ABSTRACT
A high effective specific activity (HESA) formulation of a biotin-containing (99m)Tc ligand [RP488: dimethyl-Gly-Ser-Cys(Acm)-Lys(Biotin)-Gly] conveniently prepared from solid phase was compared to a typical low effective specific activity (LESA) solution formulation to demonstrate improved targeting to streptavidin in an in vitro assay and in an in vivo rat model. RP488 was coupled to a maleimide-functionalized polyethylene glycol resin via a thiol ether linkage and labeled with (99m)Tc-gluconate at room temperature, followed by elution of the HESA (99m)Tc-RP488 in saline (minimum specific activity approximately 1000 TBq/mmol by amino acid analysis). Both HESA and LESA (99m)Tc-RP488 labeled at > 90% purity. In vitro, HESA (99m)Tc-RP488 incubated with streptavidin-agarose was bound quantitatively, but there was competition from addition of increasing amounts of cold RP488. In rats, radiotracer uptake was evident at the site of implantation of streptavidin-agarose beads for the HESA dose, less uptake of low effective specific activity (LESA) material, and no appreciable uptake in the control rats of the LESA or HESA dose. The target-to-background ratio for HESA (99m)Tc-RP488 was 5.4 times that of the control. The solid-phase technology offers a convenient way to prepare high specific activity receptor-targeting (99m)Tc radiopharmaceuticals.
Subject(s)
Biotin/chemistry , Biotin/chemical synthesis , Organotechnetium Compounds/chemical synthesis , Radiopharmaceuticals/chemical synthesis , Streptavidin/chemistry , Technetium/chemistry , Animals , Biotin/analogs & derivatives , Biotin/metabolism , Chelating Agents , Chromatography, High Pressure Liquid , Indicators and Reagents , Isotope Labeling , Organotechnetium Compounds/metabolism , Radiopharmaceuticals/metabolism , Rats , Rats, Sprague-Dawley , Technetium/metabolism , Tissue DistributionABSTRACT
A solid-phase technetium chelation chemistry was developed as a means of preparing (99m)Tc radiopharmaceuticals at high effective specific activity (HSA). Three peptidic N(3)S (99m)Tc ligands [mercaptoacetyl-Gly-Gly-Gly (MAG3), picolinyl-Ser-Cys-Gly-Thr-Lys-Pro-Pro-Arg (RP063), and dimethyl-Gly-Ser-Cys-Gly-Thr-Lys-Pro-Pro-Arg (RP128)] were used. The free thiol of Cys in each was attached to a series of commercially available amine-functionalized supports in a two-step process. The amine groups on the solid supports were converted to maleimide groups followed by the attachment of the (99m)Tc chelators through a thiol ether linkage with Cys. The optimized loading of the supports ranged 6-122 micromol/g support as determined by amino acid analysis. Each of the peptide-loaded supports (50-100 mg) was placed in either glass syringe vessels or disposable chromatography columns. Labeling with [(99m)Tc]pertechnetate (200-800 MBq) in the presence of stannous gluconate was achieved at room temperature for 30-60 min or in a 100 degrees C water bath for 10 min. Up to 80% of the activity was eluted from the column with saline to give products with purity up to 99.8% as determined by HPLC. Amino acid analysis indicated as little as 100 pmol of peptide present in the (99m)Tc products, demonstrating that extremely high effective specific activity can be achieved without the need for purification.
Subject(s)
Chelating Agents/chemistry , Isotope Labeling/methods , Oligopeptides/chemistry , Organotechnetium Compounds/chemical synthesis , Radiopharmaceuticals/chemical synthesis , Amino Acid Sequence , Chromatography, High Pressure Liquid , Ligands , Organotechnetium Compounds/isolation & purification , Radiopharmaceuticals/isolation & purification , Reagent Kits, Diagnostic , Sodium Pertechnetate Tc 99m/chemistryABSTRACT
Two 99Tcm-labelled analogues of the chemotactic peptide ForMLF were evaluated as potential agents for imaging inflammation and infection, in the hope that they would be simple to use and would give diagnostically useful images shortly after injection. The peptides differed in the chelation site for 99Tcm and the presence of a hydrophilic spacer. The sequences of RP050 and RP056 were ForNleLFNleYK(G)G-C(Acm)-GPic and ForNleLFNleYKK(DG)GC(Acm)SPic respectively, where Pic is picolinic acid. In in vitro tests of binding to the ForMLF receptor on polymorphonuclear neutrophils and potency for release of myeloperoxidase, RP056 was similar in potency to ForMLF, whereas RP050 was 10 times more potent. When administered in 5-nmol doses to rats, RP050 produced less extensive neutropenia than ForMLF, whereas RP056 produced very little neutropenia. Following labelling by ligand exchange from tartrate or glucoheptonate at 100 degrees C and purification using a C-18 solid-phase extraction cartridge, 4-MBq doses were administered to rats bearing infectious (Escherichia coli) or sterile (zymosan) inflammation sites in the thigh. The inflammation-to-normal muscle ratios at 30 min after injection were 3.9 +/- 0.4 for RP050 and 4.7 +/- 0.3 for RP056 (mean +/- S.E.M., n = 4), and the ratios were maintained for up to 3 h. These peptides are promising agents for imaging inflammation and infection.
