Nucleic Acid Therapeutics

Small interfering RNA (siRNA) represents a transformative therapeutic class that enables precise gene silencing through RNA interference (RNAi). N-acetyl galactosamine (GalNAc)-conjugated siRNAs have achieved remarkable clinical success with six FDA-approved therapeutics targeting liver diseases. Following subcutaneous administration, GalNAc–siRNAs rapidly accumulate in hepatocytes via asialoglycoprotein receptor (ASGPR)-mediated endocytosis and are sequestered within endolysosomal compartments, creating an intracellular depot that sustains therapeutic effects for weeks to months. While RNAi-mediated degradation and nuclease metabolism contribute to siRNA clearance, the detection of full-length, active siRNA in circulation long after dosing suggests alternative clearance mechanisms exist. This review examines the pharmacokinetic properties and intracellular trafficking of GalNAc–siRNA, with particular focus on underexplored clearance pathways from hepatocytes. We discuss potential mechanisms including endosomal recycling, efflux via exosomes and lysosomal exocytosis that may facilitate siRNA redistribution from tissues to circulation. Understanding these clearance mechanisms could enable correlation of plasma and tissue drug concentrations, reduce preclinical animal use, inform clinical dosing strategies and guide the design of next-generation siRNA therapeutics with optimized tissue residence time and enhanced therapeutic durability.

Aptamers are single-stranded synthetic oligonucleotides that bind noncovalently to targets with high affinity and selectivity. They are generated through an in vitro selection process known as Systematic Evolution of Ligands by Exponential Enrichment. Aptamers show significant promise in cancer diagnostics and therapeutics due to their specificity, versatility, and tunable biochemical properties. Aptamers can be used to target cellular biomarkers, which are detectable molecules that can indicate specific physiological states of cells, organs, and organisms, making them valuable indicators for diseases. Clinical trials on therapeutic aptamers are providing further insight into their potential use. Leukemia is a hematological malignancy characterized by the uncontrolled proliferation of abnormal blood-forming cells, primarily affecting the bone marrow and peripheral blood. This review will highlight the applications of aptamers in relation to the diagnosis and treatment of leukemia. Novel research on leukemia-related aptamers will be presented along with future directions for aptamer-based diagnostic and therapeutic methods.

Oligonucleotide therapeutics are emerging as a promising modality for targeting disease-associated RNAs. Phosphorothioate (PS)-containing oligonucleotides have gained prominence due to their enhanced stability and pharmacodynamic properties. However, current manufacturing practices afford a mixture of Rp and Sp stereoisomers, and this distribution has been linked to changes in product efficacy. Understanding the sensitivity of analytical methods to changes in this quality attribute has therefore become critically important. Here, we used a suite of analytical techniques—ultraviolet (UV) thermal denaturation, circular dichroism (CD), and nuclear magnetic resonance (NMR) spectroscopy—to evaluate the PS diastereomer distribution using Tegsedi, a Food and Drug Administration-approved PS-containing antisense oligonucleotide, and with other synthetic inotersen samples having varied PS diastereomer distributions. While UV and CD techniques showed limited sensitivity, NMR excelled in detecting small changes in the PS diastereomer distribution. The univariate metric of ³¹P integration was shown to be insufficient for this quality metric evaluation; application of principal component analysis to both 1D ³¹P and 2D ¹H,¹³C spectra revealed distinct PS changes that arose from the different activators used during manufacturing. This comprehensive evaluation highlights the necessity of advanced analytical techniques in ensuring the quality and consistency of PS-containing oligonucleotide therapeutics.