Bram Maas

Detection and grading of soft tissue sarcomas of the extremities with 18F-3'-fluoro-3'-deoxy-L-thymidine

Purpose: The aim of the study was to investigate the feasibility of 18F-3′-fluoro-3′-deoxy-l-thymidine positron emission tomography (FLT-PET) for the detection and grading of soft tissue sarcoma (STS). Experimental Design: Nineteen patients with 20 STSs of the extremities were scanned, using attenuation corrected whole-body FLT-PET. Standardized uptake values (SUVs) and tumor:nontumor ratios (TNTs) were compared with histopathological parameters using French and Japanese grading systems. Results: Mean SUV, maximal SUV, and TNT could differentiate between low-grade (grade 1; n = 6) STS and high-grade (grade 2 and 3; n = 14) STS according to the French grading system (P = 0.001). Mean SUV, max SUV, and TNT correlated with mitotic score, MIB-1 score, the French and Japanese grading system (• = 0.550–0.747). Conclusions: FLT-PET is able to visualize STS and differentiate between low-grade and high-grade STS. The uptake of FLT correlates with the proliferation of STS.

Synthesis and evaluation of radiolabeled antagonists for imaging of beta-adrenoceptors in the brain with PET

Five potent, lipophilic beta-adrenoceptor antagonists (carvedilol, pindolol, toliprolol and fluorinated analogs of bupranolol and penbutolol) were labeled with either carbon-11 or fluorine-18 and evaluated for cerebral beta-adrenoceptor imaging in experimental animals. The standard radioligand for autoradiography of beta-adrenoceptors, [125I]-iodocyanopindolol, was also included in this survey. All compounds showed either very low uptake in rat brain or a regional distribution that was not related to beta-adrenoceptors, whereas some ligands did display specific binding in heart and lungs. Apparently, the criteria of a high affinity and a moderately high lipophilicity were insufficient to predict the suitability of beta-adrenergic antagonists for visualization of beta-adrenoceptors in the central nervous system.

First Clinical Results of (d)-18F-Fluoromethyltyrosine (BAY 86-9596) PET/CT in Patients with Non–Small Cell Lung Cancer and Head and Neck Squamous Cell Carcinoma

(d)-18F-fluoromethyltyrosine (d-18F-FMT), or BAY 86-9596, is a novel 18F-labeled tyrosine derivative rapidly transported by the l-amino acid transporter (LAT-1), with a faster blood pool clearance than the corresponding l-isomer. The aim of this study was to demonstrate the feasibility of tumor detection in patients with non–small cell lung cancer (NSCLC) or head and neck squamous cell cancer (HNSCC) compared with inflammatory and physiologic tissues in direct comparison to 18F-FDG. Methods: 18 patients with biopsy-proven NSCLC (n = 10) or HNSCC (n = 8) were included in this Institutional Review Board–approved, prospective multicenter study. All patients underwent 18F-FDG PET/CT scans within 21 d before d-18F-FMT PET/CT. For all patients, safety and outcome data were assessed. Results: No adverse reactions were observed related to d-18F-FMT. Fifty-two lesions were 18F-FDG–positive, and 42 of those were malignant (34 histologically proven and 8 with clinical reference). Thirty-two of the 42 malignant lesions were also d-18F-FMT–positive, and 10 lesions had no tracer uptake above the level of the blood pool. Overall there were 34 true-positive, 8 true-negative, 10 false-negative, and only 2 false-positive lesions for d-18F-FMT, whereas 18F-FDG was true-positive in 42 lesions, with 10 false-positive and only 2 false-negative, resulting in a lesion-based detection rate for d-18F-FMT and 18F-FDG of 77% and 95%, respectively, with an accuracy of 78% for both tracers. A high d-18F-FMT tumor–to–blood pool ratio had a negative correlation with overall survival (P = 0.050), whereas the 18F-FDG tumor–to–blood pool ratio did not correlate with overall survival. Conclusion: d-18F-FMT imaging in patients with NSCLC and HNSCC is safe and feasible. The presented preliminary results suggest a lower sensitivity but higher specificity for d-18F-FMT over 18F-FDG, since there is no d-18F-FMT uptake in inflammation. This increased specificity may be particularly beneficial in areas with endemic granulomatous disease and may improve clinical management. Further clinical investigations are needed to determine its clinical value and relevance for the prediction of survival prognosis.

Contribution of neuroinflammation to changes in [11C]flumazenil binding in the rat brain: Evaluation of the inflamed pons as reference tissue

INTRODUCTION [11C]Flumazenil is a well-known PET tracer for GABAA receptors and is mainly used as an imaging biomarker for neuronal loss. Recently, GABAA receptors on immune cells have been investigated as target for modulation of inflammation. Since neuronal loss is often accompanied by neuroinflammation, PET imaging with [11C]flumazenil is potentially affected by infiltrating immune cells. This may also compromise the validity of using the pons as reference tissue in quantitative pharmacokinetic analysis. This study aims to evaluate whether inflammatory processes in the brain can influence [11C]flumazenil uptake and affect the outcome of pharmacokinetic modeling when the pons is used as reference tissue. METHODS The herpes simplex encephalitis (HSE) rat model is known to cause neuroinflammation in the brainstem. Dynamic [11C]flumazenil PET scans of 60-min, accompanied by arterial blood sampling and metabolite analysis, were acquired at day 6-7days post-infection of male Wistar rats (HSE, n=5 and control, n=6). Additionally, the GABAA receptor was saturated by injection of unlabeled flumazenil prior to the tracer injection in 4 rats per group. PET data were analyzed by pharmacokinetic modeling. RESULTS No statistically significant differences were found in the volume of distribution (VT) or non-displaceable binding potential (BPND) between control and HSE rats in any of the brain regions. Pre-saturation with unlabeled flumazenil resulted in a statistically significant reduction in [11C]flumazenil VT in all brain regions. The BPND obtained from SRTM exhibited a good correlation to DVR - 1 values from the two-tissue compartment model, coupled with some level of underestimation. CONCLUSION Reliable quantification of [11C]flumazenil binding in rats can be obtained by pharmacokinetic analysis using the pons as a pseudo-reference tissue even in the presence of strong acute neuroinflammation.

Improved GMP-compliant multi-dose production and quality control of 6-[18F]fluoro-L-DOPA

Background6-[18F]Fluoro-L-3,4-dihydroxyphenylalanine (FDOPA) is a frequently used radiopharmaceutical for detecting neuroendocrine and brain tumors and for the differential diagnosis of Parkinson’s disease. To meet the demand for FDOPA, a high-yield GMP-compliant production method is required. Therefore, this study aimed to improve the FDOPA production and quality control procedures to enable distribution of the radiopharmaceutical over distances.FDOPA was prepared by electrophilic fluorination of the trimethylstannyl precursor with [18F]F2, produced from [18O]2 via the double-shoot approach, leading to FDOPA with higher specific activity as compared to FDOPA which was synthesized, using [18F]F2 produced from 20Ne, leading to FDOPA with a lower specific activity. The quality control of the product was performed using a validated UPLC system and compared with quality control with a conventional HPLC system. Impurities were identified using UPLC-MS.ResultsThe [18O]2 double-shoot radionuclide production method yielded significantly more [18F]F2 with less carrier F2 than the conventional method starting from 20Ne. After adjustment of radiolabeling parameters substantially higher amounts of FDOPA with higher specific activity could be obtained. Quality control by UPLC was much faster and detected more side-products than HPLC. UPLC-MS showed that the most important side-product was FDOPA-quinone, rather than 6-hydroxydopa as suggested by the European Pharmacopoeia.ConclusionThe production and quality control of FDOPA were significantly improved by introducing the [18O]2 double-shoot radionuclide production method, and product analysis by UPLC, respectively. As a result, FDOPA is now routinely available for clinical practice and for distribution over distances.

UPLC (R)-RAD the new standard in quality control of PET radiopharmaceuticals

Aim: Inflammatory bowel disease (IBD) is defined as a chronic relapsing idiopathic inflammation of the gastrointestinal tract. The two main clinical forms of this disease family are Crohn’s Disease (CD) and Ulcerative Colitis (UC). IBD affects an estimated 3.6 million individuals in Europe and North America. To date it is thought that IBD is the result of continual activation of the mucosal immune system. In order to better understand this disease family an in-house developed animal model was implemented and characterized with [18F]FDG (used to illustrate the increased glucose consumption associated with inflammatory processes) and also with TSPO 18 kDa radioligand [18F]DPA-714, an established radiotracer for the study of inflammation within the central nervous system. Materials and Methods: Colonic inflammation was induced in male Wistar rats weighing between 200-250 g by rectal administration of trinitrobenzenesulfonic acid (TNBS) at 4cm from the anal orifice. Control animals were administered, 0.9% aq. sodium chloride analogously. A Siemens Inveon PET/CT tomograph, dedicated to small animals, was used to acquire [18F]FDG images on day 7 post TNBS administration and [18F]DPA714 images the following day. Rats were then sacrificed by an i.v. injection of pentobarbital, and then the lower intestine was extracted and analyzed by immunohistochemistry to determine macrophage infiltration and the presence of TSPO. Results: PET image analysis clearly shows an important accumulation of both radiotracers within the intestinal walls of treated animals in comparison to control animals. Mean levels of [18F]FDG uptake in treated and control animals were 1.20 ± 0.56 %ID/cc and 0.43 ± 0.18 %ID/cc, respectively. Comparable results were found when using [18F]DPA-714, with mean level of uptake in treated and control animals of 1.21 ± 0.62 %ID/cc and 0.46 ± 0.23 %ID/cc, respectively. Immunohistochemistry analysis revealed a higher presence of macrophages in TNBS treated animals. Expression of TSPO was largely increased in the treated animals, when compared to the controls animals, and mainly localized in macrophages cells. Conclusion: Preliminary results seem to indicate that [18F]DPA-714 is an adapted tracer for the study of inflammation of IBD in our animal model. Beyond this, data demonstrating that [18F]DPA-714 could be used to characterize and quantify the level of inflammation during the disease evolution, within the TNBS treated animals, will also be presented. OP366 Surface displayed SNAP-tag as a novel tool for study of Grampositive bacterial infections. B. Mills, V. Steele, J. C. A. Luckett, R. O. Awais, P. Duncanson, V. Griffiths, A. Cockayne, M. Xu, I. Correa, A. C. Perkins, P. Williams, P. Hill; School of Molecular Medical Sciences, University of Nottingham, Nottingham, UNITED KINGDOM, Radiological and Imaging Sciences, University of Nottingham, Nottingham, UNITED KINGDOM, School of Biological and Chemical Sciences, Queen Mary University of London, London, UNITED KINGDOM, New England Biolabs, Inc, Ipswich, ME, UNITED STATES, School of Biosciences, University of Nottingham, Nottingham, UNITED KINGDOM. Introduction: The design of specific probes for in vivo molecular imaging of microbial infections remains one of the greatest challenges to overcome before useful, functional data can be obtained. An increasingly attractive approach for probe design is to express a ligand-binding protein within a cell, which may then covalently bind specific synthetic ligands with attached imaging moieties. One such labelling system is the commercially available SNAP-tag. SNAP-tag specifically and covalently binds O2-benzylguanine (BG) compounds, which may have fluorophores or other functional elements attached at the 4’ position of their benzyl ring. We have designed a BG ligand labelled with Tc, suitable for SPECT imaging. We propose to utilise this technology for the imaging of Staphylococcal infection in vivo with the view to investigate bacterial pathogenicity and to visualise the effect potential antimicrobials may have on bacterial load. Methods: The SNAP-tag gene was codon optimised for expression in the Gram positive bacterium Staphylococcus aureus and fused with an N-terminal spa secretion leader sequence and a Cterminal spa cell-wall anchoring domain. The N-terminal fusion directs the expressed SNAP-tag towards the cell exterior where the C-terminal domain is recognised by the cell-wall sorting enzyme sortase A, covalently anchoring SNAPtag in such a way that the ligand binding domain decorates the cell surface. A novel 99m Tc-HYNIC–NH-BG ligand for SPECT imaging was prepared by coupling BG to HYNIC and radiolabelling with NaTcO4 in the presence of tricine as co-ligand. Radiochemical yields >99% were obtained. nanoSPECT-CT imaging will be used to assess the functional data produced by using SNAP-tag expressing S. aureus cells in in vivo infection models. Results: We have demonstrated that SNAP-tag was expressed and exported to the cell wall where it was covalently anchored. Deletion of the sortase A enzyme prevented attachment of the SNAP-tag to the cell wall, as determined by Western blot. Once situated within the cell wall, SNAP-tag was functional and able to specifically bind cell-impermeable fluorescent BG ligands and our synthesised precursor HYNIC-NH2-BG ligand, as determined by confocal microscopy and fluorometry assay. Pilot in vivo studies for fluorescence optical imaging and nanoSPECT-CT imaging with the novel 99m Tc-HYNIC-NH-BG ligand are currently under development to visualise S. aureus infections in mouse models. Conclusions: This approach should allow a higher sensitivity to be achieved when investigating bacterial infections in real time compared to current molecular imaging techniques, thus allowing bacterial virulence and the potential effects of new antimicrobials to be assessed. OP367 Dual imaging of lipopolysaccharides (LPS) by SPECT-CT and Confocal Microscopy. M. Moreau, V. Duheron, B. Collin, W. Sali, C. Bernhard, C. Goze, T. Gautier, J. Pais de Barros, V. Deckert, F. Brunotte, L. Lagrost, F. Denat; ICMUB UMR CNRS 6302, Dijon, FRANCE, INSERM UMR866, Dijon, FRANCE, Centre Georges-François Leclerc, Dijon, FRANCE, Centre Hospitalier Universitaire, Dijon, FRANCE. Introduction: Lipopolysaccharides (LPS) or endotoxins are found inserted in the outer membrane of Gram-negative bacterias. Their appearance in blood stream triggers a massive secretion of pro-inflammatory cytokines in mammals. A controlled response allows the neutralization and elimination of LPS, whereas an excessive inflammatory response leads to severe circulatory and respiratory defects. It is the endotoxemic shock or septic shock that can leads to death. Many approaches are used to study LPS, including labeling with radiochemicals (3H, 125I, 99mTc or 51Cr) or with fluorophores (FITC, Alexa488, Bodipy). Bimodality is attracting more and more interest in the field of molecular imaging since the combination of two different techniques may provide complementary information, thus improving the accuracy of diagnosis. Combining nuclear modalities (PET or SPECT) with optical imaging is of particular interest, and the similar sensitivities of the two techniques allows to fuse the signaling moieties into a unique molecule, called monomolecular multimodality imaging agent (MOMIA), ensuring a same biodistribution of the two probes. Method: A recently described bimodal probe, namely DOTA-Bodipy-NCS, has been covalently attached to LPS. The integrity of the LPS after labeling procedure was checked by SDS-PAGE electrophoresis and βhydroxymyristate titration (BHM). Pro-inflammatory activity of LPS was assessed by quantification of cytokines released by differentiated THP-1 cells. This bioconjugate was then radiometallated for SPECT-CT biodistribution imaging. Results: DOTABodipy-LPS was metallated with 111In to yield a high specific activity (600 MBq.mg1), with a radiochemical purity >98 % after purification. Biodistribution of the radiolabeled compound was then evaluated in vivo in WT mice by SPECT-CT imaging. Radiolabeled LPS is rapidly eliminated from the bloodstream and accumulates in spleen and liver. Liver slices were then analyzed by confocal microscopy, and specific fluorescent signals in the cytoplasm of hepatocytes were detected, confirming the accumulation of 111In-DOTA-Bodipy-LPS in the liver. Conclusion: These results demonstrate the efficiency of the conjugation process of our bimodal probe. It made it possible to perform both non-invasive SPECT and ex vivo fluorescence imaging of LPS biodistribution, underlining its liver uptake for further detoxification. The 111In-DOTA-Bodipy-LPS probe arises here as a relevant tool to identify key components of LPS detoxification in vivo paving the way to therapeutic issues in the field of sepsis. Acknowlegement: Support was provided by the CNRS, the University of Burgundy, the Conseil Régional de Bourgogne. O P _ M o nd ay S178 Eur J Nucl Med Mol Imaging (2013) 40 (Suppl 2):S89–S567Affibody molecules constitute a class of small (7 kDa) scaffold proteins that can be engineered to have excellent tumor targeting properties. High reabsorption in kidneys complicates development of ...