PET imaging of infection
The location of pathogen induced inflamed tissue can be found using inflammation imaging with PET, which is sufficient in many clinical situations, but it would also be useful to distinguish between inflammation caused by for example normal wound heeling and post-surgical infection, and between sterile and infectious loosening of joint replacements. At present the role of PET in the discrimination between infection and non-microbial inflammation, let alone identification of pathogens, is limited, but developing rapidly. Several PET radiotracers have been developed for imaging bacteria (Auletta et al., 2019; Mota et al., 2020), viruses, and other pathogens.
Bacterial cell walls contain molecular structures that can be targeted by specific antibodies, antibody fragments, or affibodies. β-D-glucans on some bacteria and fungi (de Sousa Lacerda et al., 2016), and lipoteichoic acid (LTA) on Gram-positive bacteria (Pickett et al., 2018) are examples of such targets.
Mannans, containing mannose, are an important constituent of cell walls of yeasts and fungi. Cells involved in inflammatory response express mannose receptor (CD206), offering a target for inflammation imaging.
Certain bacteria can produce extracellular polymeric substance, and stay embedded in this matrix which provides them protection from the immune system and antibiotics (Otto, 2018), and may prevent the binding of imaging agents targeted to bacterial surface structures. These "biofilms" aggregate on surfaces such as catheters, artificial heart valves, joint prostheses, and bone. Life-threatening sepsis is often caused by Staphylococcus aureus biofilms.
Phage-display selected peptides with an affinity towards S aureus biofilm have been labeled with 68Ga (Nielsen et al., 2016). In pig osteomyelitis model, no uptake was seen in the lesions (Afzelius et al., 2020).
Antimicrobial peptide (AMP), or host defence peptide, is a general term for over thousand molecules identified so far that could be described as natural microbicides, since they are produced by eukaryotic cells, also in humans, to target prokaryotic cells. AMPs are electrostatically attracted to the negatively charged bacterial cell walls, destroying the membranes or inhibiting intracellular processes. Although bacterial mass in infected tissues is small, the large surface-to-mass ratio of bacteria enables imaging applications.
Ubiquicidin and its fragments have been radiolabelled, and shown promise in animal models in PET and SPECT imaging. [68Ga]Ga-NOTA-UBI(31-38) could detect muscle infection in animal model and in humans (Bhatt et al., 2018). [68Ga]Ubiquicidin did not accumulate in osteomyelitis lesions in porcine model (Afzelius et al., 2020).
Exposed phosphatidylserine (PS) in apoptotic cells is the natural signal to macrophages to start phagocytosis. Many intracellular pathogens exploit the hosts response to apoptosis by exposing PS on their surface or by cloaking themselves in host cell derived PS-containing vesicles, to facilitate binding, entry, and immune response evasion (Rice et al., 2016). Apoptotic mimicry is utilized by many viruses, such as hepatitis B, HIV, and dengue; and some parasitic micro-organisms, such as trypanosomatids. Also certain bacteria, such as Listeria monocytogenes, emerge from infected macrophages packaged in PS-coated vesicles, thus gaining access to healthy macrophages via phagocytosis.
Phosphatidylserine is a used as a target in PET imaging of apoptosis, and the PET radiopharmaceuticals developed to target PS may be useful on detecting these pathogens. Obviously, PS radioligands cannot discriminate between apoptotic mimicry and normal tissue apoptosis.
Not only phosphatidylserine but also other anionic phospholipids can trigger the same response. Anionic surface charge is a common feature of the bacterial cell envelope, achieved by phosphate-containing lipopolysaccharides (LPS), techoic acids, or phospholipids such as phosphatidylglycerol (PG) and cardiolipin (CL). Escherichia coli, Staphylococcus aureus, and Streptococcus pyogenes are examples of these bacteria.
Siderophore ("iron carrier") is a general term for small molecules secreted by most micro-organisms for Fe3+ acquisition and storage (Petrik et al., 2017). Most fungi and bacteria can utilize siderophores released by other micro-organism, even if they themselves have lost the ability to produce siderophores. Since the chemistry of Ga3+ is very similar to Fe3+, 68Ga-labeled siderophores and [68Ga]Ga-citrate have been used to target bacterial and especially fungal infections. Siderophores such as [68Ga]TAFC, [68Ga]FOXE, [68Ga]DFO-B, and [68Ga]PVD-PAO1 could be used as specific infection imaging, in comparison to non-specific 68Ga-citrate or 68Ga-colloids (Petrik et al., 2015, 2018, 2020, and 2021; Misslinger et al., 2021).
Maltodextrin transporter is only found in bacteria, providing a specific target for bacterial imaging (Ning et al., 2011). First-generation radioligands targeting this transporter, fluorine-18 labelled maltohexose (Ning et al., 2014) and 6-[18F]fluoromaltose (Gowrishankar et al., 2014) could accurately distinguish infection from sterile inflammation, but suffered from suboptimal pharmacokinetics and had poor signal-to-noise ratios. Next-generation radioligand 6''-[18F]fluoromaltotriose is superior compared to those. It has been shown to distinguish bacterial infection from sterile inflammation (Gowrishankar et al., 2017), and to detect infective endocarditis in mouse model (Wardak et al., 2020).
Unlike mammals, bacteria and fungi can synthesize folate. para-aminobenzoic acid (PABA, 4-aminobenzoic acid) is an intermediate in the synthesis, and the natural substrate for dihydropteroate synthase. Mammalian cells do not express this enzyme and PABA is rapidly metabolized and excreted into urine, while in bacteria PABA is rapidly incorporated into folate. DHPS inhibitors (sulfa drugs) have been used as antimicrobial drugs since 1930s.
Labelled PABA analogues [11C]PABA and 2-[18F]F-PABA can distinguish between infection and sterile inflammation in mouse, rat, and rabbit models and could be used to monitor drug response (Mutch et al., 2018; Zhang et al., 2018; Ordonez et al., 2022). In animals and especially in humans, both [11C]PABA and 2-[18F]F-PABA are very rapidly acetylated into form that bacteria cannot be incorporated into folate. Acetylation could be slowed down by pre-administering PABA (Ordonez et al., 2022). Replacement of the aromatic amine in 2-[18F]F-PABA with a nitro group resulted in a prodrug 2-[18F]F-EN, which is converted into 2-[18F]F-PABA inside bacteria (Li et al., 2020). Additionally the renal clearance can be slowed down by esterification of the aromatic carboxyl group resulting in 2-[18F]F-ENB. Both 2-[18F]F-EN and 2-[18F]F-ENB gave better results in animal infection model than 2-[18F]F-PABA (Li et al., 2020).
[18F]FIAU and [124/125I]FIAU were initially developed for imaging of cells transfected with reporter gene (HSV1 thymidine kinase). Cells which express the enzyme will phosphorylate the radiopharmaceutical, leading to trapping of the isotope label. These radiotracers are also substrates for the endogenous thymidine kinase in most bacteria, possibly enabling in vivo visualization of bacteria and the effect of antimicrobial therapy (Diaz Jr et al., 2007; Boerman et al., 2012; Pullambhatla et al., 2012; Jang et al., 2012; Peterson et al., 2013) but radiotracers may need to be optimized to provide better image quality (Zhang et al., 2016; Cho et al., 2020).
Sorbitol is a metabolic substrate for certain strains of gram-negative bacteria. 2-[18F]-fluorodeoxysorbitol ([18F]FDS) has been shown in mice studies to specifically accumulate in tissues with active infection, but not in inflamed tissue (Li et al., 2008; Weinstein et al., 2014; Yao et al., 2016; Ordonez et al., 2017; Li et al., 2018). [18F]FDS can be used to assess BBB disruption induced by focused ultrasound (Hugon et al., 2021).
[18F]FDS is easy to produce from [18F]FDG by a one-step reduction, it is metabolically stable, and rapidly excreted to urine via kidneys (Wakabayashi et al., 2016; Werner et al., 2019). [18F]FDS is suitable for human studies from a radiation dosimetry perspective (Zhu et al., 2016).
D-amino acids (DAAs) can be consumed by micro-organisms but not by most mammalian cells, and therefore some labelled DAAs have been investigated for infection imaging with promising results; these include d-methionine (Neumann et al., 2017; Stewart et al., 2020; Polvoy et al., 2022), d-alanine (Parker et al., 2020), and d-glutamine (Renick et al., 2021; Co et al., 2022).
Fluoroquinolones target a highly conserved bacterial type II topoisomerase (DNA gyrase and topoisomerase IV), causing DNA fragmentation and in sufficiently high doses leads to cell death.
The uptake of 99mTc- and 18F-labelled ciprofloxacin have been developed for visualisation of lesions with bacterial infection. However, nonspecific binding of [18F]ciprofloxacin is relatively high and it is also retained in granulocytes. Its uptake is affected by general inflammatory responses such as increased perfusion and vascular leakage, and it is not suited as a bacteria-specific infection imaging agent (Langer et al., 2005). Uptake of also gallium-68 labelled ciprofloxacin derivatives seem to be non-specific (Gouws et al., 2022).
[18F]Fleroxacin, [18F]trovafloxacin, and [18F]lomefloxacin have been studied in animal and human studies, with disappointing results (Gouws et al., 2022).
Trimethoprim inhibits bacterial dihydrofolate reductase (DHFR). DHFR reduces dihydrofolic acid to tetrahydrofolic acid, which is a precursor in thymidine synthesis pathway and required in DNA synthesis. Trimethoprim is highly selective towards bacterial DHFR over human DHFR.
Carbon-11 and fluorine-18 labelled trimethoprim derivatives have been developed, and shown to specifically detect bacterial inflammation over sterile inflammation in animal models (Sellmyer et al., 2017a and 2017b). Trimethoprim-resistant bacteria strains generally have a wild-type copy of DHFR, and could be imaged with [11C]trimethoprim (Lee et al., 2022). [11C]Trimethoprim PET could also detect infectious lesions in humans (Lee et al., 2022).
Isonicotinid acid hydrazine (INH, isoniazid) is a prodrug selectively targeting mycobacteria, since it disrupts synthesis of mycolic acid. INH is metabolically trapped inside mycobacteria.
PT70 and PT119 are INH derivatives which directly inhibit the key enzyme of mycolic acid synthesis pathway. These have been labelled with fluorine-11 to study the pharmacokinetics of the drugs (Wang et al., 2014 and 2015).
Antiviral drugs could be labelled and used in virus-specific PET and SPECT imaging (Bray et al., 2010). Examples of labelled antivirals include [11C]oseltamivir (Tamiflu) (Seki et al., 2014), and [18F]FPMPA (analogue of antiretroviral tenofovir) (Di Mascio et al., 2009).
- Inflammation imaging
- Leukocytes and platelets
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Updated at: 2023-02-08
Created at: 2015-08-18
Written by: Vesa Oikonen