68GA-NOTA-UBI AND 68GA-DOTA-UBI AS RADIOPHARMACEUTICALS FOR THE DIAGNOSIS OF INFECTIOUS PROCESSES: PRECLINICAL STUDIES AND TRANSLATION TO CLINICAL APPLICATION

Abstract

Introduction/Justification

Infectious diseases are the second leading cause of mortality worldwide. In this context, considerable efforts are being made to develop radiopharmaceuticals that enable the accurate diagnosis of bacterial infections. A new field of research is focusing on antimicrobial peptides, such as Ubiquicidin. The 29-41 fragment (Thr-Gly-Arg-Ala-Lys-Arg-Arg-Met-Gln-Tyr-Asn-Arg-Arg) UBI(29-41) labeled with radionuclides has proved to be an important tool for the specific diagnosis of infectious processes.

Objectives

To present the radiochemical and "in vitro" studies of UBI(29-41) with the chelating agents 1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA) and 1,4,7,10- tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) for labeling with gallium-68 (68Ga) and to validate with a clinical application.

Materials and Methods

After 68GaCl3 elution, the NOTA-UBI and DOTA-UBI reactions were performed at 85oC for 5 min and 95oC for 15 min, respectively. In both cases, the purification was carried out using a Sep-Pak C18 filter and radiochemical control by Thin-Layer Chromatography (TLC) and High-Performance Liquid Chromatography (HPLC). The partition coefficient (Log P) and serum protein binding were determined, as well as the stability in saline and serum up to 90 min. After these studies, assays were carried out to determine the binding of the radiopharmaceuticals to "Staphylococcus aureus" bacteria. For the clinical study, the patient selected had a diagnosis of chronic osteomyelitis in the distal tibia, a positive blood culture for "Staphylococcus aureus" and a possible infectious/inflammatory process at the site identified by magnetic resonance imaging. The patient was injected intravenously with 281 MBq of 68Ga-DOTA-UBI to confirm the presence of an infectious process by PET-CT and the images were obtained 1 h post-administration. Resultados: The radiochemical purity (RP) of the radiopharmaceuticals after purification was 99.28±0.28% and 99.78±0.06% for 68Ga-NOTA-UBI and 68Ga-DOTA-UBI, respectively. Log P was -3.57±0.20 for 68Ga-NOTA-UBI and -3.63±0.17 for 68Ga-DOTA-UBI. Stability studies up to 90 min showed RP of 98.13±0.78% and 99.75±0.08% in saline; and 79.94±5.10% and 94.69±1.14% in serum, for 68Ga-NOTA-UBI and 68Ga-DOTA-UBI, respectively. The percentage of serum protein binding, evaluated at 30 and 60 min, was 59.90±1.21% and 53.45±2.16% for 68Ga-NOTA-UBI and 60.22±2.96% and 44.06±1.88% for 68Ga-DOTA-UBI, respectively. The binding of radiopharmaceuticals to "Staphylococcus aureus" revealed a direct relation to the amount of bacteria in the culture. Clinical images showed intense uptake of the radiopharmaceutical in the entire remnant of the talus and in the adjacent soft tissues of the left ankle. After scan, the secretion collected from the surgical site was cultured and the presence of "Staphylococcus aureus" was confirmed.

Conclusion

The radiochemical and "in vitro" assays showed that the radiopharmaceuticals studied presented similar characteristics with the potential to be implemented in clinical practice. The clinical study showed that the UBI(29-41) fragment radiolabeled with 68Ga can be used as a potential biomarker for infectious processes, according to the availability of the chelating agent.