PURPOSE: With the increase of especially hospital-acquired infections, timely and accurate diagnosis of bacterial infections is crucial for effective patient care. Molecular imaging has the potential for specific and sensitive detection of infections. Siderophores are iron-specific chelators recognized by specific bacterial transporters, representing one of few fundamental differences between bacterial and mammalian cells. Replacing iron by gallium-68 without loss of bioactivity is possible allowing molecular imaging by positron emission tomography (PET). Here, we report on the preclinical evaluation of the clinically used siderophore, desferrioxamine-B (Desferal®, DFO-B), radiolabelled with 68Ga for imaging of bacterial infections. METHODS: In vitro characterization of [68Ga]Ga-DFO-B included partition coefficient, protein binding and stability determination. Specific uptake of [68Ga]Ga-DFO-B was tested in vitro in different microbial cultures. In vivo biodistribution was studied in healthy mice and dosimetric estimation for human setting performed. PET/CT imaging was carried out in animal infection models, representing the most common pathogens. RESULTS: DFO-B was labelled with 68Ga with high radiochemical purity and displayed hydrophilic properties, low protein binding and high stability in human serum and PBS. The high in vitro uptake of [68Ga]Ga-DFO-B in selected strains of Pseudomonas aeruginosa, Staphylococcus aureus and Streptococcus agalactiae could be blocked with an excess of iron-DFO-B. [68Ga]Ga-DFO-B showed rapid renal excretion and minimal retention in blood and other organs in healthy mice. Estimated human absorbed dose was 0.02 mSv/MBq. PET/CT images of animal infection models displayed high and specific accumulation of [68Ga]Ga-DFO-B in both P. aeruginosa and S. aureus infections with excellent image contrast. No uptake was found in sterile inflammation, heat-inactivated P. aeruginosa or S. aureus and Escherichia coli lacking DFO-B transporters. CONCLUSION: DFO-B can be easily radiolabelled with 68Ga and displayed suitable in vitro characteristics and excellent pharmacokinetics in mice. The high and specific uptake of [68Ga]Ga-DFO-B by P. aeruginosa and S. aureus was confirmed both in vitro and in vivo, proving the potential of [68Ga]Ga-DFO-B for specific imaging of bacterial infections. As DFO-B is used in clinic for many years and the estimated radiation dose is lower than for other 68Ga-labelled radiopharmaceuticals, we believe that [68Ga]Ga-DFO-B has a great potential for clinical translation.

Department of Analytical Chemistry Faculty of Science Palacky University Olomouc Czech Republic

Department of Microbiology Faculty of Medicine and Dentistry Palacky University and University Hospital Olomouc Czech Republic

Department of Nuclear Medicine Medical University Innsbruck Anichstrasse 5 A 6020 Innsbruck Austria

Institute of Microbiology of the Czech Academy of Sciences v v i Prague Czech Republic

Institute of Molecular and Translational Medicine Faculty of Medicine and Dentistry Palacky University CZ 77900 Olomouc Czech Republic

See more in PubMed

Jain SK. The promise of molecular imaging in the study and treatment of infectious diseases. Mol Imaging Biol. 2017;19:341–347. PubMed PMC

Roberts RR, Scott RD, Hota B, Kampe LM, Abbasi F, Schabowski S, et al. Costs attributable to healthcare-acquired infection in hospitalized adults and a comparison of economic methods. Med Care. 2010;48:1026–1035. PubMed

Zimlichman E, Henderson D, Tamir O, Franz C, Song P, Yamin CK, et al. Health care-associated infections: a meta-analysis of costs and financial impact on the US health care system. JAMA Intern Med. 2013;173:2039–2046. PubMed

Haque M, Sartelli M, McKimm J, Abu BM. Health care-associated infections - an overview. Infect Drug Resist. 2018;11:2321–2333. PubMed PMC

Li B, Webster TJ. Bacteria antibiotic resistance: new challenges and opportunities for implant-associated orthopedic infections. J Orthop Res. 2018;36:22–32. PubMed PMC

Santajit S, Indrawattana N. Mechanisms of antimicrobial resistance in ESKAPE pathogens. Biomed Res Int. 2016;2475067. PubMed PMC

Renner LD, Zan J, Hu LI, Martinez M, Resto PJ, Siegel AC, et al. Detection of ESKAPE bacterial pathogens at the point of care using isothermal DNA-based assays in a portable degas-actuated microfluidic diagnostic assay platform. Appl Environ Microbiol. 2017;83:e02449–e02416. PubMed PMC

Mulani MS, Kamble EE, Kumkar SN, Tawre MS, Pardesi KR. Emerging strategies to combat ESKAPE pathogens in the era of antimicrobial resistance: a review. Front Microbiol. 2019;10:539. PubMed PMC

Lambregts MMC, Bernards AT, van der Beek MT, Visser LG, de Boer MG. Time to positivity of blood cultures supports early re-evaluation of empiric broad-spectrum antimicrobial therapy. PLoS One. 2019;14:e0208819. PubMed PMC

Ordonez AA, Sellmyer MA, Gowrishankar G, Ruiz-Bedoya CA, Tucker EW, Palestro CJ, et al. Molecular imaging of bacterial infections: Overcoming the barriers to clinical translation. Sci Transl Med. 2019;(11):eaax8251. PubMed PMC

Signore A, Glaudemans AW. The molecular imaging approach to image infections and inflammation by nuclear medicine techniques. Ann Nucl Med. 2011;25:681–700. PubMed

Bunschoten A, Welling MM, Termaat MF, Sathekge M, van Leeuwen FW. Development and prospects of dedicated tracers for the molecular imaging of bacterial infections. Bioconjug Chem. 2013;24:1971–1989. PubMed

Sasser TA, Van Avermaete AE, White A, Chapman S, Johnson JR, Van Avermaete T, et al. Bacterial infection probes and imaging strategies in clinical nuclear medicine and preclinical molecular imaging. Curr Top Med Chem. 2013;13:479–487. PubMed

Lawal I, Zeevaart J, Ebenhan T, Ankrah A, Vorster M, Kruger HG, et al. Metabolic imaging of infection. J Nucl Med. 2017;58:1727–1732. PubMed

Sollini M, Lauri C, Boni R, Lazzeri E, Erba PA, Signore A. Current status of molecular imaging in infections. Curr Pharm Des. 2018;24:754–771. PubMed

Mota F, Ordonez AA, Firth G, Ruiz-Bedoya CA, Ma MT, Jain SK. Radiotracer development for bacterial imaging. J Med Chem. 2020;63:1964–1977. PubMed PMC

Petrik M, Zhai C, Haas H, Decristoforo C. Siderophores for molecular imaging applications. Clin Transl Imaging. 2017;5:15–27. PubMed PMC

Drechsel H, Jung G. Peptide siderophores. J Pept Sci. 1998;4:147–181. PubMed

Hider RC, Kong X. Chemistry and biology of siderophores. Nat Prod Rep. 2010;27:637–657. PubMed

Page MGP. The role of iron and siderophores in infection, and the development of siderophore antibiotics. Clin Infect Dis. 2019;69:S529–S537. PubMed PMC

Cassat JE, Skaar EP. Iron in infection and immunity. Cell Host Microbe. 2013;13:509–519. PubMed PMC

Skaar EP. The battle for iron between bacterial pathogens and their vertebrate hosts. PLoS Pathog. 2010;6:e1000949. PubMed PMC

Fadeev EA, Luo M, Groves JT. Synthesis, structure, and molecular dynamics of gallium complexes of schizokinen and the amphiphilic siderophore acinetoferrin. J Am Chem Soc. 2004;126:12065–12075. PubMed

Fani M, André JP, Maecke HR. 68Ga-PET: a powerful generator-based alternative to cyclotron-based PET radiopharmaceuticals. Contrast Media Mol Imaging. 2008;3:67–77. PubMed

Velikyan I. Continued rapid growth in (68) Ga applications: update 2013 to June 2014. J Labelled Comp Radiopharm. 2015;58:99–121. PubMed

Petrik M, Umlaufova E, Raclavsky V, Palyzova A, Havlicek V, Haas H, et al. Imaging of Pseudomonas aeruginosa infection with Ga-68 labelled pyoverdine for positron emission tomography. Sci Rep. 2018;8:15698. PubMed PMC

Maina T, Konijnenberg MW, KolencPeitl P, Garnuszek P, Nock BA, Kaloudi A, et al. Preclinical pharmacokinetics, biodistribution, radiation dosimetry and toxicity studies required for regulatory approval of a phase I clinical trial with (111)in-CP04 in medullary thyroid carcinoma patients. Eur J Pharm Sci. 2016;91:236–242. PubMed PMC

Stabin MG, Sparks RB, Crowe E. OLINDA/EXM: the second-generation personal computer software for internal dose assessment in nuclear medicine. J Nucl Med. 2005;46:1023–1027. PubMed

Gordon O, Ruiz-Bedoya CA, Ordonez AA, Tucker EW. Jain SK. Molecular imaging: a novel tool to visualize pathogenesis of infections in situ mBio. 2019;10:e00317–e00319. PubMed PMC

Chakravarty R, Goel S, Dash A, Cai W. Radiolabeled inorganic nanoparticles for positron emission tomography imaging of cancer: an overview. Q J Nucl Med Mol Imaging. 2017;61:181–204. PubMed PMC

Man F, Gawne PJ, de Rosales RTM. Nuclear imaging of liposomal drug delivery systems: a critical review of radiolabelling methods and applications in nanomedicine. Adv Drug Deliv Rev. 2019;143:134–160. PubMed PMC

Cook BE, Membreno R, Zeglis BM. A dendrimer scaffold for the amplification of in vivo pretargeting ligations. Bioconjug Chem. 2018;29:2734–2740. PubMed PMC

Rosebrough SF. Plasma stability and pharmacokinetics of radiolabeled deferoxamine-biotin derivatives. J Pharmacol Exp Ther. 1993;265:408–415. PubMed

Mathias CJ, Lewis MR, Reichert DE, Laforest R, Sharp TL, Lewis JS, et al. Preparation of 66Ga- and 68Ga-labeled Ga(III)-deferoxamine-folate as potential folate-receptor-targeted PET radiopharmaceuticals. Nucl Med Biol. 2003;30:725–731. PubMed

Yokoyama A, Ohmomo Y, Horiuchi K, Saji H, Tanaka H, Yamamoto K, et al. Deferoxamine, a promising bifunctional chelating agent for labeling proteins with gallium: Ga-67 DF-HSA: concise communication. J Nucl Med. 1982;23:909–914. PubMed

Rangger C, Haubner R. Radiolabelled peptides for positron emission tomography and endoradiotherapy in oncology. Pharmaceuticals. 2020;13:22. PubMed PMC

Vosjan MJ, Perk LR, Visser GW, Budde M, Jurek P, Kiefer GE, et al. Conjugation and radiolabeling of monoclonal antibodies with zirconium-89 for PET imaging using the bifunctional chelate p-isothiocyanatobenzyl-desferrioxamine. Nat Protoc. 2010;5:739–743. PubMed

Heskamp S, Raave R, Boerman O, Rijpkema M, Goncalves V, Denat F. 89Zr-Immuno-positron emission tomography in oncology: state-of-the-art 89Zr radiochemistry. Bioconjug Chem. 2017;28:2211–2223. PubMed PMC

Moerlein SM, Welch MJ, Raymond KN, Weitl FL. Tricatecholamide analogs of enterobactin as gallium- and indium-binding radiopharmaceuticals. J Nucl Med. 1981;22:710–719. PubMed

Hoffer PB, Samuel A, Bushberg JT, Thakur M. Desferoxamine mesylate (Desferal): a contrast-enhancing agent for gallium-67 imaging. Radiology. 1979;131:775–779. PubMed

Chandra R, Pierno C, Braunstein P. 111In Desferal: a new radiopharmaceutical for abscess detection. Radiology. 1978;128:697–699. PubMed

Ioppolo JA, Caldwell D, Beiraghi O, Llano L, Blacker M, Valliant JF, et al. 67Ga-labeled deferoxamine derivatives for imaging bacterial infection: preparation and screening of functionalized siderophore complexes. Nucl Med Biol. 2017;52:32–41. PubMed

Petrik M, Haas H, Schrettl M, Helbok A, Blatzer M, Decristoforo C. In vitro and in vivo evaluation of selected 68Ga-siderophores for infection imaging. Nucl Med Biol. 2012;39:361–369. PubMed PMC

Petrik M, Zhai C, Novy Z, Urbanek L, Haas H, Decristoforo C. In vitro and in vivo comparison of selected Ga-68 and Zr-89 labelled siderophores. Mol Imaging Biol. 2016;18:344–352. PubMed PMC

Wandersman C, Delepelaire P. Bacterial iron sources: from siderophores to hemophores. Annu Rev Microbiol. 2004;58:11–47. PubMed

Marvig RL, Damkiaer S, Khademi SMH, Markussen TM, Molin S, Jelsbak L. Within-host evolution of reveals adaptation toward iron acquisition from hemoglobin. mBio. 2014;5:e00966–e00914. PubMed PMC

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