Basics and Principles of Radiopharmaceuticals for PET/CT

European Journal of Radiology 73 (2010) 461–469

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European Journal of Radiology

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Review Basics and principles of radiopharmaceuticals for PET/CT

W. Wadsak a, M. Mitterhauser a,b,∗ a Department of Nuclear Medicine, Medical University of Vienna, Austria b Department of Pharmaceutical Technology and Biopharmaceutics, University of Vienna, Austria article info abstract

Article history: The presented review provides general background on PET radiopharmaceuticals for oncological appli- Received 1 December 2009 cations. Special emphasis is put on radiopharmacological, radiochemical and regulatory aspects. This Accepted 15 December 2009 review is not meant to give details on all different PET tracers in depth but to provide insights into the general principles coming along with their preparation and use. Keywords: The PET tracer plays a pivotal role because it provides the basis both for image quality and clinical Radiopharmaceutical interpretation. It is composed of the radionuclide (signaller) and the molecular vehicle which determines Tracer the (bio-)chemical properties (e.g. binding characteristics, metabolism, elimination rate). Radiopharmacology Radiochemistry © 2010 Published by Elsevier Ireland Ltd.

This section is intended to provide general background on PET resulting in abnormal function it can probably be visualized long radiopharmaceuticals for oncological application. Special empha- before morphological manifestation. sis is put on radiopharmacological, radiochemical and regulatory Basically, there are three major disciplines that have to inter- aspects. This review is not meant to give details on all different PET act and collaborate closely to enable the successful application of tracers in depth but to provide insights into the general principles PET/CT in a clinical setting: medical physics, radiopharmaceuti- coming along with their preparation and use. cal sciences and clinical imaging (see Fig. 1). The protagonists of these disciplines – each covering its specific field in depth – have 1. The role of radiopharmaceuticals in PET/CT to understand the basic principles and comprehend the different scientific language. Hence, understanding of radiopharmaceutical Radiopharmaceuticals, already attributed to as drugs in 1960 issues is pivotal to understanding imaging on a molecular level! [1], are radiolabelled molecules designed for in vivo application. The PET tracer plays a central role because it provides the basis These radiopharmaceuticals consist of two parts: (1) a molecular both for image quality and clinical interpretation. The radionuclide structure determining the fate of the radiopharmaceutical within with its specific physical properties is the working basis for sig- the organism (pharmacokinetics and pharmacodynamics) and (2) a nal detection, transduction and computational translation. On the radioactive nuclide being responsible for a signal detectable outside other hand, understanding the (bio-) chemical properties of the of the organism for subsequent visualization with nuclear medi- molecular vehicle (e.g. binding characteristics, metabolism, elim- cal methods. Since these imaging methods are directly based on ination rate) is essential for molecular modelling and dosimetry. radiolabelled molecules they can truly be called “molecular imaging” From a clinical point of view, basic understanding of radiochemical modalities. preparation (e.g. formation of potential by-products, interference Since radiopharmaceuticals are chemically indistinguishable of labelled and unlabelled contaminants with the target site, spe- from their non-radioactive counterparts the organism does not cific radioactivity) is also necessary for diagnosis using PET/CT. make a difference in using the radiolabelled derivatives as surro- Radiopharmaceuticals are a very rare and special species. And gates in all its biochemical processes. Thus, radiopharmaceuticals radiopharmacists (and medical radiochemists) are also a rare can be used to directly visualize these functional processes in vivo. species within their disciplines. But it needs these specially trained If, for example, there is a pathological change on the molecular level experts to guarantee continuous availability and safety of these special pharmaceuticals. Medical radiochemistry and radiopharmacy were – and still are – treated as orchid areas within their

∗ scientiﬁc home faculties; it needs special interest and freakish Corresponding author at: Waehringer Guertel 18-20, A-1090 Vienna, Austria. dedication in the preparation of drugs on a sub-nanomolar scale. Tel.: +43 1 40400 1557; fax: +43 1 40400 1559. E-mail address: [email protected] (M. Mitterhauser). However, during the last decades a lot of emphasis has been put URL: http://www.radiopharmaceutical-sciences.net (M. Mitterhauser). into the establishment of a certiﬁed education and its recognition in

Table 1 Important PET nuclides and their most common ways of preparation.

Nuclide Production Half-life

F-18 (F−) 18O(p,n)18F 110 min 20 18 F-18 (F2) Ne(d,␣) F 110 min C-11 14N(p,␣)11C 20 min N-13 16O(p,␣)13N 10 min O-15 14N(d,n)15O 2 min Ga-68 68Ge/68Ga-Generator 68 min Rb-82 82Sr/82Rb-Generator 1.3 min

The vehicle molecules either interact directly with the afore- mentioned targets and processes – they can be substrates for enzymes, agonists or antagonists for receptors and transporters – or take part directly in metabolic processes. In a pathophysiological state, these targets and/or processes Fig. 1. Schematic illustration of the interaction of the three major disciplines may be changed significantly compared to their normal state and involved in PET/CT, highlighting the central role of the PET tracer. functionality. Therefore, interaction of the vehicle molecules may also be changed considerably. For example, in many tumours the Europe. Thus, nowadays “certified freaks” are available to perform expression rate of receptors, transporters, enzymes and antigens all kinds of radiopharmaceutical tasks in routine and R&D. is modified and these alterations may serve as a suitable predictor for stage and allocation of these tumours. To achieve a personal- 2. What is a PET radiopharmaceutical ized diagnosis from outside the body without invasive intervention, suitable interactions between vehicle and target site alone are not Principally, a PET radiopharmaceutical consists of two compo- enough. It needs for an additional signaller stably attached to the nents (see Fig. 2): vehicle. Thus, traceability of the pharmaceutical’s fate and pathway is guaranteed. (1) A molecular structure (vector, vehicle, ligand) and (2) a positron emitting radionuclide. 2.2. PET radionuclides

For a stable connection of these two parts, a linker may be chem- For PET, these signallers attached to the vehicle molecules are ically necessary. The vehicle defines the biological characteristics positron emitting nuclides. The most important PET nuclides are and is responsible for chemical and biochemical interactions within summarized in Table 1. the living organism. The positron emitting radionuclide provides a All widely used PET nuclides are short lived with limited avail- detectable signal enabling coincidence measurements of annihi- ability. On the other hand, wide and feasible availability of the lation radiation within a dedicated PET device such as a PET/CT radionuclide is a prerequisite for successful application on a routine scanner. basis. Therefore, only fluorine-18 and gallium-68 are used in a clinical setting in PET sites without on-site cyclotron, so far. On the one 2.1. Vehicle molecules hand, registered F-18 labelled radiopharmaceuticals are distributed by commercial vendors and can be used directly. On the other hand, The vehicle molecules have to provide a high degree of speci- gallium-68 generators can be simply installed locally and their easy ficity and selectivity towards the target site. These targets can be handling does not require specific technical equipment. Concerning e.g.: the other PET nuclides, generally, the production has to be performed in a medical cyclotron on-site. This demands for dedicated • selected receptor systems, equipment and specially trained and qualified personnel. • antigens, Possible ways of use are different for different types of radionu- • enzymes, clides. As examples, Fig. 3 illustrates the main pathways of • transporters, radiochemical conversions for fluorine-18 and carbon-11. • specific metabolic alterations, The selection of the PET radionuclide has to be based on the • such as up-regulated conditions, following considerations: • hypo-oxygenation of tissue, • different energy demand of cells, (1) Availability of the radionuclide; • changes in gene and protein expression, (2) physical characteristics of the radionuclide; • differences in vascularisation and perfusion. (3) radiochemical issues; (4) radiopharmacological issues.

Besides the issues already discussed above, even in PET centres with on-site production facility, the very short half-life of radionuclides such as oxygen-15 (2 min) and nitrogen-13 (10 min) limits their clinical applicability. Therefore, only a handful PET-centres worldwide is using these radionuclides on a daily routine basis. Overall, the half-life of the radionuclide should be long enough for successful radiolabelling, and the time frame of the imaging Fig. 2. Schematic design of a PET radiopharmaceutical and its interaction with the target site. procedure. W. Wadsak, M. Mitterhauser / European Journal of Radiology 73 (2010) 461–469 463

Hence, all radiolabelling steps have to be performed in a fully (lead) shielded environment (hot cells) using special forceps, tele- tongs or robotic manipulators. • For reproducible outcome, fully automated synthesis modules are used that can be remotely controlled from outside the shielded hot cells. • The amount of carrier (stable isotope of the radionuclide) influ- ences the chemical reactions leading to an indistinguishable, non-radioactive product that “dilutes” the final radiopharmaceutical. The amount of carrier also determines the achievable specific radioactivity which is measured in activity per mass unit (e.g. GBq/␮mol). Such carrier can be introduced into the radiosynthesis on purpose (due to production necessities) or by unwanted “contamination” of the reactants and solvents. In 12 case of carbon-11, even the C-CO2 always present in air may contribute significantly to a reduction in specific activity. Never- theless, the theoretically achievable maximum specific activity of carbon-11 is higher (341,140 GBq/␮mol) than of fluorine-18 (63,270 GBq/␮mol). Interestingly also in reality, specific activities of carbon-11 labelled radiopharmaceuticals are usually slightly higher although the “pollution” of reaction solutions and synthesizer environment with carbon carrier is unavoidable [2]. • Moreover, instability of the radiolabelled targeting agent due to radiolytical processes has to be considered when working with solutions with high radioactivity concentrations. This is an important issue when shipping high activities of [18F]FDG considering that nowadays commercially available FDG-synthesizers are able to produce activities in the range of 500 GBq in a final volume of approximately 20 mL! Also, during preparation and purifica- tion of the PET tracer there are steps involved leading to high Fig. 3. Main pathways of medicinal radiochemistry using F-18 and C-11. probability of radiolysis, e.g. excessive heating and evaporation to dryness under vacuo. Therefore, alternative methods under mild conditions are usually applied in medicinal radiochemistry, 2.2.1. Specific radiochemical considerations such as heating under microwave conditions, reduction of resid- Medicinal radiochemistry comprises the activation of the ual solvents by solid phase extraction (SPE), use of microchemical radionuclide, the radiolabelling procedure itself and the purifica- methods. Hence, a delicate balance has to be sought between tion of the product. Since in general radionuclides are obtained high starting activities (leading to satisfactory final activities) and from the cyclotron (or generator) in a chemical form that is not reasonable amount of radiolysis. predisposed for direct labelling reactions, primary activation steps are absolutely necessary. For example, F-18 is normally obtained as an aqueous [18F]fluoride solution from the target that is chemically 2.2.2. Specific radiopharmacological considerations inactive and has to be transformed into a more reactive chemical The radionuclide allows only for the allocation and quantifi- species. This is achieved by azeotropic drying with acetonitrile in cation of the annihilation events without taking the integrity of presence of an aminopolyether as phase transfer catalyst. The acti- the radiolabelled molecule into account. Therefore, it could well vated radionuclide is subsequently coupled with or incorporated be the case that the radionuclide (or a small part of the molecule into the predesigned predecessor of the vehicle molecule (=pre- including the signalling radionuclide) is cleaved metabolically from cursor). This labelling reaction often requires elevated temperature the intact parent molecule. Then, the signal that is detected from or microwave assistance and is performed in dedicated synthesiz- outside would be wrongly assigned and quantification would be ers. In contrast to traditional chemistry, it is usual to waive relative systematically biased: yields in favour of reduction of reaction time. One has to bear in mind, that in case of short-lived radionuclides higher relative con- • Carbon-11 is always bound covalently within the vehicle version yields are easily outbalanced by the longer reaction times. molecule, most often as a [11C]methyl group attached to an Therefore, the absolute yields (in terms of amount of radioactiv- amine, hydroxyl or carboxyl moiety. Since the carbon-11 label ity) are usually higher when applying short reaction times (i.e. replaces a carbon-12 atom no changes due to the radionuclide 1–20 min). Other differences to conventional chemistry are: can be observed. Organisms are unable to distinguish between the original compound and the [11C]labelled analogue. • The stoichiometry between the reaction partners (i.e. the acti- • Fluorine-18 is also attached covalently to the parent molecule but vated radionuclide and the precursor) is not given! The precursor usually has to be added to the original structure since they do not is present in a vast excess compared to the radionuclide. Imagine bear a fluorine atom at first. This leads to unpredictable changes in a shipload of precursor reacts with only one single sugar cube...! the compound’s characteristics. This could, on the one hand, ham- • Reactions that fail under normal chemical circumstances are per its further use due to reduced affinity, stability or selectivity sometimes successful in radiochemistry and vice versa. or, on the other hand, even ameliorate the in vivo behaviour. • Due to the minimal mass of reactants (often less then 1 mg), the • Radiometals such as gallium-68 can only be fixed to the vehicle use of special miniature scaled equipment is necessary. using a metal–ligand-complex. This always leads to signifi- • Working with high levels of radioactivity (several Gigabec- cant changes in the steric and electronical configuration and, querels, GBq) requires potent measures for radiation protection. subsequently, to probable changes in pharmacokinetics and 464 W. Wadsak, M. Mitterhauser / European Journal of Radiology 73 (2010) 461–469

sification, either. We decided to apply the attribution according to the uptake mechanism in the following section. There are several different mechanisms through which PET tracers display their way of action leading to a signalling contrast between tumour tissue and background activity. These mechanisms comprise specific interactions at the tumour cell’s surface such as antigen–antibody coupling or receptor mediated binding, metabolic trapping and supply-dependent processes. Often even the classification of PET radiopharmaceuticals and the alignment to specific mechanisms is a demanding task. In some Fig. 4. Availability vs. changes in in vivo behaviour. cases, exact uptake mechanism is still a matter of debate; in other cases, uptake is confounded by the combination of many distinct interactions; and sometimes even the definition of a specific class pharmacodynamics of the radiopharmaceutical. For example, is still disputed amongst scientists. A good example is the term despite significant efforts throughout many years, there is only “proliferation”: strictly spoken, proliferation is cell growth. Each one single central receptor-targeting complex labelled with a viable cell cohort grows and as a matter of fact, cell growth needs radiometal that found its way into clinical application, namely energy, oxygen and nutrients, it is surveilled by receptors and hor- 99mTc-TRODAT [3,4]. Furthermore, metal–ligand interactions mones and is regulated by enzymes and so on. Following this strict within a complex are normally much weaker than covalent bind- definition, almost all PET tracers could be named “proliferation ing. markers”! But in contrast, radiopharmacology understands prolif- • As already discussed in the previous section, the specific eration markers only as radiopharmaceuticals, actively taking part radioactivity is an important parameter influencing the imaging in the main processes of proliferation themselves such as growth outcome. One always has to bear in mind that having a limited of cell membranes or increased DNA replication rate. But even then number of target sites (B ) and a saturable process of bind- max there is some scope of discretion (see Section 3.4). ing between the radiopharmaceutical and the target (receptors, transporters), low specific activity (i.e. high number of compet- 3.1. PET tracers mainly reflecting energy utilization ing unlabelled molecules) of the PET tracer would lead to low or even undetectable signal. As a conclusion, the lower the density of As already mentioned before, [18F]FDG is the by far most widely the target sites the higher the required specific activity! Notably, used PET tracer worldwide. This is due to its versatility—most the specific radioactivity should be as constant as possible within pathological conditions are paired with alterations in glucose man- a (quantified) clinical trial and should always be included as a agement. Therefore, applications in therapy monitoring, tumour co-variable in statistical analysis of the data. staging, myocardial energy turnover and many different neurolog- ical diseases are found in clinical routine. This fact is also reflected in As illustrated in Fig. 4, the availability and the possible a vast number of available publications—entering “FDG AND PET” in changes in in vivo behaviour often point in the opposite direc- MEDLINE yields 8972 hits! (as of August 2009). Additionally, FDG- tions. For example, gallium-68 is readily available through a PET is unfortunately often equated with PET as such by clinicians radionuclide-generator system but leads to the highest degree denying all other available PET radiopharmaceuticals! of unpredictiveness in its in vivo behaviour. On the other hand, The first preparation of the radiolabelled FDG compound was carbon-11 labelled radiotracers are able to directly represent the achieved as early as 1978 by Ido et al. [5–7]; the real breakthrough authentic (unchanged) molecule. was enabled in 1986 by Hamacher et al. [8] allowing for the produc- The choice of the used radionuclide will therefore always be a tion of high amounts of the tracer and further improved by Füchtner compromise between availability and economical considerations, et al. [9]. This work was the kickoff for the worldwide success story physical properties, radiolabelling possibilities and radiopharma- PET as such. Due to the enormous importance of FDG, the nowadays cological issues. primarily used way of preparation, quality control and FDG’s way of action are described in more detail in the annex section. The major 3. Specific PET radiopharmaceuticals drawback of FDG is its unspecific way of uptake: in many cases, inflammation or other benign processes are indistinguishable from There is a series of PET tracers that found their way into clinical tumour entities; no assignment to specific tumour origins is pos- routine so far. There are 3 major areas of application: (1) oncology, sible; tumours embedded in tissue with high energy turnover and (2) cardiology and (3) neurology. Amongst these fields of interest, high background activity (e.g. brain, muscles, bladder) allow only oncology accounts by far for the most applications. for limited delineation. The classification of PET tracers can be done by many different Of course, there are also other PET tracers available reflect- ways: ing energy utilization, such as radiolabelled fatty acids [10–12] or [11C]acetate. 1. According to the radionuclide used (C-11, F-18, Ga-68, etc.). Tracer Specific remarks References 2. According to the main field of application (oncology, cardiology, neurology, etc.). [18F]FDG THE working horse of PET. Routine applications [5–9,13–16] 3. According to the uptake mechanism (active transporter, receptor in oncology, neurology and cardiology. [11C]Acetate Primarily applied in cardiology; nowadays [17–24] binding, metabolic trapping, etc.). often used in imaging of prostate cancer 4. According to the frequency of use or status of development (rou- (staging and therapy control). tine, phase I, preclinical status, etc.). 5. According to the target site (glucose transporter, somatostatin 3.2. Amino acids SST2 receptor, CD20 antigen, etc.). Amino acids are essential for the survival of cells and mainly But unfortunately, none of the given classifications covers the used for protein synthesis. Since tumours often show increased whole field of PET tracers and does not allow for conclusive clas- protein synthesis rates, enhanced uptake of amino acid tracers in W. Wadsak, M. Mitterhauser / European Journal of Radiology 73 (2010) 461–469 465 tumour tissue can be observed. In many cases, amino acids are Tracer Specific remarks References taken up by specific transporters through the cell membranes (e.g. 18 l [ F]FLT Gold standard. Uptake reflects [42,72–77] -amino acid transporters, LAT) which might be up-regulated in thymidine kinase activity. tumour tissue or are substrates of specialized enzymes, also up- 5-[18F]FU Initially developed as an [79–83] regulated in pathological states (e.g. tyrosine kinase, TK). anti-metabolite, it reflects RNA Moreover, amino acids serve as precursor molecules for biogenic synthesis. [11C]Thymidine Authentic pyrimidine base. Short [84–88] amines that are used in many important functions including cell half-life. signalling (e.g. neurotransmission) and hormone action. [11C]Choline Choline derivatives are substrates for [40,89–92] Radiolabelled amino acids are in many cases altered in their cell membrane build-up (as chemical structure (addition of the radiolabel to the original amino phosphatidyl choline). 18 acid) and therefore might also be altered regarding their uptake [ F]F-Choline Different fluoro-cholines are available, [93–96] differently labelled. Longer-lived mechanism. alternatives for authentic [11C]choline and [11C]acetate. Tracer Specific remarks References [18F]FDOPA First introduced for the diagnosis of [25–33] 3.5. Specific target interactions (enzymes, transporters, receptors) Parkinson’s disease. Now, also used in neuroendocrine tumour imaging. Specific targets on the tumour cells are used as binding sites for [11C]Methionine Authentic tracer (no structural [30,33–42] alteration). Short half-life. Mainly used different classes of radiopharmaceuticals. Peptide receptors, trans- in brain tumour imaging. porter proteins, enzymes or antigens do serve as binding sites. As [18F]FET Longer-lived alternative to [30,37,43–46] these binding sites are only available in a limited number and there- 11 [ C]methionine. Discussed to be able fore binding is saturable, high specific activity, affinity and sub-type to distinguish between inflammation selectivity are prerequisites. and residual tumour tissue. [11C]AMT Used only in clinical trials so far. [47–51] Tracer Specific remarks References

68Ga-DOTATOC Octapeptide with DOTA chelator [97–102] selectively binding to those somatostatin receptors that are 3.3. PET tracers for hypoxia up-regulated in neuroendocrine tumours. 68Ga is easily available through a radionuclide generator. Also: In many cases, the therapeutic effect of cyctostatics and external DOTANOC and DOTA-TATE are used as irradiation can be reflected by the extent of hypoxic areas within generation 2 tracers. 18 ␣ ␤ the tumour. Visualization of the extent of hypoxia is therefore [ F]RGD Tripeptide; targets selectively the v 3 [79,103,104] receptor core. On the edge to routine essential for fine-tuning of the therapy regimen leading to a more application. personalized approach. [18F]FES Labelled estradiol derivative; used for [105–108] The uptake mechanism of these PET tracers is based on enzymes the diagnosis of estrogen receptor up-regulated in hypoxic state (e.g. nitrooxidases) dependent on positive tumours (e.g. breast). 18 ␤ available oxygen. This leads to a positive uptake and contrast in [ F]FETO Suicide substrate for 11 -hydroxylase [109–114] predominantly expressed in the oxygen deficient regions. adrenal cortex. Used only in clinical trials so far. Also: [11C]metomidate, Tracer Specific remarks References [11C]etomidate.

[18F]FMISO The first used hypoxia marker in PET. [52–59] Now, the gold standard amongst 3.6. Other important PET tracers tracers for hypoxia imaging. 18 [ F]FAZA Chemical derivative of FMISO bearing a [60–64] [18F]Fluoride is the second most often applied in oncology play- arabinosyl-backbone. So far, only used in clinical trials. ing an important role for the detection of bone metastases and [18F]EF-5 So far, only used in clinical trials. [65–67] primary osteoblastic tumours. Although in clinical use for more 64Cu-ATSM Rather exotic radionuclide with longer [68–71] than 40 years its uptake mechanism to bone is still a matter of half-life. Now, on the step towards debate. Recent studies substantiate the evidence that uptake is clinical routine. associated with crystallization of hydroxyapatite in osteoblastic processes. In vitro studies revealed that reversibility of binding to hydroxyapatite and bone is negligible. Nevertheless, [18F]fluoride has reached broad acceptance in the nuclear medical community 3.4. Proliferation markers and adds significant value to diagnostic imaging [115–118] (Fig. 5).

As already discussed in the introduction of Section 3, defini- 4. Regulatory aspects and quality assessment tion of proliferation markers is still disputed. Nevertheless, some PET tracers entitled “proliferation markers” are available and also By law in most countries, PET radiopharmaceuticals are drugs. widely used. Thereof, FLT is the most prominent tracer. But since This implicates that a series of regulatory and legal aspects have it is not integrated into DNA [78], its uptake definitely does not to be followed prior to release for human application or clinical correspond directly to DNA synthesis rate. Therefore, its value routine. Additionally, in 99% of the applications, radiopharmaceuti- – especially in comparison with FDG – is still disputed in the cals are administered intravenously. Hence, these parenterals have nuclear medical community. On the other hand, [11C]thymidine is to follow even stricter regulations taking into account the precau- an authentic surrogate for the original DNA base, thymidine, and tions for this specific class of drugs (glass types, rubber materials for is therefore used directly in DNA synthesis. But its short half-life stoppers, sterility assessment et cetera). The short half-life of PET is not compatible with tracing the incorporation of the radiotracer radionuclides in general does not make these tasks any easier. For into DNA and therefore limit its applicability. this reason, there is a specific monograph in the European Pharma- 466 W. Wadsak, M. Mitterhauser / European Journal of Radiology 73 (2010) 461–469

5. Conclusion and outlook

Radiopharmaceuticals for PET are a very special category of pharmaceutical drugs. This has consequences both in preparation and regulatory treatment. The accessibility of the short-lived radionuclides commonly used in PET applications is limited and their use in medicinal radiochemistry is restricted to specially equipped facilities. Following the radiolabelling procedure, the possible changes in the in vivo behaviour of PET tracers have to be taken into account. Unfortunately, the better the availability of the used radionuclide is, the more drastic appears the change in the respective radiopharmaceutical. Overall, PET tracers are the heart of molecular imaging, pumping life into PET/CT. Since PET radiopharmaceuticals have a short history there is still a lot to do and the future of radiopharmaceutical sciences is bright. Fig. 5. Main targets for radiopharmaceuticals on the cell surface. Appendix A. copoiea dealing with radiopharmaceutical preparations as a whole A.1. Radiosynthesis of [18F]FDG [119]. Furthermore, there are more than 50 specific monographs for radiopharmaceuticals, 11 of which addressing PET tracers (2005), with growing numbers every year. These monographs give detailed instructions regarding preparation, precursors, tests of identity, purity, chemical purity, radionuclidic purity, radiochemical purity, measurement of radioactivity and even labelling of the containers. There are methods suggested but quality control can be performed using other methods if validated against the given methods. The short half-life of certain radionuclides is acknowledged in the monographs by allowing “parametric” release of the product in certain cases. That means that the final product may be released by a competent employee before finishing the tests concerning for example sterility, endotoxins, chemical purity (residual solvents) and radionuclidic purity. This implicates that the methods used for the preparation of the radiopharmaceutical have to be safe and robust. Therefore, the implementation of a thorough system for quality management is a must! In Europe, there are several stan- 1. Activation of cyclotron produced [18F]fluoride and phase transfer dards of quality that could find application with varying depth and catalysis using the Aminopolyether Kryptofix 2.2.2. in presence complexity. But until now, all these codes derive from the industry of potassium carbonate. standards. Hence, they do not pay deference to the specific issues 2. Nucleophilic substitution of the triflate leaving group by the regarding short-lived radiopharmaceuticals, such as single batch activated [18F]fluoride (S 2 reaction); attack from the back- production for a very limited number of applications, radiation pro- N side → therefore the precursor molecule is a mannose derivative tection combined with product safety and small markets for the (1,3,4,6-tetraacetyl-d-mannose 2-triflate). engaged companies or even academical institutions. Thus, there 3. Basic hydrolysis (sodium hydroxide) for deprotection of the are serious attempts coming from the radiopharmacy committee acetyl-protecting groups. of the European Association of Nuclear Medicine (EANM) to imple- ment specific regulations on specific “Good radiopharmaceutical practices” (GRPP). Until now, the draft is under peer assessment by the EMEA (European Medicines Agency). GRPP is deduced from GMP (Good Manufacturing Practice; industrial standard for large scale production in many areas) but takes the special aspects in the preparation and quality assessment of radiopharmaceuticals into account. Besides, within Europe the national authorities have quite an area of discretion in their interpretation of the pertinent guide- lines and directives published by the European Commission (EC). For instance, in some countries, there is the necessity for full registration of the radiopharmaceutical even for academic (in- house) preparation; in other countries, small-scale productions do not require marketing authorization. Hence, a common regulation within Europe is longingly expected and would facilitate inter- sectoral and trans-national cooperation. Such a regulation should base on a fair standard; over-regulation would endanger or delay the development and clinical application of novel specific radiopharmaceuticals and in the long run lead to discrimination of the European scientific community in this field. Fig. A1. GE FASTlab FDG synthesizer within a lead shielded hot cell environment. W. Wadsak, M. Mitterhauser / European Journal of Radiology 73 (2010) 461–469 467

Subsequently, the crude product is purified via solid phase the glucose molecule in position 6 and subsequent isomerisation extraction cartridges (SPE), formulated for intravenous injection of the gluco-pyranose derivative to yield its fructo-furanose pen- and sterile filtrated under aseptic conditions. dent. This second step needs a hydroxylic function in position 2 Fig. A1. to enable cyclisation (the oxygen in position 2 attacks the carbon atom in position 5). Then, the further steps of the glycolysis are A.2. Quality control of [18F]FDG conducted to finally yield carbon dioxide and water.

QC parameter Method Evaluated parameter Threshold

Radiochemical purity HPLC [18F]FDG +[18F]FDM >95% [18F]FDM <9.5% [18F]ﬂuoride <5% TLC [18F]FDG >95% [18F]ﬂuoride <5% partially hydrolyzed species <5%

Radionuclidic purity Gamma spectroscopy Gamma spectrum 511 keV (1022 keV) Long lived species <0.1% Activity measurement Physical half-life ±10 min

Chemical purity HPLC [19F]FDG <50 ␮g/mL GC Residual solvent: Acetonitrile <410 ppm TLC Kryptoﬁx 2.2.2. <2.2 mg/V*

Physical parameters pH electrode pH 4.5–8.5 Osmometer osmolality 230–450 mosmol/kg

Microbiological testing LAL Endotoxines <1 EU/ml Media ﬁlls Sterility <10−6 *V = maximum applicable volume.

A.3. Scheme of the “metabolic trapping” of [18F]FDG

The catabolic path of glucose for gaining energy follows glycolysis within the cell. It starts with activation (phosphorylation) of 468 W. Wadsak, M. Mitterhauser / European Journal of Radiology 73 (2010) 461–469

Since the FDG molecule comprises a fluoride-moiety instead of [28] Ishiwata K, Shinoda M, Ishii SI, Nozaki T, Senda M. Synthesis and evaluation the hydroxylic group in position 2 the second step is not able to take of an F-18-labeled dopa prodrug as a PET tracer for studying brain dopamine place and therefore no further catabolization is possible. Moreover, metabolism. Nucl Med Biol 1996;23(3):295–301. [29] Dolle F, Demphel S, Hinnen F, et al. 6-[18F]Fluoro-l-dopa by radiofluoro- the de-phosphorylation (reversing step 1) is also energetically hin- destannylation: a short and simple synthesis of a new labeling precursor. J dered in most cells. This leads to accumulation of FDG within the Label Compd Radiopharm 1998;41:105–14. cell only dependent on glucose transport into the cell and kinase [30] Langen KJ, Jarosch M, Muhlensiepen H, et al. Comparison of fluorotyrosines and methionine uptake in F98 rat gliomas. Nucl Med Biol 2003;30:501– activity in step 1. This is often referred to as “metabolic trapping”. 8. [31] Saier MH, Daniels GA, Boerner P, et al. Neutral amino acid transport systems References in animal cells: potential targets of oncogene action and regulators of cellular growth. J Membr Biol 1988;104:1–20. [32] Jager PL, Vaalburg W, Pruim J, et al. Radiolabeled amino acids: basic aspects [1] Briner WH. Radiopharmaceuticals are drugs. Mod Hosp 1960;95:110–4. and clinical applications in oncology. J Nucl Med 2001;42:432–45. [2] Welch MJ, Redvanly CS, editors. Handbook of radiopharmaceuticals radio- [33] Becherer A, Karanikas G, Szabo M, et al. Brain tumour imaging with PET: a chemistry and applications (1st issue). 2002. p. 43. comparison between [F-18]fluorodopa and [C-11]methionine. Eur J Nucl Med [3] Meegalla SK, Plössl K, Kung M-P, et al. Kung HIE synthesis and characterization Mol Imag 2003;30(11):1561–7. of Tc-99m labeled tropanes as dopamine transporter imaging agents. J Med [34] Comar D, Cartron JC, Maziere M, Marazano C. Labeling and metabolism of Chem 1997;40:9–17. methionine-methyl-C-11. Eur J Nucl Med 1976;1(1):11–4. [4] Kung HF, Kim H-J, Kung M-R, Meegalla SK, Plössl K, Lee H-K. Imaging of [35] Bolster JM, Vaalburg W, Elsinga PH, Wynberg H, Woldring MG. Syn- dopamine transporters in humans with technetium-99m TRODAT-1. Eur J thesis of Dl-[1-C-11]methionine. Appl Rad Isotopes 1986;37(10):1069– Nucl Med 1996;23:1527–30. 70. d [5] Ido T, Wan CN, Casella V, et al. Labeled 2-deoxy- -glucose analogs—F-18- [36] Langstrom B, Antoni G, Gullberg P, et al. Synthesis of l-[methyl- d d labeled 2-deoxy-2-fluoro- -glucose, 2-deoxy-2-fluoro- -mannose and C-14- C-11]methionine and d-[methyl-C-11]methionine. J Nucl Med d 2-deoxy-2-fluoro- -glucose. J Label Compd Radiopharm 1978;14(2):175–83. 1987;28(6):1037–40. [6] Gallagher BM, Ansari A, Atkins H, et al. Radiopharmaceuticals.27. F-18-labeled [37] Weber WA, Wester HJ, Grosu AL, et al. O-(2-[F-18]fluoroethyl)-l- d 2-deoxy-2-fluoro- -glucose as a radiopharmaceutical for measuring regional tyrosine and l-[methyl-C-11]methionine uptake in brain tumours: ini- myocardial glucose-metabolism invivo—tissue distribution and imaging tial results of a comparative study. Eur J Nucl Med 2000;27(5): studies in animals. J Nucl Med 1977;18(10):990–6. 542–9. [7] Ido T, Wan CN, Fowler JS, Wolf AP. Fluorination with F2—convenient synthesis [38] Mitterhauser M, Wadsak W, Krcal A, et al. New aspects on the preparation d of 2-deoxy-2-fluoro- -glucose. J Org Chem 1977;42(13):2341–2. of [C-11]methionine—a simple and fast online approach without preparative [8] Hamacher K, Coenen HH, Stöcklin G. Efficient stereospecific syn- HPLC. Appl Radiat Isot 2005;62(3):441–5. 18 d thesis of no-carrier-added 2-[ F]-fluoro-2-deoxy- -glucose using [39] Quincoces G, Penuelas I, Valero M, et al. Simple automated system for simul- aminopolyether supported nucleophilic substitution. J Nucl Med 1986;27(2): taneous production of C-11-labeled tracers by solid supported methylation. 235–8. Appl Radiat Isot 2006;64(7):808–11. [9] Füchtner F, Steinbach J, Mäding P, Johannsen B. Basic previous term hydrolysis [40] Cheung MK, Ho CL. A simple, versatile, low-cost and remotely operated 18 next term 2-[ F]fluoro-1,3,4,6-tetra-O-acetyl-image-glucose in the prepara- apparatus for [C-11]acetate, [C-11]choline, [C-11]methionine and [C-11]PIB 18 tion of 2-[ F]fluoro-2-deoxy-Image-glucose. Appl Radiat Isot 1996;47:61–6. synthesis. Appl Radiat Isot 2009;67(4):581–9. [10] Schelbert HR, Henze E, Sochor H, Grossman RG, Huang SC, Barrio JR, Schwaiger [41] Ceyssens S, Van Laere K, de Groot T, et al. [C-11]Methionine PET, histopathol- M, Phelps ME. Effects of substrate availability on myocardial C-11 palmitate ogy, and survival in primary brain tumors and recurrence. Am J Neuroradiol kinetics by positron emission tomography in normal subjects and patients 2006;27(7):1432–7. with ventricular dysfunction. Am Heart J 1986;111(6):1055–64. [42] Murayama C, Harada N, Kakiuchi T, et al. Evaluation of D-F-18-FMT, F-18- [11] Schelbert HR. PET contributions to understanding normal and abnormal car- FDG, L-C-11-MET, and F-18-FLT for monitoring the response of tumors to diac perfusion and metabolism. Ann Biomed Eng 2000;28(August (8)):922–9. radiotherapy in mice. J Nucl Med 2009;50(2):290–5. [12] Bergmann SR. Imaging of myocardial fatty acid metabolism with PET. J Nucl [43] Wester HJ, Herz M, Weber W, et al. Synthesis and radiopharmacol- Cardiol 2007;14(3 Suppl.):S118–24. ogy of O-(2-[F-18]fluoroethyl)-l-tyrosine for tumor imaging. J Nucl Med [13] Wahl RL. Principles of cancer imaging with fluorodeoxyglucose. In: Wahl RL, 1999;40(1):205–12. Buchanan JW, editors. Principles and Practice of Positron Emission Tomogra- [44] Wang MW, Yin DZ, Cheng DF, Li GC, Wang YX. Preparation of F-18 labeled phy. Williams & Wilkins, Lippincott; 2002. amino acid O-(2-[F-18]fluoroethyl)-l-tyrosine using indirect and direct label- [14] Sols A, Crane RA. Substrate specificity of brain hexokinase. J Biol Chem ing methods. J Radioanal Nucl Chem 2006;270(2):439–43. 1954;210:581–95. [45] Rau FC, Weber WA, Wester HJ, et al. O-(2-[F-18]fluoroethyl)-l-tyrosine (FET): d [15] Pacák J, Cerny M. History of the first synthesis of 2-deoxy-2-fluoro- -glucose a tracer for differentiation of tumour from inflammation in murine lymph 18 d the unlabeled forerunner of 2-deoxy-2-[ F]fluoro- -glucose. Mol Imaging nodes. Eur J Nucl Med Mol Imaging 2002;29(8):1039–46. Biol 2002;4:353–4. [46] Heiss P, Mayer S, Herz M, Wester HJ, Schwaiger M, Senekowitsch- [16] Delbeke D, Martin WH. Metabolic imaging with FDG: a primer. Cancer J Schmidtke R. Investigation of transport mechanism and uptake kinetics 2004;10(4):201–13. of O-(2-[18F]fluoroethyl)-l-tyrosine in vitro and in vivo. J Nucl Med [17] Pike VW, Eakins MN, Allan RM, Selwyn AP. Preparation of [1-C-11]-labeled 1999;40(8):1367–73. acetate—an agent for the study of myocardial-metabolism by positron emis- [47] Chaly T, Diksic M. Synthesis of no-carrier-added alpha-[C-11] methyl-l- sion tomography. Int J Appl Rad Isotopes 1982;33(7):505. tryptophan. J Nucl Med 1988;29(3):370–4. [18] Kihlberg T, Valind S, Langstrom B. Synthesis of [1-C-11], [2-C-11], [1-C-11](H- [48] Mzengeza S, Venkatachalam TK, Diksic M. Asymmetric radiosynthe- 2(3)) and [2-C-11](H-2(3))acetate for in-vivo studies of myocardium using sis of alpha-[C-11]methyl-l-tryptophan for pet studies. Nucl Med Biol pet. Nucl Med Biol 1994;21(8):1067–72. 1995;22(3):303–7. [19] Dimitrakopoulou-Strauss A, Strauss LG. PET imaging of prostate cancer with [49] Chakraborty PK, Mangner TJ, Chugani DC, Muzik O, Chugani HT. A C-11-acetate. J Nucl Med 2003;44(4):556–8. high-yield and simplified procedure for the synthesis of alpha-[C-11]methyl- [20] Mitterhauser M, Wadsak W, Krcal A, et al. New aspects on the preparation of L-tryptophan. Nucl Med Biol 1996;23(8):1005–8. [C-11]acetate - a simple and fast approach via distillation. Appl Rad Isotopes [50] Chugani DC, Muzik O. Alpha[C-11]methyl-l-tryptophan PET maps brain sero- 2004;61(6):1147–50. tonin synthesis and kynurenine pathway metabolism. J Cerebr Blood Flow [21] Wachter S, Tomek S, Kurtaran A, et al. C-11-Acetate positron emission tomog- Metab 2000;20(1):2–9. raphy imaging and image fusion with computed tomography and magnetic [51] Lundquist P, Hartvig P, Blomquist G, Hammarlund-Udenaes M, Langstrom resonance imaging in patients with recurrent prostate cancer. J Clin Oncol B. 5-Hydroxy-l-[beta-C-11]tryptophan versus alpha-[C-11]methyl-l- 2006;24(16):2513–9. tryptophan for positron emission tomography imaging of serotonin [22] Howard BV, Howard WJ. Lipids in normal and tumor cells in culture. Prog synthesis capacity in the rhesus monkey brain. J Cerebr Blood Flow Metab Biochem Pharmacol 1975;10:135–66. 2007;27(4):821–30. [23] Swinnen JV, Van Veldhoven PP, Timmermans L, et al. Fatty acid synthase [52] Grierson JR, Link JM, Mathis CA, et al. A radiosynthesis of fluorine-18 fluo- drives the synthesis of phospholipids partitioning into detergent- romisonidazole. J Nucl Med 1989;30:343–50. resistant membrane microdomains. Biochem Biophys Res Commun [53] Lim JL, Berridge MS. An efficient radiosynthesis of [18F]fluoromisonidazole. 2003;302:898–903. Appl Radiat Isot 1993;44:1085–91. [24] Schoder H, Larson SM. Positron emission tomography for prostate, bladder, [54] Cherif A, Yang DJ, Tansey W, et al. Rapid synthesis of 3-[18F]fluoro- and renal cancer. Semin Nucl Med 2004;34:274–92. 1-(2-nitro-1-imidazolyl)-2-propanol [18F]fluoromisonidazole. Pharm Res [25] Firnau G, Chiakal R, Garnett ES. Aromatic radiofluorination with 18F fluorine 1994;11:466–9. 18 l gas: 6-[ F]fluoro- -dopa. J Nucl Med 1984;25:1228–33. [55] Patt M, Kuntzsch M, Machulla HJ. Preparation of [18F]fluoromisonidazole [26] Adam MJ, Ruth TJ, Grierson JR, Abeysekera B, Pate BD. Routine synthe- by nucleophilic substitution on THP-protected precursor: yield depen- sis of L-[F-18]6-fluorodopa with F-18 acetyl hypofluorite. J Nucl Med dence on reaction parameters. J Radioanal Nucl Chem 1999;240: 1986;27(9):1462–6. 925–7. [27] Adam MJ, Jivan S. Synthesis and separation of 3-O-methyl-2-fluorodopa and [56] Tang G, Wang M, Tang X, Gan M, Luo L. Fully automated one-pot synthesis of 6-[F-18]-fluorodopa. J Label Compd Radiopharm 1992;31(1):39–43. [18F]fluoromisonidazole. Nucl Med Biol 2005;32:553–8. W. Wadsak, M. Mitterhauser / European Journal of Radiology 73 (2010) 461–469 469

[57] Foo SS, Abbott DF, Lawrentschuk N, et al. Functional imaging of intra-tumoral [90] Hara T, Kosaka N, Shinoura N, et al. PET imaging of brain tumor with [methyl- hypoxia. Mol Imaging Biol 2004;6:291–305. 11C] choline. J Nucl Med 1997;38:842–7. [58] Chapman JD, Franko AJ, Sharplin J. A marker for hypoxic cells in tumours with [91] Yamamoto Y, Nishiyama Y, Kameyama R, et al. Detection of hepatocellular potential clinical applicability. Br J Cancer 1981;43:546–50. carcinoma using C-11-choline PET: comparison with F-18-FDG PET. J Nucl [59] Rasey JS, Grunbaum Z, Magee S, et al. Characterization of radiolabeled fluo- Med 2008;49(8):1245–8. romisonidazole as a probe for hypoxic cells. Radiat Res 1987;111:292–304. [92] Reske SN. [C-11]choline uptake with PET/CT for the initial diagnosis of [60] Kumar P, Stypinski D, Xia H, et al. Fluoroazomycin arabinoside (FAZA): syn- prostate cancer: relation to PSA levels, tumour stage and anti-androgenic thesis, H-2 and H-3-labelling and preliminary biological evaluation of a therapy. Eur J Nucl Med Mol Imaging 2008;35(9):1740–1. novel 2-nitroimidazole marker of tissue hypoxia. J Label Compd Radiopharm [93] DeGrado TR, Baldwin SW, Wang SY, et al. Synthesis and evaluation 1999;42(1):3–16. of F-18-labeled choline analogs as oncologic PET tracers. J Nucl Med [61] Reischl G, Ehrlichmann W, Bieg C, et al. Preparation of the hypoxia imaging 2001;42(12):1805–14. PET tracer [F-18]FAZA: reaction parameters and automation. Appl Radiat Isot [94] DeGrado TR, Coleman RE, Wang S, et al. Synthesis and evaluation of 18F 2005;62(6):897–901. labeled choline as an oncologic tracer for positron emission tomography: [62] Souvatzoglou M, Grosu AL, Roper B, et al. Tumour hypoxia imaging with [F- initial findings in prostate cancer. Cancer Res 2001;61:110–7. 18]FAZA PET in head and neck cancer patients: a pilot study. Eur J Nucl Med [95] Hara T, Kosaka N, Kishi H. Development of [18F]-fluoroethylcholine for cancer Mol Imaging 2007;34(10):1566–75. imaging with PET: synthesis, biochemistry, and prostate cancer imaging. J [63] Picchio M, Beck R, Haubner R, et al. Intratumoral spatial distribution of Nucl Med 2002;43:187–99. hypoxia and angiogenesis assessed by F-18-FAZA and I-125-Gluco-RGD [96] Vallabhajosula S, Kostakoglu L, Kothari PJ, et al. Pharmacokinetics and autoradiography. J Nucl Med 2008;49(4):597–605. biodistribution of radiolabelled choline in human subjects: compari- [64] Piert M, Machulla HJ, Picchio M, et al. Hypoxia-specific tumor imaging with son of [11C]choline and [18F]fluorocholine. J Label Compd Radiopharm 18F-fluoroazomycin arabinoside. J Nucl Med 2005;46:106–13. 2005;48:pS281. [65] Kachur AV, Dolbier WR, Evans SM, et al. Synthesis of new hypoxia markers [97] Meyer GJ, Macke H, Schuhmacher J, Knapp WH, Hofmann M. Ga-68- EF1 and [F-18]-EF1. Appl Radiat Isot 1999;51(6):643–50. labelled DOTA-derivatised peptide ligands. Eur J Nucl Med Mol Imaging [66] Dolbier WR, Li AR, Koch CJ, Shiue CY, Kachur AV. [F-18]-EF5, a marker for PET 2004;31(8):1097–104. detection of hypoxia: synthesis of precursor and a new fluorination proce- [98] Breeman WAP, de Jong M, de Blois E, et al. Radiolabelling DOTA-peptides with dure. Appl Radiat Isot 2001;54(1):73–80. Ga-68. Eur J Nucl Med Mol Imaging 2005;32(4):478–85. [67] Komar G, Seppanen M, Eskola O, et al. F-18-EF5: a new PET tracer for imaging [99] Decristoforo C, Knopp R, von Guggenberg E, et al. A fully automated syn- hypoxia in head and neck cancer. J Nucl Med 2008;49(12):1944–51. thesis for the preparation of 68Ga-labelled peptides. Nucl Med Commun [68] Lewis JS, Laforest R, Buettner TL, et al. Copper-64-diacetyl-bis(N-4- 2007;28(11):870–5. methylthiosemicarbazone): an agent for radiotherapy. Proc Natl Acad Sci USA [100] Maecke HR, Hofmann M, Haberkorn U. (68)Ga-labeled peptides in tumor 2001;98(3), 1206+. imaging. J Nucl Med 2005;46(Suppl. (1)):172S–8S. [69] Vavere AL, Lewis JS. Examining the relationship between Cu-ATSM hypoxia [101] Hofmann M, Maecke H, Börner R, et al. Biokinetics and imaging with the selectivity and fatty acid synthase expression in human prostate cancer cell somatostatin receptor PET radioligand (68)Ga-DOTATOC: preliminary data. lines. Nucl Med Biol 2008;35(3):273–9. Eur J Nucl Med 2001;28(12):1751–7. [70] Lewis JS, Laforest R, Dehdashti F, et al. An imaging comparison of Cu- [102] Buchmann I, Henze M, Engelbrecht S, et al. Comparison of Ga-68-DOTATOC 64-ATSM and Cu-60-ATSM in cancer of the uterine cervix. J Nucl Med PET and In-111-DTPAOC (Octreoscan) SPECT in patients with neuroendocrine 2008;49(7):1177–82. tumours. Eur J Nucl Med Mol Imaging 2007;34(10):1617–26. [71] Skovgaard D, Kjaer M, Madsen J, Kjaer A. Noninvasive 64Cu-ATSM and PET/CT [103] Lee YS, Jeong JM, Kim HW, et al. An improved method of F-18 peptide label- assessment of hypoxia in rat skeletal muscles and tendons during muscle ing: hydrazone formation with HYNIC-conjugated c(RGDyK). Nucl Med Biol contractions. J Nucl Med 2009;50(6):950–8. 2006;33(5):677–83. [72] Grierson JR, Shields AF, Eary JF. Development of a radiosynthesis for 3- [104] Haubner R, Decristoforo C. Radiolabelled RGD peptides and peptidomimetics [18F]fluoro-3-deoxy-nucleosides. J Label Compd Radiopharm 1997;40:60–2. for tumour targeting. Front Biosci 2009;14:872–86. [73] Grierson JR, Shields AF. Radiosynthesis of 3-deoxy-3-[F-18]fluorothymidine: [105] Van de Wiele C, De Vos F, Slegers G, et al. Radiolabeled estradiol derivatives [F-18]FLT for imaging of cellular proliferation in vivo. Nucl Med Biol to predict response to hormonal treatment in breast cancer. Eur J Nucl Med 2000;27(2):143–56. 2000;27:1421–33. [74] Machulla HJ, Blocher A, Kuntzsch M, et al. Simplified labeling approach for [106] Mankoff DA, Shields AF, Krohn KA. PET imaging of cellular proliferation. Radiol synthesizing 3-deoxy-3-[F-18]fluorothymidine ([F-18]FLT). J Radioanal Nucl Clin N Am 2005;43:153–67. Chem 2000;243(3):843–6. [107] Kiesewetter DO, Kilbourn MR, Landvatter SW, et al. Preparation of four [75] Cleaver JE. Thymidine metabolism and cell kinetics. Front Biol 1967;6:43–100. fluorine-18-labeled estrogens and their selective uptakes in target tissues [76] Langen P, Etzold G, Hintsche R, Kowollick G. 3-Deoxy-3-fluorothymidine, a of immature rats. J Nucl Med 1984;25:1212–21. new selective inhibitor of DNA synthesis. Acta Biol Med Ger 1969;23:759–66. [108] Romer J, Fuchtner F, Steinbach J, et al. Automated production of 16␣- [77] Shields AF, Grierson JR, Dohmen BM, et al. Imaging proliferation in vivo with [18F]fluoroestradiol for breast cancer imaging. Nucl Med Biol 1999;26:473–9. [F-18]FLT and positron emission tomography. Nat Med 1998;4(11):1334–6. [109] Wadsak W, Mitterhauser M. Synthesis of [F-18]FETO, a novel potential 11- [78] Vallabhajosula S. 18F-labeled positron emission tomographic radiopharma- beta hydroxylase inhibitor. J Label Compd Radiopharm 2003;46(4):379–88. ceuticals in oncology: an overview of radiochemistry and mechanisms of [110] Mitterhauser M, Wadsak W, Wabnegger L, et al. In vivo and in vitro evaluation tumor localization. Semin Nucl Med 2007;37:400–19. of [F-18]FETO with respect to the adrenocortical and GABAergic system in [79] Barton DHR, Toh HT, Hesse RH, Pechet MM. Convenient synthesis of 5- rats. Eur J Nucl Med Mol Imaging 2003;30(10):1398–401. fluorouracil. J Org Chem 1972;37(2):329. [111] Wadsak W, Mitterhauser M, Rendl G, et al. [F-18]FETO for adrenocortical PET [80] Fowler JS, Finn RD, Lambrech Rm, Wolf AP. Synthesis of F-18-5-fluorouracil. imaging: a pilot study in healthy volunteers. Eur J Nucl Med Mol Imaging J Nucl Med 1973;14(1):63–4. 2006;33(6):669–72. [81] Diksic M, Farrokhzad S, Yamamoto YL, Feindel W. A simple synthesis of [112] Ettlinger DE, Wadsak W, Mien LK, et al. [F-18]FETO: metabolic considerations. F-18-labeled 5-fluorouracil using acetylhypofluorite. Int J Nucl Med Biol Eur J Nucl Med Mol Imaging 2006;33(8):928–31. 1984;11(2):141–2. [113] Bergström M, Bonasera TA, Lu L, et al. In vitro and in vivo primate evaluation of [82] Oberdorfer F, Hofmann E, Maierborst W. Preparation of F-18-labeled carbon-11-etomidate and carbon-11-metomidate as potential tracers for PET 5-fluorouracil of very high-purity. J Label Compd Radiopharm imaging of the adrenal cortex and its tumors. J Nucl Med 1999;39(6):982–9. 1989;27(2):137–45. [114] Bergström M, Juhlin C, Bonasera TA, et al. PET imaging of adrenal cortical [83] Brix G, Bellemann ME, Haberkorn U, Gerlach L, Lorenz WJ. Assessment tumors with the 11beta-hydroxylase tracer 11C-metomidate. J Nucl Med of the biodistribution and metabolism of 5-fluorouracil as monitored by 2000;41(2):275–82. F-18 PET and F-19 MRI: a comparative animal study. Nucl Med Biol [115] Mitterhauser M, Tögel S, Wadsak W, et al. Binding studies of [18F]-fluoride and 1996;23(7):897–906. polyphosphonates radiolabelled with [111In], [99mTc], [153Sm] and [188Re] [84] Christman D, Crawford EJ, Friedkin M, Wolf AP. Detection of DNA synthesis in on bone compartments: a new model for the pre vivo-evaluation of bone intact organisms with positron-emitting (methyl-11C)thymidine. Proc Natl seekers? Bone 2004;34:835–44. Acad Sci USA 1972;69(4):988–92. [116] Mitterhauser M, Toegel S, Wadsak W, et al. Binding studies of [18F]-fluoride [85] Poupeye E, Counsell RE, De Leenheer A, Slegers G, Goethals P. Synthesis and polyphosphonates radiolabelled with [99mTc], [111In], [153Sm] and of 11C-labelled thymidine for tumor visualization using positron emission [188Re] on bone compartments: verification of the pre vivo model? Bone tomography. Int J Rad Appl Instrum A 1989;40(1):57–61. 2005;37:404–12. [86] Steel CJ, Brady F, Luthra SK, et al. An automated radiosynthesis of 2-[C- [117] Toegel S, Hoffmann O, Wadsak W, et al. Uptake of boneseekers is solely asso- 11]thymidine using anhydrous [C-11]urea derived from[C-11]phosgene. Appl ciated with mineralisation! A study with [99mTc]-MDP, [153Sm]-EDTMP and Radiat Isot 1999;51(4):377–88. [18F]-fluoride on osteoblasts. Eur J Nucl Med Mol Imaging 2006;33:491–4. [87] Shields AF, Lim K, Grierson JR, Link JM, Krohn KA. Utilization of labeled thymi- [118] Toegel S, Mien LK, Wadsak W, et al. In vitro evaluation of no carrier added, car- dine in DNA synthesis: studies for PET. J Nucl Med 1990;31:337–42. rier added and cross-complexed [90Y]-EDTMP provides evidence for a novel [88] Wells P, Gunn RN, Steel C, et al. 2-[C-11]thymidine positron emission tomog- “foreign carrier theory”. Nucl Med Biol 2006;33:95–9. raphy reproducibility in humans. Clin Cancer Res 2005;11(12):4341–7. [119] Radiopharmaceutical Preparations. (Radiopharmaceutica, 5.0/0125). Euro- [89] Rosen MA, Jones RM, Yano Y, Budinger TF. C-11 choline—synthesis, purifi- pean Pharmacopoeia (Europäisches Arzneibuch), fifth ed. (5. Ausgabe cation, and brain uptake inhibition by 2-dimethylaminoethanol. J Nucl Med Grundwerk), Vienna: Official Austrian Version, Verlag Oesterreich GmbH; 1985;26(12):1424–8. 2005 pp. 823–831.