Gene defect fixed in 8 out of 20 cloned embryos, heralds future of gene fixes and enhancement

A team in China has taken a new approach to fixing disease genes in human embryos. The researchers created cloned embryos with a genetic mutation for a potentially fatal blood disorder, and then precisely corrected the DNA to show how the condition might be prevented at the earliest stages of development.

Huang’s team is also the first to edit out the mutation responsible for a ‘recessive’ disease: one caused by having two faulty copies of a gene. Because it would be difficult for researchers to find dozens of embryos that all have this rare double mutation, the team worked around this roadblock by developing embryonic clones from their patient’s skin cells.

* Researcher fix gene defect in 8 out of 20 cloned embryos but the fixed embryos did not have all cells corrected
* in future they will prevent inherited diseases by correcting the eggs and cells which would need IVF (In vitro fertilization aka test tube babies)
* If inherited diseases are fixed using IVF and CRISPR then future work could also enhance genes for increased intelligence and other characteristics

Precision editing

In the latest study, Huang’s team used ‘base editing’, a modification of CRISPR–Cas9. It guides an enzyme to specific gene sequences, but does not cut the DNA. Instead, the Cas9 enzyme is disabled and tethered to another enzyme that can swap out individual DNA base pairs. So far, this technique can convert guanine (‘G’) to adenine (‘A’), and cytosine (‘C’) to thymine (‘T’). Hundreds of genetic diseases are caused by single-base changes, or ‘point mutations’, and so editing of this sort at the embryonic stage could potentially stave off such conditions.

Huang’s team chose one mutation common in the Chinese population: a switch from an A to a G at a certain spot in the HBB gene, which can lead to β-thalassaemia, a recessive blood disorder associated with severe or fatal anaemia. Researchers generally source embryos from in vitro fertilization (IVF) clinics, but it’s rare for these facilities to have embryos with two copies of the same rare mutation. So Huang’s team found a person with the blood disorder, extracted their skin cells and used cloning techniques to develop embryos with the same genetic makeup.

The researchers reported that in 8 of 20 cloned embryos, they were able to convert the errant G back into an A in one or both copies of the gene. (Repairing only one copy might be enough to cure a recessive disease.) That rate is too low for the technique to be considered for clinical use, but the efficiency was high relative to that achieved in other gene-editing studies. “The repair rate is pretty good, and certainly promising,” says Gaetan Burgio, a geneticist at the Australian National University in Canberra. “Our study opens new avenues for therapy of β-thalassaemia and other inherited diseases,” says Huang.

But scientists caution that not all cells in the eight embryos were fixed. Such embryos are ‘mosaic’, meaning that they have a patchwork of cells with different genetic make-ups, which is potentially dangerous. “It looks like solid work, but highlights that the problem of mosaicism remains a challenge for any form of gene editing in the human embryo,” says Dieter Egli, a stem-cell biologist at Columbia University in New York City.
Unintended consequences

Some scientists also question whether Huang’s team looked thoroughly enough for unintended genetic changes, called off-target effects, that might have been caused by the base-editing procedure, although the authors reported that none were found.

Huang says future experiments will be more comprehensive, but that this first study was a successful proof of principle that the base-editing technique can be used to correct a disease mutation in a human embryo. It may be that conventional CRISPR–Cas9 cannot fix embryos when both copies are faulty, although this isn’t yet clear. In August, for instance, Mitalipov’s team reported using CRISPR–Cas9 to repair a mutation in a gene that can cause a potentially deadly heart disorder, by using the other, healthy copy of the gene as a template.

In the future, Huang says, he plans to ask for oocytes and sperm from donors who have one mutated copy of the gene — and so are unaffected by the condition, but are carriers of the disease — and use these to produce embryos. Some of those embryos would have two mutated copies, and some one, but Huang wants to edit both types. That raises the contentious idea that gene editing might be used not only to prevent severe disease, but also to eliminate the chance of people becoming carriers of the disorder. “Base editing can repair the mutant site and block it from being passed on to the next generation,” he says.

Protein and Cell – Correction of β-thalassemia mutant by base editor in human embryos

Abstract

β-Thalassemia is a global health issue, caused by mutations in the HBB gene. Among these mutations, HBB −28 (A G) mutations is one of the three most common mutations in China and Southeast Asia patients with β-thalassemia. Correcting this mutation in human embryos may prevent the disease being passed onto future generations and cure anemia. Here we report the first study using base editor (BE) system to correct disease mutant in human embryos. Firstly, we produced a 293T cell line with an exogenous HBB −28 (A G) mutant fragment for gRNAs and targeting efficiency evaluation. Then we collected primary skin fibroblast cells from a β-thalassemia patient with HBB −28 (A > G) homozygous mutation. Data showed that base editor could precisely correct HBB −28 (A > G) mutation in the patient’s primary cells. To model homozygous mutation disease embryos, we constructed nuclear transfer embryos by fusing the lymphocyte or skin fibroblast cells with enucleated in vitro matured (IVM) oocytes. Notably, the gene correction efficiency was over 23.0% in these embryos by base editor. Although these embryos were still mosaic, the percentage of repaired blastomeres was over 20.0%. In addition, we found that base editor variants, with narrowed deamination window, could promote G-to-A conversion at HBB −28 site precisely in human embryos. Collectively, this study demonstrated the feasibility of curing genetic disease in human somatic cells and embryos by base editor system.

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