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The use of zinc fingers in permanent gene therapy


Highly efficient endogenous human gene correction using zinc fingers
Aaron Klug Scientist
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A number of groups, two groups in fact: a group at Utah began using zinc finger nuclease and I'll explain those in a minute, to... to introduce mutants in drosophila. A man called Dana Carroll. Now he used the zinc finger nuclease developed by a man called [Srinivasan] Chandrasegaran who worked with Jeremy Berg. Now what he did, he made a zinc finger nuclease which was, had no specificity other than the DNA, other than the zinc finger part of recognising the DNA. This he took from the catalytic domain of restriction enzyme called FOCII which he had been studying. And he showed that this... that his targeting depended purely on the zinc finger which he used. Now the early work was done making three zinc fingers and it was used by a man called Matthew Porteus in David Baltimore's lab, to show stimulation of gene editing or gene correction in a model system using GFP protein. And they got factors of sort of modest ten, a factor of ten or something like that, 20 depending on what they did. Now, these were fairly home-made zinc fingers which he made himself. As I've said before you could write down the sequence of a zinc finger from all the published work which would bind the two sequences enough, what's the word? Redundancy that almost anything will bind. So it wasn't... it wasn't optimised in any way, not very efficient but it showed that you could stimulate... multory combination this way, or gene replacement. Gene replacement is what it is but it's not the whole gene, so gene editing is the best word or gene correction. I think editing is the best word of all but they tend to call it correction.

Another group was the group of Dana Carroll who I mentioned were they made their own zinc fingers and using Chandrasegaran catalytic domain, they made mutants of drosophila and they made fingers of various kinds... nothing was under control. Many of the mutants they did died, but those that survived they were able to select on the phenotype. So they had some success with that, making mutants in drosophila. So these are model experiments but now if you want to target into a patient you have to make sure there's nothing hanging over you... you can't start having selection, you have to do it in one go. And it must be absolutely targeted uniquely. So... what Sangamo... there's a group led by Michael Holmes, under Philip Gregory, these are the people... Holmes, Fernhoff and Gregory and they published a paper, it appeared in Nature on June the 5th this year. It was on-line in April and the paper was sent in last November. I had a hand in writing that paper, I don't appear as an author but I'm thanked for reading the manuscript. I was thanked for a critical reading of the manuscript, but critical this would be... in fact I wrote quite bits of it, but never mind. I think Nature editing. So it had quite a long history and the reason was I discovered that Nature were very... careful, there were very worried about publishing a paper that was zinc correction, so soon after the death of the French children, the editor clearly didn't understand that this was something different as well in gene addition and because they want to make the paper short they wanted to cut the preamble where this was explained and so on. But anyway, it appeared in Nature and they consulted quite a lot of other people, I know this because people have told me about this. So that's a great step forward and the paper's entitled Highly Efficient Correction of Endogenous Gene using Zinc Fingers [sic] or something rather like that, that's what it is. It's zinc finger nucleases. So it's a great triumph and what was done was this.

Born in Lithuania, Aaron Klug (1926-2018) was a British chemist and biophysicist. He was awarded the Nobel Prize in Chemistry in 1982 for developments in electron microscopy and his work on complexes of nucleic acids and proteins. He studied crystallography at the University of Cape Town before moving to England, completing his doctorate in 1953 at Trinity College, Cambridge. In 1981, he was awarded the Louisa Gross Horwitz Prize from Columbia University. His long and influential career led to a knighthood in 1988. He was also elected President of the Royal Society, and served there from 1995-2000.

Listeners: John Finch Ken Holmes

John Finch is a retired member of staff of the Medical Research Council Laboratory of Molecular Biology in Cambridge, UK. He began research as a PhD student of Rosalind Franklin's at Birkbeck College, London in 1955 studying the structure of small viruses by x-ray diffraction. He came to Cambridge as part of Aaron Klug's team in 1962 and has continued with the structural study of viruses and other nucleoproteins such as chromatin, using both x-rays and electron microscopy.

Kenneth Holmes was born in London in 1934 and attended schools in Chiswick. He obtained his BA at St Johns College, Cambridge. He obtained his PhD at Birkbeck College, London working on the structure of tobacco mosaic virus with Rosalind Franklin and Aaron Klug. After a post-doc at Childrens' Hospital, Boston, where he started to work on muscle structure, he joined to the newly opened Laboratory of Molecular Biology in Cambridge where he stayed for six years. He worked with Aaron Klug on virus structure and with Hugh Huxley on muscle. He then moved to Heidelberg to open the Department of Biophysics at the Max Planck Institute for Medical Research where he remained as director until his retirement. During this time he completed the structure of tobacco mosaic virus and solved the structures of a number of protein molecules including the structure of the muscle protein actin and the actin filament. Recently he has worked on the molecular mechanism of muscle contraction. He also initiated the use of synchrotron radiation as a source for X-ray diffraction and founded the EMBL outstation at DESY Hamburg. He was elected to the Royal Society in 1981 and is a member of a number of scientific academies.

Tags: Sangamo Therapeutics, Highly efficient endogenous human gene correction using zinc fingers, Dana Carroll, Srinivasan Chandrasegaran, Jeremy Berg, Matthew Porteus, Michael Holmes, Philip Gregory

Duration: 4 minutes, 38 seconds

Date story recorded: July 2005

Date story went live: 24 January 2008