Hemophilia. Cystic fibrosis. Alzheimer’s. β-thalassemia. Fragile X syndrome. These are all diseases that could be lessened or cured by CRISPR. CRISPR, or clustered regularly interspaced short palindromic repeats, is a prokaryotic defense system that has immense potential in gene editing, by targeting one’s DNA. While it first debuted in 1987 by a team of Japanese scientists, it truly began showing up in everyday news in 2018 when a scientist from China gene-edited babies. Is CRISPR controversial science fiction, or the future of biomedicine?
CRISPR is part of prokaryotes– specifically bacterias’–immune system. Repetitive DNA sequences (also called repeats) are the backbone of CRISPR, because between them lie spacer DNA. The spacers each correspond to DNA from a particular bacteriophage (virus) that previously infected the bacterial cell. If a phage whose DNA is preserved in a spacer tries to infect the bacterial cell again, the CRISPR spacer sequences are transcribed into RNA, then a protein known as Cas9 uses this RNA as a guide to the invader. The Cas9 then cuts up the invader’s DNA so it can no longer infect and reproduce within the bacteria. Miraculously, this natural nanotechnology can be applied to gene editing. A scientist will design a specific RNA (mimicking the CRISPR spacer sequences) that targets whatever sequence of genome DNA the scientist chooses to modify. This guide RNA will combine with Cas9 and lead it to a specific location in the genome, then the protein will cut the DNA. The repair mechanisms within the cell will then do one of two things: non-homologous end joining or homology-directed reaper. The former directly repairs the breaks in the DNA, but it is not an exact process, and may cause small mutations to be inserted. This is useful because this often inhibits the gene or destroys its function. The latter is more precise; when there is a homologous DNA strand present in the nucleus, it can be used as a template for the cell to repair the breaks– avoiding those small errors. Think of the difference between a large flat brush and a small pointed one, they both work but have different strengths.
A miniature version of CRISPR may have just opened up a whole new world of gene editing applications. Regular CRISPR is fairly large despite being microscopic, so it struggles to enter certain cells. This limits gene editing to “ex vivo” procedures that mainly occur on the inside of a petri dish– making blood disorders the main “editees”. Mammoth Biosciences has managed to shrink the Cas9 protein into a form that’s ⅓ smaller: NanoCas. The shrunken tool can be placed into a virus transport box, which is often used in gene therapy. The Mammoth team is in the testing stage, using mice and primate test subjects to edit the genes of two inherited conditions: Duchenne muscular dystrophy and high cholesterol. In the latter, the PCSK9 gene- which concerns LDL receptors– was targeted. A single injection and NanoCas displayed 60% efficiency in editing, a level close to the SaCas9 protein which is far larger.
Advances in CRISPR technology crop up everyday, but could this mini molecule truly make genetic disorders obsolete with a single poke?
