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 anti sense drug and its numerous chain-shortened metabolites. As part of the val idation of the CGE method, we compared the quantitation of unlabeled ISIS 3 521 (ISI 641A) and its chain-shortened metabolites with total radioactivity of [S-35]-ISIS 3521, ISIS 3521 was labeled on the fifth nucleotide linkage from the 5'-end with S-35 by well-established methods. Multiple tissues co llected from rats after administration of [35S]-ISIS 3521 were assayed by b oth radiolabel (liquid scintillation spectroscopy) and CGE methods. The CGE method provided accurate quantitation of the drug and its metabolites in k idney cortex and liver tissues. The correlation between methods for multipl e tissues over time was excellent with 88.5% of the measurements being stat istically equivalent. These data suggest that CGE is an accurate means of q uantitating oligonucleotide in tissue and that it compares favorably with t raditional radiochemical techniques, Clearance half-lives for total measura ble oligonucleotides were equivalent to clearance of total radioactivity in both liver and kidney with the longest clearance half-life associated with the kidney, (C) 1999 Academic Press.