title: The Effect of Bi-Terminal PEGylation of an Integrin αvβ₆-Targeted ¹⁸F Peptide on Pharmacokinetics and Tumor Uptake.
abstract: Radiotracers based on the peptide A20FMDV2 selectively target the cell surface receptor integrin αvβ6. 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-(18)F-fluorobenzoic acid ((18)F-FBA), we have shown that the introduction of poly(ethylene glycol) (PEG) improves pharmacokinetics, including increased uptake in αvβ6-expressing tumors. The present study evaluated the effect of site-specific C-terminal or dual (N- and C-terminal) PEGylation, yielding (18)F-FBA-A20FMDV2-PEG28 (4) and (18)F-FBA-PEG28-A20FMDV2-PEG28 (5), on αvβ6-targeted tumor uptake and pharmacokinetics. The results are compared with (18)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 (αvβ6+), DX3puro (αvβ6-), and BxPC-3 (αvβ6+), 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 αvβ6 targeting and pharmacokinetics. Although the C-terminally PEGylated 4 showed some improvements over the un-PEGylated (18)F-FBA-A20FMDV2 (1), it was the bi-terminally PEGylated 5 that displayed the more favorable combination of high αvβ6 affinity, selectivity, and pharmacokinetic profile. In vitro, 5 bound to αvβ6-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, αvβ6-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).
  
  
  CONCLUSIONS
  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 αvβ6+ tumor uptake and retention.
authors: Division of Hematology/Oncology, Department of Internal Medicine, University of California Davis, Sacramento, California Department of Biomedical Engineering, University of California Davis, Davis, California.
authors: Sven H
authors: SH
authors: Hausner
authors: Department of Biomedical Engineering, University of California Davis, Davis, California.
authors: Nadine
authors: N
authors: Bauer
authors: Department of Biomedical Engineering, University of California Davis, Davis, California.
authors: Lina Y
authors: LY
authors: Hu
authors: Division of Hematology/Oncology, Department of Internal Medicine, University of California Davis, Sacramento, California.
authors: Leah M
authors: LM
authors: Knight
authors: Division of Hematology/Oncology, Department of Internal Medicine, University of California Davis, Sacramento, California jlsutcliffe@ucdavis.edu.
authors: Julie L
authors: JL
authors: Sutcliffe
mesh_terms: D000818:Animals; D000951:Antigens, Neoplasm / Q000378:metabolism*; D019817:Benzoic Acid / Q000737:chemistry; D001692:Biological Transport; D045744:Cell Line, Tumor; D004355:Drug Stability; D005260:Female; D005462:Fluorine Radioisotopes; D006801:Humans; D016023:Integrins / Q000378:metabolism*; D007553:Isotope Labeling; D051379:Mice; D009369:Neoplasms / Q000000981:diagnostic imaging / Q000378:metabolism* / Q000473:pathology; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D011092:Polyethylene Glycols / Q000737:chemistry*; D049268:Positron-Emission Tomography; D011854:Radiochemistry
chemical_list: D000951:Antigens, Neoplasm; D005462:Fluorine Radioisotopes; D016023:Integrins; D010455:Peptides; C073878:integrin alphavbeta6; D011092:Polyethylene Glycols; D019817:Benzoic Acid
keywords: PEGylation; integrin αvβ6; metabolism; peptide; positron emission tomography