68Ga-labeling and in vivo evaluation of a uPAR binding DOTA- and NODAGA-conjugated peptide for PET imaging of invasive cancers

TY - JOUR

T1 - 68Ga-labeling and in vivo evaluation of a uPAR binding DOTA- and NODAGA-conjugated peptide for PET imaging of invasive cancers

AU - Persson, Morten

AU - Madsen, Jacob

AU - Østergaard, Søren

AU - Ploug, Michael

AU - Kjaer, Andreas

N1 - Copyright © 2012 Elsevier Inc. All rights reserved.

PY - 2012/5

Y1 - 2012/5

N2 - Introduction: The urokinase-type plasminogen activator receptor (uPAR) is a well-established biomarker for tumor aggressiveness and metastatic potential. DOTA-AE105 and DOTA-AE105-NH 2 labeled with 64Cu have previously been demonstrated to be able to noninvasively monitor uPAR expression using positron emission tomography (PET) in human cancer xenograft mice models. Here we introduce 68Ga-DOTA-AE105-NH 2 and 68Ga-NODAGA-AE105-NH 2 and evaluate their imaging properties using small-animal PET. Methods: Synthesis of DOTA-AE105-NH 2 and NODAGA-AE105-NH 2 was based on solid-phase peptide synthesis protocols using the Fmoc strategy. 68GaCl 3 was eluted from a 68Ge/ 68Ga generator. The eluate was either concentrated on a cation-exchange column or fractionated and used directly for labeling. For in vitro characterization of both tracers, partition coefficient, buffer and plasma stability, uPAR binding affinity and cell uptake were determined. To characterize the in vivo properties, dynamic microPET imaging was carried out in nude mice bearing human glioma U87MG tumor xenograft. Results: In vitro experiments revealed uPAR binding affinities in the lower nM range for both conjugated peptides and identical to AE105. Labeling of DOTA-AE105-NH 2 and NODAGA-AE105-NH 2 with 68Ga was done at 95°C and room temperature, respectively. The highest radiochemical yield and purity were obtained using fractionated elution, whereas a negative effect of acetone on labeling efficiency for NODAGA-AE105-NH 2 was observed. Good stability in phosphate-buffered saline and mouse plasma was observed. High cell uptake was found for both tracers in U87MG tumor cells. Dynamic microPET imaging demonstrated good tumor-to-background ratio for both tracers. Tumor uptake was 2.1% ID/g and 1.3% ID/g 30 min postinjection and 2.0% ID/g and 1.1% ID/g 60 min postinjection for 68Ga-NODAGA-AE105-NH 2 and 68Ga-DOTA-AE105-NH 2, respectively. A significantly higher tumor-to-muscle ratio (P<.05) was found for 68Ga-NODAGA-AE105-NH 2 60 min postinjection. Conclusions: The use of 68Ga-DOTA-AE105-NH 2 and 68Ga-NODAGA-AE105-NH 2 as the first gallium-68 labeled uPAR radiotracers for noninvasive PET imaging is reported, which combine versatility with good imaging properties. These new tracers thus constitute an interesting alternative to the 64Cu-labeled version ( 64Cu-DOTA-AE105 and 64Cu-DOTA-AE105-NH 2) for detecting uPAR expression in tumor tissue. In our hands, the fractionated elution approach was superior for labeling of peptides, and 68Ga-NODAGA-AE105-NH 2 is the favored tracer as it provides the highest tumor-to-background ratio.

AB - Introduction: The urokinase-type plasminogen activator receptor (uPAR) is a well-established biomarker for tumor aggressiveness and metastatic potential. DOTA-AE105 and DOTA-AE105-NH 2 labeled with 64Cu have previously been demonstrated to be able to noninvasively monitor uPAR expression using positron emission tomography (PET) in human cancer xenograft mice models. Here we introduce 68Ga-DOTA-AE105-NH 2 and 68Ga-NODAGA-AE105-NH 2 and evaluate their imaging properties using small-animal PET. Methods: Synthesis of DOTA-AE105-NH 2 and NODAGA-AE105-NH 2 was based on solid-phase peptide synthesis protocols using the Fmoc strategy. 68GaCl 3 was eluted from a 68Ge/ 68Ga generator. The eluate was either concentrated on a cation-exchange column or fractionated and used directly for labeling. For in vitro characterization of both tracers, partition coefficient, buffer and plasma stability, uPAR binding affinity and cell uptake were determined. To characterize the in vivo properties, dynamic microPET imaging was carried out in nude mice bearing human glioma U87MG tumor xenograft. Results: In vitro experiments revealed uPAR binding affinities in the lower nM range for both conjugated peptides and identical to AE105. Labeling of DOTA-AE105-NH 2 and NODAGA-AE105-NH 2 with 68Ga was done at 95°C and room temperature, respectively. The highest radiochemical yield and purity were obtained using fractionated elution, whereas a negative effect of acetone on labeling efficiency for NODAGA-AE105-NH 2 was observed. Good stability in phosphate-buffered saline and mouse plasma was observed. High cell uptake was found for both tracers in U87MG tumor cells. Dynamic microPET imaging demonstrated good tumor-to-background ratio for both tracers. Tumor uptake was 2.1% ID/g and 1.3% ID/g 30 min postinjection and 2.0% ID/g and 1.1% ID/g 60 min postinjection for 68Ga-NODAGA-AE105-NH 2 and 68Ga-DOTA-AE105-NH 2, respectively. A significantly higher tumor-to-muscle ratio (P<.05) was found for 68Ga-NODAGA-AE105-NH 2 60 min postinjection. Conclusions: The use of 68Ga-DOTA-AE105-NH 2 and 68Ga-NODAGA-AE105-NH 2 as the first gallium-68 labeled uPAR radiotracers for noninvasive PET imaging is reported, which combine versatility with good imaging properties. These new tracers thus constitute an interesting alternative to the 64Cu-labeled version ( 64Cu-DOTA-AE105 and 64Cu-DOTA-AE105-NH 2) for detecting uPAR expression in tumor tissue. In our hands, the fractionated elution approach was superior for labeling of peptides, and 68Ga-NODAGA-AE105-NH 2 is the favored tracer as it provides the highest tumor-to-background ratio.

KW - Acetates

KW - Amides

KW - Animals

KW - Cell Line, Tumor

KW - Cell Transformation, Neoplastic

KW - Chelating Agents

KW - Drug Stability

KW - Female

KW - Gallium

KW - Gallium Radioisotopes

KW - Glioma

KW - Heterocyclic Compounds, 1-Ring

KW - Humans

KW - Mice

KW - Neoplasm Invasiveness

KW - Peptides

KW - Positron-Emission Tomography

KW - Protein Binding

KW - Receptors, Urokinase Plasminogen Activator

U2 - 10.1016/j.nucmedbio.2011.10.011

DO - 10.1016/j.nucmedbio.2011.10.011

M3 - Journal article

C2 - 22172391

SN - 0969-8051

VL - 39

SP - 560

EP - 569

JO - Nuclear Medicine and Biology

JF - Nuclear Medicine and Biology

IS - 4

ER -