1. Introduction: Personalized ASO Therapies

This paper provides consensus guidelines for designing Antisense Oligonucleotides (ASOs) tailored to individual patients, known as N-of-1 therapies. ASOs are short, synthetic DNA strands that modify gene expression by altering pre-mRNA splicing, offering treatment potential for various genetic diseases.

A successful example of this approach is Milasen, an ASO developed for a single patient with Batten disease, demonstrating the feasibility of personalized genetic medicine.

2. Key Recommendations for ASO Development

The guidelines emphasize three major principles for effective ASO design:

• Standardized Experimental Designs: Ensuring consistency in testing methodologies.
• Reference ASOs for Consistency: Using validated ASO sequences to compare results across studies.
• Data Sharing: Promoting open collaboration among researchers to advance the field of personalized therapies.

3. ASO Preclinical Evaluation: Essential Steps

Proper preclinical evaluation is critical for the success of N-of-1 ASOs. The following steps ensure high efficacy and minimal off-target effects:

• Design Multiple ASO Candidates: Testing several ASO sequences to identify the most effective one.
• In Vitro Cell Models: Using patient-derived cells to evaluate splicing correction and functional improvements.
• Testing in Relevant Models: Assessing ASO effects in disease-relevant animal models.

4. Types of Exon Skipping for ASO Therapy

Different exon-skipping strategies are used in ASO therapies:

• Targeting Cryptic Exons: Suppressing unintended exons that cause protein dysfunction.
• Modifying Constitutive Exons: Restoring proper splicing in genetic disorders like Duchenne Muscular Dystrophy (DMD).
• Blocking Poison Exons: Preventing premature protein truncation in diseases like Spinal Muscular Atrophy (SMA).

5. ASO Design Considerations

Effective ASO design depends on multiple factors:

• Nucleotide Composition: Optimizing chemical modifications to enhance stability.
• Target Selection: Choosing the most effective exon for therapeutic intervention.
• Minimizing Off-Target Effects: Conducting bioinformatics analyses to ensure specificity.

6. The Future of N-of-1 ASO Therapies

To advance personalized ASO therapies, collaboration between researchers, clinicians, and regulatory bodies is essential. The study highlights the need for:

• Global Data Sharing: Improving research efficiency by pooling genetic and clinical data.
• Standardized Guidelines: Establishing best practices for preclinical testing and regulatory approval.
• Regulatory Pathways: Accelerating the development of patient-specific therapies.

7. Conclusion

The guidelines presented in this paper serve as a foundation for improving ASO therapy development. By refining experimental designs, standardizing methodologies, and fostering collaborative research, the potential for N-of-1 ASO treatments to benefit patients with rare mutations is significantly enhanced.