Hepatitis B is a tiny virus that causes hundreds of thousands of deaths from liver disease and cancer each year. The vaccine against it became the first of many milestones: it was the first viral protein subunit vaccine, the first recombinant vaccine, and the first vaccine to prevent a type of cancer.
In this episode, Jacob Trefethen and Saloni Dattani follow the trail of strange jaundice outbreaks that scientists traced to a stealthy liver virus, how scientists turned one viral surface protein into a lifesaving shot for newborns, and how it was all built upon breakthroughs in immunology.
Hard Drugs is a new podcast from Works in Progress and Coefficient Giving about medical innovation presented by Saloni Dattani and Jacob Trefethen.
You can watch or listen on YouTube, Spotify, or Apple Podcasts.
Chapters:
0:00:00 Introducing the hepatitis B vaccine
0:15:46 The mysterious trail of jaundice outbreaks
0:28:03 How a tiny virus causes cirrhosis and liver cancer
0:53:19 Maurice Hilleman’s purified hep B vaccine
1:17:36 Turning the hep B vaccine recombinant
1:29:14 The impact of hep B vaccination
1:39:27 The 19th century battle for immunology
2:01:34 How the body almost infinite antibodies
2:30:57 How subunit vaccines took over
2:45:33 Conclusion
Saloni’s substack newsletter: https://www.scientificdiscovery.dev/
Jacob’s blog: https://blog.jacobtrefethen.com/
Transcript
Saloni Dattani:
It took 90 years after Jenner developed the smallpox vaccine for scientists to make another one. For the next 50 years after that, scientists tried making many more vaccines through attenuation and inactivation. They tested them, found success multiple times, but still had very little understanding of why they worked.
Jacob Trefethen:
That is where we ended last episode, but the discovery of germs and the invention of attenuated and inactivated vaccines were just the beginning. One thing that changed over the 20th century were massive breakthroughs in immunology, the science of improving our understanding of our immune responses and how we learn from past infections. That meant that scientists could start making safer and more precise vaccines, which allowed particular scientists to develop the first hepatitis B vaccine.
Saloni Dattani:
The hepatitis B vaccine is special in many ways. It was the first protein subunit vaccine which contained just a small protein from the virus instead of the whole thing. It was then improved on to make the first recombinant vaccine, with genetic engineering technology to produce it in yeast. Third, it was the first vaccine against a cancer: liver cancer.
Jacob Trefethen:
It has also been one of the most successful vaccines, preventing hundreds of thousands of children from getting chronic hepatitis B infections each year and reducing liver cancer rates by about 85% in those who receive it.
Saloni Dattani:
In this episode of Hard Drugs, we are going to talk about all of that: how people figured out the science of immunology, how they made vaccines more precise, how they made hepatitis B vaccines, and how those vaccines have protected babies from jaundice and a deadly form of liver cancer.
[podcast jingle]
Saloni Dattani:
What’s your opinion about hepatitis? Good, bad, ugly?
Jacob Trefethen:
Hepatitis? God, I have a lot of opinions about hepatitis, if I am being honest. I mean, hepatitis is caused by many things. Do you want my opinions about hepatitis B or my opinions about hepatitis?
Saloni Dattani:
Yes, hepatitis B; so there are different letters and they correspond to different diseases.
Jacob Trefethen:
Well, my opinions about hepatitis B are that in my day job at Coefficient Giving, we have worked on trying to make new cures against hepatitis B, trying to improve animal models for drug development, trying to put hepatitis B vaccines in microneedle patches so that a baby can have a patch instead of an injection. We’ve done a fair bit on hepatitis B.
Saloni Dattani:
I want a patch.
Jacob Trefethen:
Yes, well...
Saloni Dattani:
It’s only for the babies.
Jacob Trefethen:
What would you put in your patch?
Saloni Dattani:
Everything. All the vaccines in one patch.
Jacob Trefethen:
Oh, wow. That would be... hey, that’s the future. Unfortunately, today we are just talking about history. I mean, I basically think hepatitis B is it is the hepatitis virus of many that leads to the most deaths. It’s hard to quite attribute how many, but somewhere in the region of 750,000, maybe 800,000; some estimates go as low as 500,000, but hundreds of thousands around the world.
Saloni Dattani:
Per year.
Jacob Trefethen:
Per year. I have a lot of respect begrudgingly for that virus because it is fucking tiny; it is four genes. It is astonishing. It is unbelievably compact. I mean, some of those genes literally overlap, that is how efficient it is. It also has a reverse transcription stage, like HIV does. It’s just unbelievably complicated for something that is so tiny. I am just like, bloody hell. Anyway. It forms its own chromosome inside your liver cells. Anyway, I did not know I had this many feelings about hepatitis B.
Saloni Dattani:
Hepatitis B has also been in the news recently because in the US, the Vaccine Advisory Committee just voted to stop universally recommending hepatitis B vaccination at birth. It’s possible they limit the vaccines even further, potentially keeping them only for teenagers or adults. That’s another reason that I think this is a really interesting and important topic to talk about.
Currently, in many countries around the world, hepatitis B vaccines are universally recommended at birth to all newborns. There are some exceptions, though. In various countries in Africa, they do not provide it at birth. That is generally because it’s just difficult to do that logistically, because many births happen outside of health facilities. The vaccine requires cold chain storage, and instead, the recommendation is to give vaccines around six weeks after birth to babies. In Europe, many countries only provide- or only recommend the vaccine to children of mothers who test positive for hepatitis B, but they have a lot of screening and high vaccination rates for those newborns.
This is interesting as well because of the epidemiology, how hepatitis B spreads, and how that transmission occurs nowadays in different countries. In poorer countries, there are a lot of infections that happen after birth in early childhood, and kids catch the virus from their siblings or other kids that they’re playing with.
In richer countries, the prevalence of hepatitis B is generally very low, and that’s generally because vaccination has been so successful so far. Most infections nowadays are from mother to child, not from other kids.
I think one reason that I find this concerning is that if there is more vaccine hesitancy against hepatitis B because of this change, it could mean that even mothers who test positive for hepatitis B would be unwilling to get their kids vaccinated. A second is that it would be very bad if other countries, especially poorer countries, follow suit.
Early vaccination is important because infections in babies and young children are much more likely to turn into long-term infections, which can lead to liver disease and then cancer. The reason for that is that infants and kids have less developed immune systems, and they cannot control the virus as well as adults can. It slips past their immune system and establishes itself in the liver for years or decades. If it was me, I would recommend universal vaccination, but that does not mean that it’s mandatory; it just means that it’s free and recommended.
Jacob Trefethen:
Yeah, and the case that people in favor of this change in the US would make is that if some mothers already know that they are negative for hepatitis B and will not pass on hepatitis B in childbirth to their child, they should be allowed to skip vaccination. Their kid will be at more risk from other sources, but at birth at less risk.
The case against, from a lot of the public health community is, well, we know that this vaccine is safe and works. It has already driven down, you know, 90-plus percent, 99% maybe in the US of hepatitis B cases because it works so well. Implementing more complicated guidelines where people have to judge for themselves when they are not health experts will mean that more kids are at risk of hepatitis B.
Yeah, I think what you said is absolutely right: if this is step one to restricting access further, that is especially concerning. That is also true for other countries where they have not yet seen the big drops in hepatitis B that the US has, as a result of the vaccine, that the risk for kids is higher.
We recorded most of this episode before this recent news from the US because hepatitis B vaccines happen to be the best next part of the story to tell of vaccine development. We wanted to talk about them as a vaccine using modern methods to become even more safe than some of the vaccines we talked about in the last episode.
Saloni Dattani:
So let’s get into the topic of hepatitis B, the vaccine, and how it was made.
[podcast jingle]
Jacob Trefethen:
Firstly, what is it?
Saloni Dattani:
What is it? What is hepatitis? Hepatitis is a liver infection. That is what it means: hepa- is liver, -itis is inflammation or infection. What happens when someone gets a hepatitis infection? Well, there are two types. There are acute infections, which are not cute. They are basically a short-term infection, reaction, that lasts weeks to months, and people feel very tired; they throw up a lot. They also could have yellowing of their skin and of the whites in their eyes. Most people recover from that.
Then there’s chronic hepatitis, which means that the virus stays around in your body. It’s hanging out for a long time, usually years, and it causes inflammation in your liver, causes your liver to get scarred, and over time, that can lead to cancer.
Jacob Trefethen:
Long-time listeners may recall that the fifth episode in our AI series, we talked about hepatitis antigens you could go after with AI designed proteins. It came up there before, but I think the thing you just said about it being a chronic infection is crucial to the link with cancer. And the issue with hepatitis C infections as well, if you have a chronic infection, then it’s sticking around, it’s spitting off new viral particles somewhat regularly, and they are going to start integrating with your own DNA sometimes, which can lead to... problems!
Saloni Dattani:
Right. We mentioned hepatitis B, and you mentioned hepatitis C, and there are many different kinds of hepatitis. There is A, B, C, D, E, and are there more? Maybe there’s an F as well.
Jacob Trefethen:
I only know those five, and those five actually, again, with my Coefficient Giving, we have worked on four of the five. Any guesses which one we have ignored?
Saloni Dattani:
A.
Jacob Trefethen:
Correct, yes.
Saloni Dattani:
I was right!
Jacob Trefethen:
A has got a great vaccine and spreads through food contamination and all that. Of those, it is kind of amazing how different they are.
Saloni Dattani:
What I did not know until doing research for this episode is that they are caused by completely different viruses. They’re just completely genetically unrelated. I remember reading about this and I was like, what the heck is going on? Why do they all have the same name then?
Jacob Trefethen:
They are named so badly. They just happened to be involved with the liver. Well, I mean, my one defence is that hepatitis D, or hepatitis delta, is related to hepatitis B.
Saloni Dattani:
Oh, okay.
Jacob Trefethen:
In the sense of you actually can’t get- it’s a real tiny, tiny, tiny, tiny, tiny virus, Delta. And you can’t even get infected with it unless you have hepatitis B, and so it only hops on in.
Saloni Dattani:
It’s a hanger-on.
Jacob Trefethen:
It’s a hanger-on. I mean, viruses themselves are making use of your cell machinery because they are no good on their own. Hepatitis delta is like, I am actually no good without another virus. I mean, that is pretty no good about it, but it’s dangerous if you do get it.
Saloni Dattani:
Well, hepatitis A causes a short-term acute infection. B and C both cause liver cancer. What does D do?
Jacob Trefethen:
Same again as B.
Saloni Dattani:
Also liver cancer?
Jacob Trefethen:
Yeah.
Saloni Dattani:
What about E?
Jacob Trefethen:
E is pretty, well, firstly, with the liver cancer ones, also often it is just cirrhosis where you do not necessarily get cancer, but you get so much liver damage that it causes- it can lead to death, that is over many years. E is one that I wish there was more epidemiology on, in the sense that I do not think it is well known how widespread E is around the world, but it is more often acute, so similar to A, where if you have infections at a particular water source, E is hanging around in some water source, then you might get transmission in a particular community that has a big explosive outbreak.
Saloni Dattani:
Huh. Yeah, I didn’t know about that. I wonder if people are going to identify more after this. Especially if they actually have no genetic relationship to each other and it is just different viruses that can infect the liver.
Jacob Trefethen:
Absolutely. Do you remember which one came up last episode?
Saloni Dattani:
No?
Jacob Trefethen:
Hepatitis C, because, do you remember the injections in Egypt?
Saloni Dattani:
Oh, yes, yes, yes, yes, yes.
Jacob Trefethen:
To cure schistosomiasis. Those introduced hepatitis C in the ‘60s and ‘70s to a lot of people through sterilized needles- or not sufficiently sterilized. And that was before hepatitis C was discovered.
Saloni Dattani:
Right, right.
Jacob Trefethen:
Who knows what else is down there in our liver?
Saloni Dattani:
Right. That is very scary. They are all different infections of the liver. They have different sources or ways that they transmit. We are going to focus in this episode on hepatitis B. Hepatitis B, as we said, has a short-term infection, and then it can also lead to a chronic infection. It is mostly spread through blood, right? Mother to child, or through shared needles, or blood products. That is actually how it was discovered and how people figured out that it was a virus at all.
People had seen the symptoms of hepatitis for thousands of years. Hippocrates, over 2,000 years ago, described the symptoms of hepatitis. People were wondering what caused it. At the time they said, humoral imbalance, and I can see why you would think that. It’s also really difficult to figure out what the cause is because often the illness that you get happens months after you get infected by the virus, so it’s hard to make the connection and figure out what is the source of an infection.
There are situations in which there are outbreaks of hepatitis. Historically, that would have been in wars – so you would potentially have blood contamination through injury and things like that. But also there would have been outbreaks sometimes after the use of medical products, such as blood transfusions, but also the use of, as you said, unclean sterilized syringes. When people were doing vaccination campaigns against smallpox or against yellow fever or against other diseases, sometimes, in some contaminated lots of the vaccine, it led to small outbreaks. You would have many people who suddenly develop jaundice around the same time. The only shared thing they have is that they all got a particular medical product at some point months ago.
There is this interesting quote that I was reading about this that said, “Vaccination against smallpox, the glass syringe, injectable antimicrobial drugs, passive immunization, the yellow fever vaccine, and blood transfusion have all been of enormous benefit to humanity, but each of these medical advances has left a trail of jaundice in its wake.”
Jacob Trefethen:
Oh my gosh, is not that crazy to think about?
Saloni Dattani:
Yeah, but thankfully it’s not that common because it’s only- typically in the past, it would have just been a few small percentage of people who had it circulating in their blood and that could contaminate those products and lead to outbreaks.
Jacob Trefethen:
Well, just as a PSA to listeners, if you, for any reason, do use needles — so people who inject drugs with needles are some of the most at risk of hepatitis C — so you have to be really careful because you do not want to introduce microbes into your blood system.
Saloni Dattani:
But also if, like me, you just like getting vaccinated.
Jacob Trefethen:
So long as you are not getting vaccinated in the 19th century, I think you will be okay.
Saloni Dattani:
That’s true. There is also another theory of the cause of hepatitis, and that is by Rudolf Virchow, the father of pathology. He thought that hepatitis was caused by overeating; that overeating inflamed the intestines, and it closed the bile ducts, and that led to a lot of problems. The only reason he basically thought this was that he had heard about an autopsy of one person who had jaundice and whose bile duct was closed and swollen by mucus.
Jacob Trefethen:
N of 1, okay.
Saloni Dattani:
He’s like, problem solved. Now I know what causes the disease.
Jacob Trefethen:
Wow. Okay. Was he right?
Saloni Dattani:
He was not right, but people believed... this theory led to a particular name of hepatitis before it was called that, catarrhal jaundice, that it was related to the intestine. And that name lasted for 80 years. People just thought it was related to the intestine for a very long time because of him.
Jacob Trefethen:
We cannot be too mad, because the current names of the diseases / viruses, hepatitis A, B, C, D, E, as just discussed, are still terrible.
Saloni Dattani:
But I’m also like, well, probably you would have been able to at least observe these outbreaks of jaundice. When you saw them, you could have done more than one autopsy, and you could have done some more pathology research, and figured out that it was the liver that was the most damaged. And that is what people eventually figured out over the 1920s and ‘30s.
Also in World War I, pathologists who were looking at soldiers who were killed in the war, but also suffered from jaundice, saw that they had a lot of abnormalities in their liver tissue. There were a lot of cells that were infiltrating their liver tissue and inflaming it. They concluded that the bile duct jaundice or the catarrhal jaundice was actually due to liver inflammation. They resolved this old debate and threw out his theory.
Jacob Trefethen:
Okay. Pew. Trash can! Okay. Now, the question I have is what is causing that inflammation, Saloni?
Saloni Dattani:
What is causing that inflammation? People do not know. People do notice sometimes there are outbreaks. The outbreaks occasionally happen months after a vaccination campaign, but only in certain places, only with certain lots of contaminated vaccine. Sometimes they happen when people are treated for diseases, so syphilis patients that are treated with antibiotics are sometimes developing jaundice months later. Doctors are like, well, it doesn’t seem like it is because of the drug, because the illness is only happening months later.
Jacob Trefethen:
These are old school antibiotics with a needle, you mean? Not oral.
Saloni Dattani:
Yes. This would have been things like salvarsan, which I think is the first antibiotic. This jaundice is probably not caused by syphilis because their other symptoms are improving by the time they have developed jaundice, which is months later. They are like, hmm, what’s going on?
Jacob Trefethen:
Something else here, yeah.
Saloni Dattani:
People keep studying, and they’re like, okay, let us try to deliberately infect people and see what happens. They try to do different things to inactivate whatever is in the serum, and they figure out that it is something that is smaller than a bacterium; it gets through bacterial filters. It can also survive heating at 56 degrees for 30 minutes.
Jacob Trefethen:
Really? That is impressive. Wait, on the temperature thing, I think that is 56 degrees celsius, I assume you mean.
Saloni Dattani:
Yes, yes.
Jacob Trefethen:
Not Fahrenheit, not Kelvin. Okay, got it. Yes.
Saloni Dattani:
It is caused by something that can get through bacterial filters, can survive heating for half an hour. It can also be transmitted by blood of people who have symptoms.
Jacob Trefethen:
Wait, so I have a question for you, Saloni, which is, do you think you could survive 30 minutes in 56 degree heat?
Saloni Dattani:
I don’t think I could, and I do not mean like physiologically, but I think psychologically I would just leave after two minutes. You know what? I like normal temperatures.
Jacob Trefethen:
Yeah. Yeah. That is... I do too, actually. Okay.
Saloni Dattani:
I was once in a sauna, and I just got bored. I was like, you know what? No more of this.
Jacob Trefethen:
Everyone else is like, “I paid 50 pounds for this.” You’re like, “This is boring. I want to get back to my books.”
Saloni Dattani:
So it’s really difficult to figure out what exactly is causing hepatitis. It’s not a bacterium; it’s probably too small for that. But it’s somehow resistant to a bunch of different types of chemical processes. It survives for heat for half an hour. And it’s really hard to grow in cell culture. And it’s really hard to cause jaundice in other animals by injecting infected serum with them.
Jacob Trefethen:
What year are we in? What year?
Saloni Dattani:
This is the 1930s.
Jacob Trefethen:
Okay. 1930s were when the electron microscope came out, and we discovered viruses-
Saloni Dattani:
True.
Jacob Trefethen:
So did they know yet the virus concept or is...
Saloni Dattani:
1931, I think, was the first electron microscope, but I think they just could not figure out the specific pathogen. I think that would have still been kind of hard.
Jacob Trefethen:
Got it. I’ve got a guess as to why it’s not working in other animals.
Saloni Dattani:
Why?
Jacob Trefethen:
I mean, working is a horrible word, but it is not causing disaster in other animals. Now we know hepatitis B only really forms a chronic infection in humans, right?
Saloni Dattani:
Doesn’t it also infect chimpanzees? I think I read that.
Jacob Trefethen:
Yes. And woodchucks, but that is a whole different story.
Saloni Dattani:
Yeah, and woodchucks. Yeah, I saw that as well.
Jacob Trefethen:
How much wood would a woodchuck chuck if a woodchuck had hepatitis?
Saloni Dattani:
Hepatitis is causing long-term infections. It hides in the liver. It causes liver damage, scarring, and then liver cancer. This was not known for a very long time, but people did see that there seemed to be different types of hepatitis. Some of them are infectious through the blood, through the serum, and some of them seem to come after serum-related things.
There are others that don’t, and there are others that seem to be spreading through contaminated food or water or something else. They have not discovered hepatitis C at this point, so that is out of the question.
They then separate these into two, and they call hepatitis A “infectious hepatitis” and hepatitis B “serum hepatitis.”
Jacob Trefethen:
That’s a terrible name.
Saloni Dattani:
They are like, hepatitis A usually seems to be spreading in outbreaks through contamination.
Jacob Trefethen:
Right, yes.
Saloni Dattani:
And that can pass from person to person. But hepatitis B seems to happen mostly after blood transfusions, and people develop jaundice weeks or months after those, so that is “serum hepatitis.”
Jacob Trefethen:
Okay. Okay.
Saloni Dattani:
Kind of makes sense.
Jacob Trefethen:
I mean, I think I could, it is going to make me sound bad, but I think I could give hepatitis A via serum, and I could give B via infection.
Saloni Dattani:
Yeah, yeah, but it’s the typical pattern, I guess.
Jacob Trefethen:
Yeah, I am with you.
Saloni Dattani:
Now we know that they are caused by totally different virus families. Hepatitis A is caused by an RNA virus of the Picornavirus family, which is the same family as poliovirus. Hepatitis B is a DNA virus in the Hepadnavirus family, which is very directly named after, I guess, hepatitis and DNA.
Jacob Trefethen:
Hepadna?
Saloni Dattani:
Yeah.
Jacob Trefethen:
Cool. I mean, it is double-stranded DNA, right? I mean, I think it’s a little strand that hangs off, but yeah, it’s a little circle basically.
Saloni Dattani:
And as you mentioned before, it also has this very weird replication strategy where it turns its DNA into RNA and then back into DNA and then double-stranded DNA.
Jacob Trefethen:
So it reverse transcribes. So I’m a little virus that then enters the bloodstream via a needle, and then I am going to find my way to the liver. It probably won’t take very long at all. And then I’m going to enter a liver cell, and I’m going to unfurl and unpackage myself into the cytoplasm of that liver cell. I’m in the nucleus, I form a little mini-chromosome of my own, which is crazy. Then that mini chromosome uses all your little machinery in the nucleus, your histones. It’s forming this nice little circle-of-eight kind of thing.
Saloni Dattani:
Aw.
Jacob Trefethen:
I know, it’s cute until it’s not. And then it’s so stable, it can stick around for decades, hence the chronic infection. And your body’s going to start transcribing it again, and once it transcribes it, it’s going to leave the nucleus. It’s in the cytoplasm, it’s going to sort of self-assemble. Maybe that’s when it does its reverse transcription, I’m not sure, and then it’s going to form a little packet - boop! - and then leaves as a new virus.
Saloni Dattani:
But it also, it reverse transcribes- so it turns its RNA back into DNA and then it uses its polymerase to make another DNA strand, and so that makes it a double-stranded DNA again.
Jacob Trefethen:
Oh, right. Yeah.
Saloni Dattani:
What the heck is going on, though? That is so complicated.
Jacob Trefethen:
That should not be allowed.
Saloni Dattani:
Why so many steps?
Jacob Trefethen:
Yeah. Why so many steps for something so small?
Saloni Dattani:
The other thing about the life cycle, or its strategy, is that the way that it infects the liver is by binding to your liver cells - on the surface of your liver cells. People make antibodies against this surface protein it uses to attach - the hepatitis B surface antigen - which we will come back to.
People are making antibodies that stick to this viral surface protein, prevent it from attaching to our liver cells, and that would normally prevent an infection if it cannot bind to the liver cells. But guess what? The virus has a strategy to fight back.
The way that it does that is by making so many of that same viral surface protein. Basically, the idea is that all of these excess surface proteins will soak up antibodies from the blood that you have developed against it, and allow the virus to attach itself to our liver cells. There’s a quote from a book that I was reading, which will come up again, called ‘Vaccinated’ by Paul Offit. He says, “Hepatitis B virus is so committed to this method of survival that people infected with the virus have about 500 quadrillion particles of viral surface proteins circulating in their bodies during an infection.”
Jacob Trefethen:
That’s... that’s a lot. I’m no mathematician.
Saloni Dattani:
That is five with 17 zeros after it.
Jacob Trefethen:
That’s kind of crazy to think about.
Saloni Dattani:
It is just like, “Oh, guess what? You think you can attack me? Can you attack 500 quadrillion-?” I guess that’s what you do if you only have seven proteins, you only have four genes. You are like, “Guess what? You can’t fight so many-
Jacob Trefethen:
Yeah, it is like one horse-sized duck versus a hundred-
Saloni Dattani:
500 quadrillion. Would you rather get infected by one tetanus bacterium or 500 quadrillion hepatitis B surface antigens?
Jacob Trefethen:
Yeah. Well, you have given me two great options, so thank you. I’ll take the Hep B. I’ll take the Hep B.
Saloni Dattani:
Really? I would have taken the tetanus.
Jacob Trefethen:
Let’s see who makes it out.
Saloni Dattani:
None of us, probably. So hepatitis B, how does it cause disease? One is, it manages to evade our immune response by making 500 quadrillion hepatitis B surface antigens.
Jacob Trefethen:
As I think of Hep B getting into your nucleus of your liver cells, which is why it stays so chronic, because it gets so stable down there, I think of two forms of it. The main one is what is called cccDNA, covalently closed circular DNA. There’s a second form, though, which is called integrated DNA, where basically sometimes it does slip in and similar to HIV that sort of gets integrated into your DNA, it will get into your main chromosomes. Oh my gosh, imagine trying to get that out. That’s... difficult. I think that that leads to some co-genetic activity.
Saloni Dattani:
I think that was basically as far as I got. The virus’s DNA integrates into your liver cells’ genomes, and then the insertions could be in places that are important for protecting your cells against cancer- or cancerous mutations. But I think it is the long-term inflammation. Basically, the virus is sticking around in your liver cells for a very long time, your immune system keeps trying to control the infection, but it never manages to fully clear it, because I guess it’s integrated itself. And this creates long-term inflammation and repeated cycles of cell death and regrowth.
I guess the liver is also the only organ that we know of that can regenerate itself almost fully. There are just many different cycles of this inflammation, and those cycles raise the chance of harmful mutations as there’s regrowth of liver cells. So, over a lot of time, over many years or decades, the combination of these things — the chronic inflammation, the cycles of cell replacement, and the insertion of its viral DNA, and also just various other crazy things it does with its HBX protein, right?
Jacob Trefethen:
Thank you for bringing up HBX. Yes! HBX, part four of our AI series, I was trying to make binders against. I don’t know the most recent literature to be sure of, but I think how that could be implicated in cancer is that HBX is somehow getting your body to relax its usual way of controlling oncogenes.
Saloni Dattani:
It affects the signalling of your cell.
Jacob Trefethen:
Yes, definitely.
Saloni Dattani:
Yeah.
Jacob Trefethen:
It is the transcriptional signalling. Well, it’s a little bit beyond our pay grade, but we might need to get an HBX expert on a future episode.
Saloni Dattani:
Oh. We’re going to do another episode on this?
Jacob Trefethen:
Great.
Saloni Dattani:
Hold on to your hats. This isn’t over. So all of this stuff eventually causes liver cancer, specifically in your hepatocytes, is that right? Hepatocellular carcinoma.
Jacob Trefethen:
HCC will not let me be.
Saloni Dattani:
So why is this important to prevent? Because it causes liver cancer, but also because it infects babies, right? It causes jaundice in babies. Most cases of hepatitis B are acquired by infants from their mothers at the time of birth.
Jacob Trefethen:
There’s something I don’t understand the driver of, but I’ve heard that if you get infected when you are an infant or a baby, you have a very high chance of forming a chronic infection, maybe over 90%. If you get infected as an adult, you have a very low chance of forming a chronic infection, maybe under 10%. I’m not sure why that is, but that means the risk per infection changes a lot.
Saloni Dattani:
Right, I’m also not sure what it is. If I had to guess, it would be that adults have various protective factors in their humour, in their serum that destroy hepatitis B virus, at least for some people, at least that’s what I would guess. Or we have other immune cells that manage to kill it, and that doesn’t happen in babies. Around 90% of babies infected at birth or in the first year of their life go on to develop chronic hepatitis, and that compares to five to 10% of adults who catch the infection.
15 to 40% of infants who develop that chronic infection will then die prematurely from liver failure or cirrhosis, which is severe scarring of the liver, or from liver cancer if they’re not treated for it. That’s really bad because there weren’t vaccines for a very long time, and we didn’t really have a good idea of what was causing that liver cancer until, I think, the 1970s and 80s.
Jacob Trefethen:
You mean that those babies will very often die of liver cancer, but as adults, not as babies?
Saloni Dattani:
Right.
Jacob Trefethen:
That’s a high percentage. It’s surprising to me how liver cancer overall is very high up there and it’s mostly caused by viruses. There’s almost an opportunity there: if you can get rid of the viruses, then maybe we actually can reduce cancer a lot more than some other cancers that might be even trickier.
Saloni Dattani:
Very true. There are a bunch of cancers that we can get rid of by getting rid of infections. One is liver cancer. Then there’s cervical cancer, which is caused by human papillomavirus. There’s stomach cancer, which is predominantly caused by Helicobacter pylori, often spread through contaminated food. So we should just get rid of all of these pathogens and then we won’t have a bunch of cancers, isn’t that cool?
Jacob Trefethen:
That is really cool.
Saloni Dattani:
I was interested in the fact that people figured out this was related to liver cancer, and as I mentioned, that was not clear until the early 1980s, around 1981. There were various lines of evidence that suggested it. People doing lab experiments or animal experiments — which I presume would have been with chimpanzees, I guess — showed hepatitis B caused liver cancer, probably because of the long-term liver damage caused by the virus. There were case-control studies where if you looked at people who had liver cancer, they had a much higher prevalence of hepatitis B antigens.
Then in the 1970s, the CDC researcher called Palmer Beasley carried out a longitudinal study in Taiwan where he tested adults for whether they had the hepatitis B surface antigen, which is a marker of them having the infection, and followed them up for several years. The people who had this chronic infection marker went on to develop liver cancer at much higher rates than people who were not infected.
And as we’ll come to later on, there’s even stronger evidence that hepatitis B causes liver cancer, because when the vaccine was rolled out, you could see a massive drop in liver cancer rates years later. This was done in a randomized trial in China, where they randomly gave the vaccine to some towns and not other towns. They gave them to all newborns in certain towns but not others, and they had an 85% reduction in liver cancer 30 years later in the towns where the newborns got vaccinated.
Jacob Trefethen:
Yeah, it’s a pretty stark result. How did we actually get to those vaccines?
Saloni Dattani:
So we have lots of hints that hepatitis B - the serum one - is caused by a virus. We know it can get through the filters, blablabla. The person who discovers the marker of hepatitis, or this hepatitis B surface antigen, is a guy called Baruch Blumberg, or I think Barry Blumberg is his nickname. He figured that out in the 1960s, but he didn’t actually know that it was related to hepatitis B. He was a geneticist studying in Africa I think, looking at elephantiasis, which is a condition where people have very swollen limbs after... that’s a parasitic infection, I think. Elephantiasis... parasitic worms. Lymphatic filariasis.
Jacob Trefethen:
Oh yes, yes. Lymphatic filariasis.
Saloni Dattani:
I think elephantiasis is the old name, and the idea is that people’s limbs have swollen so much that they look like they have turned into elephants.
Jacob Trefethen:
Not to be confused with encephalitis.
Saloni Dattani:
... which is inflammation of the brain.
Jacob Trefethen:
Absolutely.
Saloni Dattani:
So he was studying elephantiasis and noticed that different people in Africa had very different susceptibilities to it, and he figured out at some point, I don’t actually remember how, that this was because of polymorphisms in their protein. They had genetic differences that made some people more susceptible than others. There are many examples of polymorphisms related to different risks of diseases; another is sickle cell disease, where some people are protected from malaria and other blood related infections because they have a different hemoglobin protein so their blood cells look more like sickles than bouncy cells.
He was trying to figure out if there were any other examples of diseases where there were polymorphisms that could affect people’s susceptibility to those diseases. So what did he do? He’s trying to find other protein polymorphisms, and the way you can try to find protein polymorphisms is to see if people develop antibodies against other people’s proteins that are not their own.
Jacob Trefethen:
Yeah, just like the blood types earlier.
Saloni Dattani:
Exactly. So he thought, if someone has received many different blood transfusions, they would probably have antibodies to other people’s proteins.
Jacob Trefethen:
Very good point, yes.
Saloni Dattani:
Very good idea. He looked at people who had had at least 25 blood transfusions in the 1960s. He found a man who had hemophilia in New York City. So he had these samples and he tried to see basically, what this person who had hemophilia would react to.
He found that man had antibodies in his blood to a protein found from someone halfway across the world, an Australian aborigine, and he was like, oh wait, this is reacting to a very, very different person’s serum. He called this protein that it was reacting to the “Australia antigen.”
Later on, he found that the same antigen was present in a lot of people with leukemia, and then also a lot of children with Down syndrome, and he didn’t really know why at the time, I think he was quite confused by it. The reason we now know is that it’s because they were much more likely to have been infected by hepatitis B virus, and that the Australia antigen is... the hepatitis B surface antigen.
Jacob Trefethen:
Oh... Whoa! So wait, so this guy, Blumberg, was looking for genetic polymorphisms, not looking for viruses?
Saloni Dattani:
He was looking for protein polymorphisms. Because it was the 1960s, genetic sequencing was really hard.
Jacob Trefethen:
He wasn’t looking for viruses?
Saloni Dattani:
No.
Jacob Trefethen:
He was just out there. He went, I want to go for someone who’s had many blood transfusions. If you have hemophilia, you’re often going to have to have many blood transfusions. He wanted to see if they have antibodies that react against particular proteins. He found one and it reacted against a very distant protein. He was like, wait, hold on a second, hold on a second, hold on a second.
Saloni Dattani:
He’s like, why’s that going on?
Jacob Trefethen:
He found that antigen elsewhere, he found that antigen in-
Saloni Dattani:
a sample from an Australian aborigine.
Jacob Trefethen:
But then in addition, people with leukemia...
Saloni Dattani:
...and children with Down syndrome.
Jacob Trefethen:
And then it turns out that’s because there’s a common source!
Saloni Dattani:
Yeah. The levels of this antigen seem to vary a lot across the world and in different demographics.
And there’s actually a different guy who figures out what it is. There’s a virologist called Alfred Prince, and he figured out that that antigen was part of hepatitis B virus. He, again, was looking at blood samples from people who had received transfusions. What he did, was he looked at their samples before and after they received the transfusions. In 1968, he found a patient who had serum hepatitis, or hepatitis B, and the early samples of their blood didn’t have this Australia antigen, but the later samples did.
Jacob Trefethen:
Wow! Okay, gosh.
Saloni Dattani:
So he’s like, this antigen has appeared after they have developed serum hepatitis. So he concluded that the antigen was part of the virus, which is now called hepatitis B.
Jacob Trefethen:
Well done! That’s, hey, that’s clever.
Saloni Dattani:
I’ll mention some of these names later on, but the next step is to think, well, maybe we can develop vaccines. We now have figured out this one particular viral antigen that people develop antibodies to. Why don’t we develop vaccines with it? You know, in the previous episode, we talked about attenuated vaccines and inactivated vaccines. Why not try one of those routes to develop a hepatitis B vaccine?
Jacob Trefethen:
That seems key to me, because I don’t want to give just any antigen, I want to give an antigen that actually prompts an immune response that then stops the infection.
Saloni Dattani:
Right, that’s what the next person does, they try it out. There’s a scientist called Saul Krugman, who you might have heard of, or maybe not, and he is a first cousin of Albert Sabin — this is going to come up later. He did some experiments that are now considered controversial, actually also slightly after the time.
He was working with a school of intellectually disabled children. They often, the children with Down syndrome, had higher rates of hepatitis B infections. He was trying to figure out if people with hepatitis B infections had the virus in their blood, so he took blood from someone with a hepatitis B infection, he let it clot, and then he injected it into the veins of other healthy children in that same school. Almost all of them got sick with hepatitis, that confirmed that the virus was, in fact, in the blood.
It’s considered very controversial now, because at the time people didn’t know it could cause chronic infections, and also the fact that the children were disabled, but he did take their parents’ consent.
Jacob Trefethen:
But no children can themself fully consent.
Saloni Dattani:
Right. So he did figure out that the virus was in their blood, so now he’s like, let’s see if we can develop a potential vaccine.
He took the infectious serum, diluted it in water, and heated it. He hoped this would kill the virus but keep the hepatitis B surface protein intact. Then he injected that heated serum into the kids, I think at the same school. He gave some of them two doses, some of them one, and then tried again infecting them and seeing if they were protected. And it worked! It protected all of the kids who had received two doses of this heated serum vaccine, and he then showed that was because of the surface protein, so you could use that surface protein to make a vaccine.
Jacob Trefethen:
It is amazing, in that era of vaccinology- the difference back then between the bug itself and the vaccine was not as much. These things were just way more dangerous, my god. Now if you volunteer for a vaccine trial, you’re not getting injected with a virus.
Saloni Dattani:
Right, right. You’re not going to get tested to see if you’re protected by getting injected with the virus itself... unless you’re in a challenge trial.
Jacob Trefethen:
Indeed, indeed.
Saloni Dattani:
The other question I had was, why not make whole vaccines? Why not use the entire virus? But it turns out hepatitis B is really difficult to culture in cells, right?
Jacob Trefethen:
Oh, okay, yeah. That does make it much harder because you want to inject a big volume so that you get enough response.
Saloni Dattani:
So it doesn’t grow very well and it depends on very specific liver cells and liver cell conditions, and that’s very difficult to do in the lab. And you couldn’t also produce it outside of the body in a different living animal; there’s no animal model except chimpanzees and woodchucks, which we found out much later. So we can’t make a traditional vaccine, we’re instead going to try making a hepatitis B surface antigen vaccine, a protein subunit vaccine. And do you know who’s going to make it?
Jacob Trefethen:
I think it’s going to be... Sabin!
Saloni Dattani:
No.
Jacob Trefethen:
Okay... I think it’s going to be... Blumberg?
Saloni Dattani:
No, it’s Maurice Hilleman.
Jacob Trefethen:
Hilleman! Hilleman, Hilleman, Hilleman. Of course, I could have guessed that. He made approximately all of the vaccines.
Saloni Dattani:
Yeah, he made 40 vaccines, so if I asked you for a random one, you might as well have guessed Maurice Hilleman and you’d have been right most of the time.
So in summary, Baruch Blumberg found the Australia antigen, Alfred Prince showed that this Australia antigen was actually a surface protein on the hepatitis B virus - which is the hepatitis B surface antigen - which is produced in 500 hundred quadrillions in an infection. Then there’s another scientist called Saul Krugman who showed that if you inactivate this serum hepatitis, you can create a vaccine, and that it can spread through the blood. And now it’s time finally to develop a vaccine, and it’s time for Maurice Hilleman to develop it.
Jacob Trefethen:
Enter stage left.
Saloni Dattani:
So how did Hilleman produce this vaccine? What do you think?
Jacob Trefethen:
What year are we in?
Saloni Dattani:
We are... I think he started this project in 1968.
Jacob Trefethen:
So I bet he went, well, we know where the protein is, it’s in infected people’s blood. I’m going to guess he went there and took it.
Saloni Dattani:
There was literally that kitchen sink type vaccine that Saul Krugman had made by just heating it up.
Jacob Trefethen:
But he went more systematic, somehow?
Saloni Dattani:
He did! First, he said, let me get some plasma, let’s get some serum from the groups who are most likely to have hepatitis B infections. So he goes to New York City and gets serum from groups who are most likely to have infections at that time, which are gay men and drug users who live in The Bowery in New York City.
Now he needs to find a way to purify their serum and get only hepatitis B surface antigen; he doesn’t want to have anything else because some of them are injecting themselves with drugs. He really wants to make sure it’s not contaminated with something else and also that it’s not contaminated with other microbes - other viruses, other microbes.
He has two plans. The first plan is to make a big continuous flow system, a purification system where you pass the serum through hot water, UV light, formaldehyde, and all of this is just happening continuously. It didn’t really work practically because you had to build that flow system before you tested it, and it was just unfeasible.
Jacob Trefethen:
I don’t know what you mean by flow, as in you have an initial stock of serum?
Saloni Dattani:
Yes, I think what I’m thinking is like a factory line and you start out with serum and it goes through all of these tubes and then it comes out at the end, purified.
Jacob Trefethen:
But you’re not saying that I then get injected with that vaccine. I, then, it’s not like I’m part of the factory.
Saloni Dattani:
No, no, no, no, no, no. You are definitely not part of the factory. So that doesn’t work. Plan B, he tries a bunch of things that he knows are going to inactivate various contaminants. So first, pepsin; pepsin is an enzyme that breaks down proteins. In this case, he’s like, it’s going to break down other proteins in the blood serum, and hopefully it doesn’t break down hepatitis B surface antigen — and correctly, it didn’t. This reduced the number of infectious hepatitis B virus particles by 100,000-fold. But that’s not good enough. He was like, ‘I need to remove every single particle of hepatitis B from this serum. I just want the hepatitis B surface antigen.’
Jacob Trefethen:
He wants the antigen, not the virus.
Saloni Dattani:
Exactly. He doesn’t want it to be able to infect people; he just wants the antigen to trigger an immune response. Okay, so he’s added pepsin. Now he adds urea, which is the protein in urine, and which destroys other proteins as well, so it’s a bit like pepsin.
Urea is really important here because he was scared that if you derive the vaccine from serum, you could be infecting people with prion diseases; he had heard of Creutzfeldt-Jakob disease, which is an infectious prion disease, and he was like, we really need to inactivate those prions, and they had figured out by then that urea broke down the prion proteins that are involved in that. So he’s like, okay, let’s introduce urea as well. That’s going to destroy the prions.
Then next step, he adds formaldehyde. Formaldehyde, as people will probably know, is used as a fixative. Formaldehyde is going to destroy other contaminating viruses; formaldehyde can destroy poliovirus and hepatitis B virus, but it doesn’t destroy the surface antigen.
Now we have these three very important steps: pepsin, urea, and formaldehyde. Somehow the hepatitis B surface antigen is really sturdy and it remains intact even after all of these steps. I think the reason is, one, there’s just so many particles of hepatitis B surface antigen that some of them are going to survive. I think probably other proteins might survive, or other antigens might survive, but you would get rid of other infectious particles. So at this point, after these three steps, there’s a quadrillion fold decrease in infectious hepatitis B virus particles.
Jacob Trefethen:
So you’re saying there’s a chance!
Saloni Dattani:
But he’s still like, you know what, what if this isn’t good enough? So he continues to do more testing and he tries to test for specific pathogens that might still be in the serum. He’s like, maybe let’s test for rabies, let’s test for polio, influenza, measles, mumps, smallpox, herpes, and the common cold. He tested for lots of different viruses and none of them remained. And this plan succeeded.
Jacob Trefethen:
What he’s doing basically is not just targeting hepatitis B virus. Those steps are going to make it hard for any virus to stay intact. But I’m glad he checked, I don’t want to... It’s good to confirm because you’re putting- I mean, you’ve really made me realise... People say this is a subunit vaccine, but really it’s an inactivated vaccine, by which I mean, he inactivated all of the dangerous stuff.
Saloni Dattani:
But if people are not developing an immune response to most of the virus, only the surface virus remains. But it’s also really interesting because by the end of this three-step procedure, the remaining plasma-derived hepatitis B vaccine is almost purely hepatitis B surface antigen. I think that’s because of the 500 quadrillion surface antigen proteins; there’s just so much of it.
Jacob Trefethen:
That’s so interesting because my intuition would be if whatever you’ve done to destroy the stuff in there has not destroyed Hep B surface antigen, I would have thought there’s some other stuff I hadn’t destroyed either.
Saloni Dattani:
Yeah, that’s what I would have thought. I think it’s maybe two things. One, it’s just quite sturdy, and then second, there’s so much of it.
Jacob Trefethen:
Got it.
Saloni Dattani:
The other thing that’s interesting is that all this stuff happened in the 1970s before people had identified AIDS. So some of his sources of serum probably had HIV, but it didn’t matter because all of the steps that he took would destroy the virus anyway.
Jacob Trefethen:
Wow.
Saloni Dattani:
There’s this quote from a microbiologist called Harvey Alter. He says: “He had done all the right things to kill the AIDS virus, even if he didn’t know it was in there.” which I think is so cool.
Jacob Trefethen:
Yeah, that’s astonishing to think about.
Saloni Dattani:
So he has now got a plasma-derived hepatitis B vaccine, at least in principle, and now he needs to test whether it works. What’s going to happen next? He has to convince people to actually take this vaccine to see if it works. Even though he’s done all of these steps to decontaminate it, people are really scared because the vaccine is ultimately derived from the blood of intravenous drug users. Then, in the 1980s, people are like, “Wait, this is from people who might have HIV.” So they’re really terrified. And he has to do human testing. This is going to be kind of shocking, and this was really surprising to me, but guess who comes into the picture now?
Jacob Trefethen:
Uh, Sabin!
Saloni Dattani:
Yes! Albert Sabin, the developer of the oral polio vaccine, for some reason, he seems to hate other people’s vaccines. In the last episode, we talked about how he hated Jonas Salk’s vaccine and he wanted to “kill the killed vaccine.” But it turns out that he also tried to block Stanley Plotkin’s rubella vaccine, because that was grown in fetal cell culture. And he was like, this is very bad. Now he tries to block Hilleman’s hepatitis B vaccine.
Jacob Trefethen:
The Plotkin thickens and the highest Hilleman to climb comes.
Saloni Dattani:
So I was reading this from this book by Paul Offit about Maurice Hilleman and his life. And I have it right here.
Jacob Trefethen:
Paul Offit.
Saloni Dattani:
Paul Offit, ‘Vaccinated: One Man’s Quest to Defeat the World’s Deadliest Diseases.’ Hilleman is sort of recalling this episode and he says: “Albert Sabin hears about it and he says that our vaccine will not be used in any human being. Sabin said that if there was a lawsuit, he would go to court to testify against us. And he would sue Saul Krugman [who was Sabin’s first cousin by the way] if there were any problems with the studies.”
Jacob Trefethen:
Ugh.. stupid.
Saloni Dattani:
So stupid! And Maurice Hilleman’s response to this — this is also a quote — “My feeling was, screw you, Albert.” And he just proceeds anyway.
So now he’s trying to find volunteers to take this vaccine. He is going to find it really hard. So what he does is convince some of Merck’s employees to try it. Maurice Hilleman works at Merck, and he’s like, you know who’s going to take this? My own team!
Jacob Trefethen:
Do you want a good performance review, this cycle?
Saloni Dattani:
So he comes to this big meeting and he says, “I need volunteers, damn it. Just decide who among you are going to take this vaccine. Give yourselves a little bit of time to regain your senses.”
Jacob Trefethen:
Wait, what? He literally said, damn it?
Saloni Dattani:
Yeah. He tries to get volunteers twice. The first time, no one volunteers. The second time, he’s really upset and he’s like, “I need volunteers, damn it.”
Jacob Trefethen:
I wonder if him being pressure-y is why I think nowadays, there’s legislation in the US that means that employees can’t take their own vaccine. Maybe it comes from this pressure, I’m not sure?
Saloni Dattani:
Could be. There is also a quote from someone who took the vaccine in Merck, a woman called Joan Staub. She was one of those who was asked to take the vaccine. She says: “Consent forms? What consent forms? We got that vaccine because we had to get it. If Hilleman told you to do something, you did it.”
Then she learned months later, after she had taken the vaccine, where it came from and that there was a possibility that it might be contaminated with HIV. And she says: “We were scared to death. I thought I was going to die. Maurice pulled us all into one room and had to explain to us over and over again about the inactivation process and that we were going to be okay.”
Jacob Trefethen:
Well, he sounds like a very trustworthy and even-keeled man, so I’m sure I would have been convinced too.
Saloni Dattani:
I think... I would have taken it, maybe.
Jacob Trefethen:
He does seem scientifically trustworthy, to be fair, even if he’s a bit pushy.
Saloni Dattani:
Yeah, he’s very pushy. He was known for being very strict and intimidating. He made his team work seven days a week. They were very dedicated and loyal to him anyway, because he protected them whenever there were layoffs in the company as a whole, and he also made sure that his department’s budget rose 10% every year.
Jacob Trefethen:
I love it. I love- yeah, that’s great.
Saloni Dattani:
So next step, some people in his company have taken the vaccine, they’ve tested it out, and they’ve then done larger trials, and then there’s the manufacturing process. The manufacturing process is not done by Hilleman’s team, but by Merck’s manufacturing division, which was controlled by unions like the Teamsters Union, the Oil, Chemical, and Atomic Workers Union. And there’s this famous story about this, which is why I’m bringing it up.
Because the manufacturing process, it was done by other people, not on his team, he didn’t have that amount of control over it anymore. At some point, someone in the manufacturing division had slightly changed the chemical inactivation process, in the hopes of increasing production. And he was furious to hear about this. There’s a quote that I think maybe you should read out because I don’t think I will sound angry enough, and it will just sound funny coming from me.
Jacob Trefethen:
This is from Maurice Hilleman in response to the unions tweaking his process: “There is no fucking test for absolute safety except to put the vaccine in fucking man,” said Hilleman. “A procedure was developed to make the fucking vaccine and was shown to make the vaccine safe. And there are always fucking people who can’t wait to make fucking brownie points by changing the process to get more yield. You have to adhere to the goddamn process. We know that the vaccine is safe, and you have to adhere to the goddamn process. What worries me is that [someone] will get a bonus if he can get more yield, so he changes the fucking process. Goddamn meatheads are everywhere.” That’s actually the original recording, I think.
Saloni Dattani:
This is actually from a recorded memo. And the “someone” that you mentioned, I think it was censored in the quote, but I think it might have been the name of the actual employee; I think he screamed this at the employees and named the person who did it, but I don’t know for sure what that person’s name was. Does he even have a Cockney accent?
Jacob Trefethen:
Maurice Hilleman, yes. He was actually a famed Cockney. I think Merck at that time was based in East London.
Saloni Dattani:
Wait, wasn’t Merck in the US? Wasn’t he in the US?
Jacob Trefethen:
Well, maybe just the R&D division then.
Saloni Dattani:
You’ve tricked me! He was born on a farm in Montana, near the plains of Montana.
Jacob Trefethen:
“There’s no fucking tests for absolute safety, except to put the vaccine in a fucking man,” said Hilleman.
Saloni Dattani:
You somehow managed to top the previous one.
Jacob Trefethen:
I’ve been practicing my Montana accent for a long time. So, Saloni, you got any more Hilleman quotes?
Saloni Dattani:
I do. We talked about how strict he was, and the fact that he developed 40 vaccines in his lifetime... might have been related to each other. I have this quote from the Paul Offit book, where Paul Offit interviews him near the end of his life to try to understand how he developed all of these vaccines.
He said that his work style was very different from everyone else’s. He said: “If I ever caught anybody delaying a set of tests because [the results] might come out on a weekend, it would be grounds for dismissal. You can imagine how that went over. They all had wives and that sort of thing. Now Merck tells [employees] that they don’t need to put in any extra time and that you have to balance your life and that you have to have enjoyment with your job; [that way] you can do a better job and have fun. It’s all just a pile of shit. What the company should be doing is kicking ass. But that’s from the old school. I was told that I had a very unusual management style.”
Jacob Trefethen:
“What the company should be doing is kicking ass!”
Saloni Dattani:
I mean, you can see how he developed 40 vaccines.
Jacob Trefethen:
Oh, yeah. Maybe we should be making 41 podcasts. We have to start kicking ass, Saloni.
Saloni Dattani:
I think the four hour episodes are a way of doing that. So he has now developed a successful hepatitis B vaccine from the plasma of people who might have had HIV and some who used drugs. A lot of people were still scared at this point, and they then ran larger trials. What was weird about this to me was that there is this conspiracy theory that his trials of the hepatitis B vaccine are the source of the AIDS epidemic. As we have learned, his very strict purification process would have killed the HIV virus anyway.
So someone else is going to make a comeback in the story now, and it’s Baruch Blumberg, the guy who discovered the Australia antigen, which is later called hepatitis B surface antigen. Why is he involved? Well, Hilleman is trying to patent his hepatitis B vaccine and he discovers, strangely enough, that someone has already patented it. So strange! So Baruch Blumberg had patented a hepatitis B vaccine containing hepatitis B surface antigen without making it.
Jacob Trefethen:
Oh my god, not one of those, no.
Saloni Dattani:
He basically has a patent that says that he’s going to make a hepatitis B vaccine through a process that removes impurities, including other infectious components, to attenuate any virus that remains, and that the vaccine would be free of all other blood components aside from the Australia antigen, and that’s it. He hasn’t actually made the vaccine at all; he just has this patent.
There’s a quote from Maurice Hilleman about this, where he says: “People in the hepatitis field were aghast at the guts of this son of a bitch. Somebody had actually issued a patent for that crap.”
So Hilleman then goes to Baruch Blumberg’s company. He now works at the Fox Chase Cancer Center. He tries to convince them to license that patent to him. Initially, when they heard his request, they said yes, but only if Baruch Blumberg can direct the whole vaccine manufacturing process! And Hilleman was like, “Are you fucking kidding me?” And he refuses, obviously, because that guy wasn’t involved at all. He manages to convince them to license the patent to him at a cost, and then he finally manages to license the vaccine and introduces it in the US in 1981.
Jacob Trefethen:
1981.
Saloni Dattani:
But there’s more. Baruch Blumberg is back. Later on, Baruch Blumberg writes a book called ‘Hepatitis B: The Hunt for a Killer Virus’ in which he claims that the hepatitis B vaccine was invented by him and that they had simply licensed Merck to develop it.
Jacob Trefethen:
Oh, no. No, no, no, no, no. No, no, no, no.
Saloni Dattani:
This story is crazy.
Jacob Trefethen:
People need to stop this.
Saloni Dattani:
Why is everyone arguing with each other? But it really is a moment worth celebrating anyway. Because his hepatitis B vaccine “derived from plasma was made from the most dangerous starting material ever used, but was probably the safest, purest vaccine ever made.” That’s according to Paul Offit, who wrote this book, and who co-invented the rotavirus vaccine.
Jacob Trefethen:
You know, I think it is a nice quote, but is obviously false, given that last episode we talked about the rabies vaccine, which is made from literal rabies.
Saloni Dattani:
That is true, but it wasn’t the safest; that wouldn’t have been the safest.
Jacob Trefethen:
True, the first half of the quote is false. The second half, I’m fine with.
Saloni Dattani:
Yes, I agree.
Jacob Trefethen:
But I have a question for you. Because 1981, that’s far enough along... that I think we have a new technology that we could maybe use here? Because in 1977, Genentech went splashy with recombinant DNA, so hold on a second, does that get involved?
Saloni Dattani:
That’s next. So Hilleman’s program to develop the hepatitis B vaccine from plasma took 13 years, and that includes the vaccine trials and whatever other development they had to do. I think by the time recombinant DNA technology became usable, that was like 1978 or ‘79 when they first cloned insulin? So yes, recombinant DNA technology is available, but it’s still pretty new. And it would have taken much longer to get through the whole pipeline.
Jacob Trefethen:
Got it.
Saloni Dattani:
But that is next. So even though he’s made this incredibly safe vaccine, people are really scared still about using it. People are very uncomfortable because it’s made from human blood. So he decided, let’s try to find another way to make it: recombinant DNA technology!
Jacob Trefethen:
Doo doo doo!
Saloni Dattani:
Do you want to do a quick recap of what that is?
Jacob Trefethen:
Sure. So the end goal, as a reminder, is to make protein, in this case, the hepatitis B surface antigen - that’s a type of protein. How do you make proteins? Well, the body makes it by DNA, transcribing to RNA, translating to proteins. So you want to copy that gimmick, and the way you’re going to copy it is you’re going to take a DNA slice that is going to code for the protein you want, and you’re going to inject it - pleurgh - essentially, into a system that’s not human. Maybe it’s a bacteria, maybe it’s yeast, some other cell that you can get it to be produced en masse. That’s the gimmick, I’ve cut out some of the detail, but that’s the gimmick.
Saloni Dattani:
Yes. So that happened in the 1970s. The first recombinant protein that was made was human somatostatin and then insulin in 1978, and that transformed diabetes treatment, as we talked about in our third episode. Next, people think, hey, let’s use this to make vaccines better!
Why do they want to make vaccines better? I think one, this purification process is quite difficult; you have to rely on getting plasma from people. It’s also various steps and you have to check for contaminants along the way. Also, the amount of yields could in some cases be quite low, whereas with recombinant technology, if you find out general ways to improve the yields, you could find ways to scale up production by quite a lot. And it’s much safer, so you aren’t that worried anymore about accidentally contaminating people with other viruses or with other blood products or drugs or things like that.
Jacob Trefethen:
Yeah, instead of getting rid of everything and getting left with only the hepatitis B antigen, the only thing you’re making in the first place is the hepatitis B antigen. Who are we talking about here? Is this still Merck or someone else?
Saloni Dattani:
This is still Merck.
Jacob Trefethen:
Ah, so the innovators’ dilemma. They decided to eat their own lunch.
Saloni Dattani:
Yes, but in this case, I think people were just still uncomfortable with using this plasma-derived vaccine. So they were now like, okay, well, we do know quite a lot about hepatitis B, so why don’t we just make a better version that everyone will take?
So now they decide to use Genentech’s skills and all of these new recombinant DNA technologies to produced- to mass-manufacture a hepatitis B surface antigen. The first recombinant DNA molecule was made in 1972 by Paul Berg. Then Stanley Cohen and Herbert Boyer, who worked at Genentech, then improved upon that process; they developed restriction enzymes, which basically cut up DNA, and then they found a way to insert that into bacterial plasmids. Then Boyer’s lab found ways to purify the enzymes and this whole system, so that they could replicate the plasmids. Basically, you can grow proteins from other organisms in bacteria or yeast in a little bacterial manufacturing tank or something like that. They had essentially invented this cloning system so you can clone genes and produce the proteins that they code for; they can use bacteria and yeast as factories to make proteins that previously were only available from blood or tissue.
So why not make loads of hepatitis B surface antigen, but make them in... so they did manage to produce it in bacterial plasmids with one of the researchers at the UCSF group who worked with Stanley Cohen. But this guy was called William Rutter.
Jacob Trefethen:
Oh yeah, Bill Rutter. Yeah.
Saloni Dattani:
You know him?
Jacob Trefethen:
Yeah, of course. He’s a famous biotech investor. I think he actually sadly passed away this year.
Saloni Dattani:
Oh. And so they get William Rutter, they recruit him, Bill Rutter, and he is working at UCSF. Bill Rutter removes the hepatitis B surface antigen from the virus and inserts it into one of the bacterial plasmids. He then makes the bacteria reproduce; they make large quantities of the hepatitis B surface antigen. But unfortunately, this doesn’t induce an immune response in animals, so now they’re going to try something else. They’re going to try yeast; they’re going to use common baker’s yeast, which must have a more specific name, but I don’t know what it is.
Jacob Trefethen:
Wait, I’m lost. Why does it matter that it doesn’t induce an immune response in animals?
Saloni Dattani:
Well, they wanted to have some indication that it would be effective. They did manage to reproduce hepatitis B surface antigen in the bacteria, but it seems to have some kind of different configuration, which means it’s not causing an immune reaction.
Jacob Trefethen:
Got it, sorry. It wasn’t absence of evidence; it was evidence of absence.
Saloni Dattani:
Yes, yes, yes. Yup. I guess one thing that I wasn’t super sure about was how come, sometimes, the recombinant DNA technology varies depending on which species you insert it into? Like how come it’s sometimes working in yeast and sometimes it’s working in bacteria? Some species produce it better than others?
Basically there was research showing that yeast cells can also produce the hepatitis B surface antigen and assemble those proteins into particles that look very similar to the ones that are made in human plasma and that are immunogenic in mice.
This is when electron microscopy is useful; you can actually see the shape of the proteins. So I have this image here, if you’re watching the video, of these very interesting looking hepatitis B surface antigen particles. They look kind of like individual cereals, like breakfast cereal.
Jacob Trefethen:
Yeah, I would say a little... maybe... corn pops?
Saloni Dattani:
Yes. Very tiny corn pops.
Jacob Trefethen:
Very tiny corn pops and probably a bit less tasty.
Saloni Dattani:
Oh my God. I wouldn’t want to eat that.
Jacob Trefethen:
Do you think they make milk go yellow?
Saloni Dattani:
They make humans go yellow.
Jacob Trefethen:
Oh! The original corn pop!
Saloni Dattani:
Basically the yeast are producing the virus’s proteins much more closely than our own cells would produce them. Viruses that depend on our own cell’s machinery will have some other modifications, after just producing the protein itself. In order to try to replicate that further modification, it can often take a long time to figure out how exactly to do that, which specific type of system to use for recombinant DNA or which optimization pathway you can use to make sure the shape and the structure match the real version.
Jacob Trefethen:
And I think that’s still true.
Saloni Dattani:
So Hilleman develops another recombinant hepatitis B vaccine by making the hepatitis B surface antigen in yeast. That, he finds, creates protective antibodies in chimpanzees and later in people. He uses this to make the next hepatitis vaccine.
Jacob Trefethen:
Woo!
Saloni Dattani:
Then it’s licensed in 1986 and it’s still used today.
Jacob Trefethen:
Wow. Okay. So we went full circle. To summarize, how did we get to this vaccine that is made with recombinant DNA and is going to protect you against hepatitis B? Well, we had to do the whole drive of immunology. What actually is a productive immune response against infections and against different infective agents?
We had to then try and isolate the parts of the hepatitis B virus, instead of the whole thing, that generate a productive immune response. In this case, just one protein, the hepatitis B surface antigen. And the wonderful interlocking of this idea of a subunit vaccine — just one thing, not the whole thing — with recombinant DNA that came in the 70s and 80s, is that now we have a way of producing proteins really easily.
And proteins are often the one part of the virus you want to use as a vaccine. So, lo and behold, we have a lot of yeast cells in a big vat churning out hepatitis B surface antigen, and that can be used as a vaccine. And you know what? I think I’ve got that vaccine.
Saloni Dattani:
Oh, really? I guess I have as well. I don’t remember getting it though, I guess I was a baby at the time. So I think in 1981, the first one was introduced in the US and then 1986, the recombinant vaccine was introduced.
How much of an impact did it have? Just to recap, we said that newborns who are infected with hepatitis B at birth or soon after have about a 90% chance of developing a chronic infection from it. And then 15 to 40% of them, who have that chronic infection, will eventually die from liver failure, cirrhosis, or liver cancer if they’re not treated.
So the impact this vaccine is going to have is actually massive. The way that we know that is from a large randomized control trial that was done in Qidong in China. There, entire towns were randomly selected to give hepatitis B vaccine to all of their newborns who were born there, and some towns were not.
Jacob Trefethen:
So a cluster randomized trial, yeah.
Saloni Dattani:
Right. We have entire towns getting it or not getting it. The reduction that you see in the vaccinated towns is an 84% reduction in the incidence of liver cancer and a 70% reduction in deaths from liver disease.
Jacob Trefethen:
So this is a decades-long study?
Saloni Dattani:
Yes. This study was done, well, they were followed up over more than 20, almost 30 years. The thing about China’s health system is that everyone born there has an ID and that you can track that ID decades later in their medical system and see what happened to them.
Jacob Trefethen:
That is a big and long trial. Most products, we do not get that kind of information.
Saloni Dattani:
But of course, we did know that it was protecting against liver disease and hepatitis infections much before that. So they knew that it was going to have this reduction, but I think they probably didn’t know how much of a reduction it would have.
I have this chart here showing the decline in hepatitis B after the vaccines were introduced. In 1987 in the US, it was recommended that all parents who have hepatitis B surface antigen in their bodies — meaning that they have been infected — should have their infants get vaccinated. Then in 1991, it was recommended that all infants actually should get vaccinated. You can see this gradual decline in hepatitis B cases after that.
I think it’s interesting because this kind of graph actually looks quite different from the other types of graphs you often see when a vaccine is introduced. There isn’t just a sudden drop after the vaccine is introduced.
I think the reason for that is, one, there’s a long delay between infection and the disease, sometimes months, sometimes years. I think the second is that generally it doesn’t spread like other diseases in an epidemic. It’s mostly spread from mother to child. So it’s not going to have this herd immunity effect; it’s not going to massively reduce the number of cases very soon, but it’s more of a generational thing. Mothers are not going to pass it on to their children if they’re chronically infected. So there’s a much slower, but still in the long term very large, decline from hundreds of thousands of cases of hepatitis B in the 1980s and 90s to only less than tens of thousands now.
The other thing that was interesting was, they have testing for hepatitis B surface antigen, so they can test if the mother has been infected. But it’s still important to move to universal vaccination to, one, just skip that process; a lot of people are just going to miss that testing procedure, so you might as well just give it to everyone, also because it’s extremely safe. And I think the other reason is that even if infants don’t get infected at birth, they can still get infected after that. That’s quite common in some places where young children often get infected from contact with other people’s blood or small wounds or injuries or shared objects that they’re using. So just getting it to all newborns instead means that they’re protected from all of these exposures, which is really cool.
Jacob Trefethen:
On hepatitis B, lots of people around the world have chronic infections. Actually, globally, probably 300 million people have chronic infections, and most people who do don’t know that they do. Just for listeners out there, if you find out you have a chronic infection, it’s not the end of the world. There are drugs you can go on to help control the infection. There’s not complete cures, and hopefully that will change in the next generation, but there are treatments that can help. So it’s definitely worth getting tested and your doctor will be able to advise when you should go on treatment for what different level of current infections.
Saloni Dattani:
Nice. What are the treatments? Are they antivirals?
Jacob Trefethen:
Yes, they are. They are nucleoside analogs, which actually some of them also came up in our HIV episode, because they can be prescribed as PrEP for HIV.
Saloni Dattani:
Oh, so they block the virus from replicating.
Jacob Trefethen:
And it’s a good reminder that a lot of progress in one area can help progress in another. We have seen that over the course of this episode where progress in immunology on hepatitis B, guess what, it’s ended up helping other viruses. Progress on recombinant DNA for the sake of insulin, guess what, it’s helped hepatitis B. So thank you, interlocking strands of science.
Another form of innovation from one area helping another is: you know how they make a malaria vaccine?
Saloni Dattani:
Yes, with the hepatitis B surface antigen.
Jacob Trefethen:
With the hepatitis B surface antigen. I’m afraid you’re just going to have to let that linger because we are not going to get all into it now, but it self-assembles into this beautiful viral particle, those corn pops we saw earlier. Now imagine if the corn pops had little malaria antigens laced through them.
The first malaria vaccine: stir up one out of every four is a malaria protein. And then they’re going to - whoo, putiputoo - have a tendency to form a nice little corn pop. The second malaria vaccine: stir up way more. Hardly any hepatitis B, more malaria. And it just happens that that happens to get attracted together into a beautiful little self-assembling little ball.
Saloni Dattani:
You know, we didn’t talk about that. We didn’t talk about why does it self-assemble into a sphere? The surface antigen has regions that are water-loving and fat-loving, which makes the proteins gather in curved shapes.
We made this huge impact with this one hepatitis B vaccine, and it’s just the first protein subunit vaccine and it’s just the first recombinant technology vaccine that’s made. Then there are many more that are made after that. That includes the human papillomavirus (HPV) vaccines, some influenza vaccines like FluBlok, which is a recombinant hemagglutinin vaccine. The shingles vaccine is also a recombinant, Shingrix. There’s a hepatitis E vaccine, which is also a recombinant vaccine. And then there’s a few more, I think meningococcal vaccines, MenB, are a recombinant.
Jacob Trefethen:
And if a vaccine is recombinant, that also means it’s a protein subunit vaccine. So we’ve gone from the whole virus, or the whole bacteria, weakened to only a part, a subunit. And that is, in general, going to be much safer.
Saloni Dattani:
Yeah. And there are many other types of vaccines alongside the protein subunit or the recombinant DNA technology ones, including polysaccharide vaccines or toxoid vaccines or viral vector vaccines, and finally mRNA vaccines. We might talk about them in future episodes... if we feel like we should.
Jacob Trefethen:
If you subscribe.
Saloni Dattani:
Yeah. So if you’re watching the video, you might notice that I’m wearing a shirt with the names of some vaccines on them. It is an acrostic, which says VACCINATED, and it has the names of many different vaccines.
Jacob Trefethen:
Stand up again, stand up again. COVID, Measles, Varicella, Meningococcus, Polio, Tetanus, Hepatitis, Pertussis, Influenza, Diphtheria. VACCINATED!
Saloni Dattani:
I love this shirt because I saw someone on Twitter wearing it as a hoodie, and they posted a picture with it. And I was like, I want that.
Jacob Trefethen:
Yeah, that was made for you. Also, I like that when you sit down, we can only see the first couple. So COVID will be in shot for people throughout, and everyone’s going to get anxiety.
Saloni Dattani:
Well, I’m sorry.
Jacob Trefethen:
So I think we should talk about how we even got here to those modern subunit vaccines, hepatitis B and the others you mentioned. This wasn’t just the story of recombinant DNA, and of microbiology and learning about those different microbes; it was also reliant on many breakthroughs in immunology - in what prompts your immune system to actually create a protective response in the future, and understanding how vaccines actually work in that way.
So that’s why many modern vaccines are quite different than the ones we talked about last episode, because of these immunology breakthroughs. Those older ones were mostly whole viruses or whole bacteria that were killed or inactivated.
Saloni Dattani:
I mean, that’s really interesting, right? Because the vaccines that we talked about, so many of them were developed without even knowing which virus they were for or what the microbe was. People developed vaccines with these systematic methods through attenuation or inactivation, but it wasn’t until breakthroughs in immunology until people figured out how they actually worked and how to make them much more precise, safe, and sometimes more effective. Sometimes as we’ll talk about, it’s difficult or it’s impossible to make vaccines with those older methods, without these newer tools that we have today.
So let’s do it. Let’s talk about how we got here. But in order to do so, we’re going to have to go all the way back to the 19th century.
Jacob Trefethen:
Okay, let’s get back in that time machine. *time machine sound effect*
Saloni Dattani:
Yeah, so as we discussed, Pasteur, Koch, and all these other people who were working on germ theory and trying to make new vaccines were doing it through attenuation and through inactivation. They were trying to force the microbes to evolve under different conditions or trying to inactivate them with chemicals or other procedures to weaken them so that we would recognize the pathogen or the weakened pathogen and be able to attack it in the future.
But while they knew how to make some vaccines, they didn’t really understand why they worked. They didn’t understand immunology, and all of that was going to be discovered in the next few decades.
So maybe we should start with what the different theories were at the time. I think I mentioned at one point that Pasteur thought vaccines worked because they depleted the body of nutrients that that specific disease needed. And if you had depleted them once through a vaccine, then the next time it wouldn’t be able to attack you again.
There are just other mysterious reasons that people thought you would be protected from a second attack. I guess at this time, people just thought that, well, there was the very popular theory of the imbalances of humors.
Jacob Trefethen:
Yes. Humoral imbalance. What’s a humor again?
Saloni Dattani:
What is a humor? A humor... Well, it’s not very funny, they’re all quite disgusting. One of the humors is blood, the other is phlegm, and then there’s yellow bile, and there’s black bile.
Jacob Trefethen:
Right. Tag yourself.
Saloni Dattani:
This was an ancient theory by Hippocrates, so thousands of years ago. I guess I can sort of see how you might think that, because sometimes people are bleeding, sometimes people cough up blood, sometimes their skin looks yellow. Sometimes their liquids, their fluids look black and they’re spitting up lots of mucus and phlegm. I don’t know, I guess I can see how you might think something’s going wrong there.
Jacob Trefethen:
I guess I see blood and phlegm way more than yellow bile and black bile. What about you?
Saloni Dattani:
I think... I mean, as we’ll talk about, I guess yellow bile maybe refers to jaundice or something, or maybe it’s like vomit. What do you think?
Jacob Trefethen:
Oh, maybe... I don’t know. I guess I do see vomit sometimes.
Saloni Dattani:
Or is it diarrhea?
Jacob Trefethen:
Black bile is old school. I feel like if I ever saw black bile 2,000 years ago, I’d be like, “That’s a demon. That’s coming from a demon.”
Saloni Dattani:
Well, that’s scary. What’s so funny about the theory of humoral imbalance is that people kind of continue to believe this for hundreds, or thousands, of years after. So even in the 19th century, a lot of scientists still basically think that all diseases are caused by this imbalance of humors.
What changes is one, I think, microscopy, and people start to study human organs and tissues and cells and stuff. And some people who are studying this, pathologists, are claiming that, “Okay, no, you know what? It’s not because of humors, it’s because of cells, and it’s the cells that are malfunctioning.”
There’s this famous German pathologist called Rudolf Virchow, and he thinks that- in 1858, he tries to challenge humoral theory, and he says all diseases are caused by the malfunction of cells. What is also notable about him is that his colleagues referred to him as the Pope of Medicine.
Jacob Trefethen:
Oh?
Saloni Dattani:
I don’t know why. I think it’s because he was involved in politics or something.
Jacob Trefethen:
Well, yeah. I mean, I feel if black bile were demons, you do want a pope to bless you away from them. But if he believes in cells, it doesn’t quite add up.
Saloni Dattani:
No, it doesn’t... So he thinks, okay, everything is caused by the malfunction of cells instead. At the time, in the mid-19th century, people are starting to learn about infectious diseases that are caused by specific microbes. So this ‘germ theory’ is starting to gain acceptance. There’s another guy who comes on the scene called Ilya Metchnikoff. Have you heard of him?
Jacob Trefethen:
I think I have, but I couldn’t tell you why.
Saloni Dattani:
So he was a zoologist living in the Russian Empire, and he was born in present-day Ukraine. He had this theory of “phagocytosis”, and you probably know what that is.
Jacob Trefethen:
Yes, I do know that one, yeah. Cytosis, that’s to do with cells, and phago is... Oh god, the derivation, I don’t know.
Saloni Dattani:
Eating.
Jacob Trefethen:
I’m trying to think of another word that has that in there, anyway. But...
Saloni Dattani:
I don’t know if any others do, I guess it’s Greek. But eating cells and killing them. He looks under the microscope and he suggests that there are cells that can eat microbes; they’re phagocytic cells. He says these phagocytic cells, which appear during inflammation after an infection, they’re actually not harmful. He says “they’re the first line of defense because they can ingest and digest invading organisms.” That was quite radical at the time.
Jacob Trefethen:
Do you ever play that computer game Agario, where you had a little blob that had to go around eating smaller blobs until you got bigger and bigger?
Saloni Dattani:
Oh, I haven’t played that one, but I feel like this is also like Snake.
Jacob Trefethen:
Yeah, it’s like Snake. So you’ve got to imagine the snake is actually good instead of bad, and it’s one of your cells. But if I’m a cell and I eat a microbe, do I get bigger? Might temporarily.
Saloni Dattani:
I think you... I think the cell does the equivalent of pooping it out.
Jacob Trefethen:
Right.
Saloni Dattani:
I don’t know, the extra stuff gets exuded.
Jacob Trefethen:
Right. So it’s sort of like a snake swallowing an egg and the egg goes really big, big, big, big, big, big. Got it.
Saloni Dattani:
So Metchnikoff says that actually inflammation is good. And people were like, “What?! No, it’s not!” And he’s also like, actually, the way that we develop immunity to things is by having our cells eat the invaders.
Let me just tell you how he thought of this. He has this quote in his autobiography. He says: “One day, when the whole family had gone to the circus to see some extraordinary performing apes, I remained alone with my microscope, observing the life in the mobile cells of a transparent starfish larva, when a new thought suddenly flashed across my brain. It struck me that similar cells might serve in the defense of the organism against intruders.”
So he has this thought that comes to him about this starfish, and he’s like, well, maybe our cells are like starfish that eat up different intruders. He tries to run an experiment to see if this actually happens. He pokes a starfish larva with a splinter, and then he looks under the microscope at what happens next. He sees mobile cells moving around and surrounding that splinter. And he says: “That experiment formed the basis of phagocytic theory to the developments of which I devoted the next 25 years of my life.” A long time.
He studies these phagocytes and their digesting of different bacteria: the anthrax bacteria, erysipelas (which I think is Streptococcus pyogenes), typhus, tuberculosis, and then many other bacterial infections. And he’s like, all of these infections are actually destroyed by cells that eat them up.
Jacob Trefethen:
I think that’s an amazing experiment because it’s so crisp, and to see the mobile cells. And it is worth just saying- when I think about cells in my body, I do think about them as pretty much fixed in place most of the time. I’m thinking of the neurons in my head, they’re not moving. The neurons in my gut, they’re not moving that much. The skin cells, well, maybe I’ll get some new ones if I put on the right lotion. But basically the cells, I don’t think move that much. But of course, the immune system needs to have cells that move.
Saloni Dattani:
Yeah, and sometimes when you look under the microscope, you actually will see these phagocytes moving around and engulfing the bacteria and eating them up. This is especially true with one of the bacteria that he loves to study, the anthrax bacteria. Anthrax, as we’ll come to later on, is quite resistant to other forms of immunity. So he basically only sees it getting eaten up by phagocytes. And he says, “Well, this is how immunity works.”
And lots of people are kind of angry about this. Some of them are the humoralists. They’re like, ‘No, you’re wrong. It’s actually the humors. The Greeks were right.’ There are other people who try to run more experiments and they aren’t really seeing the same thing as he is seeing. When they’re looking at other diseases, they’re like, the phagocytes aren’t succeeding; they’re not able to ingest the organisms. They’re not destroying them. Or they’re just secondary. They’re just Robin and Batman is the humor.
Jacob Trefethen:
Okay.
Saloni Dattani:
That’s a terrible analogy.
Jacob Trefethen:
I mean, Robin’s important, so let’s not do him injustice.
Saloni Dattani:
That’s true.
Jacob Trefethen:
That makes sense, and I can think of other reasons why the phagocytosis view can’t be right, but that’s all with the hindsight of what I now know about the immune system. If I put myself in their headspace, I’m like, okay, well, there’s definitely some invaders that are bigger than cells! You know, like if I get a tapeworm, I’m not going to be able to eat that with a cell, but maybe I will be able to develop some immune response? So I don’t know, I could think of some ways it can’t be the full story.
Saloni Dattani:
Right. I mean, I could imagine maybe the phagocytes are just slowly biting away, but no, I don’t think that’s actually going to work. And the other thing is that he made a lot of wild claims beyond these. He basically claimed that actually the reason for aging is also phagocytosis. The neurons in our brains are getting eaten up by phagocytes, and he says, our hair goes gray because phagocytes eat up the hair pigments.
So he’s making some crazy claims and he’s making some legit claims. But ultimately, he loses the argument, and the humoralists win. So there’s this big debate between the cellularists like Metchnikoff and the humoralists. The other kind of political aspect to this is that Metchnikoff is living in Paris and he’s working at the Pasteur Institute, and a bunch of his team and his students become cellularists. And then in Germany, humoralism is much more popular, so there’s actually a nationalistic element to it as well. They hate each other at that point.
Jacob Trefethen:
So German humour... Hmm... does exist after all?
Saloni Dattani:
Yeah, what’s so funny about this is that the humoralists win for various reasons. So the humoralists, they’re not just arguing about black bile and everything - they’re saying something in the blood is responsible for attacking pathogens, and they can actually find pretty good evidence of this.
If you take the serum of someone who has previously been infected with a disease, and then you introduce that serum into someone else, it protects the second person from an infection, often. That’s something that scientists find in the 1890s against diphtheria and some other diseases. So they’re like, oh, look, it’s just the blood itself that is protective. And you can also see that in vitro; you can see that sometimes the serum is able to destroy bacteria. This basically discredits the argument... quite significantly. The humoralists have won out, they’ve convinced the rest of the scientific community.
Jacob Trefethen:
No! I don’t think it’s true. I think both are true. I’m cheating. I’m thinking ahead.
Saloni Dattani:
No, no, you’re right. And it’s kind of sad because in the late 19th century, the humoralist theory had won-
Jacob Trefethen:
That’s sad. I didn’t know that.
Saloni Dattani:
-people just ignored cellular immunology for decades until basically the mid 20th century. They just focus on the serum. They just focus on antibodies and complements and things like that. They only figure out that immune cells produce antibodies in the 1930s and 40s.
Jacob Trefethen:
No! My heart.
Saloni Dattani:
So they don’t know what T-cells are until 1961.
Jacob Trefethen:
Right. Just to give the context, antibodies are not cells, they are proteins. And that is a big distinction. I can’t be too mad about people getting obsessed with antibodies because I probably would too. But at that time, they thought they were the be-all end-all.
Saloni Dattani:
Yes, so antibodies are... People know about them; they have never seen an antibody and no one will see an antibody until the 1960s, until electron microscopy. Until then, you’re basically seeing the reactions that form through antibodies. You’re seeing, for example, precipitin reactions, which is just when antibodies clump together and get mixed with their antigens and form a clump. You can see the effects of that and you can quantify the amount of antibody, but you don’t know what it looks like and you don’t know what it’s made of. So I think Paul Ehrlich at some point figures out that it’s a protein.
But how it was discovered? In the early 1900s, Jules Bordet and Maurice Arthus described these precipitin reactions. So if you mix up the serum with the antigen, then you would see an aggregate forming at the bottom of a tube, because the antibodies are cross-linking many antigens and they make this lattice and they sort of form a little precipitate. So yeah, you can see that, but you have no idea what they look like.
We now know a lot more that would have kind of helped resolve that debate a bit. So we now know antibodies are Y-shaped proteins. They’re produced by immune cells that circulate the blood and the lymph. Back then, they just knew about antibodies circulating in the blood and the lymph somehow; they know that they bind to pathogens like bacteria or toxins and they block them. They also think know about complement; they know that some microbes get covered in something that attracts or causes them to get destroyed.
Jacob Trefethen:
So wait, just too many compliments can destroy you?
Saloni Dattani:
Yes. So complements is a group of proteins that interact in the blood. They go through a series of enzymatic reactions, and after that, they can coat microbes. We now know that they can attract immune cells to destroy them. And they would have known back then that complements can create little holes in microbial membranes and kill them, that’s very useful for extracellular pathogens which are in the bloodstream. So now we know that there is this humoral aspect to immunity with antibodies and complement.
And we also know that there’s a cellular aspect as well, so there’s B cells which have antibodies on their surface and they recognize antigens, and they produce many antibodies that circulate in the serum; and then there’s T cells, which help them out, help them mature, and can also kill cells that themselves are infected. And there’s phagocytic cells, like the ones that Metchnikoff described, that actually eat up microbes and debris and other waste.
So now we know there’s kind of both of these different aspects, and there’s coordination between them as well. But back then they were like, okay, cellularism is basically done. Maybe it only is true for some situations like anthrax, but mostly it’s antibodies. Antibodies are the future.
Jacob Trefethen:
I wonder what we currently think is done that, in 50 years, is going to have a resurgence.
Saloni Dattani:
I’m going to bring this back up later, but in the 1960s, people basically thought that immunology was all solved. They were like, we know everything; there’s nothing else to discover. And I think they probably think that back then.
Jacob Trefethen:
That’s amazing. Now I want to pick up a modern immunology textbook and see how many pages are based on things published after 1960. It’s probably the vast majority.
Saloni Dattani:
Right. But let’s go back. We have Metchnikoff, and he’s like, it’s all cells. And then we have the humoralists, and they’re like, no, it’s the serum, and it’s the antibodies within them. In a way, they’re happy that it’s the serum. They’re like, “Guess what? The Greeks were right. They had so much foresight, and they knew all of this without having the specific, precise understanding. And we’ve figured out now why they were right.”
Essentially, they’re like, “Let’s just look at the serum. Let’s see what’s going on in there.” So various people are doing experiments in the serum. One of them is Paul Ehrlich, and he develops tests to quantify the amount of antibodies in the serum. Then there’s Karl Landsteiner. Have you heard of him?
Jacob Trefethen:
Again, I recognize the name, but couldn’t tell you why.
Saloni Dattani:
He discovered blood groups in 1901.
Jacob Trefethen:
Karl Landsteiner?
Saloni Dattani:
Yes, an Austrian doctor.
Jacob Trefethen:
That’s a big one.
Saloni Dattani:
It’s funny because his name isn’t that well known now, but he would have saved so many people through making blood transfusions safer and discovering the ABO blood groups. The way he discovered this was he tried to mix blood from different people and saw how red blood cells clump together in some samples, but not others. Just by doing this systematically and comparing them, he found different patterns: A, B, and O, and that revealed that people carry antibodies against the blood group antigens that they don’t have.
I have this silly story from when I was at school. We were preparing for an exam and we were very stressed out the day, the morning of the exam. A friend in our group was telling us, “You know, you all just got to be positive.” And another friend of ours was like, “Hey, that’s my blood group!”
Jacob Trefethen:
I thought you were going to say, “That’s my grade!” But for you I think it would have been A positive. What blood types make me think of is the British radio comedian, Tony Hancock. Have you heard of him?
Saloni Dattani:
No. Why does it make you- who is that? And why do you think of that?
Jacob Trefethen:
You weren’t around in the 1960s in the Midlands? Well, he had a TV show and a radio show; I only know the radio show. I used to a family friend gave me this one episode of it, The Blood Donor, which I listened to on repeat as I was falling asleep or if I was ever feeling sick. It was a comedy. The main character, Tony Hancock, goes in to donate blood. The doctor taking the blood sample starts with a little prick or a little smear. And Tony thinks that’s the whole donation done. And so he goes, “Okay, cool.” and he heads off. And the doctor goes, “No, no, no, that’s just the initial test. We’re going to take a pint.” And he says, “A pint!? That’s almost my whole arm!”
But the real reason I remember it is because the way that he then, the doctor then gets him to do the donation is by saying, “Well, you have a very rare blood type, you know?” He goes, “Oh, oh, me? Oh, yeah.” And every time you need to get someone to donate blood, you just have to tell them, “Well, you know, you have apretty special blood type.”
Saloni Dattani:
Do you donate blood? I feel faint when I get blood taken, but maybe I have to donate blood these days after all.
Jacob Trefethen:
Do you donate blood?
Saloni Dattani:
I don’t, but it’s because I don’t qualify. I’m too small.
Jacob Trefethen:
They don’t let you do it if you’re small?
Saloni Dattani:
I’m too small. I don’t know, maybe they’re like, well, if we take blood from you, that’ll be it. You’ll be depleted. There won’t be any of you left.
Jacob Trefethen:
“We’re going to take a pint.” “Oh, she’s only got about a pint and a half.”
Saloni Dattani:
Which is so sad because I would like to donate blood. I feel like it’s cool. I like getting injections and I like seeing my blood taken out.
Jacob Trefethen:
Oh, that’s crazy, Saloni. Absolutely not true for me. However, one day I hope it will be.
Saloni Dattani:
Anyway, so Karl Landsteiner discovered blood groups. Blood groups are really interesting because they’re just really tiny differences in chemical structure that lead to huge differences in our reaction. If you get transfused with an incompatible blood group, your antibodies recognize that and then create this huge reaction to it; that amount of specificity of just these tiny chemical differences lead to huge differences in a medical response are what’s really key here.
Landsteiner tries to do more experimentation and tries to figure out how specific these antibodies are. So what he does is he injects animals with foreign proteins - he injects them with serum from other species - and then they develop antibodies towards it. Then he mixes those antibodies with different protein samples, and it’s only recognizing the same exact one that it was introduced to before and not with any other related species.
This suggests that there’s extreme specificity, there’s extraordinarily subtle differences in molecular structure that antibodies can recognize, and that even proteins from very closely related species can be distinguished by the body. This means antibodies are very specific and they’re also very diverse; he knows this in the 1910s, way before anyone has any idea of why. So everyone is now wondering, why are antibodies so specific? I wonder if you could go back to the 19th century and have a guess using that amount of knowledge, what would you think is the reason?
Jacob Trefethen:
Well, it’s so hard to screen off the knowledge I already have. Because the puzzle for me is how can they be so specific given humans only have 20,000 genes? But they didn’t even know humans had 20,000 genes, so they didn’t even have my puzzle.
Saloni Dattani:
They didn’t even know about DNA back then.
Jacob Trefethen:
They didn’t even know about DNA, let’s see, yeah, it was 1940s that DNA was thought to be the conveyor of genetic information, and double helix is 1953. Going all the way back then, what would I have thought? I would have thought, “Hold on a second. You mean to tell me that I have all sorts of stuff in my blood that’s ready at any moment to bind to anything? How does it fit in there? That would be crazy.”
And now we know that actually it’s not there the whole time; you actually promote certain types from the B cells that they didn’t know about. But how the bloody hell would I- oh my gosh. I think I would have thought if these things in blood can do all this magical binding, then the whole, the main purpose of blood is antibodies. That’s probably what I would have thought. I would have been like, cause that’s, that’s crazy, it can do whatever.
Saloni Dattani:
Let me tell you what they thought, which is kind of an inverse. They basically thought that antibodies didn’t exist until the antigens were introduced. So they thought that antibodies formed from the toxin, or from the antigen.
Jacob Trefethen:
So every package contains its own cure?
Saloni Dattani:
Kind of? That’s one version of it. The other version is that it’s a mold. There’s this guy called Hans Buchner in the 1890s. With diphtheria, for example, it releases toxins, right? So he was like, the antibodies to that, which are called antitoxin, are formed directly from the toxin itself. And that became very quickly accepted, but it didn’t really make very much sense even back then.
Émile Roux, who we talked about in the last episode, basically showed that if you injected a horse with tetanus toxin, and then you kept bleeding the horse, it didn’t reduce the quantity of antibodies... even after you’d removed the entire original blood volume, eventually. So I don’t know, I feel bad for the horses, but you still managed to keep that.
Jacob Trefethen:
I’m realizing that I would have done better than these idiots. Here’s another thing I would have noticed. I think I genuinely would have known back then that people or kids who are less well-nourished get more infections. I probably would have noticed, “That has to be to do with the food. That can’t be to do with the invader.” And sure enough, we now know the food gets broken down and used by your immune system. But I would have come up with that, I think.
Saloni Dattani:
Well, I feel like you might have believed in miasma theory then because they were also like, oh, it’s all about socioeconomic differences and there’s this contagion that spreads diseases through poverty and stuff like that.
Jacob Trefethen:
Yeah, but you don’t think that people specifically would have got the food thing because, in fact, nutrition is useful.
Saloni Dattani:
Hmm. I don’t know. Maybe. But they didn’t think of that. And Paul Ehrlich, who’s very famous, in 1897 basically comes upon the right theory. In 1897, he suggests that antibodies are a natural constituent of the cell surface. They’re formed inside cells, and they have, from the beginning, a configuration that is specific to a given antigen. Then the antigen selects from all of the different antibodies available, only the ones that can interact specifically with it, and then the cell that contains that antibody starts to produce many more of those molecules that it releases into the blood. I mean, he kind of comes up to essentially what we now say.
Jacob Trefethen:
He pretty much nailed that.
Saloni Dattani:
He pretty much nailed it. But then later in the 1930s, Linus Pauling proposes a different theory, which is wrong, which is called “instructional theory”. The idea is that the antigens mold antibodies into complementary shapes, like a key pressed into soft wax. Does that make sense? So that explains why they’re so specific - because the antibody just molds around it.
Jacob Trefethen:
Right, so that is the kind of thing I could have imagined. Yeah, I would have come up with something like that. It’s the thing that I sympathize with that leads to that thought. How the heck do you get something so specific if you can get loads of different specific things? If it was just in the blood the whole time, so that’s why it has to be microscopic or there’s loads of them, or it has to be the main thing about it. Or sure, you have this thing that gets molded. The thing I don’t believe is, it came in with it the whole time. I mean, that’s just crazy.
Saloni Dattani:
Right. Why would a scary microbe contain the cure to it? And so a lot of people then like reintroduce the same theory of the complementary shapes. It doesn’t make sense, and one of the reasons we already talked about Émile Roux’s experiment, where there are lots of antibodies being produced and they’re still in the blood even after the antigen has left. Second reason is, actually the difference in quantity between those two things - at the time, they were like, at least 100,000 antibodies are produced per antigen - we now know that it’s more like millions or billions - but how is that going to happen if it’s a mold?
Saloni Dattani:
Yeah, and they wouldn’t be exactly the same, probably.
Jacob Trefethen:
Yes, that’s another good, yeah. So why on earth did he was a smart chap? Why did Linus Pauling think such a silly thing?
Saloni Dattani:
I don’t know. I think he was focused on the structural similarity, and the fact that there’s this interface between the antigen and the antibody. I think that’s what he was focused on.
But the idea of a key being pressed into soft wax reminded me of those customized hand wax sculptures that were popular when I was young, I don’t know if you’ve ever had them.
Jacob Trefethen:
I don’t know what you’re talking about.
Saloni Dattani:
So when I was growing up, a lot of my friends had these- basically, they had put their hands into a molding gel and then peeled off that molding gel and then put warm wax into the mold that they’d created, and that reforms a wax sculpture that looks like your hand. And a bunch of my friends had these in their house, and I was like, I really want one.
Jacob Trefethen:
That’s a jealousy inducer. Now, the closest I came was that when I was growing up, we had inherited a sort of sculptural toy from my dad’s parents that involved a rectangle with a lot of pins and you would place your hand and it would move back only the pins that your hand pressed against. So then if you removed your hand, you would have a sculpture that was the pin against the perspex, and I probably spent at least tens of hours, maybe hundreds of hours sort of interrogating that artistic process.
Saloni Dattani:
How old were you back then?
Jacob Trefethen:
Probably in the four to nine range, I would say, which probably means hundreds of hours, because over the course of those years, it adds up.
Saloni Dattani:
Yeah, I feel like I would- it sounds a bit like bubble wrap, you could get distracted by that.
Jacob Trefethen:
Yeah, yeah, that’s true. Although with bubble wrap, you do finish, you know, you pop them all and then it’s all over.
Saloni Dattani:
Right, you pop them all. Sadly, this is not over. There’s so much more that they’re going to learn. So Linus Pauling is wrong; people increasingly have reasons to believe that he’s wrong, and they’re searching for a new hypothesis, and in the 1940s and 50s, people come up with it. And that is a theory of selection of antibodies.
There’s an immunologist called Niels Jerne, and he introduces the idea that the body contains a vast existing library of antibodies. The antigens don’t instruct the immune system, but instead they select the antibody that already fits them. Once this interaction is complete, the antigen carries the antibody to cells that can reproduce the antibody... which is also weird, for the same reason, like, why would the antigens help your body?
They sort of moved then from this chemistry-based theory of how antibodies are working to more of a biological one. They’re like, oh, this is basically like natural selection, some kind of selection is happening.
In 1957, is the big paper that describes the type of theory that we believe in. It’s called Clonal Selection Theory that was proposed by Macfarlane Burnet in 1957. His idea was each white blood cell expresses one unique antibody, and when that antibody binds to its matching antigen, the cell gets activated and then it proliferates into a clone of identical cells that secrete that same antibody in massive quantities... which is kind of true!
Jacob Trefethen:
It’s kind of true, but how did they figure that out?
Saloni Dattani:
They had found some white blood cells that released antibodies, but they hadn’t actually figured out what those cells were, and they hadn’t distinguished between B and T cells at that point. It was just a theory that would explain the observations.
So it explained how there’s so much specificity in antibodies and how fast the response is and how many thousands, or millions, or billions of antibodies are produced by the body. So we now know that immune cells can release thousands of antibodies per second, and over a lifetime, humans generate tens of millions of unique antibody-producing cells.
Jacob Trefethen:
I will say, to me, that seems unexplained how we can do that, still, but...
Saloni Dattani:
We haven’t figured that out yet. But even though we haven’t figured that out, Niels Jerne, who initially introduced the selection theory, he declares at this point that immunology is solved. We’ve basically figured it all out now, all that’s left is some details that we need to sort out. He says that it’s solved in 1957, and in 1969, he gave a talk that was called “The Complete Solution of Immunology.” This sounds so funny.
Jacob Trefethen:
May I live to one day give a talk saying “The complete solution to X.” That’s wonderful. But just to check, so he thought it was solved after his own paper?
Saloni Dattani:
No.
Jacob Trefethen:
Or he thought it was solved after the Macfarlane person?
Saloni Dattani:
Yup.
Jacob Trefethen:
Okay got it. At least he’s complimenting someone else.
Saloni Dattani:
He’s like, “That guy solved it. We’re done now.”
Jacob Trefethen:
Were these people in America? I know Linus Pauling was.
Saloni Dattani:
I think Niels Jerne was in Denmark. Let’s see. Okay, so he’s in the US at this point, but he’s Danish.
Jacob Trefethen:
Because this sounds to me...
Saloni Dattani:
What, very American?
Jacob Trefethen:
Well, I didn’t want to put too fine a point on it, but it sounds little American, but also it’s... It is interesting that so much of the last episode is happening in England and France, I suppose, and then this episode, we had a flurry in Germany and a little bit in France, but by the time we’re in the 20th century, we’re moving across; this is America! Is immunology kind of... American, do you think? I’m going to get so much hate mail for that comment.
Saloni Dattani:
Well, definitely a lot more, but the very next step happens in Switzerland by a Japanese researcher.
Jacob Trefethen:
Aha!
Saloni Dattani:
So yes, it’s so funny to me that he’s like, “Immunology is solved.” in 1957. They haven’t even figured out what T cells are or what B cells are; they haven’t figured out how all of this diversity is created. Obviously, we just know so much more about immunology now. And back then, they were like, this is the end, we’ve found out all there is to know.
Jacob Trefethen:
They don’t know about B cells?
Saloni Dattani:
Yes. Also the 1960s, B cells.
Jacob Trefethen:
Okay. B cells, 1960s, that’s wild.
Saloni Dattani:
Because only in the 1960s do people realize that the thymus is involved in immunity. And they’re like, oh, wait, if you don’t have a thymus, you’re kind of fucked.
Jacob Trefethen:
That’s so interesting because I had thought- I think of the organs as a very 19th century thing. I think of the organs as like, well, we figured out the heart, the kidney, the liver. I’m like, okay, they sort of got that. You’re telling me that the organs used in immune responses, or at least some of them, they just did not have an idea what was going on?
Saloni Dattani:
No, they didn’t. I was reading about this in this great book called A History of Immunology. It’s by a scientist called Arthur Silverstein, and it’s very good, and it basically explains this. In the early 20th century, immunology had sort of turned into this chemistry problem rather than a biology one. People just thought, okay, there are these things in the blood, antibodies, which are protecting us, and the way that we need to understand them is through chemistry. They didn’t know how these antibodies were produced. They just don’t really appreciate how important the biological understanding is there.
Jacob Trefethen:
In 1957, they hadn’t even nailed down all the planks of the central dogma. Like, it was known that DNA was a double helix, but they hadn’t nailed down loads about how DNA to RNA to proteins and, you know, all that. So, blooming heck. Anyway.
Saloni Dattani:
Maybe it’s post-World War II. They’re riding that high, they’re like, “You know what? We’ve done it. We’ve finished it. Biology is over.”
Jacob Trefethen:
We’ve finished physics, now we’ve finished immunology.
Saloni Dattani:
So this gets us to the next bit. Antibodies are super specific, they’re super diverse. How is so much diversity created? So as you said... How does it work if there are only 19,000 or so genes in the human body? How do we produce millions of unique antibodies?
Jacob Trefethen:
I do know the answer, but let me come up with some fake answers I might have thought of before I knew the real answer. But I wouldn’t have known the total scale of just how unbelievable the combinatorics in reality can go. So I would have known it’s bigger than 19,000, though, the number of antibodies.
Saloni Dattani:
Right, and they didn’t know how many genes there were; they didn’t know that until the Human Genome Project in the 2000s!
Jacob Trefethen:
I even forgot that.
Saloni Dattani:
They didn’t actually know any of this.
Jacob Trefethen:
Would they have had any sense of the finite number of proteins?
Saloni Dattani:
I think they would have just thought like a huge amount. So it’s like very diverse, very specific. They would still have been confused.
Jacob Trefethen:
I think of so much of the wonder of the human body as the... it’s just astonishing, A) that there are only four base pairs in DNA and RNA; that’s just crazy. So that’s just like, what the hell is going on there? But it’s almost less- I can get my head around it when I think about information and computers.
Something that is residually just wild from modern knowledge is that there’s a finite number. You’ve got the two really interesting macromolecules, and now lipids - people are going to get mad at me - and carbohydrates - people are going to get mad at me - but the really interesting ones are the nucleic acids, which are doing the information storage, and you’ve got the proteins that are doing everything in the book.
So it’s just crazy that there are only 20,000 proteins or 100,000 if you count, you know, post-translational stuff. So... I’m like, what?! So I’m confused about that now. But before I knew that, I’m not sure I would have been confused at all. Because I would have been like, oh, that’s just sort of a mess of stuff down here.
Saloni Dattani:
And so maybe that’s why they thought it was solved. They were like, “yeah, well, seems possible. This doesn’t seem strange at all!”
Jacob Trefethen:
But really, what do we have to do? If you take the evolutionary lens, we have to somehow have a way of defending against arbitrary infections, because the infections are going to keep evolving, so we’re going to have to keep having a way to defend against them. So you can realize, oh, hold on a second, that is going to explode and it’s going to blow well past any number in the tens of thousands.
Saloni Dattani:
Right. And they can sequence proteins in the 1960s, I think.
Jacob Trefethen:
They can?
Saloni Dattani:
Yes. So they would be able to tell that very slight differences are going to lead to different antibody responses.
Jacob Trefethen:
Got it. Because they would have been evolving some different microbes in the lab and slight variants lead to big difference. Yeah, okay, fair enough. How did they figure it out? You’re going to have to tell me.
Saloni Dattani:
It is by a Japanese scientist called Susumu Tonegawa. He is working in the Basel Institute for Immunology in Switzerland at the time.
But it takes probably months to sequence the DNA of individual cells. So instead, he uses electrophoresis, which separates segments of DNA based on their sequence, and by doing that, he compares the DNA of B cells in embryonic mice versus adult mice - like the same mice as they get older.
He discovers that their sequences have changed; so the genes have changed somehow. The sequences have moved around, they’ve recombined, they’ve been deleted in some parts. There’s just very diverse regions of their genome that are used to create antibodies. This process, basically, is called V(D)J recombination. So the DNA of B cells are randomly spliced together, and that creates an almost infinite number of different potential antibodies.
Jacob Trefethen:
So basically it’s a trick where you are not just stuck with the genes you came with, you’re inserting some tootoo-tootoo-too so that you can create some new stuff.
Saloni Dattani:
So we start out with the genes that create antibodies and those genes, like the sequence of those genes can be randomly cut up and then spliced together. And you can introduce additional diversity into that, by joining the different segments together imprecisely, or randomly adding in new bases, or just like mutating, just randomly mutating the antibody genes afterwards. All of these things happen, and it generates tens of millions of unique antibodies throughout a lifetime, which is crazy.
Jacob Trefethen:
That’s how the big, big, big human can still defend against all these tiny, tiny, tiny microbes.
Saloni Dattani:
I have a little summary of the whole process in case it was hard to follow.
So each white blood cell, each B cell expresses a single antibody. When an antigen finds a match, clones of that white blood cell multiply. And this massive diversity isn’t encoded from birth, it’s generated continuously as the DNA inside developing immune cells is actively cut, shuffled, and stitched back together in countless combinations. This process creates tens of millions of unique antibody-producing cells, generated continuously after birth, waiting for the right antigen to come along. It’s quite beautiful if you think about it.
Jacob Trefethen:
It is. It’s astonishing, the ratchet of evolution. What an amazing trick to evolve. But you’ve only got to do it when you’re facing some absolutely unpredictable invaders. Once you’ve discovered that, I would have thought, “Well, hold on a second. Why does infectious disease even keep killing us? That’s such an amazing defense mechanism.” So what do people have to say?
Saloni Dattani:
I don’t know the answer. I don’t know what they think. Maybe they again think that immunology is solved again. I don’t know. I was going to say that it sounds kind of romantic in a way that the antibodies are just waiting for the right antigen to come along.
Jacob Trefethen:
And then they become... many more... numerous!
Saloni Dattani:
They’re like, I found my match. Guess what? Now there are millions of me!
So we now have the theory that there are antibodies in our blood that recognize foreign invaders. They’re really specific; they’re also really diverse. They’re produced by our cells and this massive diversity is continuously generated over our lifetimes.
What this now means is that you don’t need to have immunity, necessarily, against entire organisms, but about specific molecular parts of an organism; and even slight chemical changes can change that immune response.
This means that you can make what are called subunit vaccines, so you can just have specific molecules in a vaccine. And the way that they figure that out is that using all of these different antibody tests, they find that not all of the antigens are causing people to develop protection. Each pathogen tends to only have a few protective antigens that we develop antibodies to, and the others are kind of irrelevant, or they’re distracting in some way.
For influenza, for example, it has two proteins on its outer surface called hemagglutinin, which “agglutinates”, or binds to blood, and then neuraminidase, which does something else which I’ve forgotten. Those are identified as the protective antigens. For other bacteria, they find other specific antigens which are the key ones.
They’re like, “You know what? We don’t need to have the whole pathogen anymore. We don’t need to have the whole bacterium or the whole virus. We can just have a vaccine with one or a few different antigens in it; we can teach the immune system to make antibodies against it. Then it will be able to recognize those same antigens in a different infection in the future.”
Jacob Trefethen:
Let me slow down and check I got it. Because to me, it’s almost a different realm. In the last episode, we talked about microbiology, learning all about these microbes. And what you’ve just told me is all about immunology, learning about our immune response to the microbes so far, we’re learning a lot about antibodies. I’m wondering though, could you have got further with microbiology before knowing about antibodies in quite as much depth?
You’re saying take flu, take TB, whatever. We’re dealing with a microbe that’s pretty big and well, take TB, that’s a pretty big one; it’s a bacteria and it’s got a whole bunch of different proteins, carbohydrates, all sorts of stuff on it. Your body is going to react to a bunch of stuff on it.
And then what you just said about antigens is that you can take, of the thousands of genes that code for proteins in TB, maybe there’s a subset - maybe the subset could be just one, maybe it could be two, which is the leading vaccine currently being tried, M72 - it has two antigens. You have to somehow say, “Okay, I’m going to get rid of the other 300 and just going to keep these two, and that’s going to be enough.” Do I need to know about antibodies for that? Or could I have studied TB more, and done a bunch of experiments with subsets of what’s in TB? Why do I need to know about antibodies?
Saloni Dattani:
I think you need to know about antibodies if you want to test them. If you want to test a lot of different subsets, you probably want to do that in vitro; you don’t want to just keep infecting people with different ones, and then you would need a very large sample size to tell the differences between them, right? So I think that’s one reason.
I think the other is trying to figure out which specific subsets is just easier if you can visualize that, instead of telling the efficacy; you’re seeing specifically which part of the microbe is getting attached to.
There are different types of subunit vaccines that people develop. One is the protein subunit vaccine, where the vaccine just contains a few or maybe even just one protein from the pathogen; another is a polysaccharide vaccine, which is a sugar from the pathogen, often a bacterium; and then there are toxoid vaccines. Some of these are created before anyone really understands the immunology either. They do have antibody tests, but Gaston Ramon, if you’ve heard of him.
Jacob Trefethen:
Gaston Ramon?
Saloni Dattani:
He developed some toxoid vaccines against tetanus and diphtheria in the 1920s. So people have developed now, much safer and purified ways of developing vaccines.
Jacob Trefethen:
What you just said actually helps me feel more at peace with the answer to my last question. It sounds like before we knew the full story about antibodies and V(D)J recombination, we, as a society, did make a few subunit vaccines - without understanding the full theory - and we didn’t make that many. Once you had the theory, once you could see the antibodies binding, then you could start identifying antigens better, you could start really ramping up your subunit vaccine discovery and innovation. That makes sense to me.
Saloni Dattani:
I’ve convinced you now.
Jacob Trefethen:
You’ve convinced me.
Saloni Dattani:
I think the other is that there are various other reasons why you might prefer to have these subunit vaccines. One is safety. You notice, for example, that some components of bacteria are really harmful to us, even if the bacteria is killed. So with the pertussis bacteria, Bordetella pertussis, it contains 3,000 antigens or 3,000 surface proteins; some of them trigger really harmful reactions in us, they have rare side effects. If you had just two or three different antigens instead, without all of that waste material, or without all of those other antigens that could be harmful or trigger things, then you could avoid a lot of those side effects.
I think the third is that, in some cases, you’re just not going to be able to grow the whole pathogen in the lab. So some microbes are really difficult to culture, or they don’t infect any other organism apart from humans; and hepatitis B is one of these where there are very few animal models. It’s really hard to grow it in cell culture, and if you just had figured out the specific antigens that you needed, that would make it much easier to develop a vaccine.
But you are right that a lot are going to be produced without this knowledge. Those are some of the benefits of subunit vaccines, but there are also drawbacks of subunit vaccines. And you might know some of them.
Jacob Trefethen:
You know, it’s interesting you get these sort of couple examples before you understand a theory, then you get the theory and you get way more. And I like that, happened in the last episode with Edward Jenner; he didn’t even know what a microbe was.
Anyway, subunit vaccines. What are the drawbacks? Well, I can come up with some drawbacks. Here’s a drawback: if you don’t select a good antigen, that’s a drawback; maybe you got rid of a lot of the things that led to a good immune response. Or if the thing that controls a microbe in different people, is not always the same antigen. Maybe some people react to some bits, some people react to others. Let’s say you don’t generate as big an immune response, because if you get injected with a whole bacteria, your body’s going to start freaking out. If you get injected with a few little proteins, I don’t know, is that even going to juice my immune system enough? I’m not sure. So those are some ones off the top of the dome.
Saloni Dattani:
Yes, but that’s basically all of them, I think. There’s a really good example of this in pertussis, as I mentioned. The Bordetella pertussis bacterium has thousands of antigens. And although it was a really successful vaccine in terms of reducing the spread of pertussis in the 20th century, people were also like... “This is occasionally causing very scary, rare side effects.” There were a bunch of safety concerns about this vaccine; there were controversies and there were lawsuits in the US. In Japan, the controversies rose so much that the government actually just suspended the vaccine, even though it was quite successful.
So there was this international research effort, where scientists were thinking, “Okay, so that original whole bacterial vaccine was too risky for people, what if we can develop a better one that only contains a few antigens that we need for protection?”
There are these Japanese scientists, Yuji and Hiroko Sato, who are husband and wife, and they are two scientists working in Japan’s National Institute of Health, and they figure out the two antigens that they can use in a vaccine. They use the pertussis toxin and the filamentous hemagglutinin, and they purify them, and they detoxify them with formalin, and that resulting material becomes the vaccine, and it becomes the acellular version of the pertussis vaccine, and it was introduced in Japan in 1981. So that’s a really good example where these safety concerns of all of these different antigens are driving people to use different vaccines.
But at the same time, just the way that you said, it actually had lower efficacy, and people would develop an immune response but that immunity would wane after some months. So there is definitely a drawback to the new, safer vaccines that we have. And the way to improve them is by using adjuvants!
Jacob Trefethen:
Adjuvants! Yeah.
Saloni Dattani:
So Gaston Ramon, once again, in the 1920s, he discovers some of the first adjuvants. The way that he does this is by experimenting with lots of different random stuff in his house. Tapioca starch, breadcrumbs, soap flakes...
Jacob Trefethen:
Breadcrumbs and starch make sense because his name is Gaston. Gaston Roman makes me think, you know, like a buildings roman where it’s a novel of coming of age. In my head, a “Gaston Roman” is like a story about learning about delicious food. “Gaston Roman!” It’s like, “Oh, and then I really came into my own, and now I eat all of these delicious sweets.” Anyway, keep going.
Saloni Dattani:
I was thinking about Beauty and the Beast, and that guy is called Gaston, right?
Jacob Trefethen:
Yes, yeah, yeah. Maybe it is based on a little scientist.
Saloni Dattani:
Maybe that’s him. So he was basically trying to figure out if there were substances that could boost the immune response. I think he noticed that when animals were vaccinated with toxins, sometimes they develop stronger immunity if the injection site was inflamed; he was trying to find different substances that could inflame it. And so he found the first adjuvant, which were aluminum salts, I think.
Jacob Trefethen:
Aluminium! As we would say where I come from originally.
Saloni Dattani:
Yeah. Aluminium. I should say that as well. But for some reason, I have an American accent.
Jacob Trefethen:
This episode, the Americans are winning.
Saloni Dattani:
So he discovers that you can turn up the volume of the immune reaction and you can get a better immune response with an adjuvant. And various other people discover other adjuvants that improve subunit vaccines like saponin.
Jacob Trefethen:
Yes, yes, from the Chilean tree bark.
Saloni Dattani:
That’s right.
Jacob Trefethen:
This was something that still feels too magical. I sometimes wonder if the whole vaccine is just the adjuvant, it’s so wonderful and perfect. And I’m like, where do I get some tree bark? It’s some ancient wisdom in South America is that this particular tree bark will cure you of ills. And sure enough, it’s now used in more processed form, but essentially the same chemical in several vaccines. The first malaria vaccine...
Saloni Dattani:
The TB vaccine candidate, and the shingles vaccine! It reminds me of what we said in our AI episode where people in the past occasionally came upon really effective treatment. But I would prefer if someone figured out the purified form of the adjuvant so that I could just use that instead of, I don’t know, ingesting a bunch of insects along with it. So there are lots of different subunit vaccines!
[podcast jingle]
Saloni Dattani:
All right. So we’ve now reached the end of the episode. What did we learn? What are your favorite thoughts or what did this episode make you think about?
Jacob Trefethen:
Many different things. One is how many different steps it took and takes to make the hepatitis B vaccine. So it started with analyzing the outbreaks, identifying what was then called “serum hepatitis”, contrasting that with the other “infectious hepatitis”, hepatitis A; finding the Australia antigen, which was then discovered to be the hepatitis B surface antigen; purifying it, and getting rid of all of the dangerous stuff from blood, so that you could use it as a vaccine, and then eventually making it in recombinant form and focusing only on that one protein itself.
Saloni Dattani:
The other thing that I found really interesting was just how many different fields were involved in this whole process. There’s the microbiologists and the germ theorists of the 19th century, trying to identify the cause of each infectious disease they’re finding. There’s the epidemiologists, who are also tracing the causes of outbreaks and figuring out, “Hey, isn’t that strange that all of these people are developing jaundice... but the only thing that’s common between them is that months earlier, they all received a particular medical product.”
Then there’s all the immunology advances in both the theory and all the testing that happens to improve our understanding of how the immune system actually responds to a pathogen, and that helps people develop antibody tests, and it helps people test for the presence of pathogens, and also test the ability of new vaccines against them.
Then there’s the DNA and protein sequencing tools and all of that research that helps people figure out the specific hepatitis B surface antigen, and what its code is, and try to clone it in recombinant DNA technology, which itself is another big innovation that helps develop a better hepatitis B vaccine.
Jacob Trefethen:
Another thing that’s stuck with me from the episode was how it’s almost the hubris, that is almost the opposite to what you just said, which is that people throughout history think that science is done, or they think they’ve closed off a particular line of inquiry. The humoralists who had a fight with the cellularists in the 19th century; both of them had some good evidence for what was going on, and we now know both were right about some parts of their vision. But in the 1890s, people mostly declared victory for one side, the humoralists; we kind of dropped cellular immunology for decades, generations, and it’s only really since the 1960s, it’s popped back up. And now T-cells, one of the most studied things in immunology and in vaccinology! So yeah, you can really drop things for a long time. There was the example of Niels Jerne, who thought immunology itself was done in the ‘60s or the 50s. And I would not necessarily agree with him.
Saloni Dattani:
There was so much left to discover!
Jacob Trefethen:
Yeah, totally. Any other examples like that?
Saloni Dattani:
I have a bunch of other examples, but I just thought that itself was so incredible to me because he declared that immunology was solved before people figured out what B and T cells were! Obviously he wouldn’t have known that, they hadn’t been discovered yet. But it’s just a sign of... sometimes the field seems like it’s slowing down, and then there’s this whole new breakthrough that introduces way more complexity, and way more potential understanding and science that you could be working on. And I think people sometimes forget about that in the moment, because they’ve only been studying within this one particular paradigm of that field.
Jacob Trefethen:
I mean, we didn’t even have V(D)J recombination by then.
Saloni Dattani:
We didn’t even have good DNA sequencing at that point, or protein sequencing, or any of the tools to understand how much complexity there was. The other examples that I was thinking of was that Lord Kelvin thought physics was done... in the 19th century! His mind would have been blown by Albert Einstein.
Jacob Trefethen:
Oh my God, yeah. Imagine.
Saloni Dattani:
There’s also Francis Crick, one of the co-discoverers of DNA as the genetic code. He thought molecular biology was done in 1966! When I first read about these, I was like, “Isn’t that funny? What are all of these people doing research on, now then? You guys should just get a new job.” But all of this research is still ongoing.
Jacob Trefethen:
Well, I will say in their defence, they really got a lot done, and in the 50s and 60s, molecular biology did nail down the central dogma. But guess what? We still don’t know what a lot of genes are doing today! There’s a lot more to do.
Saloni Dattani:
Well, in 1966, people hadn’t developed recombinant DNA technology. They hadn’t discovered that HIV or other viruses could reverse-transcribe their genes, their RNA into DNA, right?
And the other thing that that reminded me of was that, if I remember correctly, James Watson, who was also a discoverer of DNA as the genetic code, was really against IVF. He thought that this was like an abomination and that if you developed babies in a test tube, they would come out all abnormal. So I just feel like there’s lots of stuff about molecular biology that they probably didn’t understand back then.
Jacob Trefethen:
Sounds right. Okay, what else stuck with you from this episode?
Saloni Dattani:
What else stuck with me? I think the way that people do innovation. So in the last episode, we talked about a very different style of trying to make vaccines than in this one. In those cases, you are doing loads of different experiments; you’re just seeing what works. You have no idea really why any of those vaccines are working, what the mechanisms are. You have some general methods of how to produce them, but you don’t really understand why.
All you do have is the germ theory, and the microscopes and trying to identify the causes of different diseases, then you have some culture techniques that you’re trying to improve, to grow them in the lab, and then to attenuate or to kill them and develop vaccines from that.
This was so different where here you’re like, ‘Okay, we’ve figured out what the virus is that’s causing this. Let’s try to get a specific protein from this virus and develop a vaccine from just that alone, and try to purify this vaccine and have only that.’ And we know, from immunology, that that alone is going to stimulate an immune response and be able to protect people from an infection.
This difference - between the serendipity and the empiricism versus the goal that you have in mind and trying to figure out how to get there - I thought was really interesting.
Jacob Trefethen:
Yeah, and for me, the fact that that innovation can lead not just to more vaccines, for diseases we don’t have vaccines for yet, but improving the safety of existing vaccines. This whole transition from going from taking a whole microbe, putting it in someone’s body as a vaccine versus just going after a really small and non-replicating protein or carbohydrate. What a transition. There’s probably things that other sectors can learn from that too. Innovation doesn’t just work on the frontier of new stuff, it can make better stuff and safer stuff.
Saloni Dattani:
Right. The other thing was all the feuds and the competition along the way. In the last episode, we talked about a bunch of them. We talked about Salk and Sabin’s rivalry. In this episode, we talked about the cellularists and the humoralists. I think there’s something positive about the competition in that, in trying to debunk the other people’s views, they developed lots of new ways to test these hypotheses, and they refined their theories.
But then it was also really sad that they just gave up on cellular immunity for so long. And then we talked about Albert Sabin, again, getting in the way of Maurice Hilleman and trying to- and getting ready to sue him for developing a hepatitis B vaccine. And then we talked about Baruch Blumberg, who had patented hepatitis B vaccine without making it, and then had later claimed that he had invented it himself. You know, their company initially refused to license it to Maurice Hilleman until they allowed him to be the director of the manufacturing program, which I thought was so crazy. You know, I do think all of these scientists have obviously developed incredible inventions, but at the same time, you’re just thinking, “What is going on? Why can’t you just let other people do their work?”
Jacob Trefethen:
I mean, finally, it’s just hepatitis B itself. And all of the science and technology development that we talked about over this episode, leading to something that has been used by hundreds of millions, probably billions of people at this point, and has prevented so much liver cancer - I don’t know even how to add it all up - so much cirrhosis, and saved millions of lives over the course of several decades. How hard-won that was to get there and how cheap and easy it is now to take. In fact, it fits into the background of our lives; it’s hard to remember whether I’ve gotten it. If I really make myself think, I know I have. And it’s kept me safe all these years, it’s something we can rely on in the modern world. So thank you, all the feudsters, as always, for making progress.
Saloni Dattani:
I thought it was so crazy how, when you described how tiny this little virus is, how few genes it has, how few proteins it has, and yet it causes so much damage over years or decades of a person’s life. And guess what? We can just prevent all of that with a simple shot and 85% reduction in liver cancer rates. That’s crazy. 70% reduction in liver deaths. That’s crazy. And all of that with a single shot.
Jacob Trefethen:
19th century, 20th century... what’s coming next, Saloni?
Saloni Dattani:
What’s coming next? You’ll have to wait and see. We’re now at the end of the story of one of the most successful vaccines that’s been taken by hundreds of millions of people, potentially billions of people, and it saved millions of lives. But this was just the start of protein subunit vaccines. Many, many more came after it. And then we got recombinant DNA technology. We got many other types of vaccines too, like vector vaccines and mRNA vaccines. And we don’t even know what the future holds... because, unlike immunology, vaccinology isn’t solved yet.
Jacob Trefethen:
And all of the future can be given to another episode. But for now, we hope you enjoyed this episode. And we hope that, if so, you rate us on Spotify and Apple or wherever you’re listening to this, and share the podcast with everyone you know, your friends, your viruses, your bacteria, and more! And your liver, for sure. Your liver’s going to love this one. And hope you enjoy listening too.
Saloni Dattani:
See you next time.
Jacob Trefethen:
Bye!
References
Books:
Paul Offit (2007) Vaccinated: One Man’s Quest to Defeat the World’s Deadliest Diseases
Arthur M Silverstein (2009) A history of immunology
Ronald W Ellis (1993) Hepatitis B Vaccines in Clinical Practice
Sally Smith Hughes (2011) Genentech: The beginnings of biotech
Articles:
Timothy M. Block et al. (2016) A historical perspective on the discovery and elucidation of the hepatitis B virus https://doi.org/10.1016/j.antiviral.2016.04.012
Naijuan Yao et al. (2022) Incidence of mother-to-child transmission of hepatitis B in relation to maternal peripartum antiviral prophylaxis: A systematic review and meta-analysis https://doi.org/10.1111/aogs.14448
Jill Koshiol et al. (2019) Beasley’s 1981 paper: The power of a well-designed cohort study to drive liver cancer research and prevention https://pmc.ncbi.nlm.nih.gov/articles/PMC5866222/
William J. McAleer et al. (1984) Human hepatitis B vaccine from recombinant yeast https://doi.org/10.1038/307178a0
Chunfeng Qu et al. (2014) Efficacy of Neonatal HBV Vaccination on Liver Cancer and Other Liver Diseases over 30-Year Follow-up of the Qidong Hepatitis B Intervention Study: A Cluster Randomized Controlled Trial https://doi.org/10.1371/journal.pmed.1001774
Anthony R Rees (2020) Understanding the human antibody repertoire https://doi.org/10.1080/19420862.2020.1729683
