Single base editing: new gene editing technique with higher precision

Mutations in the human genome are often the underlying cause for genetic diseases. It is thus understandable that many researchers focus their efforts on finding tools to edit or “fix” these errors in the genetic code. CRISPR/Cas-9, one of the latest and most promising techniques, is able to alter or delete whole genes. Researchers describe it as a type of “scissors” (bit.ly/n_Enz). However, CRISPR can have unfavorable or off-target effects (bit.ly/Si_BE), for example by altering a gene not intended for change or by failing to change the intended gene. Moreover, CRISPR involves double-stranded DNA cuts and then relies on the cell’s own machinery to reassemble the DNA by filling the gap where the cut was made with the correct DNA sequence. This too can be prone to errors.

The good news is that researchers from the Broad Institute of MIT and Harvard have now established a gene editing technique that is more precise: it rewrites individual errors in single base-pairs of the genetic code, “instead of cutting and replacing whole chunks of DNA” (bit.ly/CR_new). This new way to edit DNA and RNA is called base editing and it is intended to fix point mutations involved in human disease. Before we dive into the mechanisms of this base editing technique let us go through some brief background information. DNA is composed of individual units called “bases”. The four existing bases are A, T, C and G, which base pair (A-T and C-G) to form the DNA double-stranded helix. A point mutation occurs when just one base in the DNA strand gets substituted, deleted, or inserted. Such point mutations account for 32,000 of the 50,000 changes in the human genome associated with diseases (bit.ly/CR_new).

Researchers led by David Liu, a Harvard chemistry professor and member of the Broad Institute, have now developed a way to convert one DNA base into another. When a base in one strand is converted, the cell is tricked to fixing the complementary base in the other strand to ensure that correct base pairing occurs. Specifically, this gene editing technique changes an A-T to a G-C base pair, which had previously not been possible (only G-C to A-T changes had been feasible). This is particularly useful, as mutations in which a G-C is mutated to an A-T base pair account for almost 50% of the 32,000 single point mutations associated with genetic diseases in humans. (bit.ly/Si_BE).

But how exactly does this new gene-editing technique work? Liu and his team developed a molecular machine called a gene editor, made up of an atom-rearranging enzyme that is able to molecularly change the A (adenine) base into inosine, which is read and copied by the cell’s protein-building machinery as G (guanine). The gene editor also has a “Cas9 nickase” part that puts a nick into the complementary strand of DNA, allowing the cell’s DNA repair machinery to replace the original T (paired to A) with C. Thus A-T is replaced with G-C. Guide RNA is used to make sure that the gene editor is directed to the correct spot in the DNA.

This gene-editing technique that fixes single point mutations indeed sounds like a very efficient and clean procedure. The researchers have already applied it to cell cultures derived from patients to fix the mutation responsible for causing hereditary hemochromatosis (bit.ly/CR_new), a disorder where the body absorbs excess iron from food. Over time, the iron levels can build up and cause liver cancer and diseases, as well as diabetes, heart or joint disease. In another study, Liu and his co-workers also used this technique to introduce a new mutation to suppresses sickle-cell anemia. Both of these studies turned out to be a success, as no off-target effects or unintentional DNA insertions or deletions were identified (bit.ly/CR_new).

Though this new technique sounds extremely promising and could potentially also be useful for NGLY1 deficiency, aspects such as safety, efficiency and delivery methods have to be further revised, before it can be actually used to edit the human genome.