Influenza GOF

Research
Yoshihiro Kawaoka, DVM, PhD
University of Wisconsin-Madison

Types of GOF research

GOF research resulting in the generation of viruses that:
1. Do not exist in nature
e.g., H5N1 viruses that are airborne-transmissible in ferrets
Very few GOF research of concern

2. Are more pathogenic/transmissible than the starting viruses, but still

comparable to or less pathogenic/transmissible than those existing in nature
e.g., Low-pathogenic H5N1 viruses with mutations found in natural
isolates that improve replication in mammalian cells
Many
3. Are more pathogenic/transmissible than the starting viruses in animal
models, but do not appear to be a major public health concern
e.g., high-growth A/PR/8/34 virus with increased pathogenicity in mice
Many

In my presentation:

Why we need GOF research

The benefits of GOF research
- Specific examples
- Conceptual justification

Risks of GOF research of concern

Why are we concerned?
While the probability of an accidental or deliberate release of a humantransmissible virus from a single lab is arguably small, but not zero,
the more groups performing GOF virology, the greater the overall risk.
The generation of viruses through GOF research in labs with lowlevel containment facilities or scrutiny increases the risk of
accidental release.
the risks are finite and small, but of catastrophic proportions if ever
there was a breakdown of biosafety or biosecurity.
Wain-Hobson, EMBO Mol Med 2013
Wain-Hobson, Frontiers in Public Health 2014

Alternatives to GOF research of concern

alternative scientific approaches are not only less risky, but
also more likely to generate results that can be readily
translated into public health benefits.
Lipsitch and Galvani, PLoS Med 2014

1. Loss-of-function research
2. Use of low pathogenicity viruses

3. Phenotypic analyses

Alternatives to GOF research of concern

alternative scientific approaches are not only less risky, but
also more likely to generate results that can be readily
translated into public health benefits.
Lipsitch and Galvani, PLoS Med 2014

1. Loss-of-function research
researchviruses
does not always provide answers.
2. Loss-of-function
Use of low pathogenicity
3. Phenotypic analyses

LOF research
An example in which loss-of-function research fell short

1918 virus

LOF mutations

1918 virus
LOF mutant

Receptor specificity
HA-D190E

Transmission

No transmission

Tumpey et al. Science, 2007

GOF research of concern

A new phenotype required for ferret transmission was identified by GOF
research, but not by LOF research.
Virus with
wild-type H5 HA

GOF mutations

GOF mutant

Growth in
mammalian cells

No transmission

PB2-E627K

Receptor specificity
HA-N224K, HA-Q226L, HA-N158D

HA
HAstability
stability

Transmission

HA-T317I
HA-T317I

Imai et al. Nature, 2012

Alternatives to GOF research of concern

1. Loss-of-function research
2. Use of low pathogenicity viruses

3. Phenotypic analyses

Alternatives to GOF research of concern

1. Loss-of-function research
2. Use of low pathogenicity viruses

3. Phenotypic analyses

Highly pathogenic
avian influenza viruses

Low pathogenic
avian influenza viruses

10

10

H5N1 highly pathogenic

Infectious units/ 10
g lung
3
10
1

Seasonal H1N1

10 avian influenza
Hours post-infection
Highly pathogenic
viruses differ from low pathogenic viruses
in their kinetics of virus
tissue
24 30
36 48tropism.
60 d3 d5 d7
0 3replication
6 9 12 18and

Highly pathogenic
avian influenza viruses

Low pathogenic
avian influenza viruses

10

10

H5N1 highly pathogenic

Infectious units/ 10
g lung
3
10

Seasonal H1N1

10
Data obtained
withHours
lowpost-infection
pathogenic viruses
can be misleading.
0 3 6 9 12 18 24 30 36 48 60 d3 d5 d7
1

Alternatives to GOF research of concern necessary

1. Loss-of-function research
2. Use of low pathogenicity viruses

If, after careful examination, the attenuated viruses behave exactly

the same way as the wild-type highly pathogenic viruses, this would
be a promising approach.

Alternatives to GOF research of concern

1. Loss-of-function research
2. Use of low pathogenicity viruses

3. Phenotypic analyses

Alternatives to GOF research of concern

1. Loss-of-function research
2. Use of low pathogenicity viruses

3. Phenotypic analyses

Alternative approaches
Safer approaches to studying human adaptation of influenza A viruses

Modeling
Receptor specificity, Fusion activity
Replication complex
Sequence comparison
Sequence and phenotypic comparison of natural isolates
Use of seasonal influenza viruses for transmission
Host factors
Lipsitch and Galvani, PLoS Med 2014

Alternative approaches
Safer approaches to studying human adaptation of influenza A viruses

Alternative approaches alone do not provide answers to key

questions. For example, none of these alternative approaches
revealed the transmissibility of H5N1 viruses in ferrets.
The data obtained using alternative approaches do not always
correlate with data obtained from GOF experiments.

Lipsitch and Galvani, PLoS Med 2014

Alternatives to GOF research of concern

1. Loss-of-function research
2. Use of low pathogenicity viruses
3. Phenotypic analyses
We cannot rely solely on alternative approaches.

Phenotypically new traits and the molecular basis for those

traits have been identified by GOF research of concern but
not by alternative approaches.

Benefits of GOF research of concern?

the benefit of this work to public health is unclear...
Relman JID 2012

vaccine makers consider there is little in this influenza GOF

research that will help them develop more effective vaccines
(Butler, 2012; Malakoff, 2013).

there is nothing in GOF virology that will help us predict a

pandemic or help us develop more effective vaccines.
Wain-Hobson, EMBO Mol Med 2013

Benefits of GOF research of concern

Specific benefits differ depending on the GOF research of concern.

What did we learn from the ferret transmission H5N1 GOF research?
Vaccine stockpile

H5N1 Vaccine stockpiling in Japan

Vietnam + Indonesia
10 million people

Anhui
10 million people

Expired and
discarded

Qinghai
10 million people
Vietnam + Indonesia
10 million people
Anhui
10 million people

H5N1 stockpile vaccines

Qinghai
10 million people

Nearly 600 million US dollars spent in Japan

1 billion US dollars spent in the US
WHO SAGE Working Group on Influenza Vaccines and Immunizations

Vietnam + Indonesia
10 million people
Anhui
10 million people

2005 2006 2007 2008 2009 2011 2012 2013 2014 2015 2016 2017

H5N1 Vaccine stockpiling in Japan

Vietnam + Indonesia
10 million people

Anhui
10 million people

Expired and
discarded

Qinghai
10 million people
Vietnam + Indonesia
10 million people

74 million US dollars
to produce each
H5N1 stockpile vaccine
for 10 million people

Anhui
10 million people

Qinghai
10 million people
Vietnam + Indonesia
10 million people
Anhui
10 million people

2005 2006 2007 2008 2009 2011 2012 2013 2014 2015 2016 2017

What did we learn from the ferret transmission H5N1 GOF research?
Vaccine stockpile
It has been 17 years since the emergence of H5N1 viruses,
yet they have not caused a pandemic.
Questioned the pandemic potential of H5N1 viruses
H5N1 vaccine stockpiles are needed (important for policy makers)

What did we learn from the ferret transmission H5N1 GOF research?
Vaccine stockpile
It has been 17 years since the emergence of H5N1 viruses,
yet they have not caused a pandemic.
Questioned the pandemic potential of H5N1 viruses
H5N1 vaccine stockpiles are needed (important for policy makers)
Strain selection for stockpile vaccines

WHO H5N1 candidate vaccine viruses

There are 26 H5N1 vaccine candidates.

How do we select one?

*September, 2014
http://www.who.int/influenza/vaccines/
virus/characteristics_virus_vaccines/en/

Antigenically different H5N1

viruses are circulating

Transmissible virus

We now know
which H5N1
viruses have
pandemic
potential.

Useful information
for vaccine
candidate selection

Implications

Important for pandemic preparedness

Vaccine stockpile
It has been 17 years since the emergence of H5N1 viruses,
yet they have not caused a pandemic.
Questioned the pandemic potential of H5N1 viruses
H5N1 vaccine stockpiles are needed (important for policy makers)
Strain selection for stockpile vaccines
Vaccines would ideally be produced by using a virus that is
closely related to viruses of high pandemic potential.
Policy makers, not vaccine companies, select vaccine strains.
Naturally, vaccine companies will say no when asked if GOF
research of concern has helped vaccine production.

Implications

Important for pandemic preparedness

Vaccine stockpile
It has been 17 years since the emergence of H5N1 viruses,
yet they have not caused a pandemic.
Questioned the pandemic potential of H5N1 viruses
H5N1 vaccine stockpiles are needed (important for policy makers)
Strain selection for stockpile vaccines
Vaccines would ideally be produced by using a virus that is
closely related to viruses of high pandemic potential.
Risk assessment of circulating strains

Kawaoka

N158D

Fouchier
T160A
G228S
Q226L

Q226L
N224K

H110Y

T318I

Imai et al. Nature, 2012

Herfst et al. Science, 2012

Global incidence of the mutation

among H5N1 isolates in 2009-2011
Japan
40/40

100%
China
31/49

Egypt
265/352

63%
Vietnam
33/130

75%
Other
56/137

41%

25%

H5N1 viruses in Egypt

Mutation (-)

30%

Mutation(+)

Mutation(+)

70%

2009-2011 birds

100%

2009-2011 humans

Neumann et al., PLoS Pathogens, 2012

Implications

Important for pandemic preparedness

Vaccine stockpile
It has been 17 years since the emergence of H5N1 viruses,
yet they have not caused a pandemic.
Questioned H5N1 vaccine stockpile
H5N1 vaccine stockpiles are needed (important for policy makers)
Strain selection for stockpile vaccines
Vaccines would ideally be produced by using a virus that is
closely related to viruses of high pandemic potential.
Risk assessment of circulating strains
Surveillance of H5N1 viruses may be limited.
But, this is changing, e.g., H5N8 viruses in wild birds across Europe and
Asia.

Neumann et al., PLoS Pathogens, 2012

Implications

Important for pandemic preparedness

Vaccine stockpile
It has been 17 years since the emergence of H5N1 viruses,
yet they have not caused a pandemic.
Questioned H5N1 vaccine stockpile
H5N1 vaccine stockpiles are needed (important for policy makers)
Strain selection for stockpile vaccines
Vaccines would ideally be produced by using a virus that is
closely related to viruses of high pandemic potential.
Risk assessment of circulating strains
Important information!
Imai et al., 2012, Nature

Benefits of GOF research of concern

Specific benefits differ depending on the GOF research of concern
GOF research of concern allows us to obtain information that we
could not obtain by other methods unless it actually occurred in
nature.
Droplet-transmissible H5N1 and other avian viruses to which humans
lack immunity
GOF research of concern allows us to determine whether these
viruses could emerge.
If so, it also allows us to examine the mechanism(s) for such events.

The information obtained is essential for pandemic preparation.

What are the key issues on benefits that need to be

addressed in the assessments the NIH will undertake?

1. Focus on GOF research of concern

2. Recognize that for some questions, only GOF research of
concern can provide accurate answers
3. Obtain consensus from the community to perform GOF
research of concern