Quantitative Analysis of GalNAc-siRNA Conjugate Using Liquid Chromatograph Coupled with Triple Quadrupole Mass Spectrometry

Small interfering RNA (siRNA) is double-stranded RNA and can suppress gene expression by targeting and degrading messenger RNA (mRNA). siRNAs are too hydrophilic to cross the cell membrane without a transfection agent. siRNA conjugated to a trimer of N-acetylgalactosamine(GalNAc) can avidly bind to the asialoglycoproteinreceptor (ASGPR) which is predominantly expressed on liver hepatocytes. The recent research about GalNAc-conjugated siRNA confirmed that the liver pharmacokinetic (PK) are better predictor of pharmacodynamics than plasma PK. However, the quantitation of GalNAc-conjugated siRNA in liver is challenge due to the matrix effect and endogenous interference. siRNA-01 is a GalNAc-conjugated siRNA that targets messenger RNA (mRNA) in the liver. Here we present a sensitive and robust method for quantitative analysis of siRNA-01 in rat liver homogenate with a wide calibration range 10-5000 ng/mL.