Investigation of 6-[¹⁸F]-Fluoromaltose as a Novel PET Tracer for Imaging Bacterial Infection

<div><p></p><p>Despite advances in the field of nuclear medicine, the imaging of bacterial infections has remained a challenge. The existing reagents suffer from poor sensitivity and specificity. In this study we investigate the potential of a novel PET (positron emission tomography) tracer that overcomes these limitations.</p><p>Methods</p><p>6-[¹⁸F]-fluoromaltose was synthesized. Its behavior <i>in vitro</i> was evaluated in bacterial and mammalian cultures. Detailed pharmacokinetic and biodistribution profiles for the tracer were obtained from a murine model.</p><p>Results</p><p>6-[¹⁸F]-fluoromaltose is taken up by multiple strains of pathogenic bacteria. It is not taken up by mammalian cancer cell lines. 6-[¹⁸F]-fluoromaltose is retained in infected muscles in a murine model of bacterial myositis. It does not accumulate in inflamed tissue.</p><p>Conclusion</p><p>We have shown that 6-[¹⁸F]-fluoromaltose can be used to image bacterial infection <i>in vivo</i> with high specificity. We believe that this class of agents will have a significant impact on the clinical management of patients.</p></div

<i>In vitro</i> characterization of 6-[¹⁸F]-fluoromaltose.

<p>A) Uptake of 6-[¹⁸F]-fluoromaltose in the indicated strains of bacteria for 60 minutes. B) 1 hour uptake of 6-[¹⁸F]-fluoromaltose in the mammalian cell lines, MDA MB231 and HeLa and its uptake in <i>E.coli</i> in the presence of 1 mM maltose. C) Bioluminescence imaging of a macrophage cell line J774 infected with a bioluminescent strain of <i>Listeria monocytogenes.</i> D) 1 hour uptake of 6-[¹⁸F]-fluoromaltose in the bioluminescent strain of <i>Listeria monocytogenes</i> and in macrophage cell line J774 with and without intracellular <i>Listeria</i> infections.</p

Uptake of 6-[¹⁸F]-fluoromaltose in infection versus inflammation.

<p>A) Ex-vivo biodistribution of 6-[¹⁸F]-fluoromaltose in mice bearing E.coli induced myositis, 2 h and 4 h after tail-vein injection of 7.4MBq of tracer. B) Ex-vivo biodistribution of 6-[¹⁸F]-fluoromaltose in mice bearing turpentine oil induced sterile abscess, 2 h after tail-vein injection of 7.4MBq of tracer. C) Representative gram stained muscle sections with a black arrow indicating presence of <i>E.coli</i> in the infected muscle section. D) Representative H&E stained muscle sections showing neutrophil infiltration in inflamed muscle.</p

<i>In vivo</i> characterization of 6-[¹⁸F]-fluoromaltose.

<p>A) 3D color map from a PET/CT scan of a mouse bearing <i>E.coli</i> induced infection on the left thigh (red arrow) 1 hr after tail-vein injection of 7.4MBq of 6-[¹⁸F]-fluoromaltose. B) Region of interest analysis (ROIs) from PET/CT images at the indicated time points (n = 4 for each time point) * indicates statistical significance with p<0.05. C) Time activity curve showing accumulation of 6-[¹⁸F]-fluoromaltose in the infected muscle (n = 3).</p

Specificity of 6-[¹⁸F]-fluoromaltose for viable bacteria.

<p>A) A coronal slice from a PET/CT image of a mouse bearing 10⁸ CFU of viable bioluminescent <i>E.coli</i> on the right thigh (red arrow) and 10⁸ CFU of heat-inactivated <i>E.coli</i> on the left thigh, 1hr after tail-vein injection of 7.4MBq of 6-[¹⁸F]-fluoromaltose B) A transverse slice from the same mouse shown in A), with arrows indicating sites of viable and heat inactivated bacteria. C) Bioluminescent image of the mouse shown in A). D) ROI analysis from PET/CT scan of mice (n = 3). * indicates statistical significance.</p

GLUT 5 mRNA levels by quantitative real time PCR.

<p>RNA from MCF7, MDA MB 468 and MCF10A were extracted, converted to cDNA and quantified by real time PCR (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0026902#pone-0026902-g001" target="_blank">fig 1A</a>) and the products also visualized on a 1% agarose gel (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0026902#pone-0026902-g001" target="_blank">fig 1B</a>).</p

Knocking down GLUT 5 mRNA in MCF7 cells has no effect on fructose uptake.

<p>GLUT 5 specific siRNA and negative control siRNA were transfected into MCF7 cells. 48 hrs after transfection cells were simultaneously interrogated for both mRNA levels (using quantitative real time PCR) (right Y axis) and for fructose uptake (left Y axis) by incubating with C14 labeled fructose. The error bars represent S.E.M of triplicates.</p

Staining intensities from 40 breast cancer samples stained with GLUT 5-antibody.

<p>Staining intensities from 40 breast cancer samples stained with GLUT 5-antibody.</p

Cytochalasin B treatment affects fructose uptake in MDA MB 468 cells.

<p>MDA MB 468 cells were exposed to 100 µM cytochalasin B for 30 mins. Then 1 µCi of C14 fructose or glucose was added to the cells, and the uptake was continued for 10 mins in HBSS. The activity associated with the cells was then counted and normalized to activity in the medium and to total protein mass. * indicates p values <0.05. The error bars represent S.E.M of triplicates.</p

Additional file 7: of [18F]FSPG-PET reveals increased cystine/glutamate antiporter (xc-) activity in a mouse model of multiple sclerosis

xCT protein levels in CNS tissues do not differ between control and naïve mice. SC = spinal cord. Data are represented as mean xCT/Na,K-ATPase signal ± SD (n = 3–5). Significance determined by Mann-Whitney test with NS = not significant. (DOCX 65 kb