[18F]FDG
During infection, the human body responds with the migration and activation of immune cells to the site of infection. These activated immune cells markedly increase their glucose uptake. The glucose transporters on the membrane of immune cells transport [18F]FDG, a glucose analog into the cell. Once [18F]FDG is inside the immune cell it is phosphorylated like glucose but is unable to continue along the biochemical pathway for glucose utilization and remains trapped in the immune cell. In addition to this, some microorganisms like bacteria have been found to contribute to the [18F]FDG signal by actively taking up [18F]FDG [6]. Compared to other methods used in infection imaging, the process of imaging with FDG is relatively fast with the study complete within a few hours. Unlike white cell labeling, [18F]FDG does not require direct handling of blood and laborious labeling procedures. [18F]FDG can be used in patients with neutropenia where anatomical‐based imaging modalities or white cell imaging may be less helpful. [18F]FDG has some limitations including its non‐specificity that is particularly a problem in the early postoperative period when inflammation at the site of operation limits the ability of [18F]FDG to detect infection. [18F]FDG does not image the microorganism themselves but rather the downstream immune activation. Table 1 summarizes the targets of the radiopharmaceuticals used in infection.
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Tracers used in PET imaging of infection
Table 1: Biological process and target of currently available or potential PET tracers
Process Target Tracer example Stage
Activated immune cells Energy consumption [18F]FDG Clinical
Infiltration of leucocytes Radiolabeled white blood cells Clinical
Host defense Iron availability [68Ga]citrate Clinical
Iron scavenging [68Ga]siderophores Preclinical
Immune response Antimicrobial peptides Ubiquicidine peptides Clinical Bacterial cell wall Cell wall synthesis inhibition No PET tracers Preclinical
Bacterial biosynthesis Folic acid synthesis inhibition [18F]trimethoprim Preclinical
Protein synthesis inhibition [68Ga]puromycin Preclinical
RNA synthesis inhibition [11C]rifampicin Preclinical
DNA synthesis inhibition [68Ga]ciprofloxacin Preclinical Host defense Bacterial antigens 64Cu and 89Zr-radiolabeled
Antibodies Preclinical
Bacterial metabolism Carbohydrate metabolism [18F]carbohydrates Preclinical Various other processes [18F]FLT, PET-amino acids, etc. Preclinical
[18F]FDG and Different Microorganisms
[18F]FDG‐PET has been used in the evaluation of different microorganisms. For viruses, [18F]FDG has been used to stage the infection and assess HIV‐associated infections and malignancies [1, 7]. In bacterial imaging, [18F]FDG has been used to determine the site of infection when the site of infection is unknown or stage infection to determine the location of undiagnosed disease. In bacterial infections that require prolonged treatment like tuberculosis, [18F]FDG is useful in monitoring treatment of infection (Fig. 1) [8]. In fungal infections, [18F]FDG can be used to help diagnosis and guide therapy in a wide array of fungal infections [3, 9]. For parasitic infections, [18F]FDG has been used in the follow‐
up of treatment for alveolar echinococcosis [2].
Figure 1
18F‐FDG‐PET/CT scan before anti‐TB treatment and 2 months after initiation of treatment for interim assessment of treatment response.
(a) Maximum intensity projection (MIP) image before treatment (PET images only), showing extensive disease: pulmonary, cervical, axillary, mediastinal, abdominal, pelvic and inguinal lymph nodes, hepatic and skeletal metastasis to the lumbar spine and right humerus.
(b) MIP image after 2 months of anti‐TB treatment (PET images only): complete meta‐ bolic response of the pulmonary and right humeral lesions and the pelvic and inguinal lymph nodes.
Good metabolic response in the mediastinal, cervical, and axillary nodes. Active disease is still present in the lumber spine with progression of the hepatic lesions.
(c) Transverse scans showing axillary nodes before treatment (PET and integrated PET/CT images).
(d) Transverse scans showing response of axillary nodes after 2 month of anti‐TB therapy; nodal uptake diminished but still present
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Chapter ThreeFDG Imaging of Some Site‐Specific Infections Which Have Been Validated Osteomyelitis
[18F]FDG is useful in the diagnosis of osteomyelitis, especially chronic osteomyelitis. A meta‐analysis found the sensitivity and specificity of [18F]FDG‐PET in chronic osteomyelitis to be 96% and 91%, respectively [10]. [18F]FDG‐PET is particularly useful in multi‐focal osteomyelitis.
Spondylodiscitis
[18F]FDG is used for imaging spondylodiscitis where other imaging modalities may be suboptimal [11–
13]. Differentiating degenerative disease from infection is particularly challenging, and [18F]FDG‐
PET/CT is very useful for this purpose [12]. A meta‐analysis found the pooled sensitivity and specificity of [18F]FDG‐PET/CT in spondylodiscitis to be 97% and 88% [14].
Diabetic Foot
FDG‐PET is useful in diagnosing pedal osteomyelitis. One study found PET/CT had sensitivity and specificity of 81% and 93% in the diagnosis of pedal osteomyelitis in the diabetic foot [15].
Hip and Knee Prosthesis
[18F]FDG‐PET has been used in the evaluation of infected prosthesis of the hip and knee joints. Several studies found [18F]FDG‐PET useful and also established interpretation criteria [16–20]. A meta‐analysis of the diagnosis of infected knee and ankle prosthesis by [18F]FDG‐PET found a pooled sensitivity of 87% and specificity of 87% [21].
Infected Vascular Graft
Some studies have evaluated the role of [18F]FDG‐PET in the evaluation of infected vascular graft. The sensitivity, specificity, and accuracy of [18F]FDG‐PET in the diagnosis of infected vascular graft were 100%, 86%, and 97%%, respectively [22].
Infective Endocarditis
[18F]FDG‐PET/CT is used in the evaluation of infected prosthetic heart valves and is recommended by major international guidelines [23]. The sensitivity and specificity of [18F]FDG‐PET/CT in prosthetic heart valves was found to be 71% and 95%, respectively, after confounders were removed the sensitivity and specificity improved to 91% and 95% [24]. [18F]FDG‐PET/CT is not recommended in the assessment of native valve infective endocarditis; however, it is useful in detecting septic emboli from native valve endocarditis.
Cardiac Implantable Electronic Device Infectionss
[18F]FDG‐PET/CT is a useful additive tool in patients with suspected cardiac implantable electronic device infections [25]. A meta‐analysis found [18F]FDG‐PET had a pooled sensitivity and specificity of 87% and 94% for infected implantable cardiac devices [26].
Labeled White Blood Cells
During infection, chemotactic agents attract leucocytes to the site of infection. Radiolabeled WBC provides a means of imaging infection. Leucocytes have been labeled with [18F]FDG [27, 28]. The sensitivity and specificity per lesion were found to be 91% and 85%, respectively, and 86% for both sensitivity and specificity per patient [27]. The labeling efficiency of [18F]FDG labeled white cells are lower than SPECT‐based tracers, and the short half‐life of 18F is not ideal for delayed imaging which is essential for the diagnosis of infection by labeled WBCs. Leucocytes have also been labeled with 64Cu
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Tracers used in PET imaging of infection
because the long half‐life of 64Cu would permit delayed imaging to distinguish infection. The labeling efficiency and viability of the labeled leucocytes were higher than 111In‐labeled leucocyte. However, there was higher leakage from 64Cu compared to SPECT‐based labeled leucocytes [29]. The radiation exposure to the long‐lived lymphocytes from PET‐based radioisotope is a concern [30].
[68Ga]Citrate
[68Ga]citrate has been used to image osteomyelitis, tuberculosis, and soft‐tissue infections in human studies [31–33]. Iron is utilized by both mammals and microorganism for their metabolism. Gallium mimics iron, and in infection, the uptake of gallium is due to transferrin‐dependent and independent mechanisms. [68Ga]citrate like [18F]FDG lacks specificity and accumulates in sterile inflammatory process or malignancy [34]. The ease of preparation of the radiopharmaceutical and the availability of good manufacturing practice 68Ga‐generators makes the use of [68Ga]citrate appealing. The sensitivity and specificity for osteomyelitis was 100% and 76%, respectively. Larger studies with [68Ga]citrate are required to validate its use in various clinical scenarios.
Peptides
AMPs are peptides produced by the immune system of the host against microorganisms. They have low toxicity and have activity against a wide range of microorganisms. Several AMPs have been labeled for imaging infection. For PET imaging, ubiquicidin fragments have been most widely investigated.
Ubiquicidin (UBI) Fragments
UBI, a 51 amino acid peptide that binds negatively charged moieties in the cell wall of bacteria has shown promising results for imaging infection. Several fragments of UBI have been labeled with 68Ga and 18F and tested in different animal models [35–39]. The [68Ga]UBI fragments were able to distinguish infection and inflammation and had good localization of infection (Fig.2) [36]. [68Ga]UBI fragments (UBI29‐41 and UBI31‐38) have been used in humans with promising results [36, 37]. The UBI29‐41 fragment was labeled with 18F but did not have as much success in imaging infection as the 68Ga‐labeled counterpart due to significant defluorination and the lack of specific binding to Staphylococcus aureus in vivo. Larger studies with [68Ga]UBI would be necessary to validate the use of the tracer in different clinical situations.
Figure 2
PET/CT images of healthy mouse (a), mouse with sterile inflammation (b), and mouse infected with S. aureus (c). Images were acquired 2 h after injection of 68 Ga-NOTA-UBI-29-41 (1, coronal; 2, sagittal;
and 3, axial). Images correspond to frame 2 of 4. Arrows indicate inflamed or infected muscle (Mónica Vilche et al. J Nucl Med 2016; 57:622–627.)
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Chapter ThreeExperimental Radiopharmaceuticals
Peptides
Depsipeptide Derivative
A depsipeptide derivative was labeled with 68Ga by DOTA chelation and was used for in vivo evaluation of Escherichia coli in a mouse model and Staphylococcus aureus and Mycobacterium tuberculosis in a rabbit model [40, 41]. The tracer detected these infections but was not able to differentiate infection‐
associated inflammation from bacteremia.
K‐A9 Peptide
K‐A9 is a peptide that binds to Staphylococcus aureus that was selected and proposed as an imaging peptide [42]. The peptide was radiolabeled with 68Ga but was not found to be selective towards infection probably due to some factors including vascular leakiness, hyperemia, and the peptide‐
binding epitopes in dead bacteria [43].
Antibiotics and other antimicrobials
Antibiotics have been used extensively with SPECT tracers. A few have also been labeled with PET radionuclides.
Ciprofloxacin
Radiolabeled ciprofloxacin is one of the earliest antibiotics used in imaging infections. The antibiotic or its conjugates have been labeled with 68Ga and 18F [44, 45]. [68Ga]ciprofloxacin conjugates were labeled by bifunctional DOTA and NOTA chelators [44]. The [68Ga]ciprofloxacin conjugates (Fig. 3) could discriminate between Staphylococcus aureus infected sites in rat muscle from inflammation. These tracers are yet to be tested in the clinic. Ciprofloxacin was also labeled by nucleophilic 18F‐fluorination.
However, the radioactivity was not retained in the infected tissue, and the clearance from infected tissue was similar to non‐infected tissue [45].
Trimethoprim
Trimethoprim inhibits the synthesis of the nucleic acid thymidine in bacteria by inhibiting tetrahydrofolic acid. Trimethoprim has been labeled with 18F by nucleophilic fluorination and was found to show a very high uptake (>100‐fold) in live bacteria but not in other pathologies such as cancer or sterile inflammation in a mouse model. The biodistribution of this tracer has been determined in a nonhuman primate but no human studies have been carried out [46].
Puromycin
Puromycin is an antibiotic that interrupts bacteria protein synthesis by terminating the translation of ribosomes. Puromycin has been labeled with 68Ga by DOTA chelation and showed promising results for uptake in Staphylococcus aureus foci compared to sterile inflammation [47]. Puromycin has also been labeled with 18F for imaging protein synthesis [48]. This labeled antibiotic has not yet been tested for infection imaging.
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Tracers used in PET imaging of infection
Rifampicin
The anti‐mycobacterial agent rifampicin was radiolabeled with 11C. The SPECT equivalent of this radiolabeled antibiotic was evaluated as a tuberculosis imaging agent. The 11C‐labeled antibiotic was used to determine the distribution of the agent but may play a role in imaging the infection. The short half‐life of 11C however may limit its clinical application [49].
Isoniazid
The anti‐mycobacterial drug isoniazid was radiolabeled with 18F by nucleophilic substitution [50]. The tracer was used to image mice infected with Mycobacterium tuberculosis and the tracer showed good uptake at infected sites. This tracer is yet to be translated to human studies.
Pyrazinamide
Pyrazinamide, an anti‐mycobacterial agent, was labeled with 18F by a fluoride exchange reaction. The biodistribution of the 5‐[18F]pyrazinamide compound was assessed. The pyrazinamide analog did not show significant differences in infected tissue and uninfected mice model of tuberculosis. There was rapid and extensive defluorination of [18F]pyrazinamide. As a result of this, the fluorine analog of pyrazinamide may be limited as an infection imaging agent [51].
Fluconazole
Fluconazole an antifungal agent has been radiolabeled with 99mTc and was found to be an excellent tracer for detecting Candida albicans in mice [52]. The agent was also labeled with 18F. The PET tracer, however, may not be as successful in imaging candida infections as there was intense hepatic uptake due to the excretion of the [18F]fluconazole [53].
Antibody Labeling
Antibodies Against Gram‐Negative Bacteria
Polyclonal antibodies against the outer membrane protein of Yersinia enterocolitica, adhesion A, were labeled with 64Cu. The radiolabeling was achieved by chelation with 1,4,7‐triazacyclononane,1‐glutaric acid‐4,7‐acetic acid (NODAGA). The 64Cu‐labeled antibody was taken up in infected tissues in a dose‐
dependent manner. The 64Cu‐labeled antibody was able to detect infection in spleens of mice with low dose infection in contrast to FDG [54].
Antibodies Against Gram‐Positive Bacteria
Gram‐positive bacteria surface molecule lipoteichoic acid (LTA) was imaged with the anti‐LTA antibody SAC55 labeled with 89Zr after its conjugation with bifunctional chelate, p‐SCN‐Bn‐DFO. The tracer was tested in a mouse model of prosthetic joint infection and was able to distinguish infection prosthetic joints from inflammation non‐infected joint. The findings are yet to be translated to humans [55].
Antibody Against Simian Immunodeficiency Virus Envelope Protein
A monoclonal antibody against the glycoprotein Gp120 on the surface of Simian immunodeficiency virus has been labeled with 64Cu. The radiolabeling method was DOTA chelation of the modified antibody. The tracer was used to provide the location and quantification of in vivo viral replication in a nonhuman primate [56].
Aspergillus Fumigatus‐Specific Monoclonal Antibody JF5
The JF5 monoclonal antibody binds to an extracellular mannoprotein of Aspergillus fumigatus that is produced during growth of the fungi. The JF5 monoclonal antibody was radiolabeled with 64Cu by
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Chapter Threechelating with DOTA. The tracer was tested in a neutrophil‐depleted mouse model of invasive fungal infection. The tracer was able to distinguish the growth phase of Aspergillus fumigatus from other pathology including infection with other microorganisms [57].
Imaging Targeted Molecules or Metabolism Specific to Microorganisms Siderophores
Siderophores are proteins used by microorganisms to scavenge iron in their micro‐environment. 68Ga‐
labeled siderophores have been used for imaging invasive pulmonary aspergillosis in an animal model.
Two compounds triacetylfusarinine and ferrioxamine E were radiolabeled using the direct method. The
68Ga‐labeled siderophores showed high sensitivity and selectivity for rat lungs infected with Aspergillus fumigatus [58, 59].
Carbohydrate Metabolism
Maltodextrins: Maltohexose was labeled with 18F by nucleophilic substitution. [18F]fluoromaltohexose is transported into the bacteria by a maltodextrin transporter which is unique to bacteria making it a promising agent. In an in vivo study involving a rat model of an implantable cardiac device, the tracer was found to be specific and sensitive for the detection of Staphylococcus aureus [60]. The tracer had earlier been shown to be sensitive and specific for the detection of Escherichia coli in in vitro studies [61]. The tracer was found to accumulate in as few as 105 colony forming units [62].
Maltotriose: Maltotriose is also utilized by bacteria but not mammalian cells. 6‐[18F]fluoromaltotriose was radiolabeled by nucleophilic 18F‐fluorination. In preliminary in vivo studies in mice, the tracer was specifically taken up by viable Escherichia coli but not in sterile inflammation [63].
Sorbitol: Sorbitol is a substrate for Gram‐negative bacteria, Enterobacteriaceae. The tracer [18F]fluorosorbitol ([18F]FDS). The tracer which can easily be synthesized from [18F]FDG (Fig. 4) was found to be promising as a bacteria‐specific imaging agent some studies. [18F]FDS was found to be a better biomarker than [18F]FDG, in tracking bacterial lung infection using a mouse model [64]. In another study, [18F]FDS was found to accumulate in susceptible and carbapenem‐resistant drug isolates [65]. [18F]FDS was evaluated in humans for safety and no adverseeffects were observed 24 h after administration of the tracer [66]. [18F]FDS was able to detect Enterobacteraceae in mixed infections, brain infections, and mice undergoing chemotherapy [67].
Figure 3: Chemical structure of [68Ga]‐Bz‐SCN‐ciprofloxacin Figure 4: Synthesis of [18F]FDS from clinical grade [18F]FDG via a reduction step
[18F]Fluorodeoxyglucose‐phosphate (FDG‐P): Bacteria can utilize FDG‐P using universal hexose phosphate transporters which are not present in mammalian cells. Unlike FDS which targeted only
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Tracers used in PET imaging of infection
Gram‐negative bacteria, it was hoped that FDG‐P would be useful in imaging Gram‐positive bacteria with high sensitivity and bacteria selectivity. FDG‐P uptake was higher in Staphylococcal‐infected catheter implants in mice compared to uninfected implants [68].
D‐mannitol: The ability of some bacteria to utilize d‐mannitol as a source of energy has been exploited for potential bacterial imaging. The sugar was radiolabeled with 18F. The radiolabeled d‐mannitol was rapidly taken up by Gram‐positive and Gram‐negative bacteria in a myositis model in mice. The tracer was taken up by both susceptible and resistant microorganisms [65].
Fluoroacetamide‐d‐glycopyranose (FAG): Amino sugars are important components of the bacterial cell wall. This makes them a good target for imaging bacteria. An analog of an amino sugar was radiolabeled by microwave irradiation to [18F] FAG. The radiolabeled tracer was able to distinguish infection with Escherichia coli and sterile inflammation in an animal model [69].
Trehalose analogs: Trehalose is a disaccharide sugar that is used by plants and microorganism for some functions like stress protection and energy storage. A modified 18F‐labeled analog of trehalose has been produced by the chemoenzymatic conversion of [18F]FDG. The 18F‐labeled analog of trehalose was shown to be metabolized by Mycobacterium smegmatis but not mammalian cell in cellular uptake experiments. There are no clinical studies available [70].
Other Metabolites Fialuridine (FIAU)
Fialuridine is a nucleoside analog that is a building block of the nucleic acid of microorganisms.
Fialuridine is phosphorylated by thymidine kinase and remains trapped in the microorganism allowing imaging is tagged with a radioisotope. FIAU has been radiolabeled with 18F and 124I [68, 71, 72]. The [18F]FIAU was radiolabeled by nucleophilic substitution. The synthesis of [18F]FIAU is more complex and achieves lower yields than [124I]FIAU. When [124I]FIAU was used in the evaluation of prosthetic joint prosthesis the image quality and specificity was low [73].
Para‐Aminobenzoic acid (PABA)
PABA is utilized by bacteria in the synthesis of folate. In mammals, folate is obtained from the diet and mammals cannot utilize PABA. This makes PABA a good target to image infection. PABA has been labeled with 11C and 18F [74, 75]. In an animal model, the [11C]PABA was able to distinguish between infection and sterile inflammation. The [18F]PABA was tested in an animal model where it was found to be useful in imaging Staphylococcus aureus infection.
D‐Methionine
D‐Methionine was labeled with 11C and successfully used to image bacteria. D‐amino acids are used solely by bacteria for building their cell walls. The [11C] methionine rapidly accumulated in Gram‐
positive and Gram‐negative bacteria. No human study has been done on this PET tracer [76].
Prothrombin
Some Staphylococcus sp. produce staphylocoagulase which help bacteria evade detection by the immune system of their host by covering their antigen with clotted blood from the host. An analog of prothrombin was labeled with 64Cu by chelation with DOTA. This enabled the detection of Staphylococcus sp.‐induced endocarditis in mice [77].
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Chapter Three
Transferrin
Transferrin is a protein involved in the transport of iron. Transferrin was labeled with 68Ga as [68Ga]apotransferrin. The tracer was found to be able to detect Staphylococcus aureus infection in a rat model within an hour of injection [78].
Nonspecific Tracers Used in Infection to Discriminate Infection from Malignancy
[18F]fluorothymidine ([18F]FLT)
Synthesis of DNA can be imaged by [18F]FLT. During an infection, microorganisms are constantly growing with actively synthesizing nucleic acids. The growth of Staphylococcus aureus in rabbit was imaged using [18F]FLT. However, nucleic acid formation is not limited to bacteria growth and makes the tracer just as nonspecific as [18F]FDG [79, 80]. In another study using a model with Yersinia enterocolitica, [18F]FLT was not useful in assessing bacterial proliferation [81].
Other clinically available PET tracers have been used in the evaluation of different aspects a particular infection. In tuberculosis, for example, [18F]NaF used to evaluate calcification of tuberculous granulomas in mice, the complex lipid covering was evaluated by [11C]choline or [18F]fluoroethylcholine. The use of these tracers is limited to the pathology of a particular infection and lacked specificity for the infection [8].
Other clinically available PET tracers have been used in the evaluation of different aspects a particular infection. In tuberculosis, for example, [18F]NaF used to evaluate calcification of tuberculous granulomas in mice, the complex lipid covering was evaluated by [11C]choline or [18F]fluoroethylcholine. The use of these tracers is limited to the pathology of a particular infection and lacked specificity for the infection [8].