A new US study has further optimised genome editing technology which could help to cure sickle cell disease (SCD) and beta thalassaemia.
Scientists at St. Jude Children’s Research Hospital and the Broad Institute of MIT and Harvard, USA, used adenosine base editing to restart foetal haemoglobin expression in SCD patient cells. They found the technique raised the expression of foetal haemoglobin to higher, more stable, and more uniform levels, than other genome editing technologies.
The findings were published in Nature Genetics.
Adult haemoglobin contains four protein subunits — two beta-globin and two alpha-globin – and mutations in the former gene cause sickle cell disease and beta-thalassaemia.
However, there is another subunit – gamma-globin – that is expressed during foetal development instead of beta-globin.
Gamma-globin combines with alpha-globin to form foetal haemoglobin. Around birth, gamma-globin expression is normally turned off, and beta-globin is turned on.
It means genome editing technologies can introduce mutations that turn the gamma-globin gene back on, increasing foetal haemoglobin production, which can effectively substitute for defective adult haemoglobin production.
Lead corresponding author Dr Jonathan Yen, from St. Jude Therapeutic Genome Engineering, said: “We used a base editor to create a new TAL1 transcription factor binding site that causes particularly strong induction of foetal haemoglobin.
“Creating a new transcription factor binding site requires a precise base pair change — something that can’t be done using CRISPR-Cas9 without generating unwanted byproducts and other potential consequences from double-stranded breaks.”
Corresponding author Dr Mitchell Weiss, the St. Jude Department of Haematology chair, said the gamma-globin gene is a good target for base editing because there are very precise mutations that can reactivate its expression after birth.
This, he says, may provide a powerful ‘one-size-fits-all’ treatment for all mutations that cause SCD and beta-thalassemia.
The study discovered that using base editing at the most potent site in the gamma-globin promoter achieved two- to four-fold greater foetal haemoglobin (HbF) levels than CRISPR/Cas9 editing.
The teams also demonstrated these base edits could be retained in blood stem cells from healthy donors and SCD patients by putting them into immunocompromised mice.
Base editing caused fewer genotoxic events, such as p53 activation and large deletions, and it was more consistent in its edits and products. In contrast to conventional Cas9, which generates DNA alterations called indels, base editing generates precise nucleotide changes with few undesired byproducts.
Dr Weiss said: “In our comparison, we found unanticipated problems with conventional Cas9 nucleases.
“We were somewhat surprised that not every Cas9 insertion or deletion raised foetal haemoglobin to the same extent, indicating the potential for heterogeneous biological outcomes with that technology.”
The group found that individual red blood cells derived from haematopoietic stem cells treated with the same Cas9 produce a more variable amount of foetal haemoglobin compared to cells treated with base editing.
Though base editing performed well, researchers have yet to determine its safety in patients.
“It is very important to test and compare different genome editing approaches for treating SCD and beta-thalassaemia because the best ones are not known,” said Dr Weiss.
Mayuranathan T, Newby GA, Feng R, Yao Y, Mayberry KD, Lazzarotto CR, Li Y, Levine RM, Nimmagadda N, Dempsey E, Kang G, Porter SN, Doerfler PA, Zhang J, Jang Y, Chen J, Bell HW, Crossley M, Bhoopalan SV, Sharma A, Tisdale JF, Pruett-Miller SM, Cheng Y, Tsai SQ, Liu DR, Weiss MJ, Yen JS. (2023) “Potent and uniform fetal hemoglobin induction via base editing.” Nature Genetics, doi: 10.1038/s41588-023-01434-7
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