Quantitative Determination of A Sirna (Ad00370) in Rat Plasma Using Peptide Nucleic Acid Probe and HPLC with Fluorescence Detection

AIM: Toxicokinetic and pharmacokinetic studies of therapeutic oligonucleotides require validated bioanalytical methods for sensitive and specific quantification of oligonucleotide drug candidates in biological samples. RESULTS: A peptide nucleic acid (PNA) hybridization-based HPLC-fluorescence assay was developed and validated for quantification of Arrowhead Pharmaceuticals' proprietary siRNA in rat plasma samples via hybridization and anion-exchange-HPLC (AEX-HPLC) with fluorescence detection. CONCLUSION: The validated method provided a sensitive and selective approach for quantification of siRNA in biological samples at a linear quantitation range of 1-1000 ng/ml. The assay requires only 25 μl of plasma sample and shows excellent accuracy and precision even without using an internal standard, providing a useful quantification method for siRNA determination in biological matrix with limited sample volume.