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PET/CT Based In Vivo Evaluation of 64Cu Labelled Nanodiscs in Tumor Bearing Mice.

Huda P, Binderup T, Pedersen MC, Midtgaard SR, Elema DR, Kjær A, Jensen M, Arleth L - PLoS ONE (2015)

Bottom Line: This makes them promising candidates for both drug delivery purposes and as advanced imaging agents.For the radiolabelling, a simple approach for 64Cu radiolabelling of proteins via a chelating agent, DOTA, was developed.The reaction was performed at sufficiently mild conditions to be compatible with labelling of the protein part of a lipid-protein particle while fully conserving the particle structure including the amphipathic protein fold.

View Article: PubMed Central - PubMed

Affiliation: Structural Biophysics, Niels Bohr Institute, University of Copenhagen, Universitetsparken 5, DK-2100 København Ø, Denmark.

ABSTRACT
64Cu radiolabelled nanodiscs based on the 11 α-helix MSP1E3D1 protein and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine lipids were, for the first time, followed in vivo by positron emission tomography for evaluating the biodistribution of nanodiscs. A cancer tumor bearing mouse model was used for the investigations, and it was found that the approximately 13 nm nanodiscs, due to their size, permeate deeply into cancer tissue. This makes them promising candidates for both drug delivery purposes and as advanced imaging agents. For the radiolabelling, a simple approach for 64Cu radiolabelling of proteins via a chelating agent, DOTA, was developed. The reaction was performed at sufficiently mild conditions to be compatible with labelling of the protein part of a lipid-protein particle while fully conserving the particle structure including the amphipathic protein fold.

No MeSH data available.


Related in: MedlinePlus

Schematics of nanodisc radiolabelling.A: chemical reaction of DOTA-NHS ester-conjugation to MSP lysines and further radiolabelling with 64Cu2+. B: symbol explanation for C. C 1: MD simulation of ordinary nanodisc[6] (lipids in blue, MSP in red). 2: nanodisc assembled with DOTA conjugated MSP. 3: 64Cu radiolabelled nanodisc.
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pone.0129310.g001: Schematics of nanodisc radiolabelling.A: chemical reaction of DOTA-NHS ester-conjugation to MSP lysines and further radiolabelling with 64Cu2+. B: symbol explanation for C. C 1: MD simulation of ordinary nanodisc[6] (lipids in blue, MSP in red). 2: nanodisc assembled with DOTA conjugated MSP. 3: 64Cu radiolabelled nanodisc.

Mentions: To reach this goal, a simple method for 64Cu radiolabelling was developed allowing for conjugating the protein component of the nanodisc with the chelating agent 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA). This enabled binding of the radioactive isotope 64Cu (Fig 1) while at the same time involving minimal modifications of the biomolecule in order to avoid significant structural changes. An alternative DOTA-based approach has recently been proposed for the labeling of the 12 amino acid QFP peptide [15]. For the present work, we have developed a milder reaction approach that is compatible with proteins in solution and allow for labeling of the MSP, while conserving their amphipathic fold and ability to form nanodiscs. Therefore, the method is applicable to other protein systems. 64Cu was chosen as the radionuclide because of its relatively long half-life (12.7 hours) and relatively low maximum positron energy (0.66 MeV), which makes it very suitable for PET imaging. Nanodiscs were assembled with DOTA-conjugated MSPs and structurally characterized by combined small-angle x-ray scattering (SAXS) and small angle neutron scattering (SANS) confirming that the nanodisc structure indeed remained fully intact after DOTA-conjugation. The DOTA-conjugated nanodiscs were radioactively labeled with 64Cu and their in vitro stability evaluated in mouse blood plasma, showing that the 64Cu-labeled nanodiscs were sufficiently stable over time to be relevant for in vivo studies. Finally, 64Cu-labelled nanodiscs were intravenously administered to 10 tumor bearing nude mice and the biodistribution analyzed by PET and gamma counting of tissues of interest. This showed that the nanodiscs accumulate intensively in the tumor tissue as well as in the kidneys.


PET/CT Based In Vivo Evaluation of 64Cu Labelled Nanodiscs in Tumor Bearing Mice.

Huda P, Binderup T, Pedersen MC, Midtgaard SR, Elema DR, Kjær A, Jensen M, Arleth L - PLoS ONE (2015)

Schematics of nanodisc radiolabelling.A: chemical reaction of DOTA-NHS ester-conjugation to MSP lysines and further radiolabelling with 64Cu2+. B: symbol explanation for C. C 1: MD simulation of ordinary nanodisc[6] (lipids in blue, MSP in red). 2: nanodisc assembled with DOTA conjugated MSP. 3: 64Cu radiolabelled nanodisc.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4488450&req=5

pone.0129310.g001: Schematics of nanodisc radiolabelling.A: chemical reaction of DOTA-NHS ester-conjugation to MSP lysines and further radiolabelling with 64Cu2+. B: symbol explanation for C. C 1: MD simulation of ordinary nanodisc[6] (lipids in blue, MSP in red). 2: nanodisc assembled with DOTA conjugated MSP. 3: 64Cu radiolabelled nanodisc.
Mentions: To reach this goal, a simple method for 64Cu radiolabelling was developed allowing for conjugating the protein component of the nanodisc with the chelating agent 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA). This enabled binding of the radioactive isotope 64Cu (Fig 1) while at the same time involving minimal modifications of the biomolecule in order to avoid significant structural changes. An alternative DOTA-based approach has recently been proposed for the labeling of the 12 amino acid QFP peptide [15]. For the present work, we have developed a milder reaction approach that is compatible with proteins in solution and allow for labeling of the MSP, while conserving their amphipathic fold and ability to form nanodiscs. Therefore, the method is applicable to other protein systems. 64Cu was chosen as the radionuclide because of its relatively long half-life (12.7 hours) and relatively low maximum positron energy (0.66 MeV), which makes it very suitable for PET imaging. Nanodiscs were assembled with DOTA-conjugated MSPs and structurally characterized by combined small-angle x-ray scattering (SAXS) and small angle neutron scattering (SANS) confirming that the nanodisc structure indeed remained fully intact after DOTA-conjugation. The DOTA-conjugated nanodiscs were radioactively labeled with 64Cu and their in vitro stability evaluated in mouse blood plasma, showing that the 64Cu-labeled nanodiscs were sufficiently stable over time to be relevant for in vivo studies. Finally, 64Cu-labelled nanodiscs were intravenously administered to 10 tumor bearing nude mice and the biodistribution analyzed by PET and gamma counting of tissues of interest. This showed that the nanodiscs accumulate intensively in the tumor tissue as well as in the kidneys.

Bottom Line: This makes them promising candidates for both drug delivery purposes and as advanced imaging agents.For the radiolabelling, a simple approach for 64Cu radiolabelling of proteins via a chelating agent, DOTA, was developed.The reaction was performed at sufficiently mild conditions to be compatible with labelling of the protein part of a lipid-protein particle while fully conserving the particle structure including the amphipathic protein fold.

View Article: PubMed Central - PubMed

Affiliation: Structural Biophysics, Niels Bohr Institute, University of Copenhagen, Universitetsparken 5, DK-2100 København Ø, Denmark.

ABSTRACT
64Cu radiolabelled nanodiscs based on the 11 α-helix MSP1E3D1 protein and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine lipids were, for the first time, followed in vivo by positron emission tomography for evaluating the biodistribution of nanodiscs. A cancer tumor bearing mouse model was used for the investigations, and it was found that the approximately 13 nm nanodiscs, due to their size, permeate deeply into cancer tissue. This makes them promising candidates for both drug delivery purposes and as advanced imaging agents. For the radiolabelling, a simple approach for 64Cu radiolabelling of proteins via a chelating agent, DOTA, was developed. The reaction was performed at sufficiently mild conditions to be compatible with labelling of the protein part of a lipid-protein particle while fully conserving the particle structure including the amphipathic protein fold.

No MeSH data available.


Related in: MedlinePlus