AMA on CRISPR

[Evaluate]

An AMA (ask me anything) happened on CRISPR which had some interesting discussions, some very science-y.  It was done by Dr. Paul Knoepfler.  Unfortunately herpes wasn't brought up but it talked about the current trials being done on the eye.  I wasn't too please with some of the responses I saw though (not necessarily from the doctor, but from other contributors), because it said the eye was essentially more simple than another body part because it is self contained.  Perhaps I'm missing something in the discussion though.

https://www.reddit.com/r/science/comments/3vs7te/science_ama_series_im_dr_paul_knoepfler_back/

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One of the major problems with using viruses as vectors is that we still can't really direct where they insert genetic material. Viruses like HIV and Herpes Simplex are relatively random in terms of their insertion sites, which means that they could very easily disrupt another critical gene in some of your cells by inserting their genome within the reading frame of that gene. Or, they could insert it in an untranslated region, meaning that the proteins they encode would never see the light of day.

We've somewhat worked around that by using a specific type of Adenovirus which has a far more specific insertion site in the human genome, but it's still not perfect.

Even if we could get a retrovirus to propagate genetic material throughout our entire system, there's still a good chance that the virus would simply revert to a virus which is harmful. It's often difficult to separate the dangerous parts of viruses from what makes them function as viruses. By retaining the properties we need to make them self-replicate and infect our cells, we're also bound to inadvertently keep (or be forced to keep) some basic viral properties, which could end up causing disease.

So basically we still don't really know what we're doing yet, especially with whole-genome modification. It's something worth looking in to, but it's far FAR away from clinical applications.

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Further, there are new efforts such as crowdfunding to make CRISPR kits available to the general public. George Church has said that he thinks someone could set up a CRISPR lab in their garage for $2,000.

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Remember though that the eye is a very special organ when it comes to gene therapy. It is immuno-privileged and much easier to access for would be vectors. I would temper expectations for the future of gene therapy based on only results obtained in the eye.

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This is a great point about the uniqueness of the eye. Other tissues and organs would be far more difficult.

 

Edited December 8, 2015 by Evaluate

[dont quit!]

The only viral vectors that really make sense at this point are based on adeno-associated virus (AAV), which has been successfully used in gene therapy trials in humans. The big advantage of AAV is that you can get very high levels of virus—up to 10 billion infectious units per milliliter—and the Bloom lab has clearly shown, using an AAV that expresses green fluorescent protein (gfp), that he can infect essentially every single neuron in the trigeminal ganglia where HSV-1 establishes latency.   http://sites.duke.edu/cullenlaboratory/current-research/herpes-research/update-on-hsv-research/        In Cullens own words

Edited December 8, 2015 by dont quit!

[Evaluate]

Thanks for the reply. Cullen believes they have a replication-incompetent vector, which would stop the virus from continuing to replicate in the body. Do you know if it's possible for the virus to mutate back to being replicative?

Also, is there an issue to overcome with application? For example, the posts above suggest the eye is immino-privileged (which Google says stops it from eliciting an inflammatory immune response). I'm not sure how important this is in the process of curing herpes. Does this matter?

 

 

[dont quit!]

19 minutes ago, Evaluate said:

Thanks for the reply. Cullen believes they have a replication-incompetent vector, which would stop the virus from continuing to replicate in the body. Do you know if it's possible for the virus to mutate back to being replicative?

Also, is there an issue to overcome with application? For example, the posts above suggest the eye is immino-privileged (which Google says stops it from eliciting an inflammatory immune response). I'm not sure how important this is in the process of curing herpes. Does this matter?

 

 

Im not an expert but I doubt it. Its like blowing through a motor and transmission in a cheap Ford and being repaired by your drunk uncle who is blind from one eye. But again, I'm no expert. From what i understand the DNA gets clipped by the CRISPR-CAS 9 which is guided by sgRNA; the DNA gets repaired but should mutate the extranuclear viral DNA; when this happens it should inactivate. Picture a scar in the viral DNA that simply doesnt work anymore. I can see some discomfort occurring while getting treatment but those are questions that will be answered during clinical trials.

Edited December 8, 2015 by dont quit!

[dont quit!]

Also HSV is hidden to the immune system while in latency so it probably wont cause inflammation.

[Evaluate]

I'm trying to read between the lines.  The AMA was heavily based on gene editing for GMO humans, which I assume is much more complex than ridding a virus.  Also not everyone who posted on the AMA is an expert, so I'm taking it for face value.  Brings up some interesting points though.

[dont quit!]

11 minutes ago, Evaluate said:

I'm trying to read between the lines.  The AMA was heavily based on gene editing for GMO humans, which I assume is much more complex than ridding a virus.  Also not everyone who posted on the AMA is an expert, so I'm taking it for face value.  Brings up some interesting points though.

Most of their opinions are in actually replacing a gene not clipping DNA inactive. Thats the difference i.e. HIV which is a retro virus that embeds its dna into the cells dna.

[Evaluate]

The one individual lumped HIV and HSV into the same category, although he isn't a recognized contributor on r/science.  I'm not sure if he assumes HIV and HSV are both retroviruses or if there are is another similarity he is referring to with random insertion sites.

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Viruses like HIV and Herpes Simplex are relatively random in terms of their insertion sites, which means that they could very easily disrupt another critical gene in some of your cells by inserting their genome within the reading frame of that gene.

 

[Sanguine108]

On 12/8/2015 at 5:16 PM, dont quit! said:

The only viral vectors that really make sense at this point are based on adeno-associated virus (AAV), which has been successfully used in gene therapy trials in humans. The big advantage of AAV is that you can get very high levels of virus—up to 10 billion infectious units per milliliter—and the Bloom lab has clearly shown, using an AAV that expresses green fluorescent protein (gfp), that he can infect essentially every single neuron in the trigeminal ganglia where HSV-1 establishes latency.   http://sites.duke.edu/cullenlaboratory/current-research/herpes-research/update-on-hsv-research/        In Cullens own words

Well, that's the naked trigeminal nerve in a petri dish exposed to AAV...

I mean, if there was NOTHING to be worked out and everything works so smoothly then I don't understand why they haven't released data on their potential next experiment to expose the trigeminal nerve of rats/mice to this and see how well it does.    I understand things take time but I got hsv in the beginning of April and have been following this thinking that it'd be such an easy thing to just do and publish.   Maybe it has already been done and he has spoken about it at the latest conference he was at.   Either way, I'd love to know because what's left of my hope for something better would increase dramatically.   It's ok though.  I'm just having some issues and and tired of it all. <3

 

[Herbfresh]

Didn't they JUST save a young girls life by using CRISPR to cure an incurable and rare form of cancer?

CRISPR sounds cool, and potentially offers us a cure very very soon? 

[Sanguine108]

11 hours ago, Herbfresh said:

Didn't they JUST save a young girls life by using CRISPR to cure an incurable and rare form of cancer?

CRISPR sounds cool, and potentially offers us a cure very very soon? 

That's cool.  do you have a link?

I wouldn't mind if they started curing these chronic viral infections like hsv so downstream garbage like cancer wouldn't be a thing.  But whatever.  I'm a little bitter at the whole way things are and would like for a deeper change.

[Herbfresh]

1 hour ago, Sanguine108 said:

That's cool.  do you have a link?

I wouldn't mind if they started curing these chronic viral infections like hsv so downstream garbage like cancer wouldn't be a thing.  But whatever.  I'm a little bitter at the whole way things are and would like for a deeper change.

Sure thing! 

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In what’s being billed as a medical first, scientists in London have used a new kind of biological scissors in a last-ditch effort to cure a child with otherwise untreatable leukemia. They edited genes in immune cells to make them hunt down and destroy the malignant blood cells that threatened the year-old girl’s life.

The experimental procedure was performed in June at Great Ormond Street Hospital, physicians there announced on Thursday. The hospital said it was the first time that a genome-editing technology called TALEN had been used to treat a patient.

TALENs is one of three genome-editing tools that scientists and companies are racing to test in humans as possible treatments for everything from cancer to cystic fibrosis, sickle cell disease, and Huntington’s disease.

 

 

http://www.statnews.com/2015/11/05/doctors-report-first-use-gene-editing-technology-patient/

CRISPR created T Cells that destroyed the cancer cells. It worked. So I don't see why they can't use it to create designer T Cells that recognise HSV proteins and essentially be a functional cure. I hope we can be cure in within the next few years! We could all be happy and normal again! 

The tech is here! And it seems to actually work! I too am very bitter. I want a cure, very very soon. If CRISPR technology is so cheap and easy to replicate, I hold strong hope that it will indeed be used to functionally cure HSV. Can you think of any reason it wouldn't?

Hell, we could crowd source our money, build a crisp lab, and hire a doctor to calibrate it. CRIPR lab can be made for $2000. That's nothing. If a bunch of us sufferers pooled together, and hired a scientist to handle the technical aspect, we could be cured! I know its far fetched at the moment. Just nice to know relief might be close. 

 

 

Edited December 28, 2015 by Herbfresh

[Herbfresh]

There are companies right now, that offer gene editing services. That's insane! If its possible to modify T Cells to attack HSV, then we should be cured pretty soon. Wow, the more I read about it, and the fact its right her, right now, this very moment, available to anyone with the money to use it, the more I feel like we are on the cusp of potentially being cured. 

This is what I was waiting for, some crazy new tech advancement that means to hell with waiting 30 years for a new vaccine that may or may not work. 

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At GeneCopoeia, we provide products and services for both CRISPR and TALEN. Our genome editing begins with expert design of TALEN and CRISPR vectors in both non-viral and lentiviral formats. In addition, we offer donor plasmid design for HR mediated applications. Plasmid design is followed by construction and delivery of ready-to-use, sequence verified plasmid DNA. Further, we provide more sophisticated genome editing services, such as target site validation, and use of genome editing technology for stable cell line and transgenic mouse line generation. We also offer scientific consulting services, with which you can take advantage of our extensive knowledge and experience in the field. We will help you choose the option most appropriate for your experiments, by informing you of the differences described in this document as well as others, and work with you to devise sound genome editing strategies.

 

What do the scientists make of all this? Is this the biggest development, breakthrough in a cure for us? I am no scientist, so i would like for someone to tell me why gene editing would not be able to create killer T Cells that attack HSV. Seems like that should be even easier than curing Leukaemia. 

Is it time we get excited? 

[Sanguine108]

I don't remember what immune cell reaches nervous tissue... But by and large, the nerves of the body are off limits to the immune system.  T cells don't see the nerve roots. (generally speaking).

That's why labs like Cullen and Bloom are using a neurotrophic viral vector (AAV) because you need a virus that goes to the same places that HSV goes and have that vector unload the CRISPR and CAS9 TALENs whatever to do the editing.

Edited December 29, 2015 by Sanguine108

[Herbfresh]

1 hour ago, Sanguine108 said:

I don't remember what immune cell reaches nervous tissue... But by and large, the nerves of the body are off limits to the immune system.  T cells don't see the nerve roots. (generally speaking).

That's why labs like Cullen and Bloom are using a neurotrophic viral vector (AAV) because you need a virus that goes to the same places that HSV goes and have that vector unload the CRISPR and CAS9 TALENs whatever to do the editing.

 

I see, so Cullen and Bloom are going for a complete cure that will clear all the HSV out. That is fantastic. What do you think of what they are doing? It appears to be incredibly promising, right?

The thing is, if they used CRISPR to edit T Cells so they cannot be high jacked by HSV, or edit the T Cell to easily recognise HSV, wouldn't this be a functional cure? HSV might still be present in some nerve cells, but it would pretty much not matter since they would not be able to reproduce. But if they can get to the source even better. 

I just hope this all doesn't take too long. 

Edited December 29, 2015 by Herbfresh

[Sanguine108]

Well, I'm reserving my excitement for if and when I see results for a rat/mouse infected with HSV and then their method works perfusing the entire nerve and doing what it says it does from the isolated, in the petri dish.

As far as I know, and I know very little... But one of the ways HSV evades the immune response is HSV prevents MHC (major histocompatibility Complex) from expressing 'abnormally'.  MHCs are surface proteins for cells that tell the immune system (like T cells) that the cell is healthy or sick.   When a virus or bacteria afflict a cell then MHCs express themselves and some immune cell comes by to read it, if it's abnormal then tags it for destruction/apoptosis/or whatever.   

But maybe it would do something that isn't clearly obvious right now.  All the theory is just theory.   You have theorheticians and empirical scientists.  A lot of theory is garbage.  But, a lot of theory is a starting point for the empirical scientist.  

Anyhow, another evasive aspect of HSV is the surface proteins on the virus.   So you have these research groups removing surface proteins to see if that will help the immune system recognize the free floating virus better. 

[brookeb300]

I'm sorry I'm like this but I guess I just need to vent.  If I was still clean and disease free and I knew what I know today I would never in a million years date someone with herpes.  I wouldn't even really want to be friends with them.  sorry just being honest.

[LillianPanos]

Really! keep the stigma going

 

45 minutes ago, brookeb300 said:

I'm sorry I'm like this but I guess I just need to vent.  If I was still clean and disease free and I knew what I know today I would never in a million years date someone with herpes.  I wouldn't even really want to be friends with them.  sorry just being honest.

 

[Sanguine108]

6 hours ago, brookeb300 said:

I'm sorry I'm like this but I guess I just need to vent.  If I was still clean and disease free and I knew what I know today I would never in a million years date someone with herpes.  I wouldn't even really want to be friends with them.  sorry just being honest.

A resurrected thread from last year!   ...While I'm a bit more compassionate than that, I do understand where you're coming from.  Before having hsv, I knowingly had friends with herpes but it took a fucking lying snake of one that gave me herpes.  As much as I try to forgive and explore believing in acceptance I still relapse into anger, frustration and sadness.   I think those that don't have lingering, persistent symptoms are very fortunate and they shouldn't naively extrapolate their experience with HSV to others that are actually suffering.  Of course, we have the token, delusional one who thinks everyone's pains and whatnot are 100% unrelated to herpes.  

In the end, I'd avoid piece of shit people. They're spread out among all the crowds, not just those with hsv.

Today, I have a sore on my lip with this crawling/tingling lingering, heavy back/sacral/lumbar(spine) pain.   Going through my days, I want to die. I want it over with.  I would never take my own life but welcome an event that would end this one.  So, fuck herpes! I'd never wish this upon anyone.  I'm sorry if anyone thinks this enhances the "stigma".  There are plenty of people who don't give a damn about the stigma and, if the stigma is a mental block, then that's all that it is to them.   If you're one of those who actually suffer with symptoms then let it out.   This is a herpes support and whatnot forum.  

As far as the stigma... People obviously have personal issues going on and/or have been historically hurt if they are "triggered" by the "stigma".   The only times I've ever thought of this "stigma" is when people bring it up.  I see people complain and think of their suffering, not their pro-stigmatic agenda.   My mind is more on preventing others from enduring this type of physical suffering.   But I guess that's a part of the stigma.  People suffering from their own mental or physical pain are inadvertently offending others' with stigma goggles on.

Take the stigma goggles off so you can be more supportive.

[Terrell]

6 hours ago, brookeb300 said:

I'm sorry I'm like this but I guess I just need to vent.  If I was still clean and disease free and I knew what I know today I would never in a million years date someone with herpes.  I wouldn't even really want to be friends with them.  sorry just being honest.

If you have hsv1 oral you are a herpes infected person so u shouldn't wanna be around yourself because 90% has it.

[fixme1]

so the 5 people i work with who get oral hsv shouldnt be dated if we go by brookes reply. we know you are upset but at the end of the day everyone is human, if we go by that rule. you might aswell discriminate people with other diseases or illnesses

[cantdoit]

I feel the same as Brooke except that I would never not be friends who someone who had hsv but I would not choose to date someone who had hsv.  I would date someone who has cancer however. Hsv is an incurable Std which is why if I knew someone had it I wouldn't take the chance of contracting it. 

[brookeb300]

Idk I haven't had it that long.  It's a lot worse than I thought it would be.  I wish I could go back and not get it.   But I know I was too harsh.

[Lisajd]

1 hour ago, brookeb300 said:

Idk I haven't had it that long.  It's a lot worse than I thought it would be.  I wish I could go back and not get it.   But I know I was too harsh.

What are your symptoms.  @cantdoit why would you date someone with cancer which can also be incurable

[brookeb300]

Idk crappy symptoms a lot of the time.  lol cancer isn't contagious at least.