In vivo base editing rescues Hutchinson–Gilford progeria syndrome in mice

Abstract

Summary Hutchinson-Gilford progeria syndrome (HGPS) is typically caused by a dominant-negative C•G-to-T•A mutation (c.1824 C>T, G608G) in LMNA , the nuclear lamin A gene. This mutation causes RNA mis-splicing that produces progerin, a toxic protein that induces rapid aging and shortens lifespan to ~14 years 1 – 4 . Adenine base editors (ABEs) perform targeted A•T-to-G•C base pair conversion with minimal byproducts and without requiring double-strand DNA breaks or donor DNA templates 5 , 6 . Here, we describe the use of an ABE to directly correct the pathogenic HGPS mutation in cultured progeria patient-derived fibroblasts and in a mouse model of HGPS. Lentiviral delivery of ABE to patient-derived fibroblasts results in ~90% correction of the pathogenic allele, mitigation of RNA mis-splicing, reduced progerin levels, and correction of nuclear abnormalities. Unbiased off-target DNA and RNA analysis did not detect off-target editing activity in treated patient-derived fibroblasts. In transgenic mice homozygous for the human LMNA c.1824 C>T allele, a single retro-orbital injection of adeno-associated virus 9 (AAV9) encoding the ABE resulted in substantial, durable correction of the pathogenic mutation (~20-60% across various organs 6 months post-injection), restoration of normal RNA splicing, and reduction of progerin protein. In vivo base editing rescued vascular pathology, preserving vascular smooth muscle cell counts and preventing adventitial fibrosis. A single ABE AAV9 injection at P14 improved animal vitality and greatly extended median lifespan from 215 to 510 days. These findings support the potential of in vivo base editing to treat HGPS, and other genetic diseases, by directly correcting the root cause of disease.

Molecular and Cellular Mechanisms Driving Cardiovascular Disease in Hutchinson-Gilford Progeria Syndrome: Lessons Learned from Animal Models

“…The G608G BAC mouse model, developed in 2006, expresses human lamin A, lamin C, and progerin together with endogenous mouse lamin A and lamin C and was the first HGPS animal model to recapitulate some aspects of the HGPS-associated vascular phenotype [ 42 ]. Heterozygous G608G BAC animals have a normal lifespan and do not present consistent abnormalities in bone, muscle, skin, or heart [ 42 ], whereas homozygous mice show musculoskeletal and skin anomalies, lack subcutaneous fat, and die prematurely [ 43 , 63 ]. However, both heterozygous and homozygous G608G BAC mice develop a marked vascular phenotype in the aorta, carotid artery, and iliac artery, including progressive loss of VSMCs, thickening of the adventitia and medial layer, and vessel calcification in older animals.…”

Boon and Bane of DNA Double-Strand Breaks

“…The DSBs can be generated by Cas nucleases which are guided by an RNA complementary to the target site, thus enabling site specific mutagenesis, usually via small deletions or insertion that inactivate a gene of interest. Recently, genes can be edited by precise base edition without induction of DSBs [ 26 ]. Alternatively, when additional (partially) homologous donor sequences are supplied which become integrated into the break via HDR, gene replacement with a desirable allele, or introduction of novel genetic information can be achieved (for review: [ 27 , 28 , 29 , 30 , 31 ]).…”

“…Both approaches are useful for basic research (testing and modification of gene functions) and for breeding purposes (to prevent negative or promote positive traits in crops and livestock). Therapy of gene-mediated human diseases is also envisaged [ 26 ]. Finally, there are attempts to generate by targeting DSBs at distinct loci meiotic cross-overs [ 32 ], specific chromosome rearrangements to recapitulate or reverse evolutionary rearrangements [ 33 , 34 ], or de novo rearrangements (e.g., [ 35 ]) which might initiate speciation, as well as to domesticate de novo, e.g., orphan crops (for review [ 36 ]).…”

Rescue of STAT3 Function in Hyper-IgE Syndrome Using Adenine Base Editing

“…In addition to these results, the successful use of ABEs in cellular and animal models of other monogenic diseases, such as Xeroderma pigmentosum, tyrosinemia, Duchenne muscular dystrophy, and Hutchinson/Gilford progeria syndrome, confirms that ABEs are a promising tool for the treatment of monogenic diseases lacking a curative therapy option such as STAT3-HIES. [47][48][49][50] However, additional studies concerning the safety of ABEs are required. In this regard, developments such as the ABE variant ABEmaxAW or alternative forms of delivery, for example, ABE RNP instead of ABE encoding plasmid, might further improve the safety profile of the treatment modality.…”