Automated synthesis of an 18F-labelled pyridine-based alkylating agent for high yield oligonucleotide conjugation

Abstract

Alkylating agents have been shown to be very promising for the radiolabelling of oligonucleotides with fluorine-18. In this report we describe the fully automated synthesis of 2-bromo-N-[3-(2-[18F]fluoropyridin-3-yloxy)propyl]acetamide ([18F]FPyBrA) utilizing a modular synthesis unit. Reaction conditions for the coupling of this pyridine-based alkylating agent at the 5′ end of a fully phosphorothioated random 20-mer DNA sequence were optimized to achieve very high radiochemical yields (>90%) and a maximum specific activity of 5–6 GBq/μmoL. The potential for rapid purification by solid phase extraction without need of chromatographic isolation of the radiolabelled oligonucleotide presents an overall benefit for the application of oligonucleotides in preclinical studies and potential clinical applications.

18F‐Labeling of Sensitive Biomolecules for Positron Emission Tomography

“…For example, N -(4-fluorobenzyl)-2-bromoacetamide ( [18 F]FBBA, 69 ), [63] and 2-bromo- N -[3-(2- [18 F]fluoropyridin-3yloxy)propyl]acetamide ( [18 F]FPyBrA, 70 ) [64] were used in thiol-alkylation reactions of α -bromo ketone derivatives (Figure 12A). In addition, fluorine-18 labeled maleimide analogues have been developed for reaction with thiols.…”

“…Although [ 18 F]SFB could also be employed in a cysteine (cys) thiol acylation to afford fluorine‐18 labeled peptides through conjugation with the 4‐fluorobenzoyl moiety, alternative strategies have been developed with better selectivity to thiol over amino‐targeting prosthetic groups. For example, N‐( 4‐fluorobenzyl)‐2‐bromoacetamide ([ 18 F]FBBA, 69 ), and 2‐bromo‐ N ‐[3‐(2‐[ 18 F]fluoropyridin‐3yloxy)propyl]acetamide ([ 18 F]FPyBrA, 70 ) were used in thiol‐alkylation reactions of α‐bromo ketone derivatives (Figure A). In addition, fluorine‐18 labeled maleimide analogues have been developed for reaction with thiols .…”

In Vitro Characterization of Two Novel Biodegradable Vectors for the Delivery of Radiolabeled Antisense Oligonucleotides

“…For the preparation of the 18 F-labeled prosthetic group, 2-bromo- N -[3-(2-[ 18 F]-fluoropyridin-3-yloxy)propyl]acetamide ([ 18 F]FPyBrA), the N -Boc-protected nitro-precursor, [3-(2-nitropyridin-3-yloxy)propyl] carbamic acid tert -butyl ester was synthesized according to Kuhnast et al15 The synthesis was adapted for the fully automated preparation in a commercially available modular synthesis system (Modular Lab, Eckert & Ziegler, Berlin, Germany) 16. The preparation of [ 18 F]FPyBrA including online normal phase high performance liquid chromatography (HPLC) purification was performed within 65 min, achieving 10-20% decay corrected yield with 18 F starting activities of 3-5 GBq.…”

18F-labelling innovations and their potential for clinical application

“…As an example, a plug-and-play radiosynthesis platform accompanied by computer software based on modular subunits can easily and flexibly be configured to implement a diverse range of radiosynthesis protocols [194]. Using such systems, the standard procedures ( 18 F-labelling, hydrolysis and purification) have been implemented for many clinically relevant 18 F-tracers [103,105,191,[195][196][197] or 18 Flabelled building blocks for labelling of macromolecules [198][199][200] and build-up syntheses [145].…”