B1 - Metabolic Diseases (including Diabetes)
169: In Vivo Correction of Metabolic Pathogenic Variants via Base Editing and Prime Editing: Toward the Equitable Treatment of Hepatic Inborn Errors of Metabolism
Type: Oral Abstract Session
Presentation Details
Session Title: Gene Transfer and Genome Editing for Inherited Metabolic Disorders
Phenylketonuria (PKU), an autosomal recessive disorder caused by pathogenic variants in the phenylalanine hydroxylase (PAH) gene, is the most common inborn error of metabolism. PKU results in the buildup of blood phenylalanine (Phe) to neurotoxic levels. Current dietary and medical treatments are chronic and reduce, rather than normalize, blood Phe levels. The most frequently occurring PAH variant in PKU patients is the c.1222C>T (R408W) variant. Using real-world data from a PKU cohort at an academic medical center, we found that most patients with at least one R408W allele (n = 36) experience chronic, severe Phe elevations and do not meet Phe monitoring guidelines—highlighting the high unmet medical need arising from the challenges of lifelong adherence to dietary and medical therapy. Motivated by these findings, we are developing “one-and-done” CRISPR editing therapeutics for PKU. We have established a pipeline to rapidly screen and optimize base editing or prime editing strategies in vitro and deploy them in vivo with either lipid nanoparticles (LNPs) or adeno-associated viral (AAV) vectors on a timescale of months. In initial work, we created six humanized PKU mouse models for the six most frequent PAH variants and, in parallel, prime-edited variant-bearing hepatocyte cell lines. In homozygous or compound heterozygous PKU mice, we observe complete and long-term durable normalization of blood Phe levels (>90% reduction) as soon as 48 hours after treatment, with whole-liver corrective PAH editing as high as >50% with either base editing or prime editing. We have developed therapeutic leads for multiple PAH variants to permanently normalize blood Phe levels and definitively treat PKU in these patients and are on track for early-phase clinical trials within a few years. Despite these encouraging results, we are cognizant of the risk of “mutational discrimination” in unduly focusing on more common diseases and on high-prevalence variants skewed towards specific ancestry groups. Accordingly, we have extended our pipeline to develop and validate a corrective therapeutic for any variant in any patient with a hepatic inborn error of metabolism. We have begun to apply this workflow in real time to patients with devastating, ultra-rare inborn errors of metabolism, namely the urea cycle disorders and organic acidemias, tackling any variant—even n-of-1 variants—identified via universal newborn screening. The first four patients have (1) citrullinemia type 1, with homozygosity for an ASS1 transition variant (G>A), (2) argininosuccinic aciduria, with homozygosity for an ASL transition variant (A>G), (3) propionic acidemia, with homozygosity for a PCCA 1-bp deletion, and (4) CPS deficiency, with compound heterozygosity for a CPS1 partial gene deletion and a transversion variant (A>C). To address these specific variants in parallel, we have developed (1) a high-throughput lentivirus-based multiple-variant prime editing screening platform in human hepatocytes and (2) a multi-humanized mouse with a string of genomic sequences harboring each of the four variants inserted into the Rosa26 safe harbor locus, allowing for in vivo testing of lead editing solutions for all four variants in a single mouse model, which is now underway. This augmented, scalable pipeline represents a critical step towards our goal of rapidly devising and deploying a personalized editing treatment for any patient in need.
Plain Language Summary
We are addressing the challenges of Phenylketonuria (PKU), the most common genetic disorder of metabolism causing harmful phenylalanine buildup. Current treatments are not fully effective, and PKU patients with the most common c.1222C>T (R408W) variant experience severe Phe elevations, failing to meet Phe monitoring guidelines. Motivated by this, we are developing ""one-and-done"" CRISPR editing therapies. We have established a rapid pipeline for base and prime editing, achieving promising results in humanized mouse models, with plans for clinical trials in a few years. Recognizing potential bias, we extended the approach to rare disorders. Initial success in treating citrullinemia, argininosuccinic aciduria, propionic acidemia, and CPS deficiency sets the stage for a scalable, personalized editing treatment for any patient with hepatic inborn errors.
Dominique Brooks1, Madelynn Whittaker1, Ping Qu1, William Peranteau2, Mohamad-Gabriel Alameh2, Rebecca Ahrens-Nicklas2, Kiran Musunuru1, Xiao Wang1
1University of Pennsylvania, Philadelphia, PA,2Children's Hospital of Philadelphia, Philadelphia, PA"
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