The search for the next anti-influenza drug

Influenza
iStock/Christoph Burgstedt

Dr Stephen Cusack, from the European Molecular Biology Laboratory (EMBL) in Grenoble,  speaks to SciTech Europa Quarterly about the research group and their latest findings.

The group of Dr Stephen Cusack at the European Molecular Biology Laboratory (EMBL) in Grenoble, uses X-ray crystallography and cryo electron-microscopy (cryoEM) to study the structural biology of protein-RNA complexes involved in RNA virus replication, innate immunity and cellular RNA metabolism. They have, for the first time, observed different functional states of the influenza virus polymerase as it is actively transcribing. These results, published in Nature Structural & Molecular Biology, provide valuable information for the next generation of anti-influenza drugs. When a virus infects and enters a host cell, the genomic material in the virus is both replicated to produce multiple copies of itself and transcribed into viral messenger RNA (mRNA). The viral mRNA can be read by the host cell’s protein production machinery, tricking it into making viral proteins.

The viral proteins package the copies of the viral genome to make progeny viruses that are released from the cell to infect new hosts. An enzyme called a polymerase is responsible for both the transcription and replication of the viral genome and is therefore crucial to the successful propagation of the virus. The influenza virus, whose genetic material is of RNA rather than DNA, is no exception to this modus operandi. Stephen Cusack and his research group at EMBL Grenoble started to work on influenza polymerase more than 20 years ago. In 2014, the group published the first crystal structures of the complete polymerase machine. SciTech Europa Quarterly speaks with Cusack about the Cusack research group and their latest findings.

What is the work and role of this research group?

The goal of my research group is to understand the molecular mechanism of replication of influenza virus and related viruses. One of our focuses is on the influenza polymerase, the key viral enzyme that replicates and transcribes the viral RNA genome using unique mechanisms. We use the techniques of structural biology, such as X-ray crystallography and cryo-electron microscopy, to determine atomic resolution structures of the influenza polymerase in different functional states. The information gained can be used to help develop anti-viral drugs. In addition to this, we have worked on the vertebrate innate immune response to infection by RNA viruses, in particular how the innate immune system detects the presence of viral RNA and then triggers an anti-viral response.

What was the reason why 20 years ago you decided to look into influenza polymerase?

I originally worked on the influenza virus surface proteins (those involved with cell entry and exit) but in the mid-90s I decided to take up the challenge to work on the influenza polymerase, the key replication/transcription machine of the virus. However, as that progress was very slow since the techniques of protein production and structure determination were not up to tackling such a large and complex structure.

It was not until 10 years later that we finally made progress and determined the structures of several fragments of the polymerase. Two of these, the so-called ‘cap-binding’ and ‘endonuclease domains’, were particularly interesting and they immediately became important drug targets. Indeed, EMBL spun-off a company called Savira pharmaceuticals, based in Vienna, to develop anti-influenza drugs based on our work. As it turned out, other companies also joined the race in competition with us and one of them, Shionogi, marketed the first endonuclease inhibitor as a new type of anti-influenza drug in 2018 in Japan and the USA (called Xofluza).

How far away are you from completing the next generation of anti-influenza drugs?

The problem with anti-virals is that there is always a possibility of the virus developing resistance, so to keep one step ahead you always needs to be working on the next generation of drugs. Fortunately, the influenza polymerase has many different target sites where inhibitors can bind and stop the enzyme functioning, not just the cap-binding and endonuclease domains.

Our recent work, published in NSMB, gives the first description of how the heart of the polymerase works, the catalytic site where RNA synthesis occurs. This work gives new insight into targeting directly the RNA synthesis active site, which has been done successfully with hepatitis C virus for instance. However, developing the next generation of anti-influenza drugs based on this knowledge is probably at least a decade away.

What would you say has been the biggest challenge so far? How have you overcome this?

The biggest challenge was the determination of the structure of the complete influenza polymerase. This we achieved in 2014 and depended on switching to study influenza B virus polymerase instead of influenza A, both of which infect humans. Influenza B polymerase turned out to be easier to produce in the laboratory in the large amounts needed for structural analysis. The next challenge after that was to find ways of determining structures of the polymerase in action. This we achieved in our latest NSMB publication, where we exploited the recent massive advances in cryo electron microscopy to be able to obtain snapshots of actively transcribing polymerase.

Looking towards the future, what’s next for this study?

We are now completing a molecular ‘movie’ of the complete cycle of transcription by influenza polymerase, showing how it acts like a tiny machine with lots of moving parts as it progressively synthesises RNA. Then we want to move on to understand how this all really works inside the infected cell since we know that other viral proteins and cellular proteins are involved, adding a whole new level of complexity.

 

Stephen Cusack

Head of EMBL Grenoble

cusack@embl.fr

Tweet at @embl

https://www.embl.org/

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