Yttrium-86 Labelling of Oligonucleotide Derivatives

AIM:

With the application of radioactive-labelled aptamers aiming at tumour-specific target structures, a new approach for tumour imaging should be made to increase resolution, sensitivity and specificity in tumour detection. Spiegelmers are synthetic molecules with custom-made properties. They are high-affinity L-enantiomeric oligonucleotide ligands that display high resistance to enzymatic degradation compared with D-oligonucleotides. DOTA is a commonly used chelator for the trivalent radio metals ⁸⁶Y, to form highly stable in vitro and in vivo complexes. In this report, the DOTA-functionalization of a 12-mer L-RNA [Sequenz: 5’-Aminohexyl UGA CUG ACU GAC-3’, MW 3975] (synthesized with a primary amine via an alkyl linker) and a first approach in yttrium-86 labelling is described.
MATERIAL & METHODS: p-SCN-bz-DOTA was used as bifunctional chelator (BFC) to form a stable thiourea-bond with the primary amine of the modified L-RNA. The coupling reaction was carried out in a 0.6 M sodium bicarbonate buffer, pH 8.4 with a 26-fold excess of BFC. The reaction control takes place by IP-RP-HPLC. The preparation achieved a total profit of 97 % of the spiegelmer-derivative DOTA-bz-spiegelmer. The DOTA-bz-spiegelmer was separated from the non-transformed BFC by size exclusion chromatography (SEC). The complexation took place in a 0.5 M ammonium acetate buffer, pH 6.4-7.0 by incubating at 90 °C for 60 min. The transchelation was carried out in 0.5 M ammonium acetate buffer, pH 6.4-7.0 by incubating at 25 °C for 60 min in the presence of 0.1 µmol DTPA. Results: The total labelling efficiency of the DOTA-bz-spiegelmer with 86Y was up to 58%. The non-detachable bound to the DOTA-L-RNA ⁸⁶Y activity was greater than 28%. Experiments with native L-RNA showed, that up to 30% of the RNA bound ⁸⁶Y activity could transchelated to DTPA.
CONCLUSION: DOTA can be coupled to 5’-hexylamine functionalised L-RNA over a thiourea bond and labelled with the trivalent PET-nuclide ⁸⁶Y. The polyanionic nature of the oligonucleotides should be taken into account for non-specific binding of trivalent metal cations. Sponsored by FP6 integrated project BioCare contract no.: 505785