Although a rapid and accurate diagnosis is crucial to the management of patients suspected of bacterial infection, the currently available radiopharmaceuticals are not capable of distinguishing between sterile inflammation and bacterial infections. Our goal is to develop an infection-specific PET/SPECT radiopharmaceutical for eventual use in clinical practice. In Phase I, we will evaluate two independent approaches. In a covalent approach, we will conjugate a radionuclide chelator for PET/SPECT imaging with a Zn-DPA targeting moiety that is known to selectively target the negatively charged bacterial envelope, to provide a novel small molecule nuclear imaging agent. In an alternative, non-covalent radiolabeling approach, we will use streptavidin (SA) as a linker between the biotinylated Zn-DPA targeting motif and a biotinylated chelator to form an imaging agent which may have improved bacterial lesion accumulation over the covalent approach due to: (i) its slower pharmacokinetics because of increased size, and (ii) its potential to bind up to three DPA groups for affinity enhancement.
Our Specific Aims i nclude: 1) Synthesize and characterize DOTA- DPA-(1 Zn) for the covalent conjugation approach, DOTA/SA/DPA-(1 Zn) for the non-covalent approach and radiolabel the DOTA containing agents with the PET isotope 68Ga, as well as the SPECT isotope 111In. 2) Serum stability assays and in vitro evaluation of the covalent [68Ga /111In-DOTA-DPA-(1 Zn)] and non- covalent [68Ga/111In-DOTA/SA/DPA-(1 Zn)] agents to S. pyogenes. Specific binding to bacteria will be evaluated by measuring binding to bacteria with increasing concentrations of unlabeled DOTA-DPA-Zn. Thereafter, labeled bacteria will be evaluated in 37o C serum environments to determine the stability of both radionuclide within the chelate and the stability of both agents to the bacteria. 3) Evaluate the covalent and non-covalent approaches in infection and inflammation mouse models for evidence of specific accumulations. SKH1 hairless mice will be injected in the thigh with live S. pyogenes to provide the bacterial infection model or lipopolysaccharide to provide the inflammation model. We will evaluate the agents radiolabeled with 68Ga as well as 111In in the mouse models using small animal PET and SPECT/CT cameras respectively. In all cases, the location and extent of infection will be monitored by co-injecting PSVue(R) 794 (a fluorescent bacteria targeting probe) and imaging on a small animal optical camera. Agents will be evaluated for their pharmacokinetics, their accumulation in the target, their target thigh/contralateral normal thigh accumulation, evidence of specific infection imaging and sensitivity of detection. At sacrifice, full biodistributions of each radiolabel will be done to supplement the imaging results. Key benchmarks for Phase I will be to obtain using either 68Ga/ 111In-DOTA- DPA-Zn or 68Ga/111In-DOTA/SA/DPA-Zn an infected thigh/normal thigh ratio of greater than 5 within a 10 h (68Ga) or 24 h (111In) period, obtain a statistically higher accumulation in the infected thighs compared to the inflammation thighs, and obtain an estimate of the lower limits of detection in the infection model.
Bacterial infection is one of the major causes of morbidity and mortality not only in developing countries but globally. Early diagnosis of infection and an ability to distinguish between bacterial infection and sterile inflammation is critical to the effective management of these patients. However, despite the efforts of many international imaging groups, there is currently no validated bacterial imaging agent that can distinguish infection from sterile inflammation. Obviously, the development of such an agent would greatly advance our ability to detect, localize, and quantify infections, to prescribe the appropriate treatment and to follow the patient throughout the treatment. In this project we propose to evaluate two novel approaches aimed at developing a new radiopharmaceutical which would allow noninvasive imaging of bacterial infections with the sensitivity that nuclear imaging approaches promise and would also allow infectious and inflammatory abscesses to be distinguished.