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Gene Editing: Freely You Have Received, Freely Give

This year has seen significant advances in the search for human gene editing of Mendelian disease. Back in April, a team from three major institutions in Seoul reported in Nature Biotechnology on the use of a recent CRISPR variation that does single-base-pair editing. Their proof-of-concept was the “Himalayan mutation“, an A-to-G switch in the tyrosinase gene that affects mouse pigmentation, and they got exactly the color mice that they were expecting. With this result in hand, they went on to a mutation that produces Duchenne muscular dystrophy: a premature stop codon in the Dmd gene was converted back to the codon for glutamine, allowing the gene to code for the missing dystrophin protein again. Intramuscular injections of the gene-editing system in AAV vectors (which had to be split due to the size of the needed proteins) caused about 17% of the muscle fibers to produce dystrophin, well above the level believed needed for therapeutic benefit.

And this week two papers have come out in Nature Medicine with further results. One is from Zürich (the university and the ETH) on correction of phenylketonuria (the well-known PKU) in adult mice. The second is from a team at Penn and Children’s Hospital in Philadelphia, also in mice, but targeting two different genes in utero. Both of these groups appear to have been very successful, and these results make it more likely than ever that we’re going to be able to reverse single-nucleotide genetic diseases in human beings.

The Zürich group also (like the Korean one) used an AAV viral vector targeting a base switch in the phenylalanine hydroxlase enzyme, again with an intein-split base editor system to get around the cargo capacity of that viral technique. (Split inteins allow you to “reassemble” a longer protein from two pieces; there are quite a few variations on the idea, and it’s a whole topic in itself). Intravenous treatment of the mutant mice with this system appears to have worked: the mice show reduced levels of phenylalanine in the blood, and mRNA from the liver shows corrected transcripts (in a dose-dependent fashion). The rate of correction increases over the weeks after injection – the highest dose group, at the 14-week time point, shows 63% corrected mRNA. The mice themselves start to put on weight normally and to grow normally pigmented fur (reversing two other consequences of the PKU mutation). A check for ten closely homologous DNA regions showed that none of them had been inappropriately edited.

Meanwhile, the Penn paper also edited hepatocytes, but this time before the mice had even been born. They used Ad adenoviral vectors rather than AAV (they didn’t go to the trouble of working out the split-intein system, and the Ad vector can carry more freight), so this is a delivery system may have to be modified for human use. But the proof-of-concept in mice certainly worked: they introduced a mutation into PCSK9 to try to bring on the low-cholesterol phenotype that’s noted in natural mutations in this gene. And that’s exactly what they got – interestingly, with this vector the prenatal-treated mice showed lasting effects, whereas a postnatal treatment group showed gradually decreasing ones. But the gene editing again seems to have been specific, and no effects were seen in the mothers, either. They also tried correcting hereditary tyrosinemia type 1 (a mutation in the Fah gene), and succeeded in this as well: normal mice resulted, with increasing correction over time as the normalized cells expanded compared to the impaired ones, and no evidence of the other predicted likely off-target edits.

Taken together, these studies have been extremely successful, from what I can see. The mouse systems studied here are directly analogous to the human diseases; you could hardly ask for stronger animal-model evidence that these techniques should be applicable in human patients. And the selectivity and overall lack of side effects are very impressive. There are a great many genetic disorders that fall into just the category that has been addressed here: point mutations that can be corrected by bringing in the guide-RNA targeted deaminase that does the base switch in these systems. Many of these are metabolic errors that can be addressed in the liver (where these two new papers are working). Eventually, despite the high barrier to trying it in the clinic, the prenatal route may well be the way to go: there are far fewer immune complications in treating the developing fetus, the diseases can be caught early by amniocentesis screening, and then can be corrected as early as possible in development.

This is going to be a fast-moving field, and in some cases it may push aside some earlier CRISPR human-therapy attempts. One way or another, this idea is heading into people, and it’s going to be doing so as quickly as scientifically and ethically possible. I hope that there aren’t any unexpected complications (because those could well be bad ones), but for now, the way looks as wide-open as anything like this will ever be. Here we go. It’s time to stop playing the genetic hands we’ve been dealt, and heal the sick.

16 comments on “Gene Editing: Freely You Have Received, Freely Give”

  1. loupgarous says:

    Given the deaths that have occurred during clinical safety trials of viral-vectored gene therapy for cystic fibrosis, caution is indicated – but what good news! If CRISPR’s downside (off-target base pair changes) can be eliminated or reduced, this is the sort of research which makes your previous with for a Nobel Prize in biology very sensible. Molecular biology and genetic engineering have now come into their own, and we may be able to replace numerous not entirely satisfactory or safe drug interventions by editing patients’ genomes!

    1. Spike says:

      loupgarous – can you provide the source of your statement “Given the deaths that have occurred during clinical safety trials of viral-vectored gene therapy for cystic fibrosis, caution is indicated”. It seems that you are implying that the gene therapy killed the patient. I don’t recall hearing that

      1. UudonRock says:

        There were trials in the early 2000’s on viral vector gene therapy for X-SCID patients that did result in the occasional case of T-Cell leukemia. This was directly therapy related since these subjects had no T-Cells to speak of prior to treatment. The potential damage is likely related to unknown harmful mutations that may occur should the viral vector inject their carried genes into healthy calls or the wrong location in the DNA. More recent CART T therapies that utilize a viral vector can on occasion cause cytokine-release syndrome. This has proven fatal in a small number of cases and has been looked at by the FDA. It’s hard to say this is related to the viral vector, but it is a directly caused by the therapy. I don’t know much about the cyctic fibrosis trials but here’s a link to an X-SCID trial subject adverse reaction.

      2. loupgarous says:

        My fault for relying on unreliable wetware, Spike. I was thinking of Jesse Gelsinger, who, developed a fever and blood clots throughout his body within hours of adenovirus-vectored cell therapy for partial ornithine transcarbamylase (OTC) deficiency, and died soon thereafter. Tragically, while Gelsinger had to take 22 pills a day and restricted to a very strict diet, he was not in danger of dying before enrolling in the study – he hoped for successful treatment, but his hopes were ultimately for those with OTC deficiency not as fortunate as he was.

        How I came a-cropper was probably thinking of the trials of adenovirus vectors in gene therapy for cystic fibrosis. My apologies, and thanks for the corrections.

  2. Harrison says:

    I have been cautiously following this expecting it to take a decade or more to reach human trials, but it does seem likely it will happen sooner.

    I wonder how long before this is applied to apolipoprotein E e4/4 mice? It’s by far the largest genetic risk factor for Alzheimer’s disease, it’s just a single amino acid substitution (Cys to Arg at position 112).

    1. Ian Malone says:

      I wonder how long before this is applied to apolipoprotein E e4/4 mice? It’s by far the largest genetic risk factor for Alzheimer’s disease, it’s just a single amino acid substitution (Cys to Arg at position 112).

      Not sure. Things like Duchenne are loss of function mutations, if they can get the right protein produced by 17% of the muscle fibres then that makes a difference. For mutations where the isoform is actually harmful then you’re looking at reducing the amount produced, and 17% reduction doesn’t sound as good. ApoE e4 vs e3 is somewhere in the middle, as the protein does various things, e4 is less good at clearing amyloid beta, it’s pretty straightforward that more clearance is probably better, but it may also play a role in inflammation and to fix that maybe you have to get all (or most) of it. e4/e3/e2 are only risk factors in the end, and we’re still trying to demonstrate disease modification from going after amyloid.

      What might be more hopeful are things like progranulin in FTD or some of the familial / autosomal dominant AD mutations.

  3. Eugene says:

    Over the years I have read enough Science Fiction to know where this is heading. An effective treatment for deleterious mutations and a way to insert advantageous mutations. How far are we from Pre-natal gene editing boutiques?

    1. Martin says:

      As an avid Science fiction reader myself, I thought we were always going to end there… Whether legal or illegal: if the former, for many, if the latter, only for the happy (read: filthy rich) few.
      As soon as it became apparent that we might understand the genome, it was only a matter of when, not if, we would have “designer babies”, since it would be unethical not to try to cure diseases that way, and the underlying principles are the same for both application.

    2. Ken says:

      Don’t forget the other group of stories, where the viral editors are released deliberately or accidentally and spread as a plague.

  4. a says:

    Who’s gonna bring this stuff to market? Time to go shopping for stocks.

    (No seriously, who?)

  5. drocto says:

    This is definitely an ethical minefield, but potentially a much needed relief to people suffering from these terrible disorders.

  6. dams says:

    Soon mouse will be a disease free species… #Mickeywilloutliveallofus

    1. Dr. L. A. Brat says:

      Maybe the NHI budget is better directed towards efforts to mousify man?

  7. alex says:

    I appreciate the great advances that have occurred in the medical field. i would be more happy if these advances are commectricailized to help more people. i think i can easily donate DNA if it can help someone/

  8. Biotechie says:

    Actually despite the Hollywood sci-fci scenarios, there are not many instances where gene editing of embryos makes sense. This is because preimplantation genetic diagnosis (PGD) can be used to screen out embryos with a propensity for disease. Here is one article laying this out but interested readers should go to the NAS or Nuffield Council in the UK to find out more.

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