Characterization and evaluation of (64)Cu-labeled A20FMDV2 conjugates for imaging the integrin αvβ 6.
OBJECTIVE
The integrin αvβ6 is overexpressed in a variety of aggressive cancers and serves as a prognosis marker. This study describes the conjugation, radiolabeling, and in vitro and in vivo evaluation of four chelators to determine the best candidate for (64)Cu radiolabeling of A20FMDV2, an αvβ6 targeting peptide.


METHODS
Four chelators were conjugated onto PEG28-A20FMDV2 (1): 11-carboxymethyl-1,4,8,11-tetraazabicyclo[6.6.2]hexadecane-4-methanephosphonic acid (CB-TE1A1P), 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), 1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA), and 4,4'-((3,6,10,13,16,19-hexazazbicyclo[6.6.6]ico-sane-1,8-diylbis(aza-nediyl))bis(methylene)dibenzoic acid (BaBaSar). All peptides were radiolabeled with (64)Cu in ammonium acetate buffer at pH 6 and formulated to pH 7.2 in PBS for use. The radiotracers were evaluated using in vitro cell binding and internalization assays and serum stability assays. In vivo studies conducted include blocking, biodistribution, and small animal PET imaging. Autoradiography and histology were also conducted.


RESULTS
All radiotracers were radiolabeled in good radiochemical purity (>95 %) under mild conditions (37-50 °C for 15 min) with high specific activity (0.58-0.60 Ci/μmol). All radiotracers demonstrated αvβ6-directed cell binding (>46 %) with similar internalization levels (>23 %). The radiotracers (64)Cu-CB-TE1A1P-1 and (64)Cu-BaBaSar-1 showed improved specificity for the αvβ6 positive tumor in vivo over (64)Cu-DOTA-1 and (64)Cu-NOTA-1 (+/- tumor uptake ratios-3.82 +/- 0.44, 3.82 ± 0.41, 2.58 ± 0.58, and 1.29 ± 0.14, respectively). Of the four radiotracers, (64)Cu-NOTA-1 exhibited the highest liver uptake (10.83 ± 0.1 % ID/g at 4 h).


CONCLUSIONS
We have successfully conjugated, radiolabeled, and assessed the four chelates CB-TE1A1P, DOTA, NOTA, and BaBaSar both in vitro and in vivo. However, the data suggests no clear "best candidate" for the (64)Cu-radiolabeling of A20FMDV2, but instead a trade-off between the different properties (e.g., stability, selectivity, pharmacokinetics, etc.) with no obvious effects of the individual chelators.
Department of Biomedical Engineering, University of California Davis, Davis, CA, USA.
Lina Y
LY
Hu
Nadine
N
Bauer
Leah M
LM
Knight
Zibo
Z
Li
Shuanglong
S
Liu
Carolyn J
CJ
Anderson
Peter S
PS
Conti
Julie L
JL
Sutcliffe
D000818:Animals; D000951:Antigens, Neoplasm / Q000378:metabolism*; D001345:Autoradiography; D056831:Coordination Complexes / Q000138:chemical synthesis / Q000737:chemistry; D003301:Copper Radioisotopes / Q000652:urine; D005260:Female; D007091:Image Processing, Computer-Assisted; D016023:Integrins / Q000378:metabolism*; D008819:Mice, Nude; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry; D049268:Positron-Emission Tomography; D011849:Radioactive Tracers; D044967:Serum / Q000000981:diagnostic imaging; D014018:Tissue Distribution; D014057:Tomography, X-Ray Computed
D000951:Antigens, Neoplasm; D056831:Coordination Complexes; D003301:Copper Radioisotopes; D016023:Integrins; D010455:Peptides; D011849:Radioactive Tracers; C073878:integrin alphavbeta6