VACCINES

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Keith Chappell is a laconic man with sideburns that wouldn’t be
out of place in a hipster brewery. His tone is quiet and calm, even as
he discusses the news that derailed 11 months of intense research
to try and solve the global problem that has changed our lives.
Molecular clamp “You have your heart and mind set on, ‘We’re going to save
some lives – we’re going to help the world get back to normal
technology may prove to
through this devastating pandemic.’ And to have that hope taken
be the future of vaccines,
away from us was incredibly hard to deal with,” he says.
but in the fight against
COVID-19 it was An associate professor at the University of When he returned to Australia in 2011, he began

famously abandoned at Queensland, Chappell has spent much of his career

trying to find ways to stabilise viral surface proteins.
to search for a stabilising method that is both versatile
and quick. Together with Paul Young and Daniel
RADOSLAV ZILINSKY/GETTY IMAGES

the 11th hour. MANUELA He began studying flaviviruses – viruses transmitted

to humans by mosquitoes and ticks that cause the most
Watterson, he created the molecular clamp technology.
In January 2020, the research team received

CALLARI reports. prevalent viral infections worldwide – during his

PhD at UQ. Then he spent three years at the Instituto
Salud Carlos III in Madrid working on stabilising
the Respiratory Syncytial Virus (RSV) surface fusion
protein, the target for a potential RSV vaccine.
funding from the Coalition for Epidemic Preparedness
to boost the technology and develop new vaccines to
help stop the world’s next epidemic.
Only days later, we woke up to find an unknown
virus – then named SARS-CoV-2 – quickly spreading

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VACCINES

IN FOCUS:
Antibodies
VIRAL INFECTION When a virus infects us, our
across the globe, killing thousands. The UQ team proteins, which stick out of the virus surface, attach to
partnered with global biotech company CSL and the ACE2 receptors in human cells. These receptors immune system responds
rapidly got to work trying to apply their patented are attached to our cells’ membrane and play a role in by generating antibodies
clamp technology to the new virus. The vaccine regulating blood pressure. When the key-like spike to clear the infection,
was one of Australia’s most promising, but after protein unlocks the cell, the virus can then insert its Spike protein while establishing a
strenuous work and encouraging early results, its genetic material into the cell to replicate, making us “memory” for a rapid
trial was abandoned when numerous participants unwell. second response to
returned false-positive HIV tests.  Molecular clamp technology works as a scaffold repeat infections.
Virus RNA Virus membrane
It appeared that the team’s innovative molecular that holds the spike protein in the right shape for the
clamp technology was to blame. But could this Keith Chappell, immune system to make antibodies. 
technology still be the future of vaccines? co-leader, Traditional vaccines contain either inactivated
UQ vaccine team or live but attenuated viruses. Inactivated vaccines Unstable synthetic copies A simple way to make
Vaccines the Australian way – such as some types of flu vaccines – contain of surface protein a vaccine is to produce
Bacteria can replicate on its own, but a virus must use viruses treated with heat, radiations or chemicals, copies of the spike
the mechanisms of the human cells to proliferate. In such as formalin or ß-propiolactone, so they cannot proteins. In SARS-CoV-2
the case of SARS-CoV-2, protein stalks called spike replicate, but can still trigger an immune response. these are often unstable
Similarly, live, attenuated vaccines, like measles and either fall apart or
and polio vaccines, contain viruses weakened in change structure so much
the lab. The virus is still viable and stimulates an that they can produce
immune response, but it cannot cause disease.  ineffective antibodies
Effective and that do not mimic the
Both come with risks. “You have to make sure that
ineffective antibodies surface protein – and can
your pathogen is genuinely inactivated so that you don’t
unintentionally infect people, but it still maintains prove harmful.
Kym Hoger (left), the immunogenicity that will elicit a neutralising
quality manager at response,” says Daniel Wrapp, a postdoctoral fellow at
the University of the University of Texas in the US.
Queensland’s National “One issue scientists have run into when trying to Coronaviruses employ
Biologics Facility inactivate or attenuate viruses to use them in vaccines their spikes to infiltrate
(NBF), helps to analyse is that they can unintentionally cause greater infection living cells. When the
metabolites during upon exposure to the genuine pathogen,” he explains. conditions are right, the
production of the SARS- That happened in 1966, when a vaccine against Host cell interior
virus enters.
CoV-2 molecular clamp RSV, which infects children before they turn two, was
vaccine. Martina Jones tested in the US. The trial had dreadful consequences. Virus RNA entering
(below), manager of the Many children still caught the virus, some suffered the host cell
NBF, prepares vaccine worse symptoms than usual, and two toddlers died
samples for testing. due to enhanced symptoms. A safe vaccine for RSV
has still not been found.
It appears that the formalin scientists used to
inactivate the virus changed the shape of the antigen,
HOW MOLECULAR CLAMP TECHNOLOGY WORKS
suggesting that the failure was primarily a result of
the immunisation triggering the creation of poorly This
designed antibodies. Molecular stabilisation
Using a subunit of the virus, such as the SARS- clamps help ensures the
CoV-2 spike protein, avoids the problems of a the synthetic vaccine is able
pathogen replicating in the human body, which is why spike protein to accurately
many of the research teams working on COVID-19 remain stable, teach the
vaccines target the spike rather than dealing with the compressing immune
whole virus. it top and system to
But to do this, scientists have to figure out how to bottom and recognise
COURTESY OF UNIVERSITY OF QLD.

preserve the shape of the spike protein as it shows on preventing it the protein
the surface of SARS-CoV-2.  from falling present on the
The spike protein is anchored to the virus surface apart or virus surface,
through a region that traverses the viral membrane. changing and produce
When the spike fuses with the ACE2 receptor, it structure. the correct
undergoes a significant change, refolding into a highly antibodies.
stable post-fusion form. Similarly, the isolated spikes Molecular clamps Effective antibodies

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VACCINES
VACCINE VELOCITY
In addition to being easily scalable, the significance of the molecular clamp
technology lies in the speed at which a vaccine can be created once any virus’
genome has been sequenced.
New virus Viral protein
genomic genetic code
sequence is received from a
identified biotechnology
company
Day 1 Days 4–7
– in pre-fusion form – are naturally unstable and
rearrange into the stable post-fusion state, which is a
very different shape.
Targeting the post-fusion form could cause the
immune system to produce antibodies designed for
the wrong enemy. They would not protect us from the
virus and could enhance the disease, as happened with
the RSV vaccine.
“It’s really important to be able to maintain
pre-fusion [form],” says Fasséli Coulibaly from the
Department of Biochemistry and Molecular Biology
at Monash Biomedicine Discovery Institute. “And
that’s exactly what the clamp is able to do.”
To make the synthetic version of the spike protein,
the UQ team used genetic technology to create a
gene that encodes the spike protein and a fragment
of an HIV protein known as gp141, replacing the
transmembrane domain to hold the spike in shape
and stop the transformation to its post-fusion form.
The stitched-together gene sequence is delivered
into Chinese hamster ovary cells. These are placed
into bioreactors for about 10 days, at the end of which
the code is translated into the polypeptide. The HIV
fragment naturally self-assembles, clamping the spike
The vaccine does not infect people because it
does not contain any genetic material of HIV.
“The worst thing that could happen is that it
might actually protect you from HIV infection.”
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VACCINES
Eve Radunz (right,
foreground) and Mallory
Daleris run analytics
on the vaccine. Radunz
(below) prepares
equipment for the
vaccine’s purification.
Viral protein code is integrated “Clamped” “Clamped” protein
with clamp code to create a gene protein is is analysed to
sequence. The stitched-together purified confirm structure.
sequence is delivered into ovary Ready for pre-
cells for protein production clinical trials
Days 9–12 Days 14–19 Days 15–21
At the 57-day mark, study participants were
tested for a number of effects, including HIV
reactivity. Gradually, the results started returning,
into its native structure and giving the same shape and evidence mounted: patients were testing positive
the spike would typically have when attached to the to HIV. The immune system had made antibodies
surface of the virus. against SARS-CoV-2, but it had also made antibodies
“At the time, I knew the clamp was a very solid against the clamp. When an HIV diagnostic test finds
technology,” says Damian Purcell, Professor of those antibodies, it registers the patient positive for
Virology at the University of Melbourne and head of HIV as a correlation.
the Department of Microbiology and Immunology “When SARS-CoV-2 hit, there were just so many
at the Peter Doherty Institute. Purcell, who studies issues, problems and unknowns that we had to get
HIV, has helped the UQ team develop methods to around. We were really run off our feet, and this was
measure the quality and the breadth of the immune really down low on our priority list,” says Chappell. 
response elicited by the vaccine against SARS-CoV-2. “Everything was done right, from my point of
He explains that 50 years of HIV research has vaccine was ready to test. After preclinical studies view, and I think that’s the opinion of the [scientific]
led scientists to discover that the HIV envelope testing safety and dosing in animals, phase one human community, as far as I could tell from my colleagues,”
spike’s soluble form – the HIV antigen – can be clinical trials were launched in July. The vaccine was says Coulibaly. “If anything, it’s actually a good
manufactured by introducing mutations around the safe and elicited a high immune response. demonstration that the system is robust and detects
gp141 region to stabilise the structure. Purcell has What was unclear then was whether the small things that weren’t likely, and we stopped the
encoded these mutations in the HIV vaccine that his HIV fragment would also elicit an immune response. problem before vaccines.”
team is currently trialling, which shows promising The UQ team knew that was a possibility and The vaccine does not infect people with HIV
results in animals. informed all participants of the risk. But because because it does not contain any genetic material of
The UQ group found that they could further HIV tests are built to detect human antibodies, there Daniel Wrapp, HIV, and thus the virus cannot replicate. “The worst
(DIAGRAM) USCDC. NICK DAVID, LEONTURA/GETTY IMAGES.
modify HIV gp141 and stitch it onto the base of many was no way to test cross-reactivity in animal studies postdoctoral fellow thing that could happen is that it might actually
different virus surface proteins that are similar to the before clinical trials. University of Texas protect you from HIV infection as well,” adds Purcell.
coronavirus spikes.  “The human immune system is extremely good But the widespread use of this vaccine would
Over the past decade, Chappell and his at recognising material that it deems ‘foreign’,” says interfere with HIV testing and demand new HIV
collaborators have tested the clamp on several viruses, Wrapp. “If you inject a non-human protein, then it diagnostics. To add to this, the risk to public
COURTESY OF UNIVERSITY OF QLD
including RSV, influenza, Ebola, Nipah, Lassa fever is pretty much inevitable that your immune system confidence in vaccines was high.
and MERS coronavirus, so were confident that it will eventually raise antibodies against that protein.” “It would have been incredibly difficult to explain
would work well for this SARS coronavirus. The But because the clamp weighs only about 10% of to 100% of the population that no, this is not HIV,
approach was ready, it was fully funded and was “the the whole polypeptide, the widespread assumption it’s a small fragment of one particular protein,” says
best lead candidate we had in Australia,” says Purcell. among scientists was that it wouldn’t register. Chappell. “There is still a huge amount of stigma
The team started work on SARS-CoV-2 at the end “I think if you talked to me – and probably others – attached to those three letters, and it’s disappointing.”
of January 2020. Within just three weeks, the first we’d say the chances were very low,” says Purcell. On 11 December, 2020 the trial was abandoned.
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VACCINES
Being prepared
The idea of sticking spike proteins together to lock
them into the pre-fusion state isn’t new. “That’s
something people have been working on for quite
some time,” says Chappell.
“It sort of came out of necessity,” says Wrapp.
“When working with these proteins in the lab, a lot
of them are so unstable that when we try to express
them, we are getting no pre-fusion whatsoever.”
Wrapp is a postdoc fellow in McLellan Lab, a
research group at the University of Texas that works
on protein stabilisation. The group has engineered
two mutations called proline mutations that, when
added to the spike, inhibit the transition from pre- to
post-fusion form.
Almost 10 years ago, they started studying a
virus called HKU1, which causes mild respiratory
symptoms similar to the common cold. Using a
technique called cryo-Electron Microscopy (crEM)
– which can observe molecular structure at an
atomic level, using extremely cold temperatures to
stop molecules vibrating – they solved the HKU1
spike protein’s atomic structure and could engineer
the two stabilising proline mutations. They then
transferred the mutations into the structure of the
MERS coronavirus spike.
Studying the atomic structure of a spike protein
from a new virus can take months, but thanks to the
previous painstaking work done on closely related
viruses, the McLellan Lab team were able to quickly
engineer those stabilising mutations into SARS-
CoV-2, a new pathogen that no one had heard of
before.
“You need to have a lot of basic discovery science
on an ongoing basis – to have things in the bag of
FORM AND
FUNCTION
The “train tracks” shown are
lipid membranes. The top
layer represents the target cell
membrane, and the bottom
layer represents the virus with
spike in pre-fusion form, at   A .
Because SARS-CoV-2 has a lipid
membrane, it requires a protein
spike to fuse the membrane
of the virus with the target
cell     B . This fusion changes the
shape of the spike   C , which
creates antibodies that are
ineffective against the virus.
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VACCINES
The clamp technology is easily
scalable, allowing it to rapidly produce
large amounts of the antigen protein
needed for the vaccine.
information that you can deploy when the chips are
down,” says Purcell. 
In the past months, the McLellan team has
worked closely with the National Institutes of
Health – the primary agency of the US government
responsible for biomedical and public health
research. Their proline mutations are now
incorporated into the Pfizer and Moderna vaccines,
Martina Jones (right, at
which have been approved in some parts of the
left) and Mallory Daleris,
world, and into the NovaVax and Johnson & Johnson
of the NBF, perform
vaccines, both currently under clinical trials.
analytics on the SARS-
While the proline mutations worked well for
CoV-2 molecular clamp
COVID-19 vaccines, the need for a technology
vaccine.
readily applicable to a broad range of pathogens is
notably urgent. Human activity and environmentally
destructive practices are creating the perfect
conditions for zoonotic pathogens like SARS-CoV-2
to emerge. The risk of a global pandemic occurring in
“I wholeheartedly believe that we’ve
the future will continue.  shown that the technology works and that
“The big benefit of the molecular clamp is that
we don’t need to know the structure ahead of time,” it’s safe ... We’ve learnt a lot over the past
Daniel Watterson Chappell says. “We’ve shown that the technology few months. We’ll be back, I guess.”
researcher works for a whole range of virus families. It’s broadly
UQ vaccine team applicable.” 
The technology is also easily scalable, allowing it to
rapidly produce large amounts of the antigen protein “It’s incredibly disappointing that this vaccine
needed for the vaccine, and it enables the production candidate is not being progressed,” says Chappell,
but he is confident there are ways to solve the HIV
test cross-reactivity. 
The UQ researchers are back in the lab to restart
from where they began in January 2020. They had
been tweaking the clamp to remove parts of HIV
(WRAPP) UNIVERSITY OF TEXAS. (OTHERS) COURTESY OF UNIVERSITY OF QLD
(DIAGRAM) DANIEL WATTERSON. (IMAGES) COURTESY OF UNIVERSITY OF QLD
of proteins otherwise difficult to express and gp141 known to elicit significant antibody responses.
reduces the amount of protein needed for each Now, they are trying to find ways to reduce the clamp
dose of vaccine. The UQ team has also developed antigenicity even more, to hide it from the immune
a purification mechanism so that any vaccine system completely.
candidate is purified using the same methodology. The team is also running through a broad panel
“That’s a huge advantage,” says Chappell. of similar bundled proteins to find candidates that
   C “It’s an exciting technology because it gives can replace the HIV clamp while achieving the same
you a generic platform for something you can stability levels.
adapt quite easily to different antigens and “I wholeheartedly believe that we’ve shown that
different diseases,” adds Coulibaly. the technology works and that it’s safe,” says Chappell.
Never before have so many different new “We do need to go back to square one, but we’ve learnt
  A    B technologies been developed in such a short time. a lot over the past months. We’ll be back, I guess.”
Innovative vaccine technologies, such as the
molecular clamp and the mRNA approach used MANUELA CALLARI is a Sydney-based freelance
in other vaccines being rolled out, are going to be science writer specialising in health. This is her first
crucial in dealing with future pandemics.  feature for Cosmos.
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