A non-nucleotide-based linking method for the preparation of psoralen-derivatized methylphosphonate oligonucleotides.

Abstract

A method is reported for conjugating an analog of 4'-(aminomethyl)-4,5',8- trimethylpsoralen to methylphophonate oligonucleotides. This method enables the psoralen moiety to be coupled to the phosphonate backbone between any two desired bases in a sequence. When hybridized to a target mRNA, the psoralen moiety can be directed toward a uridine base and, in turn, can undergo a photo-addition reaction with the target under UV irradiation at 365 nm. Several different non-nucleotide-based amino-linker reagents have been prepared for incorporation into methylphosphonate oligonucleotides by standard phosphonamidite chemistry. In addition, an N-hydroxysuccinimide activated ester analog of 4'-[(3-carboxypropionamido)methyl]-4,5',8- trimethylpsoralen has been synthesized for conjugation to the amino-linker moieties. Using this approach, we have prepared a number of psoralen-methylphosphonate-oligonucleotide conjugates which are complementary to the chimeric bcr/abl mRNA associated with chronic myelogenous leukemia. Solution hybridization studies with a 440-base subfragment of the bcr/abl RNA have shown that the psoralen moiety does not adversely affect duplex stability. Polyacrylamide gel electrophoresis analyses have demonstrated that the psoralen-oligonucleotide conjugates undergo photo-addition to the RNA in a sequence-specific manner. Optimal photo-addition occurs when the psoralen moiety is inserted adjacent to one or more adenine residues in the oligonucleotide sequence, particularly between adenine and thymine (5'-3'). This internal labeling approach greatly increases the number of potential target sites available for photo-cross-linking experiments.

• Authors: M A Reynolds, T A Beck, R I Hogrefe, A McCaffrey, L J Arnold Jr, M M Vaghefi
• Journal: Bioconjugate chemistry (Bioconjug Chem) 1992 Sep-Oct Vol. 3 Issue 5 Pg. 366-74 ISSN: 1043-1802 [Print] United States
• PMID: 1420436 (Publication Type: Journal Article)

Chemical References

• Cross-Linking Reagents
• Furocoumarins
• Oligonucleotides
• Organophosphorus Compounds
• RNA, Messenger
• methylphosphonic acid
• aminomethyltrioxsalen
• Trioxsalen

Topics

• Base Sequence
• Chimera (genetics)
• Cross-Linking Reagents (chemical synthesis)
• Furocoumarins (chemical synthesis)
• Leukemia, Myeloid (genetics)
• Molecular Sequence Data
• Oligonucleotides (chemical synthesis)
• Organophosphorus Compounds (chemical synthesis)
• Photochemistry
• RNA, Messenger (chemistry, genetics)
• Structure-Activity Relationship
• Trioxsalen (analogs & derivatives, chemical synthesis)