Radiotracers based on the peptide A20FMDV2 selectively target the cell surface receptor integrin α<inf>v</inf>β<inf>6</inf>. This integrin has been identified as a prognostic indicator correlating with the severity of disease for several challenging malignancies. In previous studies of A20FMDV2 peptides labeled with 4-<sup>18</sup>F-fluorobenzoic acid (<sup>18</sup>F-FBA), we have shown that the introduction of poly(ethylene glycol) (PEG) improves pharmacokinetics, including increased uptake in α<inf>v</inf>β<inf>6</inf>-expressing tumors. The present study evaluated the effect of site-specific C-terminal or dual (N- and C-terminal) PEGylation, yielding <sup>18</sup>F-FBA-A20FMDV2-PEG<inf>28</inf> (4) and <sup>18</sup>F-FBA-PEG<inf>28</inf>-A20FMDV2-PEG<inf>28</inf> (5), on α<inf>v</inf>β<inf>6</inf>-targeted tumor uptake and pharmacokinetics. The results are compared with <sup>18</sup>F-FBA-labeled A20FMDV2 radiotracers (1-3) bearing either no PEG or different PEG units at the N terminus. Methods: The radiotracers were prepared and radiolabeled on solid phase. Using 3 cell lines, DX3puroβ6 (α<inf>v</inf>β<inf>6</inf>+), DX3puro (α<inf>v</inf>β<inf>6</inf>-), and BxPC-3 (α<inf>v</inf>β<inf>6</inf>+), we evaluated the radiotracers in vitro (serum stability; cell binding and internalization) and in vivo in mouse models bearing paired DX3puroβ6-DX3puro and, for 5, BxPC-3 xenografts. Results: The size and location of the PEG units significantly affected α<inf>v</inf>β<inf>6</inf>targeting and pharmacokinetics. Although the C-terminally PEGylated 4 showed some improvements over the un-PEGylated <sup>18</sup>F-FBA-A20FMDV2 (1), it was the bi-terminally PEGylated 5 that displayed the more favorable combination of high α<inf>v</inf>β<inf>6</inf>affinity, selectivity, and pharmacokinetic profile. In vitro, 5 bound to α<inf>v</inf>β<inf>6</inf>-expressing DX3puroβ6 and BxPC-3 cells with 60.5% ± 3.3% and 48.8% ± 8.3%, respectively, with a significant fraction of internalization (37.2% ± 4.0% and 37.6% ± 4.1% of total radioactivity, respectively). By comparison, in the DX3puro control 5 showed only 3.0% ± 0.5% binding and 0.9% ± 0.2% internalization. In vivo, 5 maintained high, α<inf>v</inf>β<inf>6</inf>-directed binding in the paired DX3puroβ6-DX3puro model (1 h: DX3puroβ6, 2.3 ± 0.2 percentage injected dose per gram [%ID/g]; DX3puroβ6/DX3puro ratio, 6.5:1; 4 h: 10.7:1). In the pancreatic BxPC-3 model, uptake was 4.7 ± 0.9%ID/g (1 h) despite small tumor sizes (20-80 mg). Conclusion: The bi-PEGylated radiotracer 5 showed a greatly improved pharmacokinetic profile, beyond what was predicted from individual N- or C-terminal PEGylation. It appears that the 2 PEG units acted synergistically to result in an improved metabolic profile including high α<inf>v</inf>β<inf>6</inf>+ tumor uptake and retention.
- Integrin α<inf>v</inf>β<inf>6</inf>
- Positron emission tomography
ASJC Scopus subject areas
- Radiology Nuclear Medicine and imaging