Metabolism Studies of Unformulated Internally [³H]-Labeled Short Interfering RNAs in Mice

Abstract

Absorption, distribution, metabolism, and excretion properties of two unformulated model short interfering RNA (siRNAs) were determined using a single internal [ Syndrome antigen B) siRNA, respectively). After an initial rapid decrease, concentrations of total radiolabeled components in dried blood decreased at a much slower rate. A nearly complete mass balance was obtained for the [ 3 H]SSB siRNA, and renal excretion was the main route of elimination (38%). The metabolism of the two model siRNAs was rapid and extensive. Five minutes after administration, no parent compound could be detected in plasma. Instead, radiolabeled nucleosides resulting from nuclease hydrolysis were observed. In the metabolism profiles obtained from various tissues, only radiolabeled nucleosides were found, suggesting that siRNAs are rapidly metabolized and that the distribution pattern of total radiolabeled components can be ascribed to small molecular weight metabolites.

Biodistribution and Metabolism Studies of Lipid Nanoparticle–Formulated Internally [3H]-Labeled siRNA in Mice

“…The half-life values were significantly longer than after i.v. administration of the unformulated [ 3 H]-SSB siRNA, as recently reported (Christensen et al, 2013), where 5-minute p.d. no parent guide strand was observed.…”

“…The sensitivity of MS imaging allowed specific detection of cationic lipid DLin-KC2-DMA in sections from all time H Label. The fraction of the administered radioactive dose converted to tritiated water was initially estimated using urine and blood as matrices, as previously reported (Christensen et al, 2013). Concentration-time course of tritiated water in body water resulted in apparent elimination half-life values of 108 and 115 hours in urine and blood, respectively (Supplemental Fig.…”

“…14-18Mer oligonucleotide metabolites were identified by a corresponding peak in the radiochromatogram versus a control sample, and have to be considered as proposals only. Consult Christensen et al (2013) (incl. Table S4 Supplemental Data) for metabolite nomenclature, assignment, structural elucidation, and identification.…”

“…In addition, siRNAs can also have specificity issues, which can result in off-target silencing or stimulation of the innate immune system. As such, systemic delivery of unformulated siRNA seems more unlikely to be successful (Christensen et al, 2013). For future applications of siRNAs as therapeutics, delivery systems are anticipated to hold the key to their success.…”

“…Nevertheless, the choice of labeling approach has previously been shown to influence the pharmacokinetic behavior of large molecules, such as for siRNAs (Christensen et al, 2013). There are several radiolabels that have been used for this approach: 3 H, 18 F, 32 P, 64 Cu, 111 In.…”