Arabinofuranose-derived positron-emission tomography radiotracers for detection of pathogenic microorganisms

Article abstract of DOI:10.1002/jlcr.3835

PURPOSE: Detection of bacteria-specific metabolism via positron emission tomography (PET) is an emerging strategy to image human pathogens, with dramatic implications for clinical practice. In silico and in vitro screening tools have recently been applied to this problem, with several monosaccharides including l-arabinose showing rapid accumulation in Escherichia coli and other organisms. Our goal for this study was to evaluate several synthetically viable arabinofuranose-derived ¹⁸F analogs for their incorporation into pathogenic bacteria. PROCEDURES: We synthesized four radiolabeled arabinofuranose-derived sugars: 2-deoxy-2-[¹⁸F]fluoro-arabinofuranoses (d-2-¹⁸F-AF and l-2-¹⁸F-AF) and 5-deoxy-5-[¹⁸F]fluoro-arabinofuranoses (d-5-¹⁸F-AF and l-5-¹⁸F-AF). The arabinofuranoses were synthesized from ¹⁸F^- via triflated, peracetylated precursors analogous to the most common radiosynthesis of 2-deoxy-2-[¹⁸F]fluoro-d-glucose ([¹⁸F]FDG). These radiotracers were screened for their uptake into E. coli and Staphylococcus aureus. Subsequently, the sensitivity of d-2-¹⁸F-AF and l-2-¹⁸F-AF to key human pathogens was investigated in vitro. RESULTS: All ¹⁸F radiotracer targets were synthesized in high radiochemical purity. In the screening study, d-2-¹⁸F-AF and l-2-¹⁸F-AF showed greater accumulation in E. coli than in S. aureus. When evaluated in a panel of pathologic microorganisms, both d-2-¹⁸F-AF and l-2-¹⁸F-AF demonstrated sensitivity to most gram-positive and gram-negative bacteria. CONCLUSIONS: Arabinofuranose-derived ¹⁸F PET radiotracers can be synthesized with high radiochemical purity. Our study showed absence of bacterial accumulation for 5-substitued analogs, a finding that may have mechanistic implications for related tracers. Both d-2-¹⁸F-AF and l-2-¹⁸F-AF showed sensitivity to most gram-negative and gram-positive organisms. Future in vivo studies will evaluate the diagnostic accuracy of these radiotracers in animal models of infection.

Full text of DOI:10.1002/jlcr.3835

Received: 20 December 2019
DOI: 10.1002/jlcr.3835
Revised: 6 February 2020
Accepted: 26 February 2020
R E S E A R C H A R T I C L E
Arabinofuranose-derived positron-emission tomography
radiotracers for detection of pathogenic microorganisms
Mausam Kalita1 | Matthew F.L. Parker1 | Justin M. Luu1
Megan N. Stewart1 | Joseph E. Blecha1 | Henry F. VanBrocklin1
|
|
Michael J. Evans1 | Robert R. Flavell1 | Oren S. Rosenberg2
Michael A. Ohliger1,3 | David M. Wilson1
|
1Department of Radiology and Biomedical
Abstract
Imaging, University of California San
Francisco, San Francisco, California
PURPOSE: Detection of bacteria-specific metabolism via positron emission
tomography (PET) is an emerging strategy to image human pathogens, with
dramatic implications for clinical practice. In silico and in vitro screening tools
have recently been applied to this problem, with several monosaccharides
including L-arabinose showing rapid accumulation in Escherichia coli and other
organisms. Our goal for this study was to evaluate several synthetically viable
arabinofuranose-derived 18F analogs for their incorporation into pathogenic
bacteria. PROCEDURES: We synthesized four radiolabeled arabinofuranose-
derived sugars: 2-deoxy-2-[18F]fluoro-arabinofuranoses (D-2-18F-AF and
L-2-18F-AF) and 5-deoxy-5-[18F]fluoro-arabinofuranoses (D-5-18F-AF and L-
5-18F-AF). The arabinofuranoses were synthesized from 18F- via triflated,
peracetylated precursors analogous to the most common radiosynthesis of
2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG). These radiotracers were screened
for their uptake into E. coli and Staphylococcus aureus. Subsequently, the sensi-
tivity of D-2-18F-AF and L-2-18F-AF to key human pathogens was investigated
in vitro. RESULTS: All 18F radiotracer targets were synthesized in high radio-
chemical purity. In the screening study, D-2-18F-AF and L-2-18F-AF showed
greater accumulation in E. coli than in S. aureus. When evaluated in a panel of
pathologic microorganisms, both D-2-18F-AF and L-2-18F-AF demonstrated sen-
sitivity to most gram-positive and gram-negative bacteria. CONCLUSIONS:
Arabinofuranose-derived 18F PET radiotracers can be synthesized with high
radiochemical purity. Our study showed absence of bacterial accumulation for
5-substitued analogs, a finding that may have mechanistic implications for
related tracers. Both D-2-18F-AF and L-2-18F-AF showed sensitivity to most
gram-negative and gram-positive organisms. Future in vivo studies will evalu-
ate the diagnostic accuracy of these radiotracers in animal models of infection.
2Department of Medicine, University of
California San Francisco, San Francisco,
California
3Department of Radiology, Zuckerberg
San Francisco General Hospital, San
Francisco, California
Correspondence
: David Wilson, Department of Radiology
and Biomedical Imaging, University of
California, San Francisco, 505 Parnassus
Ave, San Francisco, CA 94143, USA.
Email: david.m.wilson@ucsf.edu
Michael Ohliger, Department of
Radiology and Biomedical Imaging,
University of California, San Francisco,
1001 Potrero Ave., San Francisco, CA
94110, USA.
Email: michael.ohliger@ucsf.edu
Oren Rosenberg, Department of Medicine,
University of California, San Francisco,
513 Parnassus Ave., San Francisco, CA
94143, USA.
Email: oren.rosenberg@ucsf.edu
Funding information
National Institute of Biomedical Imaging
and Bioengineering, Grant/Award
Number: NIH R01EB024014, NIH
R01EB025985; U.S. Department of
Defense, Grant/Award Number: DOD
A132172; UCSF
K E Y W O R D S
arabinofuranose, imaging, infection, positron emission tomography, sugars
J Label Compd Radiopharm. 2020;1–9.
wileyonlinelibrary.com/journal/jlcr
© 2020 John Wiley & Sons, Ltd.
1

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KALITA ET AL.
1 | INTRODUCTION
metabolites. For example, in gram-positive bacteria Clos-
tridium tetani, D-arabinose-5-phosphate (A5P) isomerase
catalyzes D-ribulose-5-phosphate conversion into D-arabi-
nose-5-phosphate.20 Beyond transport, biotransforma-
tion, and phosphorylation, there are additional
mechanisms of bacteria-specific probe retention for L and
D-arabinose-derived analogs. For example, the L-arabi-
nose-binding protein (ABP) derived from the E. coli peri-
plasm can bind both α- and β- anomers of L-arabinose.
The ABP acts as a primary receptor for L-arabinose accu-
mulation by bacteria.21–24
We hypothesized that different structural isomers of
18F-labeled arabinofuranose would incorporate differen-
tially into bacteria. In the context of high bacterial accu-
mulation, these radiotracers could potentially be used as
PET probes to distinguish acute bacterial infection from
radiologic mimics. The radiotracers described in this
report could also potentially be applied to imaging fungi
and mammalian cells that metabolize arabinofuranoses.
In this work, we developed effective radiosyntheses of
four arabinofuranose-derived PET radiotracers, namely
Bacterial infection is a major health threat in the United
States and worldwide. According to the Center for Dis-
ease Control and Prevention (CDC), more than 2 million
people get infected and approximately 23 000 die of bac-
terial infections each year in United States, with special
risks posed by hospital-acquired and antibiotic-resistant
microorganisms.1 Accurate and rapid diagnosis of infec-
tion will reduce patient morbidity through effective ther-
apeutic intervention. In addition to traditional laboratory
and sampling methods, diagnostic imaging tools are fre-
quently used in the work-up of infection and include
computed tomography (CT), single photon emission CT
(SPECT), magnetic resonance imaging (MRI), and
hyperpolarized 13C magnetic resonance (HP MR) spec-
troscopy. While essential, these methods frequently suffer
from low diagnostic accuracy due to their inability to dif-
ferentiate infection from inflammation, cancer, and rheu-
matologic disease and tedious data acquisition.2–5 In light
of these concerns, several groups including ours hope to
develop faster, more accurate, and higher resolution posi-
tron emission tomography (PET) tools for imaging bacte-
rial infection. Newer radiotracers derived from maltose,
para-aminobenzoic acid (PABA), D-amino acids, bacterial
siderophores, sorbitol, and other small-molecules have
targeted bacteria-specific metabolism, with the potential
for widespread clinical dissemination.6–12 The sorbitol-
derived radiotracer 2-deoxy-2-[18F]fluoro-sorbitol ([18F]
FDS), a reduced product of [18F]FDG, has been studied
extensively in elegant preclinical models and more
recently in patients.13–15 This radiotracer shows high
uptake in key strains of gram-negative bacteria including
2-deoxy-2-[18F]fluoro-D-arabinofuranose
2-deoxy-2-[18F]fluoro-L-arabinofuranose
5-deoxy-5-[18F]fluoro-D-arabinofuranose
(D-2-18F-AF),
(L-2-18F-AF),
(D-5-18F-AF),
and 5-deoxy-5-[18F]fluoro-L-arabinofuranose (L-5-18F-
AF). These radiotracers were subsequently screened for
bacterial incorporation in vitro.
2 | EXPERIMENTAL
2.1 | Syntheses of 19F arabinofuranose
standards
multidrug
resistant
Escherichia
coli,
Klebsiella
pneumoniae, and Yersinia enterocolitica.
2.1.1 | 5-deoxy-5-fluoro-D-
As shown by [18F]FDS, exploiting the metabolic
requirements of pathogenic bacteria for various sugar
alcohols can drive the discovery of new PET tracers
targeting bacterial infection. In silico and in vitro screen-
ing methods for bacteria-specific sugars, employing com-
mercially available β-emitting 14C and 3H molecules,
have identified numerous other candidate probes includ-
ing arabinose-derived structures. Specifically, [1-14C]L-
arabinose showed avid and selective accumulation in
E. coli.16 In gram-negative bacteria, both D- and L-
arabinose are converted into ribulose by the action of
arabinose isomerase (Figure 1). Next, ribulokinase phos-
phorylates the oxygen at the 5-position generating
ribulose-5-phosphate. Finally, the action of ribulose-
5-phosphate-4-epimerase results in xylulose-5-phosphate,
which enters the pentose phosphate pathway (PPP).17–19
In several bacteria, additional metabolic pathways may
be employed to incorporate D- and L-arabinose and their
arabinofuranose and 5-deoxy-5-fluoro-L-
arabinofuranose (D-5-F-AF and L-5-F-AF)
For detailed descriptions of syntheses for all new com-
pounds, please refer to the Supporting Information. The
compound D-5-F-AF was synthesized according to a pre-
viously published procedure.25 The compound L-5-F-AF
(5L-5, see B.1. in Supporting Information) was synthe-
sized using an analogous method.
2.1.2 | 2-deoxy-2-fluoro-D-
arabinofuranose and 2-deoxy-2-fluoro-L-
arabinofuranose (D-2-F-AF and L-2-F-AF)
Compound D-2-F-AF was synthesized according to a pre-
viously published procedure.26 (2L-3, see B.2. in
Supporting Information).

KALITA ET AL.
3
FIGURE 1 Potential mechanisms
of arabinofuranose incorporation by
bacteria. A, Structures of 18F D- and L-
arabinofuranose analogs studied and, B,
metabolism of D- and L-arabinofuranose
highlighting the metabolic fates of the
1-5 positions of these molecules.
Arabinose isomerases (E.C. 5.3.1.3 and
5.3.1.4 for D and L forms respectively)
convert D-(and L-) arabinoses into D-(and
L-) ribuloses, which are subsequently
phosphorylated at 5-position hydroxyl
group catalyzed by ribulose kinase (E.C.
2.7.1.47 and 2.7.1.16 for D and L forms
respectively). Next, ribulose-5-phosphate
epimerase (E.C. 5.1.3.1 and 5.1.3.4 for D
and L forms respectively) yields 2-D-
xylulose-5-phosphate, which finally
enters the pentose phosphate pathway.
Of note, both 2- and 5-18F substitution
yields 18F structures that are poor
substrates for arabinose isomerase and
ribulose kinase respectively; radiotracer
incorporation for these analogs is likely
mediated by transport and arabinose
binding. DMSO, dimethyl sulfoxide; RT,
room temperature
2.2 | Radiosyntheses of 18F
arabinofuranoses
groups were deprotected in a C-18 plus short column via
1 N NaOH. Excess 18F ion was removed by passing the
final compound through C-18 plus short column followed
by 2× (AG11-A8-long aluminaN) columns. RadioTLC
(95:5 CH3CN:H2O) confirmed the synthesis of D-5-18F-AF
and L-5-18F-AF (see Figures S1B and S2B for radioTLC).
The Rf value of the final products was consistent with the
Rf value of the cold standard (5L-5).
2.2.1 | D-5-18F-AF and L-5-18F-AF
These radiotracers were synthesized in two steps (see C.2.
in Supporting Information). First, the cyclotron derived
18F in [18O]H2O was passed through a QMA anion
exchange column eluted with 1 mL solvent (500 μL
water + 500 μL acetonitrile) K2CO3 (2 mg) and kryptofix
K222 (12 mg). This mixture was dried under nitrogen gas
and vacuum with multiple additions of anhydrous aceto-
nitrile (4X) (K2CO3-kryptofix K222 mix). The compound
5D-20 or 5L-20 (5 mg) was dissolved in anhydrous
Dimethyl sulfoxide (DMSO) (500 μL) and added to the
above anhydrous K2CO3-kryptofix K222 mix and heated
to 155ꢀC for 10 min. The Rf value of the 18F-labeled com-
pounds by radioTLC (1:1 hexanes:ethyl acetate) were
comparable with the Rf values of 5L-4 and its D-
counterpart standards (see Figures S1A and S2A for
radioTLC), and high-performance liquid chromatography
(HPLC) analysis of the 18F compounds with co-injected
characterized standards confirmed the identity of the
esterified D- and L-5-18F sugar alcohols (see Figures S1C-
E and S2C-E for analytical HPLC). Second, the acetate
2.2.2 | D-2-18F-AF and L-2-18F-AF
These syntheses were performed similarly and are fully
described in section C.2. of the Supporting Information.
Briefly, anhydrous 18F-K2CO3-K222 mixture was refluxed
with compound 2D-20 (or 2L-20) in dry acetonitrile
(500 μL). Both analytical HPLC and RadioTLC (1:1 hex-
ane:ethyl acetate) confirmed the 18F labeling (see Figures
S3A and S4A for radioTLC). In both cases, the 18F-labeled
intermediates co-eluted with the standard D- (or L-) 2-F-
OBz3-α-AF (see Figures S3C-E and S4C-E for analytical
HPLC). The benzoyl esters were subsequently removed
on C-18 plus short column using 1 N NaOH (1 mL) for
3 min. This crude reaction mixture was neutralized with
1 N HCl and finally passed through a sequence of C-

4
KALITA ET AL.
18-AG11-A8-long aluminaN columns. The purity of the
aliquots was determined in RadioTLC (95:5 CH3CN:H2O)
(see Figures S3B and S4B for radioTLC).
2.4 | Statistical analyses
All synthetic data including radiochemical yields and %
radiochemical purities are reported as mean standard
error. Radiochemical yields are reported with and without
decay-correction for 18F (t1/2 = 110 min). in vitro data were
normalized to OD600 for sensitivity analysis to account for
differential growth rates between organisms. All statistical
analysis was performed using Microsoft Excel and Prism
8.2. Four data sets were acquired for all in vitro studies
(N = 4). Data were analyzed using an unpaired two-tailed
Student's t-test. All graphs are depicted with error bars
corresponding to the standard error of the mean.
2.3 | In vitro analyses of 18F-
arabinofuranoses
Screening studies were performed to evaluate 18F
arabinofuranose tracer accumulation by E. coli and
Staphylococcus aureus. E. coli and S. aureus were grown
aerobically in luria broth (LB) for 16 hr with agitation of
111 rpm. The cultures were pelleted at 3400 rpm for
5 minutes and resuspended in an equivalent amount of
Ham's F12 media (Gibco). Following a 1/16 dilution, cul-
tures were then incubated with
1
μCi of 18F
3 | RESULTS AND DISCUSSION
arabinofuranoses (D-2-18F-AF, L-2-18F-AF) at 180 rpm for
120 min. The bacterial suspensions were transferred to
filter tubes (Corning Costar Spin-X) and centrifuged at
8000 rpm for 5 min. Phosphate-buffered saline was added
to each tube, and the cultures were centrifuged at
8000 rpm for 5 minutes. The pellet and supernatant were
separated and counted on a γ counter (Hidex Automatic
Gamma Counter). Four replicates were performed for
each bacterial strain.
A sensitivity study was subsequently performed to
evaluate 18F arabinofuranose accumulation by a panel of
disease-relevant pathogens. The bacterial strains used,
and their growth conditions are listed in Table S1. Bacte-
ria strains (except Mycobacterium marinum) were grown
aerobically in their listed media for 16 hr with agitation
of 111 rpm. M. marinum was grown aerobically for 3 days
with media replenishment every 24 hr. Culture and other
experimental methods were identical to those described
above. Again, four replicates were performed for each
bacterial strain.
3.1 | Radiosyntheses of D-2-18F-AF, L-
2-18F-AF, D-5-18F-AF, L-5-18F-AF, and their
corresponding 19F standards
To investigate bacterial incorporation in vitro, four 18F-
labeled arabinofuranose sugars were elaborated.
Detailed synthetic and radiosynthetic procedures, as
well as compound characterization are described in the
Supporting Information. The basic strategy involved
synthesis of triflated, peracetylated precursors for each
molecule, to mimic the most common synthetic
method used for [18F]FDG. For D-5-18F-AF and L-5-18F-
AF, the increased nucleophilicity of the hydroxyl at
the 5-position was used to generate orthogonally
protected precursors (Figure 2A). A previous synthesis
of D-2-18F-AF was adapted for this purpose, while an
analogous method was used for its enantiomer L-2-18F-
AF (Figure 2B). Diethylaminosulfur trifluoride (DAST)
chemistry was then used to generate all 19F standards
FIGURE 2 Synthesis of 18F-labeled
arabinofuranose-derived structures. A,
D-5-18F-AF and L-5-18F-AF
radiosyntheses B, D-2-18F-AF and L-
2-18F-AF radiosyntheses

KALITA ET AL.
5
which were characterized fully using 1H, 13C, 19F
NMR, and high-resolution mass spectrometry. Subse-
quent radiosynthesis using nucleophilic 18F was used
to synthesize the four sugars, which were isolated via
cartridge purification and characterized via radio-TLC
(Figures S1A,B; S2A, B; S3A,B; and S4A,B). The sum-
mary of characterization data is as follows: D-2-18F-AF
3.3 | The sensitivities of D-2-18F-AF and L-
2-18F-AF were evaluated in vitro using a
broad panel of important human
pathogens
Based on screening data, D-2-18F-AF and L-2-18F-AF were
studied in a panel of pathogenic bacteria. These bacteria
were chosen as representative gram-negative and gram-
positive organisms implicated in a variety of dangerous
human infections. For example, gram-negatives
K. pneumoniae and Pseudomonas aeruginosa are involved
in hospital-acquired disease while gram-positive Staphy-
lococcus epidermidis is a cause of skin and catheter-
related infections.31, 32 A reproducible, high-throughput
screening assay was developed for simultaneous evalua-
tion of 12 living microorganisms using a single 18F radio-
tracer synthesis. Both D-2-18F-AF and L-2-18F-AF were
incorporated into several gram-negative and gram-
positive bacteria as shown in Figure 5A,B. For D-2-18F-AF
the highest radioactivity was retained for Proteus
mirabilis, Salmonella typhimurium, Enterococcus faecalis
and S. epidermidis (approximately 7-8 Bq/10 million
cells). The uptake levels of D-2-18F-AF in these four bacte-
ria were similar (<3-fold difference based on
(N = 12), decay-corrected yield (13.2
synthesis (EOS) radiochemical yield (7.5
1.6)%, end of
1.2)%, and
radiochemical purity (98.0
(N = 10, decay-corrected yield (18.5
radiochemical yield (10.5 1.4)%, and radiochemical
purity (98.0
2.0)%; D-5-18F-AF (N = 3, decay-
corrected yield (2.0
2.0)%; L-2-18F-AF
2.6)%, EOS
0.2)%, EOS radiochemical yield
0.1)%, and radiochemical purity (93.0 1.8)%;
(1.0
and L-5-18F-AF (N
=
3, decay-corrected yield
0.1)%,
(2.2
0.2)%, EOS radiochemical yield (1.1
1.3)%.
and radiochemical purity (94.8
3.2 | Screening of 18F arabinofuranoses
revealed accumulation of D-2-18F-AF and L-
2-18F-AF in E. coli
Two of the most important bacterial pathogens are
E. coli and S. aureus, which cause several clinically rel-
evant infections. Specifically, gram-negative E. coli is a
frequent cause of urinary tract, intestinal, and biliary
28
infections.27,
S. aureus is a frequent gram-positive
organism cultured in skin, musculoskeletal, and blood-
30
borne infections.29,
Therefore we initially evaluated
18F arabinofuranoses in these organisms prior to a
broader survey of pathogens (Figure 3). Initial studies
showed low accumulation of all radiotracers into
S. aureus (mean uptake <2.0 Bq/10 million cells). Fur-
thermore, there was lower accumulation of L-5-18F-AF
and D-5-18F-AF in both E. coli and S. aureus (mean
uptake <2.0 Bq/10 million cells). Higher incorporation
was observed for L-2-18F-AF and D-2-18F-AF in E. coli;
similar uptake was found for both radiotracers in
E. coli, significantly higher than all probe combinations
(P < 0.05 in all cases). The accumulation of L-2-18F-AF
in E. coli was 6-fold higher (P < 0.0001) than that in
S. aureus, and 2.5-fold higher than that of its
corresponding 5-labeled counterpart (P = 0.0001). For
D-2-18F-AF, accumulation in E. coli was 4.5-fold higher
than that in S. aureus, and 2.4-fold higher than that of
its corresponding 5-labeled counterpart (P = 0.0072).
The specificity of this process for L-2-18F-AF was fur-
ther validated in blocking studies (Figure 4A,B). In
summary, these studies indicated higher accumulation
in gram-negative organisms for D-2-18F-AF and L-2-18F-
AF and motivated further sensitivity studies.
FIGURE 3 Screening of arabinofuranose-derived 18F PET
radiotracers in Escherichia coli and Staphylococcus aureus. in vitro
uptake of L-2-18F-AF in live E. coli was approximately 6-fold higher
than that in S. aureus (P < 0.0001). D-2-18F-AF accumulates in live
E. coli approximately 4-fold more than in live S. aureus
(P = 0.0002). In vitro uptake of D-5-18F-AF and L-5-18F-AF showed
<1.5 Bq/10 million bacterial cells incorporation for live E. coli and
S. aureus in all cases. There was no statistically significant
difference in accumulation, for any combination of D- and L-5-18F-
AF radiotracer and organism

6
KALITA ET AL.
FIGURE 4 Blocking studies for L-2-18F-AF. A, L-2-18F-AF accumulates in live Escherichia coli 2.5× more than in heat-killed
(P < 0.0001) and 1.75× more than in a blocking study using 10 mM unlabeled L-arabinose (P = 0.0006). This probe is retained in live E. coli
approximately 4 times more than in live S. aureus. B, L-2-18F-arabinofuranose uptake in E. coli in competition with unlabeled L-
arabinofuranose at various concentrations. Cellular uptake of the radiotracer decreases as the concentration of L-arabinofuranose increases
in the solution
incorporated radioactivity/10 million cells) to data
recently obtained for D-[methyl-11C]methionine.33 Evalu-
ation of L-2-18F-AF showed the highest uptake in
million cells. Of note, D-2-18F-AF and L-2-18F-AF were
incorporated into both gram-negative and gram-positive
species. Also significantly, there was little or no signal
retained for many important pathogens for example
S. aureus, P. aeruginosa, and L. monocytogenes.
K.
pneumoniae,
Acinetobacter baumannii,
and
M. marinum that retained between 3.5 and 4 Bq/10
FIGURE 5 Sensitivity study in a panel of bacterial pathogens. A, D-2-18F-AF showed the highest retention in Proteus mirabilis,
Salmonella typhimurium and Enterococcus faecalis. B, L-2-18F-AF showed the highest retention in several gram-positive and gram-negative
bacterial strains with Klebsiella pneumoniae, Acinetobacter baumannii and Mycobacterium marinum accumulating about 4 Bq/10 million
cells

KALITA ET AL.
7
4 | CONCLUSIONS
processes, and (b) if the tracer detects important catego-
ries of organisms that are relevant to antimicrobial
therapy (eg, [18F]FDS that is sensitive to most
Enterobacteriaceae). The radiotracers investigated in this
manuscript D-2-18F-AF and L-2-18F-AF showed promis-
ing sensitivity to several microorganisms, with no
apparent preference for gram-negative vs gram-positive
bacteria. Furthermore, there were several important
pathogens that showed low accumulation of these
radiotracers most notably S. aureus. Future studies will
investigate whether the species-specific incorporation of
D-2-18F-AF and L-2-18F-AF can be used to noninvasively
identify strains and represent an advantage in compel-
ling clinical scenarios. Importantly, these radiotracers
may also have significant application to other organ-
Molecular imaging methods targeting bacteria-specific
metabolism have outstanding clinical potential. This
potential lies in distinguishing active infection from other
disease entities that appear similar on morphologic imag-
ing (CT and MRI), allowing appropriate antimicrobial
management. With the advent of multi-modality scan-
ning, PET is particularly well suited to complement the
structural information typically obtained in the work-up
of infected patients. In developing PET tracers for infec-
tion, both specificity and sensitivity are important. A PET
tracer specifically targeting a microbial pathway is the
simplest approach; however, another strategy might be
based on relative avidity of the tracer for bacterial metab-
olism. For this reason, we considered both D- and L-
arabinofuranose-derived PET tracers.
isms and diseases; arabinofuranoses are metabolized
37
avidly by fungi,36,
and as highlighted by previous
Both D- and L-arabinofuranose-derived 18F radio-
tracers were obtained in high radiochemical purity.
Interestingly, our screening assay showed little or no
bacterial incorporation of 18F-arabinofuranoses with
substitution at the 5-position. Although mechanistic
analysis is beyond the scope of this manuscript, this
finding suggests the 5-OH is necessary for retention in
bacteria, perhaps based on interaction with arabinose-
binding proteins. In contrast the 2-substituted mole-
cules D-2-18F-AF and L-2-18F-AF demonstrated incorpo-
ration into live E. coli, which was not seen in heat-
killed bacteria or in the presence of the nonlabeled
parent sugars. The mechanisms of retention for these
two tracers are currently unknown and the basis for
future study. As highlighted by Figure 1, 18F substitu-
tion at the 2-position would render D-2-18F-AF and L-
2-18F-AF poor substrates for arabinose isomerase (E.C.
5.3.1.3 and 5.3.1.4 for D and L forms respectively). In
this case, the bacterial retention of these probes may
be mediated by their relative affinities for L-ABPs.
Additional arabinofuranose-derived 18F substrates that
could potentially undergo biotransformation would be
3-deoxy-3-[18F]fluoro-L-arabinofuranose and 3-deoxy-
3-[18F]fluoro-D-arabinofuranose. We made several
attempts to synthesize these using an identical radio-
chemical strategy, without success. Several factors can
impede a ring SN2 fluorination reaction, including ste-
ric hindrance from neighboring positions, ring confor-
mations that impede nucleophilic attack, and
work, the D-arabinose isoforms in particular may find
use in oncologic imaging.
ACKNOWLEDGEMENTS
Grant sponsors NIH R01EB024014, NIH R01EB025985,
DOD A132172, UCSF Resource Allocation Program. The
authors would also like to thank Prof Sanjay Jain and
Alvaro Ordonez (Johns Hopkins University) for their
assistance with in vitro methods.
CONFLICT OF INTEREST
The authors declare no competing financial interests.
AUTHOR CONTRIBUTIONS
DW, MO, and OR proposed and supervised the overall
project. MK, JL, and MP obtained in vitro data. MK, MS,
JB, and HVB performed the radiochemistry. MK, DW,
MO, OR, RF, and ME wrote and edited the paper.
ORCID
Henry F. VanBrocklin https://orcid.org/0000-0003-
2849-0841
David M. Wilson https://orcid.org/0000-0002-1095-
046X
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How to cite this article: Kalita M, Parker MFL,
Luu JM, et al. Arabinofuranose-derived positron-
emission tomography radiotracers for detection of
pathogenic microorganisms. J Label Compd
Radiopharm. 2020;1–9. https://doi.org/10.1002/jlcr.
3835