Hey, remember a while back when I mentioned how scientists used evolution (random mutation and natural selection) in the lab to ‘perfect’ gene therapy vectors?

Theyve just one it again! In a BIG way!

Molecular Evolution of Adeno-associated Virus for Enhanced Glial Gene Delivery.

These folks were looking for a way to get man-made viruses to deliver therapeutic genes to infected/defective cells. Now, you dont want these viruses to infect EVERY cell of a sick person, necessarily, you want to target the specific, sick cells. A golden target for these folks is glial cells– as their malfunctions have been connected to basically every neurodegenerative disease you can think of– Lou Gehrigs Disease, Parkinsons, Huntingtons, MS, hell, even migraines.

A barrier to any putative gene therapy for these diseases has been the fact we dont have adeno associated viral vectors that infect glial cells. In the lab, you can dump a LOT of virus on astrocytes, and some of the serotypes will infect them, but that just doesnt work in vivo.

Why not let evolution make you an adeno associated virus that likes to infect glial cells, and only glial cells?

STEP 1: MUTATION
The paper I talked about before? They randomized their viral vectors by shuffling bits of the structural protein around. It was a random as they could get it.

This paper, they made things even MOAR random! “… random mutagenesis, DNA shuffling, AAV peptide display, and a new semi-random loop replacement method…” AHHH! Thats some mighty fine random!

STEP 2: NATURAL SELECTION
They put this super-random mixture of viruses on primary astrocytes from human donors. Some variants wont get in at all. Some will get in kinda, and some will kick ass.

The ones that get in kinda/great will produce lots of babby viruses.

Take the babby viruses, put them on new astrocytes.

Take THEIR babby viruses, put them on new astrocytes.

Koerbe et al only had to passage their babby viruses 4-5 times to get a population of viruses that LOVED astrocytes!!

STEP 3: DOES IT WORK IRL?
Koerbe took their astrocyte-specific viruses and gave them a gene to deliver, GFP. So when they injected these viruses into rats, they could see where the viruses went by looking for green glowing cells. Result? Lots of green in the rat brains (even though human cells were used for selection).

Bonus: You dont really need to know how this is working for it to work. As in, we dont need to figure out each and every step in adeno associated viruses life cycle and selection pressures at each stage and intelligently design viral vectors that can avoid those challenges. We can let evolution do all the work for us, and we can figure out the details later!

Importantly, while a lack of detailed mechanistic knowledge of specific gene delivery barriers in this and other cases can preclude rational design strategies to improve vector properties, a broader “black box” approach to engineer efficient AAV vectors through random diversification and high-throughput selection can still succeed.

You know, like, when you had to do group projects in elementary school? And whoever was the ‘smartest’ in the group ended up doing all the work, but everyone else got a slice of the A? Evolution is smarter than us. Lets let it do all the work, and we can take credit for the new therapies it figures out.

In summary, by employing directed evolution with a diverse array of novel AAV libraries including a new peptide loop replacement library, we engineered novel AAV vectors capable of highly efficient delivery to astrocytes in vitro and importantly to astrocytes and Müller glia in vivo….
… modulating mutant superoxide dismutase-1 expression within astrocytes may reduce neuronal cell death in amyotrophic lateral sclerosis, or therapeutic gene delivery may prevent the accumulation of amyloid beta-plaques in Alzheimer’s disease. Furthermore, because Müller cells span the entire retina and surround every class of neuron present within this tissue, transduction of Müller cells would permit spread of neurotrophic factors throughout all layers of this tissue to significantly augment existing therapies for retinitis pigmentosa, age-related macular degeneration and neovascularization.

Comments

  1. #1 red rabbit
    September 3, 2009

    Too much coolness for one project. I love how the more we learn, the lazier we can get.

  2. #2 Divalent
    September 3, 2009

    Yeah, well they weaseled snuck the information into the experiment in the glial cell, which (as we all know) God an intelligent designer designed, so God an intelligent designer did it.

    Really cool.

  3. #3 clinteas
    September 3, 2009

    Don’t you in most, or at least some of these neurodegenerative diseases, rather need one or more genes switched off, rather than a new one introduced?

    Wiki gives 2 possible explanations:

    # An abnormal gene could be swapped for a normal gene through homologous recombination.
    # The abnormal gene could be repaired through selective reverse mutation, which returns the gene to its normal function.

    Hm, I dont know enough about those to comment, maybe someone could shed some light?

  4. #4 rrt
    September 4, 2009

    Did they do any work on making sure the virus only infected the target cells? I’d be interested in seeing how they went about it.

  5. #5 Sili
    September 4, 2009

    I think I was think that as well, rrt. But it sounds as though the brains lit up specifically in the rats. Presumably the GFP’d be more scattered, if more than glial cells had been infected.

    Also: hey! I resemble that remark!

  6. #6 Uncephalized
    September 4, 2009

    Cooooooooool…

    And yet another nail in the coffin for ID/creationism–they’re running out of things to claim evolution can’t do. Of course, everyone’s known they’re lying for a while and they still do it, so…

  7. #7 snoeman
    September 4, 2009

    But waitaminute, I thought that Dr. Egnor, neurosurgeon and Disco Institute flack, had incontrovertibly established that evolution has made absolutely no contributions to, or is useless for, medical science. Surely he’ll reconsider his position now, right?

  8. #8 cakeforme(mo)
    September 4, 2009

    When I was in elementary, I would suggest things to my group but they would just ignore me and say I was stupid. And in the end the teacher would say the result was fine, but it would have been better if they had done it like I suggested. Not that this would have helped me in the next group project. :(

    @ that wouldn’t work with the used virus (AAV) because it always integrates in a specific region of the genome. For Gene-targeting in the DNA like it is required in these two methods you need other transfer methods and they are to inefficient do work in vivo. But RNAi can also switch off gene function, and this can hypothetically work with that system.

    http://en.wikipedia.org/wiki/RNAi

  9. #9 anevilmeme
    September 5, 2009

    But, but, but evolution “just a theory.”

  10. #10 gillto
    September 8, 2009

    Take home lesson: evolution works in mysterious ways.

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