A nonradioisotope biomedical assay for intact oligonucleotide and its chain-shortened metabolites used for determination of exposure and elimination half-life of antisense drugs in tissue.
Rigorous extraction methods coupled with capillary gel electrophoresis (CGE) provide a basis for a nonradiolabel assay for quantitation of intact antisense drug and its numerous chain-shortened metabolites. As part of the validation of the CGE method, we compared the quantitation of unlabeled ISIS 3521 (ISI 641A) and its chain-shortened metabolites with total radioactivity of [(35)S]-ISIS 3521. ISIS 3521 was labeled on the fifth nucleotide linkage from the 5'-end with (35)S by well-established methods. Multiple tissues collected from rats after administration of [(35)S]-ISIS 3521 were assayed by both radiolabel (liquid scintillation spectroscopy) and CGE methods. The CGE method provided accurate quantitation of the drug and its metabolites in kidney cortex and liver tissues. The correlation between methods for multiple tissues over time was excellent with 88.5% of the measurements being statistically equivalent. These data suggest that CGE is an accurate means of quantitating oligonucleotide in tissue and that it compares favorably with traditional radiochemical techniques. Clearance half-lives for total measurable oligonucleotides were equivalent to clearance of total radioactivity in both liver and kidney with the longest clearance half-life associated with the kidney.
Geary
R S
RS
Isis Pharmaceuticals, 2292 Faraday Avenue, Carlsbad, California, 92008, USA.
Matson
J
J
Levin
A A
AA
D000818:Animals; D019075:Electrophoresis, Capillary / Q000379:methods; D006207:Half-Life; D007672:Kidney Cortex / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D020319:Oligodeoxyribonucleotides, Antisense / Q000493:pharmacokinetics; D009841:Oligonucleotides / Q000032:analysis*; D016376:Oligonucleotides, Antisense / Q000493:pharmacokinetics*; D051381:Rats; D013873:Thionucleotides / Q000493:pharmacokinetics
D020319:Oligodeoxyribonucleotides, Antisense; D009841:Oligonucleotides; D016376:Oligonucleotides, Antisense; D013873:Thionucleotides; C108494:ISIS 3521