Researchers at the Shanghai Frontiers Science Center of Genome Editing and Cell Therapy and YolTech Therapeutics have reported promising preclinical data for a non-viral CRISPR-based therapy for primary hyperoxaluria type 1 (PH1). The study, published in Molecular Therapy, demonstrates that a single dose of lipid nanoparticle (LNP)-delivered CRISPR-Cas9 targeting the Hao1 gene can effectively reduce oxalate levels in both mouse and humanized mouse models of PH1.
Addressing PH1 with CRISPR-Cas9
PH1 is a rare, life-threatening genetic disorder caused by mutations in the AGXT gene, leading to a deficiency in the alanine-glyoxylate aminotransferase enzyme. This deficiency results in the accumulation of oxalate in the kidneys and other organs, affecting approximately 45,000 people worldwide. Current treatments include substrate reduction therapies (SRTs) like Oxlumo and Rivfloza, which require multiple doses to maintain efficacy. AAV-delivered CRISPR-Cas9 therapies have raised safety concerns. This new approach offers a potentially safer and more effective alternative.
The research team developed a mouse model of PH1 using LNPs to deliver Cas9 mRNA and a guide RNA targeting the AGXT gene. These mice exhibited elevated oxalate excretion and calcium deposition in the kidneys, characteristic of PH1. Deep sequencing revealed no significant off-target editing in the liver.
Effective Gene Editing and Oxalate Reduction
Following the establishment of the PH1 mouse model, a single intravenous dose of LNP-CRISPR-Cas9 targeting the mouse Hao1 gene (mHao1) was administered. Analysis two weeks later showed near-complete silencing of AGXT expression and 61-75% editing efficiency of Hao1 in liver tissue. Over a 12-month follow-up, the mice exhibited reduced glycolate oxidase expression and decreased urinary oxalate levels. Long-term editing efficiency of both AGXT and Hao1 was confirmed, with Western blot and immunohistochemistry indicating reduced GO expression in treated PH1 mice compared to controls.
Translation to Humanized Model
To facilitate clinical translation, the researchers created a humanized Hao1 mouse model by injecting a modified human Hao1 sequence into mouse zygotes. These mice expressed human Hao1 in their livers, mimicking human Hao1 expression patterns. Intravenous delivery of LNP-CRISPR-Cas9 targeting human Hao1 (hHao1) in these mice resulted in dose-dependent in vivo editing efficiency (up to 59%), reduced hHao1 gene and protein expression, and decreased urinary oxalate levels.
Clinical Implications
These findings demonstrate the potential of LNP-mediated CRISPR therapy to safely and effectively alleviate PH1 symptoms. YolTech Therapeutics is currently advancing this approach with YOLT-203 in clinical trials.
