Correction of the Marfan Syndrome Pathogenic FBN1 Mutation by Base Editing in Human Cells and Heterozygous Embryos

Abstract

There are urgent demands for efficient treatment of heritable genetic diseases. The base editing technology has displayed its efficiency and precision in base substitution in human embryos, providing a potential early-stage treatment for genetic diseases. Taking advantage of this technology, we corrected a Marfan syndrome pathogenic mutation, FBN1. We first tested the feasibility in mutant cells, then successfully achieved genetic correction in heterozygous human embryos. The results showed that the BE3 mediated perfect correction at the efficiency of about 89%. Importantly, no off-target and indels were detected in any tested sites in samples by high-throughput deep sequencing combined with whole-genome sequencing analysis. Our study therefore suggests the efficiency and genetic safety of correcting a Marfan syndrome (MFS) pathogenic mutation in embryos by base editing.

Enabling large-scale genome editing by reducing DNA nicking

“…CRISPR has recently brought about a radical transformation in the basic and applied 30 biological research, leading to commercial applications and a multitude of clinical trials (31), and even to the controversial tests of human germline modifications (32)(33)(34)(35)(36). While the use of CRISPR and its myriad derivatives has greatly reduced the activation energy and technical skill required to perform genome editing several barriers limit fundamental and clinical applications: 1) The need for a custom gRNA, for each target, 2) difficult delivery, 3) inefficiencies once delivered, 4) off- 35 target errors, 5) on-target errors, 6) the cytotoxicity of DNA damage when multiplexing beyond 62 loci(16), 7) the limitation of insertion to sizes below 7.4kb(37), 8) immune reactions to Cas, gRNA and vector.…”

Attitudes of people with inherited retinal conditions toward gene editing technology

“…Recent advances in gene editing technology, particularly the efficient and relatively inexpensive CRISPR‐Cas9 system, have brought gene editing to the forefront of medical research as well as bioethical debate. The first clinical trials using somatic gene editing technology are already underway (Kaiser, ), as are early forays into gene editing in embryos (Cyranoski & Ledford, ; Ma et al, ; Zeng et al, ). However, ethical concerns about uses of this technology remain, and many have called for the voices of those who stand to be most impacted by its development, people with genetic conditions and disabilities, to be included in the conversation (Check Hayden, ; Shakespeare, ).…”

Increasing the targeting scope and efficiency of base editing with Proxy‐BE strategy

“…Base editing holds great promise for gene therapy strategies, and it has been applied to treat genetic disorders in cell culture . However, some obstacles have hindered the application of BE, such as the limitation of PAMs, G/C‐rich regions, and chromatin accessibility.…”

“…Cas9 nickase and DNA deaminase are fused, thus enabling single‐base‐resolution substitution (C•G to T•A substitution or A•T to G•C substitution) without cleaving the double‐strand DNA . Because of its unique ability to perform precise gene therapy, BE technology has been applied in animals, plants, and even human embryos, in directions ranging from disease modeling to correction of mutations . However, the application of BE has been hindered by some obstacles, such as limited protospacer adjacent motifs (PAMs), G/C‐rich regions, and chromatin accessibility .…”