How the ABCA4 Gene Model Supports Research Breakthroughs in Gene Therapy Drug Development

Gene editing and RNA therapies are reshaping the future of medicine, with recent breakthroughs making headlines worldwide. Among these, the ABCA4 gene has emerged as a critical focus, especially in the fight against Stargardt disease, a leading cause of inherited vision loss. With cutting-edge treatments like base editing, RNA exon editing, and antisense oligonucleotides entering clinical trials, ABCA4 stands at the forefront of innovation.

In this article, explore how advancements in DNA/RNA therapies are converging on ABCA4 to drive next-generation treatments for inherited retinal diseases. Discover why this gene is a key target for emerging technologies and how humanized mouse models are accelerating preclinical research. Keep reading to uncover the future of retinal disease therapy.

How ABCA4 is Revolutionizing Gene and RNA Therapies

Recent advancements in DNA/RNA editing and small nucleic acid therapies have highlighted the potential of novel treatment modalities, some of the key preclinical & clinical research developments in 2024 include:

• ADAR-mediated RNA editing: On October 17, Wave Life Sciences announced the successful clinical proof-of-concept for WVE-006, the world’s first ADAR-based RNA editing therapy, with promising data results.^[1]

• Base editing: On November 6, Beam Therapeutics will present early clinical proof-of-concept data for its base editing therapy BEAM-101 at the American Society of Hematology (ASH) annual meeting. The therapy significantly increased fetal hemoglobin (HbF) levels in patients with sickle cell disease (SCD).^[2]

• CRISPR gene editing: Research on Intellia Therapeutics' ATTR amyloidosis CRISPR-based gene editing therapy (Nexiguran ziclumeran) published on November 16 in The New England Journal of Medicine shows a sustained 90% reduction in disease markers over two years.^[3]

• RNA interference (RNAi): On November 17, RNAi therapy pioneer Alnylam announced early clinical data for its third-generation TTR-targeted RNAi drug, Nucresiran, which demonstrated a six-month efficacy with a single dose.^[4]

These advances clearly demonstrate the enormous potential of DNA/RNA-based therapies. Amid these developments, the ABCA4 gene is emerging as a key target, particularly for RNA exon-editing therapies. ABCA4’s role in Stargardt disease (STGD), the most common form of juvenile macular degeneration, positions it as a potential cornerstone for next-generation therapies. Poised as the first RNA exon editing therapy target to enter clinical trials, ABCA4 has attracted significant attention and may become a key breakthrough point for the next generation of gene and RNA therapies.

Figure 1. The world’s first in vivo RNA exon editing therapy, ACDN-01, was approved for clinical trials earlier this year.^[5]

1. ABCA4 and Stargardt Disease (STGD)

The macula is the central region of the retina, responsible for "high-resolution" color vision. Age-related macular degeneration (AMD) and monogenic macular degeneration can both lead to blindness. Among these, monogenic macular degeneration typically manifests in adolescence, is more severe, and can result in gradual loss of abilities such as reading, driving, or facial recognition, ultimately leading to the loss of central vision. The most common form of monogenic macular degeneration is Stargardt disease (STGD), affecting approximately 1 in every 6,500 people, primarily caused by biallelic loss-of-function mutations in the ABCA4 gene.^[6]

The ABCA4 gene encodes the ABCA4 membrane transporter protein located in photoreceptors and retinal pigment epithelial cells, which mainly functions to clear toxic metabolic products generated during the bleaching of rhodopsin, preventing the accumulation of these compounds in the retina.^[7-8] Mutations in the ABCA4 gene lead to abnormal accumulation of toxic metabolites such as N-retinylidene-phosphatidylethanolamine and lipofuscin A2E, which trigger the death of retinal pigment epithelial cells and photoreceptors, ultimately resulting in retinal degenerative diseases.^[7-8]

Figure 2. Localization of the ABCA4 protein in the outer segments of photoreceptor rods and its role in the visual cycle.^[8]

2. ABCA4: A Key Research Target For Novel Therapies

With ABCA4 gene mutations accounting for approximately 30% of cases of inherited retinal diseases (IRD), providing a broad patient base.^[9] Although STGD is the most common form of inherited macular dystrophy, there is currently no effective treatment. Traditional gene replacement therapies face challenges due to the large size of the ABCA4 gene (approximately 128 kb) and the minimum sequence required to encode the ABCA4 protein (6.8 kb), both of which exceed the delivery limit of AAV vectors (4.7 kb).^[10] Furthermore, there are more than 2,200 disease-associated variants in the ABCA4 gene, most of which are missense mutations, followed by mutations that affect pre-mRNA splicing.^[11] As such, humanized models require a gene editing method with large gene fragment knockin capabilities.

These characteristics make ABCA4 a key research target for novel therapies, such as gene editing and RNA-based therapies:

• Antisense oligonucleotides (AON): ProQR Therapeutics developed QR-1011, which treats STGD patients by correcting abnormal splicing.^[12-13]
• RNA exon editing: Ascidian Therapeutics' ACDN-01 can replace the first 22 exons of ABCA4 mRNA, theoretically covering about 70% of STGD patients.^[14-15]
• Base editing: Beam Therapeutics developed an efficient base editor targeting common mutations in the ABCA4 gene, demonstrating mutation correction effects in ABCA4 humanized mice, non-human primates, and human retinal tissue.^{[9, 16]}

Both QR-1011 and ACDN-01 are now in clinical trials, while other innovative therapies—including CRISPR-mediated editing, novel AAV vector strategies, and homology-independent targeted integration (HITI)—continue to advance preclinical validation. Several institutions presented preclinical validation data for various novel therapies at this year's Association for Research in Vision and Ophthalmology (ARVO) annual meeting. These therapies include gene editing (e.g. Cas9-mediated editing, transposon systems, SaKKH base editing), RNA editing (e.g. Cas13-mediated RNA base editing, ADAR-mediated RNA editing), novel AAV vector therapies (e.g. mini-ABCA4 protein, dual AAV vector strategy), antisense oligonucleotides (AON), and Homology-Independent Targeted Integration (HITI) treatment strategies.^[17-18]

Figure 3. Antisense oligonucleotide (AON) therapy QR-1011 treats STGD by inhibiting exon skipping caused by pathological mutations.^[12]

3. Application Of Humanized Mice In Preclinical Evaluation Of Gene/RNA Editing Therapies

Due to the genetic differences between humans and mice—and since gene editing, RNA editing, and antisense oligonucleotide (AON) therapies all target the human ABCA4 gene—using fully genomically humanized mouse models can significantly improve the accuracy of preclinical in vivo assessments for such therapies. For example, Beam Therapeutic's base editing therapy successfully evaluated its in vivo editing efficiency using a humanized mouse model carrying pathogenic mutations.^[16]

Figure 4. The A-to-G base editor (ABE) successfully corrected mutations in ABCA4 G1961E humanized mice in vivo.^[16]

4. Cyagen' s proprietary ABCA4 humanized model and humanized point mutation models

Cyagen has developed a combination of humanization models for preclinical research involving ABCA4:

1. Fully humanized mouse model B6-hABCA4 (product number: C001551) with whole genomic replacement of ABCA4;
2. B6-hABCA4*c.5461-10T>C humanized point mutation mouse model (product number: I001210) carrying the pathogenic mutation c.5461-10T>C.

These humanized mouse models successfully recapitulate the abnormal splicing patterns of the ABCA4 gene observed in patients with ABCA4-related Stargardt disease.

Figure 5. Gene editing strategy of B6-hABCA4 mice. The sequences from the ATG start codon to the TGA stop codon of the mouse Abca4 gene were replaced with the sequences from the ATG start codon to the TGA stop codon of the human ABCA4 gene

(1) Humanized ABCA4 Mice: B6-hABCA4

This model expresses only the human ABCA4 gene and its transcripts, while maintaining normal retinal structure and photoreceptor function.

Figure 6. The B6-hABCA4 mouse successfully expresses the human ABCA4 gene while maintaining normal retinal structure and photoreceptor function.

(2) Pathogenic ABCA4 Point Mutation Model: B6-hABCA4*c.5461-10T>C Mice

This mouse was constructed by introducing the c.5461-10T>C mutation into the B6-hABCA4 mouse model. This mutation leads to splicing abnormalities, generating aberrant transcripts with exon 39 skipping and exon 39-40 skipping, while reducing the normal ABCA4 transcript. This mutation is also the target site for the ASO therapy QR-1011. Gene expression and sequencing results show that this model successfully recapitulates the splicing defects and transcriptional patterns observed in patients, providing an important tool for studying STGD-related therapies.

Figure 7. The B6-hABCA4*c.5461-10T>C mouse recapitulates the abnormal splicing pattern of ABCA4 observed in human disease patients.

Conclusion

Cyagen’s ABCA4 humanized mouse models (B6-hABCA4, B6-hABCA4c.5461-10T>C*) provide essential tools for evaluating novel therapies targeting the gene for Stargardt disease while providing the proof of concept for applications across other diseases.

B6-hABCA4 Mouse Model (product number: C001551) 

• Expresses only the human ABCA4 gene and RNA transcript.
• Maintains normal retinal structure, vascular morphology, and photoreceptor function.
  
  

B6-hABCA4*c.5461-10T>C Mouse Model (product number: I001210)

• Introduced the c.5461-10T>C mutation into the B6-hABCA4 model.
• This mutation causes abnormal splicing, generating aberrant transcripts with exon 39 skipping and exon 39–40 skipping, while reducing the normal ABCA4 transcript.
• Successfully recapitulates the splicing defects and decreased abnormal transcript patterns seen in patients, making it an invaluable tool for preclinical studies of therapies such as AONs and base editing.
  
  

These models maintain the structural and functional integrity of the retina while enabling precise replication of disease-relevant genetic abnormalities. Together, these two models provide essential tools for the preclinical in vivo evaluation of novel therapies for Stargardt disease (STGD), such as gene editing, antisense oligonucleotides, base editing, and RNA editing.

Preclinical Ophthalmology Research Modeling & CRO Solutions

Beyond Stargardt disease, Cyagen offers a comprehensive suite of pre-developed retinal disease models and preclinical ophthalmology solutions tailored to small animal eye research needs.

Our model generation capabilities enable us to rapidly develop advanced research models, including: custom target and full-genome humanized models, humanized immune system mouse models, inducible Cre models, rare disease models, and more—catering to the diverse needs of researchers in disease studies and therapy development.

Contact us for more details or a free consultation to discuss your research needs.

Validated Genetic Ophthalmic Disease Models

References
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[2]Beam Therapeutics. "Beam Therapeutics to Present Data Across Hematology Franchise, Including First Clinical Data for BEAM-101 in Sickle Cell Disease and ESCAPE Non-human Primate Data, at American Society of Hematology (ASH) Annual Meeting." Accessed November 22, 2024. Beam Therapeutics.
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[4]Alnylam Pharmaceuticals. "Alnylam Pharmaceuticals Press Release | Nov 17, 2024 | Alnylam Announces Interim Phase 1 Data of Nucresiran (ALN-TTRsc04) Showing Rapid Knockdown of TTR that is Sust." Accessed November 22, 2024. Alnylam Pharmaceuticals.
[5]Ascidian Therapeutics. "Ascidian-IND-Acceptance-Release_FINAL.1.29.24.pdf." Accessed November 22, 2024. https://ascidian-tx.com/wp-content/uploads/2024/01/Ascidian-IND-Acceptance-Release_FINAL.1.29.24.pdf.
[6]Hanany M, Rivolta C, Sharon D. Worldwide carrier frequency and genetic prevalence of autosomal recessive inherited retinal diseases. Proc Natl Acad Sci U S A. 2020 Feb 4;117(5):2710-2716.
[7]Quazi F, Lenevich S, Molday RS. ABCA4 is an N-retinylidene-phosphatidylethanolamine and phosphatidylethanolamine importer. Nat Commun. 2012 Jun 26;3:925. 
[8]Scortecci JF, Molday LL, Curtis SB, Garces FA, Panwar P, Van Petegem F, Molday RS. Cryo-EM structures of the ABCA4 importer reveal mechanisms underlying substrate binding and Stargardt disease. Nat Commun. 2021 Oct 8;12(1):5902.
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