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Practical Recommendations for the Selection of Patients for Individualized Splice-Switching ASO-Based Treatments

Bianca Zardetto, Marlen C. Lauffer, Willeke van Roon-Mom, and Annemieke Aartsma-Rus on behalf of the Dutch Center for RNA Therapeutics, Department of Human Genetics, Leiden University Medical Center, Leiden, Netherlandsthe and The N = 1 Collaborative

April 18, 2024


Although around 6% of the world’s population is affected by rare diseases, only a small number of disease-modifying therapies are available. In recent years, antisense oligonucleotides (ASOs) have emerged as one option for the development of therapeutics for orphan diseases. In particular, ASOs can be utilized for individualized genetic treatments, addressing patients with a known disease-causing genetic variant, who would otherwise not be able to receive therapy. Careful prioritization of genetic variants amenable to an ASO approach is crucial to increase chances for successful treatments and reduce costs and time for drug development. At present, there is no consensus on how to systematically approach this selection procedure. Here, we present practical guidelines to evaluate disease-causing variants and standardize the process of selecting -of-1 cases. We focus on variants leading to a loss of function in monogenic disorders and consider which splice-switching ASO-mediated treatments are applicable in each case. To ease the understanding and application of our guidelines, we created a hypothetical transcript covering different pathogenic variants and explained their evaluation in detail. We support our recommendations with real-life examples and add further considerations to be applied to specific cases to provide a comprehensive framework for selecting eligible variants.


Rare diseases are defined as conditions affecting less than 1 in 2,000 individuals within Europe. However, with an estimated 7,000 different rare diseases [1], approximately 470 million people are affected worldwide [2]. Although a genetic origin has been identified for more than 70% of these diseases [3], less than 6% currently have approved disease-modifying treatments [4, 5]. Affected individuals are often refractory to drugs commonly used for the management of clinical manifestations [6], illustrating the urgency of developing targeted treatments.

Recent progress in RNA-based therapies has shown that antisense oligonucleotide- (ASO-) mediated treatments offer the potential to partially fill this therapeutic gap [7]. For ultrarare disorders, ASOs represent an opportunity to design a disease-modifying drug in a patient-specific fashion. As of now, 19 oligonucleotide-based therapies have been approved by either the European Medicines Agency (EMA), the Food and Drug Administration (FDA), or the Japanese Ministry of Health, Labour and Welfare [8], 10 of which are ASOs. The case of an -of-1 ASO treatment for a single patient with neuronal ceroid lipofuscinosis (milasen) has paved the way for the development of individualized treatments targeting private variants [9, 10].

With the increasing use of whole-genome sequencing (WGS) and other advanced diagnostic methods, more of these unique pathogenic variants are being identified. However, not all are amenable to treatments using ASO-based approaches. Careful consideration of each case is required to ensure that resources are focused on promising targets. There is often only a narrow window of opportunity for treatment development and initiation as patients will progress further during the development process, increasing the urgency for timely drug delivery and being specific in whom to select for targeted treatment.

Here, we provide practical recommendations for the assessment of pathogenic variants for individualized genetic treatments and explain which variants are eligible for customized approaches. We will focus on one particular type of ASOs, called splice-switching ASOs (ssASOs).

Decision tree for variant evaluation of exonic LoF variants not disrupting splicing. The aim of this decision tree is to identify exonic LoF variants that can be targeted with a ssASO via exon skipping.

Zardetto, B., Lauffer, M. C., van Roon-Mom, W., Aartsma-Rus, A., & on behalf of the N = 1 Collaborative, undefined. (2024, April 18). Practical recommendations for the selection of patients for individualized splice-switching ASO-based treatments. Human Mutation.



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