9) and were significantly restricted to the lung. For all particle types, uptake was predominant in alveolar macrophages and, to a lesser extent, lung epithelial cells. In general, particles did not induce local inflammatory responses, with the exception of microparticles bearing Arg(9). Whereas microparticles may be advantageous for short-term applications, nanosized particles constitute an efficient high-retention and non-inflammatory vehicle for the delivery of diagnostic imaging agents and therapeutics to lung airspaces and alveolar macrophages that can be enhanced by Arg(9). Importantly, our results show that minor particle modifications may significantly impact in vivo behavior within the complex environments of the lung, underscoring the need for animal modeling.

PMID: 19852512 [PubMed - indexed for MEDLINE]

PET imaging of HER1-expressing xenografts in mice with (86)Y-CHX-A''-DTPA-cetuximab.

Eur J Nucl Med Mol Imaging. 2010 Feb 13;

Authors: Nayak TK, Regino CA, Wong KJ, Milenic DE, Garmestani K, Baidoo KE, Szajek LP, Brechbiel MW

PURPOSE: Cetuximab is a recombinant, human/mouse chimeric IgG(1) monoclonal antibody that binds to the epidermal growth factor receptor (EGFR/HER1). Cetuximab is approved for the treatment of patients with HER1-expressing metastatic colorectal cancer. Limitations in currently reported radiolabeled cetuximab for PET applications prompted the development of (86)Y-CHX-A''-DTPA-cetuximab as an alternative for imaging HER1-expressing cancer. (86)Y-CHX-A''-DTPA-cetuximab can also serve as a surrogate marker for (90)Y therapy. METHODS: Bifunctional chelate, CHX-A''-DTPA was conjugated to cetuximab and radiolabeled with (86)Y. In vitro immunoreactivity was assessed in HER1-expressing A431 cells. In vivo biodistribution, PET imaging and noncompartmental pharmacokinetics were performed in mice bearing HER1-expressing human colorectal (LS-174T and HT29), prostate (PC-3 and DU145), ovarian (SKOV3) and pancreatic (SHAW) tumor xenografts. Receptor blockage was demonstrated by coinjection of either 0.1 or 0.2 mg cetuximab. RESULTS: (86)Y-CHX-A''-DTPA-cetuximab was routinely prepared with a specific activity of 1.5-2 GBq/mg and in vitro cell-binding in the range 65-75%. Biodistribution and PET imaging studies demonstrated high HER1-specific tumor uptake of the radiotracer and clearance from nonspecific organs. In LS-174T tumor-bearing mice injected with (86)Y-CHX-A''-DTPA-cetuximab alone, (86)Y-CHX-A''-DTPA-cetuximab plus 0.1 mg cetuximab or 0.2 mg cetuximab, the tumor uptake values at 3 days were 29.3 +/- 4.2, 10.4 +/- 0.5 and 6.4 +/- 0.3%ID/g, respectively, demonstrating dose-dependent blockage of the target. Tumors were clearly visualized 1 day after injecting 3.8-4.0 MBq (86)Y-CHX-A''-DTPA-cetuximab. Quantitative PET revealed the highest tumor uptake in LS-174T (29.55 +/- 2.67%ID/cm(3)) and the lowest tumor uptake in PC-3 (15.92 +/- 1.55%ID/cm(3)) xenografts at 3 days after injection. Tumor uptake values quantified by PET were closely correlated (r (2) = 0.9, n = 18) with values determined by biodistribution studies. CONCLUSION: This study demonstrated the feasibility of preparation of high specific activity (86)Y-CHX-A''-DTPA-cetuximab and its application for quantitative noninvasive PET imaging of HER1-expressing tumors. (86)Y-CHX-A''-DTPA-cetuximab offers an attractive alternative to previously labeled cetuximab for PET and further investigation for clinical translation is warranted.

PMID: 20155263 [PubMed - as supplied by publisher]

Synthesis and evaluation of 2-pyridylbenzothiazole, 2-pyridylbenzoxazole and 2-pyridylbenzofuran derivatives as (11)C-PET imaging agents for beta-amyloid plaques.

Bioorg Med Chem Lett. 2010 Jan 25;

Authors: Swahn BM, Wensbo D, Sandell J, Sohn D, Slivo C, Pyring D, Malmström J, Arzel E, Vallin M, Bergh M, Jeppsson F, Johnson AE, Juréus A, Neelissen J, Svensson S

The syntheses and SAR of new series of beta-amyloid binding agents are reported. The effort to optimize signal-to-background ratios for these ligands are described. Compounds 8, 21 and 30 displayed desirable lipophilicity and pharmacokinetic properties. Compounds 8 and 21 were evaluated with in vitro autoradiographic studies and in vivo in APP/PS1 transgenic mice. It is shown that it was possible to increase the signal-to-background ratios compared to PIB 1, as demonstrated by compounds 8 and 21.

PMID: 20153963 [PubMed - as supplied by publisher]

An improved radiosynthesis of [(18)F]AV-133: a PET imaging agent for vesicular monoamine transporter 2.

Nucl Med Biol. 2010 Feb;37(2):133-141

Authors: Zhu L, Liu Y, Plössl K, Lieberman B, Liu J, Kung HF

INTRODUCTION: Recently, a PET tracer, 9-[(18)F]fluoropropyl-(+)-dihydrotetrabenazine ([(18)F]AV-133), targeting vesicular monoamine transporter 2 (VMAT2) in the central nervous system has been reported. It is currently under Phase II clinical trials to establish its usefulness in the diagnosis of neurodegenerative diseases including dementia with Lewy bodies and Parkinson's disease. The radiolabeling of [(18)F]AV-133, nucleophilic fluorination reaction and potential effects of pseudo-carrier were evaluated by in vivo biodistribution. METHODS: The preparation of [(18)F]AV-133 was evaluated under different conditions, specifically by employing different precursors (-OTs or -Br as the leaving group at the 9-propoxy position), reagents (K222/K(2)CO(3) vs. tributylammonium bicarbonate) and solvents (acetonitrile vs. DMSO), reaction temperature and reaction time. With optimized conditions from these experiments, radiosynthesis and purification with solid-phase extraction (SPE) of [(18)F]AV-133 were performed by an automated nucleophilic [(18)F]fluorination module. In vivo biodistribution in mice on [(18)F]AV-133 purified by either HPLC (no-carrier-added) or the SPE method (containing a pseudo-carrier) was performed and the results compared. RESULTS: Under a mild fluorination condition (heating at 115 degrees C for 5 min in dimethyl sulfoxide), [(18)F]AV-133 was obtained in a high yield using either -OTs or -Br as the leaving group. However, the -OTs precursor gave better radiochemical yields (>70%, thin layer chromatography analysis) compared to those of the -Br precursor. The optimized reaction conditions were successfully implemented to an automated nucleophilic fluorination module. Labeling and purification of [(18)F]AV133 were readily achieved via this automatic module in good radiochemical yield of 21-41% (n=10) in 40 min. The radiochemical purity was larger than 95%. Biodistribution of SPE-purified product (containing a pseudo-carrier) in mice showed a high striatum/cerebellum ratio (4.18+/-0.51), which was comparable to that of HPLC-purified [(18)F]AV-133 (4.51+/-0.10). CONCLUSIONS: The formation of [(18)F]AV-133 was evaluated under different labeling conditions. These improved labeling conditions and SPE purification were successfully implemented into an automated synthesis module. This offers a short preparation time (about 40 min), simplicity in operation and ready applicability for routine clinical operation.

PMID: 20152712 [PubMed - as supplied by publisher]

The Forefront for Novel Therapeutic Agents Based on the Pathophysiology of Lower Urinary Tract Dysfunction: Bladder Selectivity Based on In Vivo Drug-Receptor Binding Characteristics of Antimuscarinic Agents for Treatment of Overactive Bladder.

J Pharmacol Sci. 2010 Feb 4;

Authors: Yoshida A, Fujino T, Maruyama S, Ito Y, Taki Y, Yamada S

We have reviewed the binding of antimuscarinic agents, used to treat urinary dysfunction in patients with overactive bladder, to muscarinic receptors in target and non-target tissues in vivo. Transdermal administration of oxybutynin in rats led to significant binding in the bladder without long-term binding in the submaxillary gland and the abolishment of salivation evoked by oral oxybutynin. Oral solifenacin showed significant and long-lasting binding to muscarinic receptors in mouse tissues expressing the M(3) subtype. Oral tolterodine bound more selectively to muscarinic receptors in the bladder than in the submaxillary gland in mice. The muscarinic receptor binding activity of oral darifenacin in mice was shown to be pronounced and long-lasting in the bladder, submaxillary gland, and lung. In vivo quantitative autoradiography using (+)N-[(11)C]methyl-3-piperidyl benzilate in rats showed significant occupancy of brain muscarinic receptors on intravenous injection of oxybutynin, propiverine, solifenacin, and tolterodine. The estimated in vivo bladder selectivity compared to brain was significantly greater for solifenacin and tolterodine than oxybutynin. Darifenacin occupied few brain muscarinic receptors. Similar findings were also observed with positron emission tomography in conscious rhesus monkeys. The newer generation of antimuscarinic agents may be advantageous in the bladder selectivity after systemic administration.

PMID: 20134113 [PubMed - as supplied by publisher]

A HER2-binding Affibody molecule labelled with (68)Ga for PET imaging: direct in vivo comparison with the (111)In-labelled analogue.

Eur J Nucl Med Mol Imaging. 2010 Feb 4;

Authors: Tolmachev V, Velikyan I, Sandström M, Orlova A

PURPOSE: Overexpression of HER2 receptors is a prognostic and predictive biomarker in breast cancer and a number of other malignancies. Radionuclide molecular imaging of HER2 overexpression may influence patient management making treatment more personalized. Earlier, (111)In-DOTA-Z(HER2:342-pep2) (ABY-002) Affibody molecule demonstrated excellent imaging of HER2-expressing xenografts in mice shortly after injection. The use of the positron-emitting nuclide (68)Ga instead of (111)In might increase both the sensitivity of HER2 imaging and accuracy of expression quantification. The goal of this study was to prepare and characterize (68)Ga-labelled ABY-002. METHODS: (68)Ga labelling of ABY-002 was optimized. In vitro cell binding and procession of (68)Ga-ABY-002 was evaluated. Biodistribution and tumour targeting of (68)Ga-ABY-002 and (111)In-ABY-002 was compared in vivo by paired-label experiments. RESULTS: ABY-002 was incubated with (68)Ga at 90 degrees C for 10 min resulting in a radiochemical labelling yield of over 95%. Capacity for specific binding to HER2-expressing cells was retained. In vivo, both (68)Ga-ABY-002 and (111)In-ABY-002 demonstrated specific targeting of SKOV-3 xenografts and high-contrast imaging. Background radioactivity in blood, lungs, gastrointestinal tract and muscle fell more rapidly for (68)Ga-ABY-002 compared with (111)In-ABY-002 favouring imaging shortly after injection. For (68)Ga-ABY-002, a tumour uptake of 12.4 +/- 3.8%ID/g and a tumour to blood ratio of 31 +/- 13 were achieved at 2 h post-injection. CONCLUSION: (68)Ga-ABY-002 is easy to label and provides high-contrast imaging within 2 h after injection. This makes it a promising candidate for clinical molecular imaging of HER2 expression in malignant tumours.

PMID: 20130858 [PubMed - as supplied by publisher]

Different Strategies for Reducing Intestinal Background Radioactivity Associated with Imaging HSV1-tk Expression Using Established Radionucleoside Probes.

Mol Imaging. 2010 Feb 1;9(1):47-58

Authors: Ruggiero A, Brader P, Serganova I, Zanzonico P, Cai S, Lipman NS, Hricak H, Blasberg RG

One limitation of HSV1-tk reporter positron emission tomography (PET) with nucleoside analogues is the high background radioactivity in the intestine. We hypothesized that endogenous expression of thymidine kinase in bacterial flora could phosphorylate and trap such radiotracers, contributing to the high radioactivity levels in the bowel, and therefore explored different strategies to increase fecal elimination of radiotracer. Intestinal radioactivity was assessed by in vivo microPET imaging and ex vivo tissue sampling following intravenous injection of 18F-FEAU, 124I-FIAU, or 18F-FHBG in a germ-free mouse strain. We also explored the use of an osmotic laxative agent and/or a 100% enzymatically hydrolyzed liquid diet. No significant differences in intestinal radioactivity were observed between germ-free and normal mice. 18F-FHBG-derived intestinal radioactivity levels were higher than those of 18F-FEAU and 124I-FIAU; the intestine to blood ratio was more than 20-fold higher for 18F-FHBG than for 18F-FEAU and 124I-FIAU. The combination of Peptamen and Nulytely lowered intestinal radioactivity levels and increased (2.2-fold) the HSV1-tk transduced xenograft to intestine ratio for 18F-FEAU. Intestinal bacteria in germ-free mice do not contribute to the high intestinal levels of radioactivity following injection of radionucleoside analogues. The combination of Peptamen and Nulytely increased radiotracer elimination by increasing bowel motility without inducing dehydration.

PMID: 20128998 [PubMed - in process]

Direct detection and quantification of abasic sites for in vivo studies of DNA damage and repair.

Nucl Med Biol. 2009 Nov;36(8):975-83

Authors: Wang Y, Liu L, Wu C, Bulgar A, Somoza E, Zhu W, Gerson SL

Use of chemotherapeutic agents to induce cytotoxic DNA damage and programmed cell death is a key strategy in cancer treatments. However, the efficacy of DNA-targeted agents such as temozolomide is often compromised by intrinsic cellular responses such as DNA base excision repair (BER). Previous studies have shown that BER pathway resulted in formation of abasic or apurinic/apyrimidinic (AP) sites, and blockage of AP sites led to a significant enhancement of drug sensitivity due to reduction of DNA base excision repair. Since a number of chemotherapeutic agents also induce formation of AP sites, monitoring of these sites as a clinical correlate of drug effect will provide a useful tool in the development of DNA-targeted chemotherapies aimed at blocking abasic sites from repair. Here we report an imaging technique based on positron emission tomography (PET) that allows for direct quantification of AP sites in vivo. For this purpose, positron-emitting carbon-11 has been incorporated into methoxyamine ([(11)C]MX) that binds covalently to AP sites with high specificity. The binding specificity of [(11)C]MX for AP sites was demonstrated by in vivo blocking experiments. Using [(11)C]MX as a radiotracer, animal PET studies have been conducted in melanoma and glioma xenografts for quantification of AP sites. Following induction of AP sites by temozolomide, both tumor models showed significant increase of [(11)C]MX uptake in tumor regions in terms of radioactivity concentration as a function of time, which correlates well with conventional aldehyde reactive probe (ARP)-based bioassays for AP sites.

PMID: 19875055 [PubMed - indexed for MEDLINE]

In Vivo Evaluation of alpha7 Nicotinic Acetylcholine Receptor Agonists [C]A-582941 and [C]A-844606 in Mice and Conscious Monkeys.

PLoS One. 2010;5(2):e8961

Authors: Toyohara J, Ishiwata K, Sakata M, Wu J, Nishiyama S, Tsukada H, Hashimoto K

BACKGROUND: The alpha7 nicotinic acetylcholine receptors (nAChRs) play an important role in the pathophysiology of neuropsychiatric diseases such as schizophrenia and Alzheimer's disease. The goal of this study was to evaluate the two carbon-11-labeled alpha7 nAChR agonists [(11)C]A-582941 and [(11)C]A-844606 for their potential as novel positron emission tomography (PET) tracers. METHODOLOGY/PRINCIPAL FINDINGS: The two tracers were synthesized by methylation of the corresponding desmethyl precursors using [(11)C]methyl triflate. Effects of receptor blockade in mice were determined by coinjection of either tracer along with a carrier or an excess amount of a selective alpha7 nAChR agonist (SSR180711). Metabolic stability was investigated using radio-HPLC. Dynamic PET scans were performed in conscious monkeys with/without SSR180711-treatment. [(11)C]A-582941 and [(11)C]A-844606 showed high uptake in the mouse brain. Most radioactive compounds in the brain were detected as an unchanged form. However, regional selectivity and selective receptor blockade were not clearly observed for either compound in the mouse brain. On the other hand, the total distribution volume of [(11)C]A-582941 and [(11)C]A-844606 was high in the hippocampus and thalamus but low in the cerebellum in the conscious monkey brain, and reduced by pretreatment with SSR180711. CONCLUSIONS/SIGNIFICANCE: A nonhuman primate study suggests that [(11)C]A-582941 and [(11)C]A-844606 would be potential PET ligands for imaging alpha7 nAChRs in the human brain.

PMID: 20126539 [PubMed - in process]

Evaluation of a 64Cu-Labeled Cystine-Knot Peptide Based on Agouti-Related Protein for PET of Tumors Expressing {alpha}v{beta}3 Integrin.

J Nucl Med. 2010 Feb;51(2):251-8

Authors: Jiang L, Kimura RH, Miao Z, Silverman AP, Ren G, Liu H, Li P, Gambhir SS, Cochran JR, Cheng Z

Recently, a truncated form of the agouti-related protein (AgRP), a 4-kDa cystine-knot peptide of human origin, was used as a scaffold to engineer mutants that bound to alpha(v)beta(3) integrin with high affinity and specificity. In this study, we evaluated the potential of engineered integrin-binding AgRP peptides for use as cancer imaging agents in living subjects. METHODS: Engineered AgRP peptides were prepared by solid-phase peptide synthesis and were folded in vitro and purified by reversed-phase high-performance liquid chromatography. Competition assays were used to measure the relative binding affinities of engineered AgRP peptides for integrin receptors expressed on the surface of U87MG glioblastoma cells. The highest-affinity mutant, AgRP clone 7C, was site-specifically conjugated with 1,4,7,10-tetra-azacyclododecane-N,N',N''N'''-tetraacetic acid (DOTA). The resulting bioconjugate, DOTA-AgRP-7C, was radiolabeled with (64)Cu for biodistribution analysis and small-animal PET studies in mice bearing U87MG tumor xenografts. In addition to serum stability, the in vivo metabolic stability of (64)Cu-DOTA-AgRP-7C was assessed after injection and probe recovery from mouse kidney, liver, tumor, and urine. RESULTS: AgRP-7C and DOTA-AgRP-7C bound with high affinity to integrin receptors expressed on U87MG cells (half maximal inhibitory concentration values, 20 +/- 4 and 14 +/- 2 nM, respectively). DOTA-AgRP-7C was labeled with (64)Cu with high radiochemical purity (>99%). In biodistribution and small-animal PET studies, (64)Cu-DOTA-AgRP-7C displayed rapid blood clearance, good tumor uptake and retention (2.70 +/- 0.93 percentage injected dose per gram [%ID/g] and 2.37 +/- 1.04 %ID/g at 2 and 24 h, respectively), and high tumor-to-background tissue ratios. The integrin-binding specificity of (64)Cu-DOTA-AgRP-7C was confirmed in vitro and in vivo by showing that a large molar excess of the unlabeled peptidomimetic c(RGDyK) could block probe binding and tumor uptake. Serum stability and in vivo metabolite assays demonstrated that engineered AgRP peptides are sufficiently stable for in vivo molecular imaging applications. CONCLUSION: A radiolabeled version of the engineered AgRP peptide 7C showed promise as a PET agent for tumors that express the alpha(v)beta(3) integrin. Collectively, these results validate AgRP-based cystine-knot peptides for use in vivo as molecular imaging agents and provide support for the general use of AgRP as a scaffold to develop targeting peptides, and hence diagnostics, against other tumor receptors.

PMID: 20124048 [PubMed - in process]

Direct detection and quantification of abasic sites for in vivo studies of DNA damage and repair.

Nucl Med Biol. 2009 Nov;36(8):975-83

Authors: Wang Y, Liu L, Wu C, Bulgar A, Somoza E, Zhu W, Gerson SL

Use of chemotherapeutic agents to induce cytotoxic DNA damage and programmed cell death is a key strategy in cancer treatments. However, the efficacy of DNA-targeted agents such as temozolomide is often compromised by intrinsic cellular responses such as DNA base excision repair (BER). Previous studies have shown that BER pathway resulted in formation of abasic or apurinic/apyrimidinic (AP) sites, and blockage of AP sites led to a significant enhancement of drug sensitivity due to reduction of DNA base excision repair. Since a number of chemotherapeutic agents also induce formation of AP sites, monitoring of these sites as a clinical correlate of drug effect will provide a useful tool in the development of DNA-targeted chemotherapies aimed at blocking abasic sites from repair. Here we report an imaging technique based on positron emission tomography (PET) that allows for direct quantification of AP sites in vivo. For this purpose, positron-emitting carbon-11 has been incorporated into methoxyamine ([(11)C]MX) that binds covalently to AP sites with high specificity. The binding specificity of [(11)C]MX for AP sites was demonstrated by in vivo blocking experiments. Using [(11)C]MX as a radiotracer, animal PET studies have been conducted in melanoma and glioma xenografts for quantification of AP sites. Following induction of AP sites by temozolomide, both tumor models showed significant increase of [(11)C]MX uptake in tumor regions in terms of radioactivity concentration as a function of time, which correlates well with conventional aldehyde reactive probe (ARP)-based bioassays for AP sites.

PMID: 19875055 [PubMed - indexed for MEDLINE]

Trypsinization severely perturbs radioiodide transport via membrane Na/I symporter proteolysis: implications for reporter gene imaging.

Nucl Med Biol. 2009 Nov;36(8):967-74

Authors: Jung KH, Paik JY, Lee YL, Lee YJ, Lee J, Lee KH

INTRODUCTION: Cell preparation procedures injurious to Na/I symporters (NIS) could deter their usefulness for reporter gene assays and in vivo cell imaging. In this study, we investigated the effects of cell collection by trypsinization on radioiodide transport and in vivo cell imaging results. METHODS: The influence of trypsinization procedures on (125)I transport was evaluated using Huh-7/NIS hepatoma cells. The effects of graded concentrations of trypsin and EDTA were assessed on Huh-7/NIS and A431/NIS lung cancer cells. Trypsin-induced NIS proteolysis was investigated by immunoblots of plasma membrane prepared from adenovirus-infected mouse liver tissue. (99m)Tc-O(4)(-) scintigraphy was performed in Balb/C nude mice at 1 and 4 h following administration of Huh-7/NIS cells collected with and without trypsin. RESULTS: (125)I Transport ability of Huh-7/NIS cells was severely impaired within minutes of standard trypsinization and further deteriorated up to 24 h after termination of treatment. This perturbation was caused by trypsin, which dose- and time-dependently induced substantial reductions of (125)I uptake in Huh-7/NIS and A431/NIS cells. Immunoblot analysis revealed significant dose- and time-dependent losses of membrane NIS protein by trypsin. NIS proteolysis was completely blocked by soybean trypsin inhibitor, and partial protection was offered by the substrates iodide and perchlorate. On (99m)Tc-O(4)(-) scintigraphy of mice, cells prepared by trypsinization were poorly visualized, whereas those collected with a nonenzymatic method showed significantly better uptake and contrast. CONCLUSION: Trypsinization leads to serious perturbations in iodide accumulating capacity through tryptic degradation of membrane NIS protein. Hence, NIS-based reporter assays and in vivo cell imaging studies may benefit from better-optimized cell cultivation and harvesting procedures.

PMID: 19875054 [PubMed - indexed for MEDLINE]

Trypsinization severely perturbs radioiodide transport via membrane Na/I symporter proteolysis: implications for reporter gene imaging.

Nucl Med Biol. 2009 Nov;36(8):967-74

Authors: Jung KH, Paik JY, Lee YL, Lee YJ, Lee J, Lee KH

INTRODUCTION: Cell preparation procedures injurious to Na/I symporters (NIS) could deter their usefulness for reporter gene assays and in vivo cell imaging. In this study, we investigated the effects of cell collection by trypsinization on radioiodide transport and in vivo cell imaging results. METHODS: The influence of trypsinization procedures on (125)I transport was evaluated using Huh-7/NIS hepatoma cells. The effects of graded concentrations of trypsin and EDTA were assessed on Huh-7/NIS and A431/NIS lung cancer cells. Trypsin-induced NIS proteolysis was investigated by immunoblots of plasma membrane prepared from adenovirus-infected mouse liver tissue. (99m)Tc-O(4)(-) scintigraphy was performed in Balb/C nude mice at 1 and 4 h following administration of Huh-7/NIS cells collected with and without trypsin. RESULTS: (125)I Transport ability of Huh-7/NIS cells was severely impaired within minutes of standard trypsinization and further deteriorated up to 24 h after termination of treatment. This perturbation was caused by trypsin, which dose- and time-dependently induced substantial reductions of (125)I uptake in Huh-7/NIS and A431/NIS cells. Immunoblot analysis revealed significant dose- and time-dependent losses of membrane NIS protein by trypsin. NIS proteolysis was completely blocked by soybean trypsin inhibitor, and partial protection was offered by the substrates iodide and perchlorate. On (99m)Tc-O(4)(-) scintigraphy of mice, cells prepared by trypsinization were poorly visualized, whereas those collected with a nonenzymatic method showed significantly better uptake and contrast. CONCLUSION: Trypsinization leads to serious perturbations in iodide accumulating capacity through tryptic degradation of membrane NIS protein. Hence, NIS-based reporter assays and in vivo cell imaging studies may benefit from better-optimized cell cultivation and harvesting procedures.

PMID: 19875054 [PubMed - indexed for MEDLINE]

Design of silicon-based misonidazole analogues and (18)F-radiolabelling.

Nucl Med Biol. 2009 Nov;36(8):895-905

Authors: Bohn P, Deyine A, Azzouz R, Bailly L, Fiol-Petit C, Bischoff L, Fruit C, Marsais F, Vera P

INTRODUCTION: Development of new (18)F-labeled tracers for positron emission tomography (PET) imaging is increasingly important. Herein, we described the synthesis of silicon analogues of [(18)F]fluoromisonidazole in order to develop new radiolabelled compounds for the detection of tumour hypoxic domain. Their stabilities and their in vivo biodistribution were evaluated. METHODS: (18)F-labeled silicon-based misonidazole analogues were synthesized by alkylating 2-nitroimidazole with alkyloxy-(3-chloropropyl)dialkyl or diarylsilane. These intermediates were labeled with [(18)F]F(-) with a mixture of K(18)F and Kryptofix (K222) in acetonitrile as standard condition. PET imaging was performed using a dedicated small animal PET scanner. RESULTS: (18)F-labeled silicon-based misonidazole analogues were easily synthesized in three steps. The hydrolytic and radiolytic stability of these new fluorosilanes depend on the steric hindrance at the silicon center. Indeed, partial uptake of dimethylfluorosilane [(18)F]2a(1-(3-(Fluorodimethylsilyl)propyl)-2-nitro-1H-imidazole) in tumor hypoxic area was observed but defluorination also appeared. Moreover, PET studies indicated that, owing to its high lipophilicity, the most stable dinaphtylfluorosilane [(18)F]2d is retained mainly by the lungs. CONCLUSION: We have described an efficient and versatile approach for the synthesis of (18)F-labeled, silicon-based misonidazole analogues. PET imaging of one of these compounds revealed that hypoxia could be detected. Controlling the biodistribution of (18)F-labeled silicon-based misonidazole analogues will require additional studies.

PMID: 19875046 [PubMed - indexed for MEDLINE]

Synthesis of fluorine-18 labeled rhodamine B: A potential PET myocardial perfusion imaging agent.

Appl Radiat Isot. 2010 Jan;68(1):96-100

Authors: Heinrich TK, Gottumukkala V, Snay E, Dunning P, Fahey FH, Ted Treves S, Packard AB

There is considerable interest in developing an (18)F-labeled PET myocardial perfusion agent. Rhodamine dyes share several properties with (99m)Tc-MIBI, the most commonly used single-photon myocardial perfusion agent, suggesting that an (18)F-labeled rhodamine dye might prove useful for this application. In addition to being lipophilic cations, like (99m)Tc-MIBI, rhodamine dyes are known to accumulate in the myocardium and are substrates for Pgp, the protein implicated in MDR1 multidrug resistance. As the first step in determining whether (18)F-labeled rhodamines might be useful as myocardial perfusion agents for PET, our objective was to develop synthetic methods for preparing the (18)F-labeled compounds so that they could be evaluated in vivo. Rhodamine B was chosen as the prototype compound for development of the synthesis because the ethyl substituents on the amine moieties of rhodamine B protect them from side reactions, thus eliminating the need to include (and subsequently remove) protecting groups. The 2'-[(18)F]fluoroethyl ester of rhodamine B was synthesized by heating rhodamine B lactone with [(18)F]fluoroethyltosylate in acetonitrile at 165 degrees C for 30min using [(18)F]fluoroethyl tosylate, which was prepared by the reaction of ethyleneglycol ditosylate with Kryptofix 2.2.2, K(2)CO(3), and [(18)F]NaF in acetonitrile for 10min at 90 degrees C. The product was purified by semi-preparative HPLC to produce the 2'-[(18)F]fluoroethylester in >97% radiochemical purity with a specific activity of 1.3GBq/mumol, an isolated decay corrected yield of 35%, and a total synthesis time of 90min.

PMID: 19783150 [PubMed - indexed for MEDLINE]