Durable cure for Pseudoxanthoma Elasticum: High-Efficiency, Precision-Targeted Gene Therapy

Lipid nanoparticles containing messenger RNA (mRNA-LNPs) encoding Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) base-editors that correct pseudoxanthoma elasticum-causing mutations.
Problem:
Pseudoxanthoma elasticum (PXE) is a genetic disorder affecting approximately 1 in 25,000 Americans. It is caused by loss-of-function mutations in the gene ABCC6. Since ABCC6 regulates the release of circulating calcium inhibitors from the liver, its dysfunction dangerously elevates systemic levels of calcium, leading to calcification in the skin, eyes, and cardiovascular system. These changes manifest clinically as inelastic skin, loss of vision and cardiovascular complications, with the latter two causing significant morbidity and mortality. Currently, there is no cure for PXE nor any targeted therapies.
Solution:
A liver-directed gene therapy that corrects ten prevalent ABCC6 mutations which cause PXE. For each mutation, the inventors identified an optimized adenine base editor (ABE) and hybrid DNA-RNA guide combination, which are delivered via mRNA-LNPs. The guide directs the ABE to the mutation site highly specifically, while the ABE efficiently edits the mutation, restoring ABCC6 function and normal calcium levels.
Technology:
The inventors generated hepatocyte cell lines expressing PXE-causing ABCC6 variants to screen editor-guide combinations in vitro. For each variant, they identified the optimal combination which demonstrated the highest correction efficiency for the mutated ABCC6. They also substituted portions of the RNA of standard guides with DNA, creating hybrid DNA-RNA guides that are more target-specific while remaining compatible with mRNA-LNP delivery. In humanized mice, the delivery of the optimized editor-guide combination via mRNA-LNPs achieved liver-specific correction of the pathogenic R1164X ABCC6 variant. This treatment restored ABCC6 expression and calcium inhibitor levels and demonstrated long-term prevention of muzzle calcification in mice.
Advantages:

Permanent correction of ten pathogenic variants of ABCC6, including the two most prevalent pathogenic variants in the Caucasian population and three most prevalent pathogenic variants in a Japanese cohort
Improved on-target editing efficiency for the two most prevalent pathogenic variants in the Caucasian population compared to previous reports
Full prevention of muzzle calcification in humanized mice model for the second most prevalent pathogenic ABCC6 variant in the Caucasian population
Hybrid DNA-RNA guides achieve higher target specificity than standard guide RNA
Scalable and generalizable development framework for efficient optimization, validation, and translational advancement of variant-specific base-editing strategies

Stage of Development:

Preclinical Development

Treatment of LNP encoding optimized editor-guide combination ABE8.8-hyb18 for ABCC6 variant R1164X restored the calcium content within the muzzle tissue of humanized homozygous R1164X mice back to wild-type levels. Quantification of calcium content of muzzle tissue from 18- week-old wild-type mice (n = 8 mice), heterozygous mice (n = 7 mice), homozygous R1164X mice treated with PBS (n = 8 mice), and homozygous R1164X mice treated with LNP (n = 9 mice). Each dot represents calcium values from one mouse. Data expressed as mean (SD). Statistical significance determined by one-way ANOVA with Sidak’s multiple comparisons test.
Intellectual Property: