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Thursday, 6 September 2018

Gene editing: off the bench and into the clinic

As the first sponsored CRISPR-Cas9 clinical study gets underway,
industry has high hopes that genome editing will eventually make gene
therapy a mainstream treatment for both inherited and non-inherited
Recent progress in gene therapy, with successful proof of concept being reported in a range of otherwise untreatable conditions and the grant of marketing authorisations for commercially-developed treatments makes it easy to forget that these successes were built on almost three decades of endeavour and regular disappointment.

Gene therapy restores aberrant or missing biological functions through the introduction of functional genes carried into the cell using engineered viruses or by zapping with electrical pulses. A more involved process is the alteration of specific nucleotide sequences within a faulty gene to restore or 
modify function- so-called “gene editing”. 

Around the turn of the millennium, certain bacterial genomes were observed to have multiple copies of repeated DNA sequences which read the same in both directions -“clustered regularly interspaced palindromic repeats” (CRISPR), which were subsequently found to be responsible for the exquisitely specific recognition of infecting viruses. In 2012, it was shown that CRISPR could be exploited to edit genes in mammalian cells, prompting an explosion in translational science, patent wrangles and an influx of venture funding. 

Reduced to its bare bones, CRISPR gene editing utilises a short piece of “guide” RNA, designed to bind to a specific sequence of DNA, complexed with an enzyme able to cut double-stranded DNA, with the bacterial enzyme “Cas9” being widely used. Once within the nucleus, the guide RNA latches onto its complementary DNA sequence, directing the enzyme to cut in the right place: the cell’s DNA repair mechanism completes the editing process. 

CRISPR-Cas9 (and other CRISPR based systems) have brought relative simplicity and low cost to gene editing, providing a powerful research tool with application across biological disciplines and a route to significant advances in agriculture and the treatment of both inherited and non-inherited disease.

American and European regulators have been understandably cautious with respect to CRISPR clinical studies. As with other forms of gene therapy, CRISPR-Cas9 editing is not without potential risk, such as unwanted alteration of DNA sequences other than the target gene sequence, or the triggering of immune responses to the Cas9 enzyme (obtained from one or other of two common skin bacteria) or through introduced RNA being recognised as being  “foreign”. 

Chinese investigators have been less troubled by the unknowns surrounding CRISPR clinical development:  first in man studies began in 2010, with around 80 subjects being treated in cancer and HIV gene editing studies by February this year. Several Chinese research groups have explored  gene editing in non-viable, and most recently, viable human embryos.  

CRISPR Therapeutics, in partnership with Vertex,  successfully addressed undisclosed FDA concerns raised in May this year and are now recruiting beta thalassemia patients for a Phase I/II gene editing study. Like sickle cell disease, beta thalassemia arises from an inherited defect in the gene encoding the oxygen-carrying protein, haemoglobin. 

Rather than directly editing the haemoglobin gene, the CRISPR-Cas9 complex (designated CTX001) is designed to alter a DNA sequence responsible for the shutdown of foetal haemoglobin production, in the hope that turning the switch back on will produce sufficient levels of functional haemoglobin. The editing process is performed ex vivo, with blood-forming stem cells being extracted from the patient, transfected with the CRISPR-Cas9 complex, expanded and then returned by infusion. 

CRISPR Therapeutics and its peers, Editas Medicine and Intellia Therapeutics have ambitions to develop gene editing therapies for a range of genetic conditions that result in metabolic dysfunction (Hurler syndrome, glycogen storage disease, alpha-1 antitrypsin deficiency, transthyretin amyloidosis) and visual impairment (Leber congenital amaurosis, Usher syndrome), as well as cystic fibrosis and Duchenne muscular dystrophy (DMD). Exonics Therapeutics recently published data indicating that their “SingleCut” CRISPR technology was able to  restore near normal levels of dystrophin expression in an animal model of DMD. 

CRISPR may prove to be more efficient and flexible means of engineering “off the shelf” (allogeneic) cancer-fighting T cell therapies, reducing the cost and complexity of current patient-specific CAR-T therapy  [ASCO Clinical Cancer Advances 2018. And the winner is...].

Pre-CRISPR era gene editing methodologies may also prove to have therapeutic application. Sangamo Therapeutics has used a zinc finger nuclease-based approach to achieve “in body” editing of a faulty gene in four individuals with Hunter syndrome, an enzyme disorder that prevents the breakdown of complex sugars, leading to harmful accumulation. A decrease in urinary complex sugar level was observed in two individuals, although not an increase in normal enzyme levels in the blood. Full interpretation of the significance of these results will take time but importantly, “in body” editing raised no safety issues. 

Clinical development of  gene editing-based therapies will continue to advance cautiously (at least in the West), but is likely to can to benefit from experience gained in viral and non-viral vector design now being exploited in other forms of gene therapy and growing regulatory familiarity with gene therapy risk-benefit assessment. 

Image credit: Credit: Jill George, NIH.

Update added 29th November 2018:

Rather than offer a perspective on the much publicised use of CRISPR to gene-edit babies (the word "abhorrent" would certainly feature in my commentary), I recommend Sharon Begley's excellent article in STAT.

Amid uproar, Chinese scientist defends creating gene-edited babies. Online 28th November 2018. http://tinyurl.com/yacrjc56

The heroes of CRISPR. Lander, ES. Cell (2016);164: 18-28.

A safety and efficacy study evaluating CTX001 in subjects with transfusion-dependent β-Thalassemia. NCT03655678. http://tinyurl.com/y72xxugk.

Gene editing restores dystrophin expression in a canine model of Duchenne muscular dystrophy. L. Amoasii et al., Science 10.1126/aau1549 (2018).

Did CRISPR really fix a genetic mutation in these human embryos? Callaway, E. Nature News, published online 8th August 2018. doi: 10.1038/d41586-018-05915-2

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