A groundbreaking study published in Nature by researchers from the Children’s Mercy Research Institute (Kansas) and the University of Kansas Medical Center introduces patient-derived organoid models for evaluating personalized therapeutics. The study presents a rapid and scalable platform for screening antisense oligonucleotides (ASOs), offering a cost-effective and accessible method that can be implemented in standard laboratories without the need for specialized equipment.

What Are Organoids and Patient-Derived Organoids

Organoids are three-dimensional (3D) multicellular structures derived from pluripotent or adult stem cells, designed to mimic the structure and function of real organs. Unlike traditional 2D cell cultures, organoids develop tissue-specific architecture, making them more accurate models for studying human diseases and drug responses. Patient-derived organoids (PDOs) are customized 3D tissue models created from a patient’s induced pluripotent stem cells (iPSCs) or biopsy-derived cells. These organoids preserve the patient’s genetic and epigenetic profile, making them valuable tools for disease modeling, drug screening, and precision medicine applications.

In disease modeling, organoids are used to investigate conditions such as neurological disorders (e.g., Alzheimer’s, Parkinson’s), cancer, and rare genetic diseases, replicating disease mechanisms more accurately than traditional models. In drug discovery and screening, they provide a valuable tool for assessing drug efficacy and toxicity, enabling more precise preclinical evaluations. Precision medicine benefits from patient-specific organoids, allowing researchers to test personalized treatments before clinical application, improving therapeutic outcomes. Additionally, regenerative medicine leverages organoids for tissue transplantation and organ repair, presenting potential solutions for degenerative conditions. In infectious disease research, organoids serve as models for studying viral infections like COVID-19, Zika, and hepatitis, facilitating the development of targeted antiviral treatments.

Advantages of organoids over animal models in preclinical studies

Traditional preclinical models, including animal studies and 2D cell cultures, have significant limitations in replicating human physiology, which slows down drug development and gene therapy testing. Animal models often fail to accurately mimic human diseases due to species differences, leading to misleading or non-translatable results in clinical trials. Additionally, animal studies are time-consuming and costly, often requiring years of research before a drug can advance to human trials. In contrast, organoids offer a human-relevant, scalable, and cost-effective alternative for disease modeling and therapeutic screening. These three-dimensional structures more accurately replicate human tissue architecture, improving the reliability of preclinical testing. Organoids also provide a faster and more cost-effective approach compared to lengthy animal studies, while enabling high-throughput screening and patient-specific testing, making them ideal for precision medicine applications. By addressing the shortcomings of traditional models, organoids are transforming preclinical research, offering a more reliable, efficient, and ethical platform for drug discovery and gene therapy development.

Advantages of patient-derived organoids for antisense oligonucleotide (ASO) screening

The advent of personalized medicine has revolutionized the treatment of genetic disorders by offering patient-specific therapeutic strategies. Antisense oligonucleotides (ASOs), short synthetic nucleic acids that modulate RNA processing, have shown promise in treating rare genetic diseases, particularly those caused by splicing errors or premature transcript termination. However, ASO therapy development is hampered by high costs, lengthy preclinical validation, and a lack of efficient screening platforms. The research team led by Scott Younger, Children’s Mercy Research Institute and the University of Kansas Medical Center , a scalable platform for generating patient-derived cellular models and demonstrate their effectiveness in the preclinical evaluation of patient-specific ASOs.

Innovative methodology: From PBMCs to functional organoids

The study presents a novel platform that streamlines the development of patient-derived organoids for ASO screening. The core innovation lies in a streamlined workflow that converts cryopreserved peripheral blood mononuclear cells (PBMCs) into induced pluripotent stem cells (iPSCs) within 3 weeks, achieving a 93% success rate when ≥300,000 PBMCs are available. By optimizing a reprogramming cocktail, combining GSK, MEK, and TGFβ inhibitors with ROCK inhibitors and growth factors, the researchers bypassed costly viral vectors and episomal plasmids. The resulting iPSCs exhibited robust pluripotency, confirmed by trilineage differentiation into ectoderm (PAX6+/NESTIN+), endoderm (FOXA2+/SOX17+), and mesoderm (BRACHYURY+/NCAM+) lineages, and maintained genomic stability over 30 passages.

Functional validation ASO delivery in iPSC-derived cardiac organoids

To assess the efficacy of ASOs in patient-derived organoids, the researchers optimized ASO delivery using cardiac troponin T (TNNT2) as a model target in cardiac organoids derived from a commercially sourced iPSC line. TNNT2 is a key protein in cardiomyocytes, and its inhibition provides a quantifiable measure of ASO activity. The study designed ASOs targeting different regions of TNNT2, including the translation start site, splice donor site, and splice acceptor site. When applied to cardiac organoids, these ASOs effectively modulated TNNT2 expression, demonstrating the platform’s ability to screen ASOs for functional impact.

Functional validation was performed using calcium imaging, which assessed the contraction patterns of cardiac organoids. Cardiac organoids treated with ASOs targeting TNNT2 exhibited disrupted calcium transients, confirming successful genetic perturbation. This approach established the feasibility of using patient-derived organoids for ASO efficacy testing.

Screening FDA-Approved ASO for Duchenne Muscular Dystrophy (DMD) Using Patient-Derived Organoids

Duchenne muscular dystrophy (DMD) is a severe genetic disorder caused by mutations in the dystrophin (DMD) gene. The study evaluated the platform’s ability to screen existing ASO therapeutics for DMD using patient-derived cardiac organoids. A patient with a structural deletion in the DMD gene (exons 46–53) was used as a model system. This mutation results in a loss of dystrophin expression, leading to progressive muscle degeneration.

This study evaluated the effectiveness of patient-derived cardiac organoids as a platform for screening existing ASO therapeutics for DMD. Using a model derived from a patient with a structural deletion spanning exons 46–53, researchers tested an FDA-approved exon 45-skipping ASO, which successfully restored dystrophin expression and improved cardiac function. These improvements were confirmed through RNA sequencing, which validated exon skipping, calcium imaging, which showed the restoration of rhythmic transients, and functional assays that demonstrated improved contraction. Furthermore, a second ASO candidate currently in clinical trials also restored dystrophin expression, reinforcing the robustness of patient-derived organoids as a reliable system for ASO evaluation.

Beyond testing existing therapeutics, the study also designed and evaluated personalized ASOs for two DMD patients (siblings) carrying a deep intronic mutation that led to abnormal splicing. The custom-designed ASOs effectively corrected the splicing defects, restored dystrophin expression, and improved calcium transients in patient-derived cardiac organoids. These findings highlight the potential of patient-derived organoids as a scalable and clinically relevant platform for developing both standardized and personalized ASO therapies for rare genetic diseases like DMD.

Importantly, ASO transfection showed no toxicity effects, and restored functional contractions for up to 21 days post-treatment. These findings establish patient-derived organoids as a scalable and clinically relevant platform for ASO drug screening.

Conclusion and Future Implications

This study marks a significant advancement in gene therapy screening, demonstrating a rapid and patient-specific organoid platform for ASO evaluation. By integrating PBMC-derived iPSCs and functional organoids, this approach provides a scalable and cost-effective alternative to animal models, accelerating preclinical ASO validation. Beyond ASO screening, this platform has broader applications in drug discovery, regenerative medicine, and high-throughput compound screening. The study also highlights the potential use of brain organoids for neurological disease modeling. However, regulatory challenges remain for the approval of personalized ASOs. As organoid technology evolves, its impact on drug development, precision medicine, and rare disease therapeutics will redefine the future of personalized healthcare.

Reference

Means, J.C., Martinez-Bengochea, A.L., Louiselle, D.A. et al. Rapid and scalable personalized ASO screening in patient-derived organoids. Nature 638, 237–243 (2025). https://doi.org/10.1038/s41586-024-08462-1

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