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Dr. Jen Phillips @ClutchScience
, 16 tweets, 4 min read Read on Twitter
Great work as usual by @edyong209 detailing the troubling missteps of the germline #CRISPR editing story of last week. Ed's #3 leaves a lot more to unpack, though, so here's a thread:
theatlantic.com/science/archiv…
Some have interpreted He’s work as a (failed) attempt to replicate the natural 32bp deletion, but I don’t think that was the goal. I think he just hypothesized that any old indel in this genetic region would get the job done and went for it. 2/
Obviously there’s a whole host of things to be concerned about here, but this approach was shockingly cavalier even within the limited scope of what’s known about loss of CCR5 function. 3/
No other natural mutations have been studied to the extent that ∆32 has been, given its prevalence in the European gene pool. This deletion results in a frameshift and a premature stop after 31 missense residues. 4/
The truncated protein is synthesized, but the membrane targeting motif is missing so it’s sequestered in the ER.

jbc.org/content/277/3/…
One of Nana’s mutations is in the same reading frame as ∆32, thus truncates in the same spot and has and most (not all) of the same missense residues after the shift. Her other allele shifts into the 2nd alt reading frame and truncates a different, shorter, run of missense. 6/
(this is all nicely illustrated by @RyderLab here) 7/
There’s one report that protein translation of the ∆32 variant is necessary for HIV resistance activity, even though the protein doesn’t make it out of the ER. 8/
ncbi.nlm.nih.gov/pubmed/17522201
We don’t know whether either of Nana’s alleles, especially the more novel of the two, would have the same stability. If not, it would not be expected to provide the same infection resistance. 9/
@edyong209 describes Lulu’s genotype correctly as having one unaffected (wild type) allele and one altered one, but what Ed didn’t go into is that Lulu’s single alteration is in-frame, which is concerning. 10/
5 amino acids are deleted, but then the code picks up again on the other side of the gap. We’d predict an almost full-length transcript that has all the right stuff at the C-terminal end for the resulting protein to traffick to the membrane. Then what? 11/
JK He hypothesized that the deleted residues would inhibit HIV binding, but we don’t know if this is true. Moreover, maybe the new shape would make it more likely to bind inappropriately to something entirely different. 12/
There are all kinds of ways it could play out inside the cells of this human being. It is a terrible travesty that all we can do is speculate about this. 13/
Nothing is 100% in medicine--or indeed, in life. There will always be uncertainties and variable risks for any given patient, but this is far, far beyond a standard risk-benefit analyses. 14/
CRISPR will be an amazing tool for treating human genetic diseases. I still believe that, and I know it's a dream shared by many of my colleagues as well as the families I work with. It's beyond devastating that the first realized application has gone so horribly awry. 15/15
Addendum: I stress that the takeaway here isn't 'so many things are unknowable, ergo we should never do this'. Standard experimental procedures with cell & animal models could have addressed every one of the issues laid out upthread. The researchers chose not to do any of them.
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