68Ga-Siderophores for PET Imaging of Invasive Pulmonary Aspergillosis: Proof of Principle

The diagnosis of invasive pulmonary aspergillosis (IPA) is difficult and lacks specificity and sensitivity. In the pathophysiology of Aspergillus fumigatus, iron plays an essential role as a nutrient during infection. A. fumigatus uses a specific and highly efficient iron uptake mechanism based on iron-complexing ferric ion Fe(III) siderophores, which are a requirement for A. fumigatus virulence. We aimed to evaluate the potential of siderophores radiolabeled with Ga-68, a positron emitter with complexing properties comparable to those of Fe(III), as a radiopharmaceutical for imaging IPA. Methods: Ga-68 radiolabeling of the A. fumigatus siderophores desferri-triacetylfusarinine C (TAFC) and desferri-ferricrocin (FC) was performed at high specific activity. Stability, protein binding, and log P values were determined. In vitro uptake in A. fumigatus cultures was tested under varying conditions. Biodistribution was studied in healthy noninfected BALB/c mice, and uptake was studied in a model of A. fumigatus infection using immunosuppressed Lewis rats. Results: High-specific-activity Ga-68 labeling could be achieved, and resulting complexes were stable in serum, toward diethylenetriaminepentaacetic acid and Fe(III) challenge. Both siderophores showed hydrophilic properties (Ga-68-TAFC, log P = -2.59; Ga-68-FC, log P = 23.17) with low values of protein binding for (68)GaTAFC (<2%). Uptake of both siderophores was highly dependent on the mycelial iron load and could be blocked with an excess (10 mu M) of siderophore or NaN3, indicating specific, energy- dependent uptake. In noninfected mice, Ga-68-TAFC showed rapid renal excretion and lowblood values (1.6 +/- 0.37 percentage injected dose per gram [% ID/g] at 30 min); in urine only intact Ga-68-TAFC was detected. In contrast, Ga-68-FC revealed high retention in blood (16.1 +/- 1.07 % ID/g at 90 min) and rapid metabolism. In the rat IPA model, lung uptake of Ga-68-TAFC was dependent on the severity of infection, with less than 0.04 % ID/g in control rats (n = 5) and 0.29 +/- 0.11 % ID/g in mildly infected (n = 3) and 0.95 +/- 0.37 % ID/g in severely infected (n 5 4) rats. PET showed focal accumulation in infected lung tissue. Conclusion: Both siderophores bound Ga-68 with high affinity, and Ga-68-TAFC, especially, showed high stability. Ga-68-TAFC displayed highly selective accumulation by A. fumigatus subspecies in vitro and in vivo. The high and specific uptake by A. fumigatus proves the potential of Ga-68-labeled siderophores for the specific detection of A. fumigatus during infection. They hold promise as new PET agents for IPA.