Correction of a pathogenic gene mutation in human embryos

Abstract

More than 10,000 monogenic inherited disorders have been identified, affecting millions of people worldwide. Among these are autosomal dominant mutations, where inheritance of a single copy of a defective gene can result in clinical symptoms. Genes in which dominant mutations manifest as late-onset adult disorders include BRCA1 and BRCA2, which are associated with a high risk of breast and ovarian cancers 1 , and MYBPC3, mutation of which causes hypertrophic cardiomyopathy (HCM) 2 . Because of their delayed manifestation, these mutations escape natural selection and are often transmitted to the next generation. Consequently, the frequency of some of these founder mutations in particular human populations is very high. For example, the MYBPC3 mutation is found at frequencies ranging from 2% to 8% 3 in major Indian populations, and the estimated frequency of both BRCA1 and BRCA2 mutations among Ashkenazi Jews exceeds 2% 4 .HCM is a myocardial disease characterized by left ventricular hypertrophy, myofibrillar disarray and myocardial stiffness; it has an estimated prevalence of 1:500 in adults 5 and manifests clinically with heart failure. HCM is the commonest cause of sudden death in otherwise healthy young athletes. HCM, while not a uniformly fatal condition, has a tremendous impact on the lives of individuals, including physiological (heart failure and arrhythmias), psychological (limited activity and fear of sudden death), and genealogical concerns. MYBPC3 mutations account for approximately 40% of all genetic defects causing HCM and are also responsible for a large fraction of other inherited cardiomyopathies, including dilated cardiomyopathy and left ventricular non-compaction 6 . MYBPC3 encodes the thick filament-associated cardiac myosin-binding protein C (cMyBP-C), a signalling node in cardiac myocytes that contributes to the maintenance of sarcomeric structure and regulation of both contraction and relaxation 2 .Current treatment options for HCM provide mostly symptomatic relief without addressing the genetic cause of the disease. Thus, the development of novel strategies to prevent germline transmission of founder mutations is desirable. One approach for preventing second-generation transmission is preimplantation genetic diagnosis (PGD) followed by selection of non-mutant embryos for transfer in the context of an in vitro fertilization (IVF) cycle. When only one parent carries a heterozygous mutation, 50% of the embryos should be mutationfree and available for transfer, while the remaining carrier embryos are discarded. Gene correction would rescue mutant embryos, increase the number of embryos available for transfer and ultimately improve pregnancy rates.Recent developments in precise genome-editing techniques and their successful applications in animal models have provided an option for correcting human germline mutations. In particular, CRISPR-Cas9 is a versatile tool for recognizing specific genomic sequences and inducing DSBs 7-10 . DSBs are then resolved by endogenous DNA repair mechanisms, prefer...

Crossing the germline: CRISPR-Cas9 and our responsibility as reproductive endocrinology and infertility physicians

“…However, some investigators only look at sites in the genome that are highly homologous to the intended site of repair as off-target effects may be enriched here. Importantly, in the recent report by Ma et al, no off-target effects were detected by WGS in any of the examined blastomeres of wild-type embryos [10]. An off-target effect was defined as one that differed from the on-target sequence by up to 7-nucleotide mismatches or up to 5-nucleotide mismatches with a 2-nucleotide DNA bulge.…”

“…In August 2017, Ma et al published an innocuously titled report in Nature called BCorrection of a pathogenic gene mutation in human embryos^in which they describe repair of a heterozygous four base pair deletion in MYBPC3, one genetic cause of hypertrophic cardiomyopathy [10]. The authors repaired 58/58 (100%) of human preimplantation embryos.…”

“…Of note, M-phase injection also allowed most (42/58, 72.4%) embryos to be restored to wild type, so they were indistinguishable from unaffected embryos [10]. The • Continued reassessment of both health and societal benefits and risks, with broad ongoing participation and input by the public • Reliable oversight mechanisms to prevent extension to uses other than preventing a serious disease or condition remaining 16/58 (27.6%) embryos were in fact corrected, though through an error-prone non-homologous end-joining (NHEJ) mechanism, which can lead to insertions or deletions Bindels^at the break site.…”

How Genome Editing could be used in the Treatment of Cardiovascular Diseases

“…Arguably, the maximum benefit would be obtained if the therapy was administered at the earliest possible time, in the zygote, since every cell in the resulting person would carry the therapeutic change and descendants of the person would carry the therapeutic change as well. This scenario is no longer a theoretical one, in light of a recent report of therapeutic genome editing in human embryos resulting from sperm cells of a man with hypertrophic cardiomyopathy due to a mutation in MYBPC3 [15]. While the corrected embryos were not intended to be carried to term, the embryos were viable and had the potential to become living people.…”

Why the way we look at things in reproductive medicine is changing

“…Perhaps we should all take pause with the recent papers on gene editing in human embryos, a milestone in human medicine that is likely to affect the practice of ARTs [6]. Notable also that only weeks later, this technology was used not to effect a corrective influence on a mutant gene but to explore a fundamental aspect in mammalian embryology having to do with how the various cell lineages of the early embryo their future identities [7].…”