Antimicrobial peptide LyeTx I mn∆K labeled with 68Ga is a potential PET radiopharmaceutical for molecular imaging of infections

Abstract

Background

Antimicrobial peptides have been radiolabeled and investigated as molecular diagnostic probes due to their propensity to accumulate in infectious sites rather than aseptic inflammatory lesions. LyeTx I is a cationic peptide from the venom of Lycosa erythrognatha, exhibiting significant antimicrobial activity. LyeTx I mn∆K is a shortened derivative of LyeTx I, with an optimized balance between antimicrobial and hemolytic activities. This study reports the first ⁶⁸Ga-radiolabeling of the DOTA-modified LyeTx I mn∆K and primarily preclinical evaluations of [⁶⁸Ga]Ga-DOTA(K)-LyeTx I mn∆K as a PET radiopharmaceutical for infection imaging.

Methods

DOTA(K)-LyeTx I mn∆K was radiolabeled with freshly eluted ⁶⁸Ga. Radiochemical yield (RCY), radiochemical purity (RCP), and radiochemical stability (in saline and serum) were evaluated using ascending thin-layer chromatography (TLC) and reversed-phase high-performance liquid chromatography (RP-HPLC). The radiopeptide's lipophilicity was assessed by determining the logarithm of the partition coefficient (Log P). Serum protein binding (SBP) and binding to Staphylococcus aureus (S. aureus) cells were determined in vitro. Ex vivo biodistribution studies and PET/CT imaging were conducted in healthy mice (control) and mice with infection and aseptic inflammation to evaluate the potential of [⁶⁸Ga]Ga-DOTA(K)-LyeTx I mn∆K as a specific PET radiopharmaceutical for infections.

Results

[⁶⁸Ga]Ga-DOTA(K)-LyeTx I mn∆K was obtained with a high RCY (>90 %), and after purification through a Sep-Pak C18 cartridge, the RCP exceeded 99 %. Ascending TLC and RP-HPLC showed that the radiopeptide remained stable for up to 3.0 h in saline solution and up to 1.5 h in murine serum. [⁶⁸Ga]Ga-DOTA(K)-LyeTx I mn∆K exhibited hydrophilic characteristics (Log P = −2.4 ± 0.1) and low SPB (ranging from 23.3 ± 0.4 % at 5 min of incubation to 10.5 ± 1.1 % at 60 min of incubation). The binding of [⁶⁸Ga]Ga-DOTA(K)-LyeTx I mn∆K to S. aureus cells was proportional to bacterial concentration, with binding percentages of 8.8 ± 0.5 % (0.5 × 10⁹ CFU^.mL⁻¹), 16.2 ± 1.4 % (1.0 × 10⁹ CFU^.mL⁻¹), and 62.2 ± 0.6 % (5.0 × 10⁹ CFU^.mL⁻¹). Ex vivo biodistribution studies and PET/CT images showed higher radiopeptide uptake at the infection site compared to the aseptic inflammation site; the latter was similar to the control group. Target-to-non-target (T/NT) ratios obtained by ex vivo biodistribution data were approximately 1.0, 1.3, and 3.0 at all investigated time intervals for the control, aseptic inflammation, and infection groups, respectively. Furthermore, T/NT ratios obtained from PET/CT images were 1.1 ± 0.1 for the control group and 1.4 ± 0.1 for the aseptic inflammation group. For the infection group, T/NT ratio was 5.0 ± 0.3, approximately 5 times greater compared to the former groups.

Conclusions

The results suggest the potential of [⁶⁸Ga]Ga-DOTA(K)-LyeTx I mn∆K as a PET radiopharmaceutical for molecular imaging of infections.