AI transcript
0:00:02 Hi and welcome to the inaugural edition
0:00:04 of the A16Z BioJournal Club.
0:00:05 I’m Hannah.
0:00:08 Our goal here is to take an interesting new research paper
0:00:10 in the field and talk about why it’s cool,
0:00:12 break down a little of the science involved,
0:00:14 and consider what the implications of this research
0:00:15 for industry might be.
0:00:17 So in our first take, A16Z General Partner
0:00:21 on the Bio Fund Jorge Conde and Deal Team Partner Andy Tran
0:00:24 chat with me about two papers recently published.
0:00:27 The first transposon encoded CRISPR/CAS systems
0:00:30 direct RNA guided DNA integration
0:00:33 was published by a group under Samuel H. Sternberg
0:00:36 at Columbia in nature of June 2019.
0:00:39 The second is RNA guided DNA insertion
0:00:42 with CRISPR associated transposases
0:00:45 with a team under Feng Zhang from the Broad Institute
0:00:48 published in science also in June 2019.
0:00:50 We talk about what these papers are all about
0:00:53 in the field of CRISPR development and beyond.
0:00:54 This mini podcast is available
0:00:57 as part of our new A16Z Bio Newsletter.
0:01:00 So if you like it and you want to hear more or read more,
0:01:05 please sign up for the newsletter at a16z.com/subscribe.
0:01:07 Let’s talk about what specifically is happening here.
0:01:10 What does, what a transposon encoded CRISPR/CAS system
0:01:13 direct RNA guided DNA integration?
0:01:14 Like, what does that actually mean?
0:01:16 Can you help me understand what was the interesting science
0:01:17 that was going on here?
0:01:18 – Yeah, so what’s really interesting
0:01:19 in the field of CRISPR lately,
0:01:22 and actually a few papers that came out in the recent times
0:01:26 developed a way to basically use these transposon machinery
0:01:28 inside the cell and use CRISPR to direct it
0:01:30 into a specific place in the genome
0:01:34 and really edit the genome without cutting it open at all.
0:01:36 So you can think of it as this scarless type
0:01:37 of genetic modification.
0:01:39 – Essentially what a transposon is,
0:01:42 is this sort of phenomenon that’s been observed
0:01:44 of where you have sort of these genes
0:01:46 that jump into the genome.
0:01:47 – It’s kind of mysterious what the jumping
0:01:48 is really used for.
0:01:50 People surmise that, you know, potentially, you know,
0:01:54 it helps, you know, the cells and organism evolved in general.
0:01:56 So what this paper shows is to utilize
0:01:58 this CRISPR machinery known as Cascade,
0:02:02 and it’s formed by, you know, CASS6, CASS7, CASS8 proteins
0:02:04 to really insert genes into the genome.
0:02:08 And this Cascade protein is directed to the chromosome
0:02:11 by using guided RNA, you know, CRISPR machinery,
0:02:15 and it then binds to this transposase-associated protein,
0:02:18 TNIQ, and it allows them to recruit
0:02:20 these transposable elements and then effectively integrate
0:02:22 the gene into the genome.
0:02:25 And this is super powerful because now we’re able to,
0:02:27 you know, add genes in the genome directly
0:02:29 and precisely without cutting it open.
0:02:32 It’s a scarless way to modify the genome.
0:02:35 – So in some ways, this is almost like if CRISPR
0:02:38 is about surgically inserting or editing DNA,
0:02:41 this is almost in many ways like plastic surgery, right?
0:02:42 It’s scarless. – Ah, right, right.
0:02:45 – It doesn’t leave a mark and therefore in some ways
0:02:48 it’s less risky from an intervention standpoint
0:02:49 on the genome side.
0:02:51 – What it really boils down to is that
0:02:53 this machinery utilizes a way
0:02:56 to really efficiently integrate genes into the cell,
0:02:58 into the genome specifically,
0:02:59 without having to cut it open at all.
0:03:02 And to the metaphor of the surgery,
0:03:04 this is really important in the whole context
0:03:06 of gene therapy as a whole, right?
0:03:08 ‘Cause when we started off in gene therapy,
0:03:09 we had these random integration
0:03:12 of these trans genes into the cell.
0:03:15 And, you know, this was a more stochastic process.
0:03:17 So think of it almost as, you know,
0:03:20 the arrival of this paper and papers like it
0:03:24 are showing us that we’ve gotten to a point
0:03:28 where we have a fully programmable ability
0:03:32 to integrate new genes or new DNA into a genome
0:03:34 without having to first open up
0:03:36 or break apart the DNA to do that.
0:03:38 – So let’s go back and actually situate it
0:03:40 into the development of the science,
0:03:42 what it represents for where we’ve gotten
0:03:44 from where we began.
0:03:45 – Yeah, so if you want to do the really,
0:03:47 the fast forward montage version of this.
0:03:49 – Yeah, the 80s montage.
0:03:51 – Yeah, exactly, so I’m sure you’ll put
0:03:52 in the pop music behind it.
0:03:55 – Right, yeah, and the overalls in the paintbrushes.
0:03:56 – He always started in the 60s.
0:03:57 – In the time motion.
0:03:58 – Yeah, let’s go.
0:04:00 – But look, I mean, if you go all the way back,
0:04:02 you know, one of the earliest sort of technologies
0:04:04 that came to the fore was the discovery
0:04:05 of something called the restriction enzyme.
0:04:07 And the restriction enzyme was this ability
0:04:10 to take this protein that could cut DNA
0:04:12 at these predetermined sites,
0:04:15 essentially open up the DNA,
0:04:19 and then you could introduce new genetic material,
0:04:20 and it would eventually integrate randomly,
0:04:23 but it would integrate into that DNA.
0:04:25 That’s what gave us the ability to do
0:04:27 or recombinant DNA technology,
0:04:29 which gives rise to the entire biotech field.
0:04:32 One of the earliest applications of biotechnology is
0:04:35 getting bacterial systems to integrate human insulin
0:04:38 so that we could coax bacteria to make that drug
0:04:41 or human insulin on our behalf, of course, to treat diabetics.
0:04:43 So that starts the whole field.
0:04:45 Now, you know, if you sort of move forward,
0:04:46 the objective always has been,
0:04:51 can you cure disease by repairing
0:04:55 or replacing what’s broken in DNA?
0:04:58 And so the whole field of gene therapy arises
0:04:59 with this idea that, you know,
0:05:02 can we introduce a corrected version
0:05:05 of a non-functional gene in a patient
0:05:06 and have it do the job
0:05:08 that the non-functional gene cannot do?
0:05:12 The first version was just put that gene
0:05:16 into a viral vector, into almost like a delivery vehicle,
0:05:18 introduce that into patient cells,
0:05:20 that gets taken in by the cells,
0:05:22 and the gene just starts to sort of do its job.
0:05:23 It integrates randomly in the genome,
0:05:24 but it does the job it needs to do.
0:05:26 Hopefully it integrates, or maybe it doesn’t,
0:05:28 but it just does the job to compensate
0:05:30 for some mutated gene.
0:05:33 Yeah, so it’s basically almost a parallel support system
0:05:35 that the cell has.
0:05:37 We just had the first gene therapies approved
0:05:39 over the course of the last couple of years.
0:05:42 If you look at the spark therapy, gene therapy drug for,
0:05:44 this rare inherited form of blindness.
0:05:47 And so that was one big advance forward.
0:05:49 Now, that is introducing a full gene
0:05:51 and just hoping it gets taken up by the cell.
0:05:52 Just kind of throw it in the mix.
0:05:54 Yeah, throw it in the mix and it has its own risk.
0:05:56 The original discovery of CRISPR-Cas9,
0:05:59 that system, the way that system works
0:06:02 is by making what’s called a double-stranded break
0:06:03 in the DNA molecule.
0:06:04 So if you think of, you know,
0:06:07 we all remember DNA from high school biology
0:06:10 as sort of the beautiful double helix.
0:06:12 So imagine sort of cutting that double helix in two
0:06:14 and making an end of it, right?
0:06:17 And then, you know, and then putting it back together.
0:06:18 That’s not riskless, right?
0:06:20 And by the way, it’s very well known
0:06:22 and documented that one of the big setbacks
0:06:25 that the gene therapy field had a couple of decades ago
0:06:27 was when they ran a clinical trial
0:06:28 at the University of Pennsylvania,
0:06:31 there was a patient named Jesse Gelsinger
0:06:32 who died after receiving the therapy
0:06:34 just because there’s a lot of risk associated
0:06:37 with introducing, you know, a viral capsid with a gene
0:06:39 into a cell system, into human being
0:06:41 where you could have a catastrophic result
0:06:41 and that patient died.
0:06:44 And that actually put a big pause on how we thought
0:06:47 about developing gene therapies for humans.
0:06:49 But that approach will have
0:06:51 and does have therapeutic potential.
0:06:52 And there are several companies
0:06:56 that are pursuing developing CRISPR-Cas9-based therapeutics.
0:06:58 As that advanced in parallel,
0:07:02 we started to see other gene or genome editing technologies
0:07:03 come to the fore.
0:07:06 The first one was one known as zinc finger nucleases,
0:07:08 which really didn’t have as much uptake
0:07:10 as one would expect
0:07:13 because it’s just very hard to deal with these proteins.
0:07:16 – How zinc fingers work is that you actually use these proteins
0:07:18 to bind onto DNA sequences.
0:07:21 And every time you want to iterate to find a new target,
0:07:22 cost tens of thousands of dollars
0:07:24 in a few months to develop one protein.
0:07:26 – So it’s enormously expensive and labor intensive.
0:07:27 – Exactly.
0:07:28 So it never really caught on like wildfire
0:07:30 in the community, right?
0:07:31 – It’s a very bespoke process.
0:07:34 It’s one of the type of thing.
0:07:36 And then there were other technologies.
0:07:38 There was Talens,
0:07:40 which was a bit better than zinc finger nucleases,
0:07:45 but really the big sort of shift in the field
0:07:47 was the arrival of CRISPR.
0:07:49 – And so when scientists discovered CRISPR,
0:07:52 they noticed that it was just always constant evolutionary
0:07:55 warfare between bacteriophages and bacteria
0:07:56 for billions of years.
0:07:58 – Bacteriophages are viruses.
0:07:58 – Yes.
0:08:03 – And so when these viruses inject their viral genome
0:08:04 into the bacteria,
0:08:06 you can think of it as the bacterial immune system,
0:08:10 whereas they’re able to sense these little snippets
0:08:11 of viral genome,
0:08:13 they would create a vaccination card from it,
0:08:14 and then they’ll put it back into this,
0:08:16 what is known as this CRISPR array.
0:08:18 And this would be like a vaccine card
0:08:20 or a vaccine database, if you will.
0:08:22 So every time it recognizes that foreign viral sequence.
0:08:23 – Right, then it knows what to do.
0:08:25 – It would know how to snip it away, right?
0:08:27 And then people have hijacked this machinery.
0:08:31 So what if we can program this outside of bacteria
0:08:32 and use it in human cells
0:08:35 and program it to sense not viral DNA,
0:08:38 but specific targets in the human genome.
0:08:42 And then then we can program and effectively target
0:08:44 anywhere we want in the cell, right?
0:08:46 – So now bring us forward to today.
0:08:48 So is this new development,
0:08:50 does it mean that the therapeutic potentials
0:08:52 are essentially less risky?
0:08:54 Or are there new possibilities
0:08:56 that we haven’t been able to do before?
0:08:56 Or both?
0:08:59 – Basically, you know, it’s really powerful
0:09:01 because when we talk about the first gen of gene therapy
0:09:03 to the second gen of CRISPR,
0:09:05 this paper really represents this third wave
0:09:07 in this scarless genomic editing.
0:09:10 This is basically genomic surgery at its finest, right?
0:09:13 You know, when we think about laparoscopic surgery
0:09:15 and all these advanced surgery tools,
0:09:16 we want to have, you know,
0:09:19 basically a scarless methodology of doing surgery.
0:09:21 This is a way to really cleanly integrate,
0:09:23 you know, genomic segments into the cell
0:09:25 without even touching, right?
0:09:28 And then this also represents an even broader tool
0:09:31 of, you know, the entire, you know, genomic toolkit landscape.
0:09:33 – The real promise on the near term side
0:09:37 is that we get to a point where you can make
0:09:40 scarless integrations of genetic material into the DNA.
0:09:42 So it’s less traumatic in that regard.
0:09:44 So if you can do that more precisely,
0:09:46 there’s less scar tissue that hopefully is, you know,
0:09:48 a better intervention altogether.
0:09:50 So that holds great promise from a potential
0:09:53 for future therapeutics based on this type of technology.
0:09:55 I think the other thing that’s worth noting
0:09:57 is that in a relatively short period of time,
0:10:01 the programmability of these kinds of systems
0:10:04 has improved dramatically.
0:10:06 So the kinds of things that we could do,
0:10:09 that we can do now with, you know, with CRISPR
0:10:11 based on these kinds of advances
0:10:15 gives us a very broad repertoire and toolkit to work with,
0:10:17 whether it’s for therapeutic applications
0:10:18 or for diagnostic applications
0:10:20 or for any other number of things.
0:10:25 When most people think about CRISPR developing CRISPR
0:10:26 for human health,
0:10:28 they’re thinking about therapeutic applications.
0:10:31 But the reality is there’s also a lot of potential
0:10:32 for diagnostic applications.
0:10:34 So going back to the original discovery
0:10:35 of the CRISPR-Cas system,
0:10:38 this was essentially the memory banker immune system
0:10:40 for bacteria to remember what viruses
0:10:41 had attacked it before
0:10:43 so they could protect themselves going forward.
0:10:47 And the way it does that is by essentially cutting
0:10:49 that viral genetic material.
0:10:52 So it’s ineffective essentially basically, you know,
0:10:56 cutting it off at its Achilles heel, so to speak.
0:10:57 And so as you can imagine,
0:11:00 if you’re hijacking that capability
0:11:01 for therapeutic purposes,
0:11:03 you could also hijack that capability
0:11:05 for diagnostic purposes.
0:11:07 And a way that that could potentially work, for example,
0:11:12 is if you know what bacteria or what virus
0:11:15 or even what, you know, mutation in DNA you’re looking for
0:11:18 in say a human sample, a blood sample
0:11:20 or urine sample or anything like that.
0:11:24 If it’s present, it will get cut by the right CAS system.
0:11:27 You program the CRISPR-Cas system to say,
0:11:31 these are the sequences of genetic material
0:11:33 that I am looking for in the sample.
0:11:34 This is basically the search engine.
0:11:36 Like you’re doing essentially a Google search.
0:11:38 – And you basically just look for the kill switch
0:11:40 to have been activated.
0:11:41 – You just look for the identifier.
0:11:42 So you basically say,
0:11:44 if I want to look in this patient’s sample,
0:11:47 let’s say I want to look for a specific bacteria
0:11:48 or a specific virus
0:11:51 or specific genetic mutation associated with disease,
0:11:53 you can say, if you find the presence
0:11:55 of any one of these sequences,
0:11:56 those are almost like the search terms,
0:11:58 cut them and when you cut them,
0:12:00 you can essentially engineer the system
0:12:02 to send out some sort of a reporter,
0:12:04 a reporter, usually it’s a visual marker.
0:12:06 And so basically if it lights up,
0:12:08 it’s because the CRISPR-Cas system cut the DNA
0:12:10 you told it to look for.
0:12:12 So that has a potential diagnostic application
0:12:13 and you could run that diagnostic
0:12:15 essentially without a lab, right?
0:12:16 ‘Cause it just happens with the biology.
0:12:19 So a whole other kind of new tool, essentially.
0:12:21 – Yeah, and I think the diagnostic applications
0:12:23 for these kinds of technologies
0:12:26 are actually pretty intriguing
0:12:28 because most of the way we do diagnostics
0:12:33 is based on developing some very specific biological
0:12:34 or chemical assays.
0:12:36 So look for something and if so,
0:12:38 have the reaction take place
0:12:40 and one that I can visualize and quantify
0:12:42 or quantitate in some way.
0:12:44 But here you’re just basically letting the biology
0:12:45 do the work for you.
0:12:47 In the CRISPR toolkit,
0:12:51 a lot of the initial applications was using
0:12:53 CRISPR-Cas9, this one nucleus.
0:12:56 And actually the diagnostic application
0:12:57 that Jorge was talking about
0:12:59 was actually using these other nucleas
0:13:01 known as Cas13 and Cas12.
0:13:04 And so all these different CRISPR proteins,
0:13:08 Cas9, 12, 13, X, Y, that we’re continuing to discover
0:13:11 has all different fundamental applications, right?
0:13:12 Even fundamentally changing
0:13:14 what these CRISPR nucleases even do.
0:13:15 It doesn’t even cut anymore.
0:13:17 It can do scarless editing.
0:13:20 And then we can even add different applications on it.
0:13:22 There’s new applications adding,
0:13:24 deaminase is just to do base editing.
0:13:28 So we can really do single base pair resolution editing.
0:13:30 There’s really the final frontier of precision
0:13:32 in terms of genomic modification.
0:13:34 And I think what’s really important is that
0:13:36 we’ve really seen this shift from
0:13:38 when it became this random bespoke science
0:13:40 and really turning into full-fledged,
0:13:42 modified engineering tool.
0:13:44 And this paper that we talk about here
0:13:47 is not only a great representation
0:13:49 of this engineering biology thesis,
0:13:53 but also a pretty huge potential step change
0:13:54 for the field as a whole.
0:13:56 – You can program this to turn genes up and down
0:13:58 as opposed to just editing them.
0:14:01 There’s even work that’s ongoing to use this technology
0:14:03 to image DNA directly,
0:14:04 which is a pretty remarkable thing
0:14:07 because since this is acting locally on DNA,
0:14:10 you can add all kinds of agents to make it imageable.
0:14:12 So therefore, you can observe chromatin
0:14:14 or genome structure directly.
0:14:16 So there’s a lot that can be done with this technology,
0:14:20 and you hear the old adage about the pickaxes for the gold.
0:14:22 And with these toolkits,
0:14:25 I mean, we are quite literally panning for gold here.
0:14:28 I mean, where these things get found,
0:14:31 they’re found in soil and in ocean vents.
0:14:32 – New York City subways.
0:14:34 – The New York City subway.
0:14:37 So these people are quite literally looking in nature
0:14:39 because nature has ingenious ways
0:14:41 to do a lot of the things that we’re trying to do
0:14:45 from an engineering biology or programmable biology standpoint.
0:14:50 And so I think it’s a remarkable moment to take pause
0:14:53 and see how far this technology has come
0:14:55 in a relatively short period of time.
0:14:58 This generation’s recombinant DNA or restriction enzymes
0:15:01 that really gave rise to the biotechnology industry.
0:15:03 I think this tool kit, this CRISPR tool kit,
0:15:06 as we’re describing it and discussing it,
0:15:07 represents sort of the next frontier
0:15:10 for what will happen in biology.
0:15:13 So an incredible development of precision
0:15:15 in what the tools can do and at the same time,
0:15:17 a huge expansion of what that will enable us
0:15:18 to do going forward.
0:15:20 Thank you guys so much for joining us.
0:15:23 Should I say on the A16Z Journal Club?
0:15:23 – No, no, please don’t say that.
0:15:24 – Yeah, okay.
0:15:34 [BLANK_AUDIO]
0:00:04 of the A16Z BioJournal Club.
0:00:05 I’m Hannah.
0:00:08 Our goal here is to take an interesting new research paper
0:00:10 in the field and talk about why it’s cool,
0:00:12 break down a little of the science involved,
0:00:14 and consider what the implications of this research
0:00:15 for industry might be.
0:00:17 So in our first take, A16Z General Partner
0:00:21 on the Bio Fund Jorge Conde and Deal Team Partner Andy Tran
0:00:24 chat with me about two papers recently published.
0:00:27 The first transposon encoded CRISPR/CAS systems
0:00:30 direct RNA guided DNA integration
0:00:33 was published by a group under Samuel H. Sternberg
0:00:36 at Columbia in nature of June 2019.
0:00:39 The second is RNA guided DNA insertion
0:00:42 with CRISPR associated transposases
0:00:45 with a team under Feng Zhang from the Broad Institute
0:00:48 published in science also in June 2019.
0:00:50 We talk about what these papers are all about
0:00:53 in the field of CRISPR development and beyond.
0:00:54 This mini podcast is available
0:00:57 as part of our new A16Z Bio Newsletter.
0:01:00 So if you like it and you want to hear more or read more,
0:01:05 please sign up for the newsletter at a16z.com/subscribe.
0:01:07 Let’s talk about what specifically is happening here.
0:01:10 What does, what a transposon encoded CRISPR/CAS system
0:01:13 direct RNA guided DNA integration?
0:01:14 Like, what does that actually mean?
0:01:16 Can you help me understand what was the interesting science
0:01:17 that was going on here?
0:01:18 – Yeah, so what’s really interesting
0:01:19 in the field of CRISPR lately,
0:01:22 and actually a few papers that came out in the recent times
0:01:26 developed a way to basically use these transposon machinery
0:01:28 inside the cell and use CRISPR to direct it
0:01:30 into a specific place in the genome
0:01:34 and really edit the genome without cutting it open at all.
0:01:36 So you can think of it as this scarless type
0:01:37 of genetic modification.
0:01:39 – Essentially what a transposon is,
0:01:42 is this sort of phenomenon that’s been observed
0:01:44 of where you have sort of these genes
0:01:46 that jump into the genome.
0:01:47 – It’s kind of mysterious what the jumping
0:01:48 is really used for.
0:01:50 People surmise that, you know, potentially, you know,
0:01:54 it helps, you know, the cells and organism evolved in general.
0:01:56 So what this paper shows is to utilize
0:01:58 this CRISPR machinery known as Cascade,
0:02:02 and it’s formed by, you know, CASS6, CASS7, CASS8 proteins
0:02:04 to really insert genes into the genome.
0:02:08 And this Cascade protein is directed to the chromosome
0:02:11 by using guided RNA, you know, CRISPR machinery,
0:02:15 and it then binds to this transposase-associated protein,
0:02:18 TNIQ, and it allows them to recruit
0:02:20 these transposable elements and then effectively integrate
0:02:22 the gene into the genome.
0:02:25 And this is super powerful because now we’re able to,
0:02:27 you know, add genes in the genome directly
0:02:29 and precisely without cutting it open.
0:02:32 It’s a scarless way to modify the genome.
0:02:35 – So in some ways, this is almost like if CRISPR
0:02:38 is about surgically inserting or editing DNA,
0:02:41 this is almost in many ways like plastic surgery, right?
0:02:42 It’s scarless. – Ah, right, right.
0:02:45 – It doesn’t leave a mark and therefore in some ways
0:02:48 it’s less risky from an intervention standpoint
0:02:49 on the genome side.
0:02:51 – What it really boils down to is that
0:02:53 this machinery utilizes a way
0:02:56 to really efficiently integrate genes into the cell,
0:02:58 into the genome specifically,
0:02:59 without having to cut it open at all.
0:03:02 And to the metaphor of the surgery,
0:03:04 this is really important in the whole context
0:03:06 of gene therapy as a whole, right?
0:03:08 ‘Cause when we started off in gene therapy,
0:03:09 we had these random integration
0:03:12 of these trans genes into the cell.
0:03:15 And, you know, this was a more stochastic process.
0:03:17 So think of it almost as, you know,
0:03:20 the arrival of this paper and papers like it
0:03:24 are showing us that we’ve gotten to a point
0:03:28 where we have a fully programmable ability
0:03:32 to integrate new genes or new DNA into a genome
0:03:34 without having to first open up
0:03:36 or break apart the DNA to do that.
0:03:38 – So let’s go back and actually situate it
0:03:40 into the development of the science,
0:03:42 what it represents for where we’ve gotten
0:03:44 from where we began.
0:03:45 – Yeah, so if you want to do the really,
0:03:47 the fast forward montage version of this.
0:03:49 – Yeah, the 80s montage.
0:03:51 – Yeah, exactly, so I’m sure you’ll put
0:03:52 in the pop music behind it.
0:03:55 – Right, yeah, and the overalls in the paintbrushes.
0:03:56 – He always started in the 60s.
0:03:57 – In the time motion.
0:03:58 – Yeah, let’s go.
0:04:00 – But look, I mean, if you go all the way back,
0:04:02 you know, one of the earliest sort of technologies
0:04:04 that came to the fore was the discovery
0:04:05 of something called the restriction enzyme.
0:04:07 And the restriction enzyme was this ability
0:04:10 to take this protein that could cut DNA
0:04:12 at these predetermined sites,
0:04:15 essentially open up the DNA,
0:04:19 and then you could introduce new genetic material,
0:04:20 and it would eventually integrate randomly,
0:04:23 but it would integrate into that DNA.
0:04:25 That’s what gave us the ability to do
0:04:27 or recombinant DNA technology,
0:04:29 which gives rise to the entire biotech field.
0:04:32 One of the earliest applications of biotechnology is
0:04:35 getting bacterial systems to integrate human insulin
0:04:38 so that we could coax bacteria to make that drug
0:04:41 or human insulin on our behalf, of course, to treat diabetics.
0:04:43 So that starts the whole field.
0:04:45 Now, you know, if you sort of move forward,
0:04:46 the objective always has been,
0:04:51 can you cure disease by repairing
0:04:55 or replacing what’s broken in DNA?
0:04:58 And so the whole field of gene therapy arises
0:04:59 with this idea that, you know,
0:05:02 can we introduce a corrected version
0:05:05 of a non-functional gene in a patient
0:05:06 and have it do the job
0:05:08 that the non-functional gene cannot do?
0:05:12 The first version was just put that gene
0:05:16 into a viral vector, into almost like a delivery vehicle,
0:05:18 introduce that into patient cells,
0:05:20 that gets taken in by the cells,
0:05:22 and the gene just starts to sort of do its job.
0:05:23 It integrates randomly in the genome,
0:05:24 but it does the job it needs to do.
0:05:26 Hopefully it integrates, or maybe it doesn’t,
0:05:28 but it just does the job to compensate
0:05:30 for some mutated gene.
0:05:33 Yeah, so it’s basically almost a parallel support system
0:05:35 that the cell has.
0:05:37 We just had the first gene therapies approved
0:05:39 over the course of the last couple of years.
0:05:42 If you look at the spark therapy, gene therapy drug for,
0:05:44 this rare inherited form of blindness.
0:05:47 And so that was one big advance forward.
0:05:49 Now, that is introducing a full gene
0:05:51 and just hoping it gets taken up by the cell.
0:05:52 Just kind of throw it in the mix.
0:05:54 Yeah, throw it in the mix and it has its own risk.
0:05:56 The original discovery of CRISPR-Cas9,
0:05:59 that system, the way that system works
0:06:02 is by making what’s called a double-stranded break
0:06:03 in the DNA molecule.
0:06:04 So if you think of, you know,
0:06:07 we all remember DNA from high school biology
0:06:10 as sort of the beautiful double helix.
0:06:12 So imagine sort of cutting that double helix in two
0:06:14 and making an end of it, right?
0:06:17 And then, you know, and then putting it back together.
0:06:18 That’s not riskless, right?
0:06:20 And by the way, it’s very well known
0:06:22 and documented that one of the big setbacks
0:06:25 that the gene therapy field had a couple of decades ago
0:06:27 was when they ran a clinical trial
0:06:28 at the University of Pennsylvania,
0:06:31 there was a patient named Jesse Gelsinger
0:06:32 who died after receiving the therapy
0:06:34 just because there’s a lot of risk associated
0:06:37 with introducing, you know, a viral capsid with a gene
0:06:39 into a cell system, into human being
0:06:41 where you could have a catastrophic result
0:06:41 and that patient died.
0:06:44 And that actually put a big pause on how we thought
0:06:47 about developing gene therapies for humans.
0:06:49 But that approach will have
0:06:51 and does have therapeutic potential.
0:06:52 And there are several companies
0:06:56 that are pursuing developing CRISPR-Cas9-based therapeutics.
0:06:58 As that advanced in parallel,
0:07:02 we started to see other gene or genome editing technologies
0:07:03 come to the fore.
0:07:06 The first one was one known as zinc finger nucleases,
0:07:08 which really didn’t have as much uptake
0:07:10 as one would expect
0:07:13 because it’s just very hard to deal with these proteins.
0:07:16 – How zinc fingers work is that you actually use these proteins
0:07:18 to bind onto DNA sequences.
0:07:21 And every time you want to iterate to find a new target,
0:07:22 cost tens of thousands of dollars
0:07:24 in a few months to develop one protein.
0:07:26 – So it’s enormously expensive and labor intensive.
0:07:27 – Exactly.
0:07:28 So it never really caught on like wildfire
0:07:30 in the community, right?
0:07:31 – It’s a very bespoke process.
0:07:34 It’s one of the type of thing.
0:07:36 And then there were other technologies.
0:07:38 There was Talens,
0:07:40 which was a bit better than zinc finger nucleases,
0:07:45 but really the big sort of shift in the field
0:07:47 was the arrival of CRISPR.
0:07:49 – And so when scientists discovered CRISPR,
0:07:52 they noticed that it was just always constant evolutionary
0:07:55 warfare between bacteriophages and bacteria
0:07:56 for billions of years.
0:07:58 – Bacteriophages are viruses.
0:07:58 – Yes.
0:08:03 – And so when these viruses inject their viral genome
0:08:04 into the bacteria,
0:08:06 you can think of it as the bacterial immune system,
0:08:10 whereas they’re able to sense these little snippets
0:08:11 of viral genome,
0:08:13 they would create a vaccination card from it,
0:08:14 and then they’ll put it back into this,
0:08:16 what is known as this CRISPR array.
0:08:18 And this would be like a vaccine card
0:08:20 or a vaccine database, if you will.
0:08:22 So every time it recognizes that foreign viral sequence.
0:08:23 – Right, then it knows what to do.
0:08:25 – It would know how to snip it away, right?
0:08:27 And then people have hijacked this machinery.
0:08:31 So what if we can program this outside of bacteria
0:08:32 and use it in human cells
0:08:35 and program it to sense not viral DNA,
0:08:38 but specific targets in the human genome.
0:08:42 And then then we can program and effectively target
0:08:44 anywhere we want in the cell, right?
0:08:46 – So now bring us forward to today.
0:08:48 So is this new development,
0:08:50 does it mean that the therapeutic potentials
0:08:52 are essentially less risky?
0:08:54 Or are there new possibilities
0:08:56 that we haven’t been able to do before?
0:08:56 Or both?
0:08:59 – Basically, you know, it’s really powerful
0:09:01 because when we talk about the first gen of gene therapy
0:09:03 to the second gen of CRISPR,
0:09:05 this paper really represents this third wave
0:09:07 in this scarless genomic editing.
0:09:10 This is basically genomic surgery at its finest, right?
0:09:13 You know, when we think about laparoscopic surgery
0:09:15 and all these advanced surgery tools,
0:09:16 we want to have, you know,
0:09:19 basically a scarless methodology of doing surgery.
0:09:21 This is a way to really cleanly integrate,
0:09:23 you know, genomic segments into the cell
0:09:25 without even touching, right?
0:09:28 And then this also represents an even broader tool
0:09:31 of, you know, the entire, you know, genomic toolkit landscape.
0:09:33 – The real promise on the near term side
0:09:37 is that we get to a point where you can make
0:09:40 scarless integrations of genetic material into the DNA.
0:09:42 So it’s less traumatic in that regard.
0:09:44 So if you can do that more precisely,
0:09:46 there’s less scar tissue that hopefully is, you know,
0:09:48 a better intervention altogether.
0:09:50 So that holds great promise from a potential
0:09:53 for future therapeutics based on this type of technology.
0:09:55 I think the other thing that’s worth noting
0:09:57 is that in a relatively short period of time,
0:10:01 the programmability of these kinds of systems
0:10:04 has improved dramatically.
0:10:06 So the kinds of things that we could do,
0:10:09 that we can do now with, you know, with CRISPR
0:10:11 based on these kinds of advances
0:10:15 gives us a very broad repertoire and toolkit to work with,
0:10:17 whether it’s for therapeutic applications
0:10:18 or for diagnostic applications
0:10:20 or for any other number of things.
0:10:25 When most people think about CRISPR developing CRISPR
0:10:26 for human health,
0:10:28 they’re thinking about therapeutic applications.
0:10:31 But the reality is there’s also a lot of potential
0:10:32 for diagnostic applications.
0:10:34 So going back to the original discovery
0:10:35 of the CRISPR-Cas system,
0:10:38 this was essentially the memory banker immune system
0:10:40 for bacteria to remember what viruses
0:10:41 had attacked it before
0:10:43 so they could protect themselves going forward.
0:10:47 And the way it does that is by essentially cutting
0:10:49 that viral genetic material.
0:10:52 So it’s ineffective essentially basically, you know,
0:10:56 cutting it off at its Achilles heel, so to speak.
0:10:57 And so as you can imagine,
0:11:00 if you’re hijacking that capability
0:11:01 for therapeutic purposes,
0:11:03 you could also hijack that capability
0:11:05 for diagnostic purposes.
0:11:07 And a way that that could potentially work, for example,
0:11:12 is if you know what bacteria or what virus
0:11:15 or even what, you know, mutation in DNA you’re looking for
0:11:18 in say a human sample, a blood sample
0:11:20 or urine sample or anything like that.
0:11:24 If it’s present, it will get cut by the right CAS system.
0:11:27 You program the CRISPR-Cas system to say,
0:11:31 these are the sequences of genetic material
0:11:33 that I am looking for in the sample.
0:11:34 This is basically the search engine.
0:11:36 Like you’re doing essentially a Google search.
0:11:38 – And you basically just look for the kill switch
0:11:40 to have been activated.
0:11:41 – You just look for the identifier.
0:11:42 So you basically say,
0:11:44 if I want to look in this patient’s sample,
0:11:47 let’s say I want to look for a specific bacteria
0:11:48 or a specific virus
0:11:51 or specific genetic mutation associated with disease,
0:11:53 you can say, if you find the presence
0:11:55 of any one of these sequences,
0:11:56 those are almost like the search terms,
0:11:58 cut them and when you cut them,
0:12:00 you can essentially engineer the system
0:12:02 to send out some sort of a reporter,
0:12:04 a reporter, usually it’s a visual marker.
0:12:06 And so basically if it lights up,
0:12:08 it’s because the CRISPR-Cas system cut the DNA
0:12:10 you told it to look for.
0:12:12 So that has a potential diagnostic application
0:12:13 and you could run that diagnostic
0:12:15 essentially without a lab, right?
0:12:16 ‘Cause it just happens with the biology.
0:12:19 So a whole other kind of new tool, essentially.
0:12:21 – Yeah, and I think the diagnostic applications
0:12:23 for these kinds of technologies
0:12:26 are actually pretty intriguing
0:12:28 because most of the way we do diagnostics
0:12:33 is based on developing some very specific biological
0:12:34 or chemical assays.
0:12:36 So look for something and if so,
0:12:38 have the reaction take place
0:12:40 and one that I can visualize and quantify
0:12:42 or quantitate in some way.
0:12:44 But here you’re just basically letting the biology
0:12:45 do the work for you.
0:12:47 In the CRISPR toolkit,
0:12:51 a lot of the initial applications was using
0:12:53 CRISPR-Cas9, this one nucleus.
0:12:56 And actually the diagnostic application
0:12:57 that Jorge was talking about
0:12:59 was actually using these other nucleas
0:13:01 known as Cas13 and Cas12.
0:13:04 And so all these different CRISPR proteins,
0:13:08 Cas9, 12, 13, X, Y, that we’re continuing to discover
0:13:11 has all different fundamental applications, right?
0:13:12 Even fundamentally changing
0:13:14 what these CRISPR nucleases even do.
0:13:15 It doesn’t even cut anymore.
0:13:17 It can do scarless editing.
0:13:20 And then we can even add different applications on it.
0:13:22 There’s new applications adding,
0:13:24 deaminase is just to do base editing.
0:13:28 So we can really do single base pair resolution editing.
0:13:30 There’s really the final frontier of precision
0:13:32 in terms of genomic modification.
0:13:34 And I think what’s really important is that
0:13:36 we’ve really seen this shift from
0:13:38 when it became this random bespoke science
0:13:40 and really turning into full-fledged,
0:13:42 modified engineering tool.
0:13:44 And this paper that we talk about here
0:13:47 is not only a great representation
0:13:49 of this engineering biology thesis,
0:13:53 but also a pretty huge potential step change
0:13:54 for the field as a whole.
0:13:56 – You can program this to turn genes up and down
0:13:58 as opposed to just editing them.
0:14:01 There’s even work that’s ongoing to use this technology
0:14:03 to image DNA directly,
0:14:04 which is a pretty remarkable thing
0:14:07 because since this is acting locally on DNA,
0:14:10 you can add all kinds of agents to make it imageable.
0:14:12 So therefore, you can observe chromatin
0:14:14 or genome structure directly.
0:14:16 So there’s a lot that can be done with this technology,
0:14:20 and you hear the old adage about the pickaxes for the gold.
0:14:22 And with these toolkits,
0:14:25 I mean, we are quite literally panning for gold here.
0:14:28 I mean, where these things get found,
0:14:31 they’re found in soil and in ocean vents.
0:14:32 – New York City subways.
0:14:34 – The New York City subway.
0:14:37 So these people are quite literally looking in nature
0:14:39 because nature has ingenious ways
0:14:41 to do a lot of the things that we’re trying to do
0:14:45 from an engineering biology or programmable biology standpoint.
0:14:50 And so I think it’s a remarkable moment to take pause
0:14:53 and see how far this technology has come
0:14:55 in a relatively short period of time.
0:14:58 This generation’s recombinant DNA or restriction enzymes
0:15:01 that really gave rise to the biotechnology industry.
0:15:03 I think this tool kit, this CRISPR tool kit,
0:15:06 as we’re describing it and discussing it,
0:15:07 represents sort of the next frontier
0:15:10 for what will happen in biology.
0:15:13 So an incredible development of precision
0:15:15 in what the tools can do and at the same time,
0:15:17 a huge expansion of what that will enable us
0:15:18 to do going forward.
0:15:20 Thank you guys so much for joining us.
0:15:23 Should I say on the A16Z Journal Club?
0:15:23 – No, no, please don’t say that.
0:15:24 – Yeah, okay.
0:15:34 [BLANK_AUDIO]
Two recent scientific journal papers show what’s possible when CRISPR moves from cutting DNA tool to a full-fledged platform — expanding its toolkit for medicine across R&D, therapeutics, and diagnostics:
”Transposon-encoded CRISPR-Cas systems direct RNA-guided DNA integration” in Nature — by Sanne Klompe, Phuc Vo, Tyler Halpin-Healy, and Samuel Sternberg (of Columbia University)
”RNA-guided DNA insertion with CRISPR-associated transposases” in Science — by Jonathan Strecker, Alim Ladha, Zachary Gardner, Jonathan Schmid-burgk, Kira Makarova, Eugene Koonin, and Feng Zhang (of the Broad Institute)
What do these two papers — both about techniques for getting rid of the need to cut the genome to edit it — make possible going forward, given the ongoing shift of biology becoming more like engineering? Where are we in the wave of the genome engineering ”developer community” building on top of CRISPR with a constantly growing suite of programmable functionalities? a16z bio general partner Jorge Conde and bio deal team partner Andy Tran chat with Hanne Tidnam about these trends — and these two papers — in this short internal hallway-style conversation, part of our new a16z Journal Club series.
This podcast is also part of our new a16z bio newsletter, which you can sign up for at a16z.com/subscribe