A team of investigators at Ionis Pharmaceuticals (USA), led by first author Dr. Mariana Bravo Hernandez, PhD, and corresponding author Holly Kordasiewicz, PhD, describe the use and characterization of oligonucleotide (ON) therapeutics as disease-modifying approaches for central nervous system disorders. The study cited (“Transient Acute Neuronal Activation Response Caused by High Concentrations of Oligonucleotides in the Cerebral Spinal Fluid”) and selected for featured “Breakthrough” status at Nucleic Acids Research was evaluated and published February 4, 2026 side-by-side with a companion study published January 6, 2026 entitled “Reversable Acute Sedation Response of Phosphorothioate Antisense Oligonucleotides Following Local Delivery to the Central Nervous System.” Dr. Kordasiewicz is also corresponding author of this companion piece.
Oligonucleotide therapeutics (OTs) delivered locally to the central nervous system (CNS) are being approved as therapies for neurological diseases. There are now more than a dozen clinical trials ongoing with intrathecally delivered OTs, and a similar number of n=1 therapies advancing into patients. Despite the huge potential of OTs in the CNS, this is not without its challenges and risk. It is therefore critical that investigators share our experiences regarding therapeutic approaches, observed toxicities and mechanistic understanding of them.
One class of safety findings that are unique to local CNS injection of OTs are transient neurological responses that occur immediately after dosing. These are often dose-limiting, and if not understood and mitigated preclinically can occur in human patients. Multiple groups have reported on these findings in rodents, but considerable inconsistencies remain in classifying them and their underlying mechanisms (as the effect of OTs in the CNS is extremely complex and requires large data sets to untangle the various mechanisms). How to interpret and translate these findings beyond rodents has not been systematically discussed in the literature.
The investigators involved in this have characterized two very distinct mechanisms of acute toxicity, and compiled all our learnings into two companion publications. The division of their results is intended to make it clear that they describe distinct mechanisms, with very different implications for CNS OT development.
In the first manuscript, the authors describe the acute sedation response observed with phosphorothioate antisense oligonucleotides (ASOs). This is sequence-dependent, phosphorothioate-dependent, translates directly across species, and is caused by binding of ASO extracellularly to neurons. The authors provide methods to identify particularly poor sequences in silico and for in vitro and in vivo screening methods, as well as design approaches that can mitigate the response and describe the underlying mechanism. They also provide the details on how this translates from rodents to non-human primates, a point of considerable interest to n=1 communities.
In the second manuscript (the Breakthrough article) the authors focus on the acute neuronal activation response. They demonstrate this is driven by binding of the charged oligonucleotides to divalent cations. As this is due to divalent cation binding, this can occur with both ASOs and siRNAs and is not largely sequence- or phosphorothioate-driven. They demonstrate this is due to the molar ratio of divalent cations to OT in the dosing solution, as pre-saturation of the OT with divalent cations mitigates the effect. Because aCSF (the typical formulation for CNS OTs) has a fixed divalent cation ratio, this finding is exacerbated in preclinical species where dosing volumes are limited and divalent cation to ASO ratio is much lower than the human equivalents. They provide evidence that this is largely a preclinical challenge, due to the low dosing volumes, and provide the methods for determining if the human dosing solutions have sufficient divalent cation to OT ratios.
In totality, in these two manuscripts the authors provide a mechanistic understanding of two unique acute neurological responses, and a comprehensive framework for quantifying and mitigating both responses. The goal is to provide a definitive resource on the subject and offer comprehensive references that define and clarify these two distinct mechanisms. These publications may aid in better and more rapid design of molecules, more consistent evaluation of the CNS safety of molecules across the field and should significantly advance the field of OTs for neurological disease while providing safer alternatives for patients.
Reviewers and editors at NAR who examined this study and its companion manuscript stated that “There’s so much interest in CNS development of nucleic acid therapeutics, and so little public data on the safety issues addressed in these studies, that I think this paper should immediately be read and studied by everyone in the space” and “from my own understanding of the issues, the authors have performed an excellent analysis of this topic which advances the field and will guide others.”
Ionis Pharmaceuticals focuses on developing RNA-targeted therapeutics, primarily using antisense oligonucleotide (ASO) technology, to treat diseases that are hard to address with conventional small molecules or antibodies. Their pipeline spans neurologic, cardiovascular/metabolic, renal, and rare genetic disorders.
Nucleic Acids Research is a publication of Oxford University Press and is fully open access. Its mission is to provide outstanding, scientist-led evaluation and dissemination of the highest caliber research across a wide range of disciplines focused on the role of nucleic acids and nucleic acid-interacting molecules in cellular and molecular biology. Breakthrough articles at NAR describe studies that solve a long-standing problem in their field or provide exceptional new insight and understanding into an area of research that will clearly motivate and guide new research opportunities and directions. They represent the top papers that NAR receives for publication and are selected by the Editors based on nominations by authors and/or reviewers, and on the subsequent recommendation of the reviewers and Editorial Board members.