Prospects for controlling future pandemics of influenza

Virus Research 162 (2011) 39–46

Contents lists available at SciVerse ScienceDirect

Virus Research journal homepage: www.elsevier.com/locate/virusres

Review

Prospects for controlling future pandemics of influenza James S. Robertson ∗ , Stephen C. Inglis National institute for Biological Standards and Control, Blanche Lane, South Mimms, Potters Bar, EN6 3QG, UK

a r t i c l e

i n f o

Article history: Available online 22 September 2011 Keywords: Influenza Vaccine Anti-virals Pandemics Surveillance Pandemic preparedness

a b s t r a c t Pandemic influenza remains one of the most serious threats to global public health and continued global vigilance to monitor emerging threats is crucial. Of the weapons available to control a pandemic, vaccination is potentially the most powerful, but there are currently serious limitations to timely availability of vaccine supply in an emergency. Many novel influenza vaccines are in development, some of which have the potential to deliver the massive quantities of vaccine that would be required in a pandemic in a short period of time. However, for the foreseeable future, it is likely that the principal vaccine that will be deployed in a pandemic will be an inactivated egg-derived vaccine of the kind that has been available for several decades. This review will focus on the practical hurdles that need to be surmounted to deliver large amounts of safe and effective pandemic vaccine to the general public. There needs to be a continued focus on improvement to the vaccine response system that will require close collaboration between influenza and vaccine experts, manufacturers, regulators and public health authorities around the world. © 2011 Elsevier B.V. All rights reserved.

Contents 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11.

Seasonal and pandemic influenza . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Past pandemics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Future prospects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Global surveillance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Reducing the impact of a pandemic: antivirals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Reducing the impact of a pandemic: vaccines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Influenza vaccines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . WHO pandemic vaccine preparedness (pre-2009) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Timelines for preparing pandemic vaccine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2009 H1N1 pandemic . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Vaccines for a future pandemic . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1. Seasonal and pandemic influenza Patterns of disease caused by influenza viruses are well recognised. Seasonal epidemics occur regularly during the winter months of the temperate zones of the northern and southern hemisphere whilst a less regular pattern of disease is found in the tropics. Very occasionally pandemics of influenza rampage globally; these occur at intervals varying from 11 to 30 or more years and can cause very high levels of mortality and morbidity. The seasonal outbreaks are a result of the accumulation of a small number

∗ Corresponding author. Tel.: +44 1707 641304; fax: +44 1707 641050. E-mail addresses: [email protected] (J.S. Robertson), [email protected] (S.C. Inglis). 0168-1702/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.virusres.2011.09.024

39 40 40 40 40 41 41 42 42 43 43 44

of mutations in the surface haemagglutinin molecule, the virus antigen against which the principal neutralising antibodies are formed. These mutations occur due to immune selection pressure (Pan and Deem, 2011) and are commonly referred to as antigenic drift. Pandemics of influenza occur when a virus appears which has a novel subtype of haemagglutinin against which the world’s population has no or very little immunity. These viruses can be a result of reassortment between a pre-existing human (seasonal) virus and a virus from the large pool of influenza viruses that exist in the animal world, especially in birds where all 16 subtypes of haemagglutinin (H1–H16) have been found compared with only three (H1, H2 and H3) in human influenza viruses. This was the cause of the 1957 pandemic (when an H2N2 virus replaced the prevailing H1N1 virus) and the 1968 pandemic (when an H3N2 virus replaced the

40

J.S. Robertson, S.C. Inglis / Virus Research 162 (2011) 39–46

prevailing H2N2 virus). Pandemic viruses can also result from species jumping, where the entire virus adapts from an animal source to replicate in and transmit amongst humans. This was believed to be the origin of the 1918 pandemic (Taubenberger and Morens, 2006). Whilst the genetic makeup of the 2009 pandemic virus has been analysed in detail and contains genome segments of swine, avian and human origin (Garten et al., 2009), the precise mechanism by which the virus became a human pandemic virus is not clear. The principal feature of pandemic viruses is that they have a novel haemagglutinin against which the human population has no or very little pre-existing immunity. This scenario is commonly referred to as antigenic shift. 2. Past pandemics Historical epidemiological data has identified probable influenza pandemics that have occurred over the centuries (Potter, 1998). In the late 19th century, sero-archaeology provided clues as to the subtype nature of the haemagglutinin in the virus causing the pandemics of 1889 (an H2 subtype) and 1898 (an H3 subtype). For the pandemics that occurred in the 20th century, the causative agent had been available for detailed study: an H1N1 virus in the 1918 pandemic, the H2N2 virus in the 1957 pandemic and the H3N2 virus in the 1968 pandemic. Interestingly, the 2009 H1N1 pandemic occurred at a period when a human H1N1 subtype virus had been in circulation for several decades albeit with a sufficiently different H1 haemagglutinin such that the world’s population had little to no immunity against the novel 2009 H1N1 pandemic virus. This new H1 haemagglutinin is derived from a swine virus and not from extensive drift of the pre-existing seasonal virus (Garten et al., 2009). Needless to say, since its appearance, the 2009 H1N1 pandemic virus has come under intense scrutiny. 3. Future prospects The threat of pandemic influenza remains ever present. The 2009 pandemic is considered to have been mild (Nishiura, 2010) with an estimated mortality in the USA approximately half that of seasonal influenza, although the case–fatality rate varied significantly amongst different age groups (Neumann and Kawaoka, 2011). However, there is no reason why future emerging pandemic virus strains should not be highly pathogenic, equalling or even exceeding the high morbidity and mortality seen in 1918. The many virus subtypes continually circulating in animals provide a rich source of haemagglutinin (HA) proteins with which humans have little or no immunological experience, and the capacity for the virus to adapt its genetic composition to accommodate new gene combinations is clear. Hence pandemic influenza is recognised by many developed countries and by WHO as one of the highest risks that they face (Cabinet Office, 2010; WHO, 2011a). In recognition of this ongoing threat, what can be done to stop or lessen the impact of future pandemics? One of the most powerful weapons in the armoury to prevent infectious disease is clearly vaccination, which has worked spectacularly well in many cases. However the influenza virus’s capacity for antigenic variation and the plethora of different viral strains circulating in animals has presented a major hurdle to the goal of developing a vaccine that could block all strains and hence eliminate the pandemic threat. This is an area of intense research and there have been some promising leads in recent years (Compans, this issue), but the prospects of success are still very uncertain and even it were to prove possible, it would take many years to bring such a vaccine to the market. So for the foreseeable future we must deal with the ongoing threat by anticipating as far as possible

what virus strains are likely to be problematic and trying to react as effectively as possible when a problem does arise. 4. Global surveillance The speed with which influenza can spread means that a new strain arising in any country can pose an immediate risk to the whole world. For this reason an extensive and well developed global surveillance system has been established to act as the world’s antennae for emergence of potentially dangerous new strains – the WHO’s Global Influenza Surveillance and Response System (GISRS, 2011). National Influenza Centres in countries all round the world collaborate with a group of specialist laboratories, the WHO Collaborating Centres in UK, USA, Australia, Japan and most recently China, to analyse continually circulating strains in humans. This activity underpins annual decisions on the optimal composition of vaccines for seasonal influenza, which many countries routinely deploy in autumn/winter to reduce the burden of respiratory illness in high risk populations. The global network also, however, provides a monitoring system for the emergence of potentially dangerous pandemic strains. Since new strains can arise in any country, excellent international co-operation is required for this to work well, and after several years of intense discussion and negotiation, WHO brokered an agreement, endorsed at the World Health Assembly in April 2011, to establish a new and more robust network for global collaboration and sharing of pandemic virus specimens amongst all countries (WHA, 2011). At the heart of this agreement is a commitment to build capacity for dealing with pandemics in developing countries in return for global co-operation. This global network concentrates primarily on virus strains isolated from humans, but of course it is possible and even likely that a new pandemic could emerge from the animal population. A parallel network of laboratories overseen by OFFLU, a collaboration between the Organisation Internationale Epizootique (OIE), the veterinary equivalent of WHO, and the UN Food and Agriculture Organisation (FAO), therefore carries out regular surveillance of animal influenza strains (OFFLU, 2011). Effective ongoing surveillance, including avian and swine influenza, is essential for maintaining vigilance against the threat of pandemic influenza. But the key question is how this intelligence can be used optimally to mitigate the impact of the disease once the threat is recognised. 5. Reducing the impact of a pandemic: antivirals The impact of a pandemic can be lessened either by reducing the number of people who become infected or by reducing the severity of infection in those who do become infected, and the two main interventional tools available are antivirals and vaccines. Other measures such as hand hygiene and social distancing may have a useful role in certain circumstances, but on the basis of current evidence, their overall impact is likely to be short-lived and limited. Several antiviral drugs have been developed to combat influenza virus infection. Amantadine and rimantadine, which operate by targeting the virus M2 protein, were developed many years ago and there is evidence for efficacy, but their side effect profile has precluded their use on a large scale (Aoki, 1998). More recently two drugs have been developed that work by inhibiting the virus neuraminidase, zanamivir (trade name Relenza), and oseltamivir (trade name Tamiflu) (McNicholl and McNicholl, 2001). Clinical trials of these drugs have shown that if administered early in infection, they can reduce the duration of symptoms significantly, and experience in the UK during the 2009 H1N1 pandemic showed that widespread use of the drug did have an impact on the severity of disease,

J.S. Robertson, S.C. Inglis / Virus Research 162 (2011) 39–46

resulting in reduced hospitalization (Nguyen-Van-Tam et al., 2010). Clinical trials also showed that the drugs could be used prophylactically to prevent virus replication and hence both disease and onward transmission (Ward et al., 2005). Once again the 2009 experience provided some evidence to back this up. During the initial stages of the outbreak in the UK, antivirals were given to immediate contacts of those showing symptoms and there was evidence of protection through this strategy (HPA, 2010). The practical difficulty with use of the drugs prophylactically during a pandemic is that it would require continual treatment throughout the entire period of threat, which might be many months or perhaps even years. The capacity of antivirals to block transmission when used prophylactically raises the possibility that they might be used extensively at the source of a pandemic to ‘stamp out’ the infection. Modelling work has suggested that this might be possible in theory (Ferguson et al., 2005), but the practical difficulties in identifying a source early enough for this strategy to have a chance of working are huge and so, whilst probably worth attempting, it would seem unlikely to succeed. Influenza is a highly transmissible disease and this coupled with modern population mobility means that we can expect the virus to spread around the world in just a few weeks. A further potential concern with the widespread and early use of antiviral drugs is the possibility that it might drive emergence of resistant viruses. It is well known that resistance to oseltamivir, for example, can result from a single mutation in the neuraminidase gene (Hurt et al., 2009), and given that the arsenal of antiviral drugs currently available is limited, any strategy that undermined the ability to treat severe cases effectively could be highly counterproductive. However, extensive use of oseltamivir during the 2009 H1N1 pandemic did not result in any significant emergence of resistant viruses (Hurt et al., 2009). 6. Reducing the impact of a pandemic: vaccines As it stands, therefore, the best option for mitigating the impact of a pandemic virus remains through vaccination to reduce the number of susceptibles. But in the absence of a ‘universal vaccine’ that protects against all possible strains, this also poses a tremendous challenge, because vaccines take many months to manufacture and administer to a large population. Where a particular virus strain has been identified in animals that poses a serious risk to humans, it may be possible to prepare effectively by preparing and stockpiling a relevant vaccine. Many countries have agreed that this would be feasible (WHO, 2007) and several countries have done this to counter the threat of H5 avian influenza (HPA, 2008; CIDRAP, 2008), which has shown itself capable of transmission to humans (though fortunately not between humans to any extent) on many occasions, often with fatal results (WHO, 2011a). However since current influenza vaccines generally only protect against closely related strains this strategy is not guaranteed to work. An alternative approach to stockpiling vaccine against specific potential disease threats could be to include additional components in seasonal influenza vaccines. Current vaccines are trivalent with their constituents specified by pharmacopoeiae and marketing authorisations although more components could, in theory, be added. This strategy could, through a routine influenza vaccine programme, build a degree of immunity in the population that might at least reduce infection severity in the event of a pandemic caused by a related strain. It has the disadvantage, however, that the immunity might not last long enough to be useful and that in most countries the vaccine is recommended only for specific risk groups. In a pandemic all of the population may be at risk. Overall, given the costs and uncertainties, stockpiling vaccine or ‘pre-pandemic’ vaccination on a large scale against even a limited number of potentially dangerous viruses would be impractical.

41

The best that can currently be done, therefore, is to devise ways of responding as quickly as possible to an emerging threat through development of tailored vaccines, the principal focus of this review. 7. Influenza vaccines The vast bulk of influenza vaccine in use today is prepared by growing the virus in embryonated hens’ eggs and subsequently inactivating it (Wood and Williams, 1998; Furminger, 1998). Although little has changed in this respect since influenza vaccines were first developed in the mid-20th century, there have been changes to the downstream manufacturing process. Virus purification has been improved by the introduction of zonal centrifugation, and a detergent step has been introduced to ‘split’ the virus particles which reduces the reactogenicity of the vaccine (Wood and Williams, 1998; Furminger, 1998). In some cases the surface antigens, the haemagglutinin (HA) and NA are further purified from the split virus preparation to generate a ‘subunit’ vaccine. Other developments in influenza vaccine have been the use of cell cultures for growing the virus instead of using eggs (Rappuoli, 2003), and the development of live attenuated vaccine strains of the influenza virus (Maassab and Bryant, 1999). Preparation of influenza vaccines is hampered by two fundamental problems: (i) due to the continuously changing antigenic nature of seasonal viruses (antigenic drift) or in the event of a pandemic (antigenic shift), the virus used to prepare vaccine needs to be updated regularly, and (ii) since wild type viruses isolated directly from clinical samples generally do not replicate well in embryonated eggs, the vaccine virus has to be modified to enhance its growth properties. Both these features constitute a major obstacle to routine seasonal vaccine production and are particularly problematic in the event of a pandemic when large quantities of vaccine are required as quickly as possible. The first of these problems is handled through GISRS, the network of WHO laboratories that conducts surveillance of influenza and characterisation of relevant viruses isolated from the community (GISRS). Under the auspices of the WHO, a panel of experts decides upon the best and most relevant strains that should be used to produce vaccine, taking into consideration the epidemiological situation, the characteristics of viruses causing disease and the ability of current human sera to react with novel viruses (WHO, 2011b). This is done routinely for seasonal influenza vaccines and ad hoc to deal with specific pandemic threats. The second hurdle is to develop a version of the strains recommended by WHO that can grow well in eggs for use in vaccine manufacture. This is also undertaken by laboratories operating within the WHO network, making use of the segmented nature of the influenza genome that allows two individual viruses to reassort their genetic elements and produce viruses with novel properties. This was first used to the advantage of vaccine manufacture by Edwin Kilbourne four decades ago (Kilbourne, 1969; Kilbourne et al., 1971). By reassorting the WHO recommended strain with a high yielding laboratory strain, a virus with the antigenic phenotype of the recommended strain but with the high growth properties of the laboratory strain can be selected from a mixed infection under appropriate selection conditions. This has been a relatively reliable method for generating high growth reassortants for use by the egg-based vaccine manufacturing industry and makes use of the high yielding donor strain A/Puerto Rico/8/34 (commonly known as PR8) which has been passaged several hundreds of times in vivo, including in ovo, consequently growing to very high titre in eggs. In some cases though, for example, for influenza B viruses where a reliable high yielding donor strain is not readily available, or where virus is required for cell-based vaccine manufacture, the derivation of a high growth reassortant based upon PR8 may not provide a growth advantage over the wild type virus.

42

J.S. Robertson, S.C. Inglis / Virus Research 162 (2011) 39–46

The reassorting method of generating high yielding vaccine viruses is highly empirical, relying on chance to generate the desired genetic composition. The use of reverse genetics (RG) provides a method of directly generating reassortants that have high yielding properties (Fodor et al., 1999; Subbarao et al., 2003). Using the RG approach, viruses with HA and NA gene segments from the recommended strain and the remainder from the high yielding parent (a ‘6:2’ gene constellation) can be created to order. It also offers the opportunity to make directed genetic changes within the genes as required (see next section). Whilst studies (Nicolson et al., 2005; unpublished) have shown that such RG viruses indeed have improved growth properties, viruses derived by random reassortment may nevertheless give superior yields (Bucher, 2011) – and experience has shown that in some cases 5:3 reassortants (5 genome segments from PR8 and 3 from the WHO strain) have better growth properties (Robertson et al., 2011). In addition the use of RG technology has intellectual property implications, whereas the classical reassortment method does not. 8. WHO pandemic vaccine preparedness (pre-2009) In 1997 in the Hong Kong SAR there were 18 cases of human infection with a highly pathogenic1 H5N1 avian influenza virus; six of the cases were fatal and others suffered serious illness (Chan, 2002). This heightened general awareness of the potential threat from pandemic influenza, although no further cases appeared in the short term. In 2003/2004 several further cases occurred in South East Asia, especially in Vietnam and Thailand, and in the ensuing years highly pathogenic H5N1 spread across Asia into Europe and Africa causing outbreaks of avian flu in domestic chickens, in other avian and terrestrial animal species, and notably in humans with a high (∼60%) level of mortality although there has been no sustained human-to-human transmission (Wikipedia, 2011; WHO, 2011c). As of 09 August 2011, there have been 564 human cases of H5N1 with 330 deaths (WHO, 2011d). Currently, the incidence of H5N1 infection is reduced compared to 2004–2006, but it continues in both poultry and humans, with associated human fatalities (WHO, 2011c). Early in 2004, in response to the re-appearance of the H5N1 virus, the WHO requested their influenza collaborating laboratories to prepare candidate vaccine viruses. However, the highly pathogenic H5N1 virus presented a unique situation: the virus was potentially lethal and required high containment to protect staff involved in the work and also minimise the risk of environmental spread. Preparing high growth reassortants by the traditional route was felt to be unworkable under these circumstances and so the alternative was to use reverse genetics to create a 6:2 reassortant vaccine virus. This approach offered a further advantage. The short stretch of basic amino acids around the point at which the full length HA protein is cleaved proteolytically to generate the HA1 and HA2 subunits is known to be a key determinant of pathogenicity and H5 viruses without this sequence do not display high pathogenicity (Rott, 1992). RG technology could therefore be used to remove the coding region for these extra amino acids and generate a virus that should be safe to use in the laboratory and for virus manufacture (Subbarao et al., 2003; Nicolson et al., 2005). Using this approach, two laboratories, St Jude Children’s Research Hospital in Tennessee and NIBSC in the UK, were able to derive potential vaccine candidates by rescuing viruses with the six internal segments from the high yielding PR8 strain and the NA and

1

Highly pathogenic avian influenza viruses are defined by the Organisation Internationale Epizootique (OIE) using an intravenous pathogenicity test in chickens (see http://web.oie.int/fr/normes/mmanual/A 00037.htm) and are associated with the presence of multiple basic amino acids at the HA cleavage site of H5 and H7 subtypes.

a modified HA from human H5N1 isolates from Vietnam (Webby et al., 2004; Nicolson et al., 2005). Tests in embryonated hens’ eggs, in tissue culture and the OIE in vivo pathogenicity test in chickens (OIE, 2005) demonstrated that these new viruses were indeed attenuated (Nicolson et al., 2005), but there remained the possibility that the virus might still be unacceptably pathogenic for mammalian species and so a new test had to be devised. The ferret is an animal model often used for the study of influenza virus infection and this was used to assess the pathogenicity of the rescued H5N1 viruses: the results showed them to be much less pathogenic than the parental H5N1 Vietnamese isolates (Nicolson et al., 2005). Following from such studies, the popularity of ferrets as a model for studying influenza disease and vaccines has increased dramatically. The candidate H5N1 vaccine virus developed at NIBSC, NIBRG-14 (Nicolson et al., 2005), has now been used extensively in clinical trials worldwide and is the basis for vaccine stockpiles in a number of countries (Sinovac, 2011; EMA, 2011a); fortuitously, it has not had to be used in earnest in a vaccination campaign. From this period onward, the WHO also recommended the development of vaccine viruses for other potential pandemic viruses giving priority to H5N1 and H7 avian strains, both of which can exist in the highly pathogenic form and both of which have documented histories of causing human infection and disease (Wong and Yuen, 2006; WER, 2011). Also on the priority list are H9N2 avian viruses which have occasionally caused mild influenza infections in humans and H2N2 which was the subtype that was the casual agent of the 1957 pandemic and to which, as time progresses, an increasing proportion of the human population has never been exposed. A number of international laboratories are now developing ‘libraries’ of candidate vaccine viruses against these strains (Robertson and Engelhardt, 2010). There is no guarantee that a member of such a library would have sufficient antigenic match against a future pandemic virus of the same subtype, but even if the match is less than perfect, the generation of some cross-reactive immune responses in vaccinees could be beneficial, perhaps by reducing disease severity. Furthermore there is evidence that the extent of cross reactivity may be increased by the use of adjuvants (Banzhoff et al., 2008), a number of which are being developed by manufacturers for use with influenza vaccines. An important aspect of preparing a pandemic vaccine for public use is licensure. Before any vaccine is deployed it needs to be shown, as far as possible, to be safe and effective, and this is done through the regulatory licensing process. It was recognised that the standard regulatory process could not operate fast enough to deal with an influenza pandemic emergency, and so the EU developed a process by which vaccine companies could prepare for licensure of their pandemic vaccines in advance; this involved the manufacture of a monovalent pandemic-like vaccine and demonstration of its clinical safety and immunogenicity (EMA, 2008). For these ‘mock’ pandemic vaccines, the clinical assessment programme is considerably less rigorous than normally would be required for a novel vaccine. The intent was that manufacturers would have developed and gained experience with their pandemic manufacturing procedure, and when a genuine pandemic occurred their vaccines could be licensed using a ‘fast track’ regulatory procedure. This process was applied in reality recently at the start of the 2009 pandemic. Whilst successful, it involved a burdensome process of continuous data evaluation for the regulators (and no doubt for the company personnel putting together the data). Lessons learnt from this experience are now being studied and applied.

9. Timelines for preparing pandemic vaccine The speed with which a pandemic vaccine can be produced is all important. Unlike any other vaccine, seasonal influenza

J.S. Robertson, S.C. Inglis / Virus Research 162 (2011) 39–46

vaccines are produced on an annual basis to take account of currently circulating strains. Producers have only 5–6 months in which to manufacture and release their products (Gerdil, 2003) and so even for these vaccines there is great pressure on all stages of the process, including generating candidate vaccine viruses (CVV). Considerable effort has therefore been invested into minimising the time required for each step, and this provides a very good platform for responding to a pandemic. In an emergency situation, however, the pressures will be extreme and so every aspect of the vaccine development from the WHO recommendation, vaccine virus development, vaccine manufacture and licensure need to be as resilient and rapid as possible. Since influenza can spread around the world in just a few weeks, it is well recognised that the likelihood of being able to produce and deliver enough vaccine in time to deal with a first wave of infection is extremely low. However there may be subsequent waves of infection upon which available vaccine could have substantial impact, saving a large number of lives. The five WHO Collaborating Centres for Influenza are responsible for collecting together influenza viruses from around the world and collating data on their characteristics. This system works very effectively providing rapid and timely intelligence on emerging problems, and well characterised isolates as a platform for the vaccine response. The derivation of a suitable CVV is a technically demanding process from which success cannot be guaranteed. Both the traditional mixed infection approach for the generation of high growth reassortants and the reverse genetics approach have been refined to the point that they take just under three weeks and there appears little further opportunity to reduce this time (Nicolson et al., 2005). During the initial development of pandemic H5N1 CVVs, considerable time was required for in vivo safety testing of CVVs. However it was agreed by WHO that an H5N1 candidate vaccine virus generated by reverse genetics and in which the pathogenic determining amino acids in the HA had been excised, could be dispatched to vaccine manufacturers prior to completion of in vivo safety tests if sufficient evidence had accrued to indicate that the virus appeared to be attenuated (WHO, 2005). In this way two to three weeks could be saved and whilst the in vivo tests were being completed, manufacturers could receive candidate viruses and develop their seed stocks under suitable containment until the safety of the viruses was confirmed. The time required for manufacturing the vaccine is often quoted as ∼3 to 4 months (WHO, 2009a). This is the minimum time to produce the first vaccine batches, however, rather than for completion of delivery of a vaccine order for a target population. In a pandemic there will be demand for hundreds of millions of doses and meeting this need will take many more months, making it still less likely that vaccine could be available to have a significant impact on a first pandemic wave. The final step in the process of vaccine manufacture is the release of finished batches, and a central element of this is testing for potency. For the current egg-derived inactivated vaccine, potency assays are performed using a single radial immunodiffusion (SRD) assay (Williams, 1993). These assays require vaccine specific reagents which also have to be prepared fresh using components of the newly emerged pandemic virus, another key pressure point in the process.

10. 2009 H1N1 pandemic It was perhaps fortuitous that the 2009 H1N1 pandemic turned out to be relatively mild compared with what many had been expecting. Although the age distribution of morbidity was similar to seasonal influenza, there was significantly greater mortality in young adults, especially in adolescents (Lemaitre and Carrat,

43

2010). How well did the vaccine development and manufacture process perform? A detailed account of the development of pandemic H1N1 candidate vaccine viruses by a small network of WHO laboratories has been published (Robertson et al., 2011). CVVs both traditional high growth reassortants and reverse genetics derived viruses were first made available within three weeks of the laboratories receiving the wild type virus recommended by WHO and these were distributed to industry up to two weeks before the WHO had declared a pandemic on 11 June 2009 (Chan, 2009; WHO, 2009b,c,d,e). Using what was determined to be the highest yielding of these initial CVVs, manufacturers were able to generate their first small batches of vaccine to enable initial clinical assessment by July 2009 (UPI, 2009). Such clinical trials were required to assess the immunogenicity of individual manufacturers’ vaccine, data that is required to determine the amount of antigen required per dose of vaccine and the number of doses that will be required to provide individuals with an acceptable level of protection. This information is vital to give public health bodies an indication of how many doses of vaccine can be produced and when. These studies also provide some early indication of the safety of the vaccines. One downside of these first 2009 pandemic vaccine viruses was that, whilst they were far superior to the wild type virus itself for vaccine production, they provided only about one third of the quantity of antigen per egg compared to what is usually expected from an H1N1 vaccine virus (Robertson et al., 2011). This was clearly an unsatisfactory situation. Thus, the WHO laboratories continued vaccine virus development and by July/August, two improved viruses were available, one a high growth reassortant, one a reverse genetics derived virus (WHO, 2009f,g). Their antigen yield now approached that more typically achieved for H1N1 viruses (Robertson et al., 2011). However, vaccine production was now well underway using the best of the initial series of CVVs and changing seed virus mid-way through a campaign presents difficulties for manufacturers. Hence these improved strains were not widely used. An important lesson learnt from this is that a vaccine virus with satisfactory growth properties needs to be available from the outset rather than midway through the campaign. The yield problem aside, the 2009 pandemic vaccine worked remarkably well (Valenciano et al., 2011; Hardelid et al., 2011). Perhaps inevitably, however, the availability of vaccine in large quantity from September/October 2009 onwards (EMA, 2009a,b) was too late for the first infection wave, which hit the northern hemisphere by June/July. Indeed by the time vaccines were manufactured, licensed, delivered and distribute, many countries were already experiencing a second wave of disease.

11. Vaccines for a future pandemic Given the investment in, and experience with egg-derived inactivated influenza vaccines, and the time that it would take to develop any serious alternative, it seems likely that they will continue to be mainstay of the pandemic response for at least the next 5 years. Cultured cell-derived vaccines have been developed, but manufacturing capacity is still very limited (Robertson, 2011) and the promise of a faster response, without the need for development of high yielding egg-adapted strain (wild type virus can be used for production, but this would require containment for a pathogenic virus) did not materialise. During the 2009 pandemic one cell-derived vaccine was marketed (EMA, 2009b). There is a continued need, therefore, to focus on improvements to the egg-based process to optimise the response, both in terms of overall output and speed. Overall output depends on the scale of manufacturing capacity around the world, but also crucially on the yield from each inoculated egg. In order to improve yields of antigen in egg-derived vaccines, studies in the authors’

44

J.S. Robertson, S.C. Inglis / Virus Research 162 (2011) 39–46

laboratory have investigated chimeric haemagglutinin structures that incorporate parts of the wild type haemagglutinin required for the vaccine and parts of the PR8 haemagglutinin. These chimeric structures contain the ectodomain of the wild type haemagglutinin and the endodomain and functional non-coding regions of the PR8 HA genome segment and have resulted in improved yield of an H5N1 vaccine virus and for a 2009 pandemic H1N1 vaccine virus (Harvey et al., 2010, 2011). It may therefore be possible in future to counter the problem of low yield experienced in 2009. Live attenuated influenza vaccines (LAIV) have potential advantages in a pandemic situation. It is possible that they might generate a broader and hence more effective immune response in a wider population (Chen and Subbarao, 2009). Clinical data indicate that they are more effective in children and adolescents in particular. It should also be possible to produce far larger numbers of vaccine doses in a shorter time, since very much less virus is needed per dose. LAIV was produced for the 2009 pandemic in the USA and the Russian Federation (WER, 2009). In the USA, LAIV comprised ∼25% of 2009 pandemic vaccine usage and there was significant use in children (Coelingh, 2011). An adjuvanted influenza vaccine has been on the European market for over ten years and has a proven safety profile with many millions of doses having been administered (Banzhoff et al., 2008). The use of adjuvants was a key feature of the 2009 pandemic vaccine response for European (EMA, 2011b) and other markets, but noticeably not in the USA where regulators took a more cautious approach to their use (Schubert, 2009). Their immunestimulating properties allowed a reduction in the quantity of virus antigen required for each dose of vaccine (antigen sparing), and hence a greater number of doses of vaccine to be prepared from a given batch of harvested antigen. There is also good evidence that adjuvanted vaccines give a broader immune response than nonadjuvanted vaccine and so can provide greater protection against drifted strains (Carter and Plosker, 2008; Leroux-Roels et al., 2008). In the 2009 pandemic, oil-in-water emulsions such as AS03 used in Pandemrix (EMA, 2011a), and MF59 used in Focetria (EMA, 2011c) were more prevalent that traditional aluminium based adjuvants. Whilst their use in the pandemic was, in general, free from safety issues, one particular adjuvanted vaccine – Pandemrix – appears to be associated with an increase in narcolepsy in children and adolescents in Finland (NIHW, 2011) and Sweden (MPA, 2011), resulting in restrictions on its use being imposed by the European Medicines Agency (EMA, 2011d). However, in response to the findings in Europe, a study in China suggests that narcolepsy results from seasonal upper respiratory tract infections and not from pandemic H1N1 vaccination (Han et al., 2011). Despite such concerns over increased reactogenicity of adjuvanted vaccines, they remain an important avenue for research (not just for influenza vaccine) and will no doubt play a key role in any future pandemic response. Influenza vaccine manufacture is concentrated in the developed world and the WHO has for several years been implementing a plan to introduce influenza vaccine manufacture into developing countries (Friede et al., 2011; GAP-II, 2011). This would help provide such areas with vaccine which they might otherwise not have access to or which they could not afford; at the same time it will help increase the global capacity of vaccine manufacture. Out of 11 countries being aided in this way, three have already achieved licensure of their first influenza vaccine (Kieny, 2011). However, the projected total capacity of these ventures remains low compared to the theoretical current global capacity of ∼850 million doses (per annum) (IFPMA, 2011). Novel influenza vaccines are the subject of a separate chapter in this issue but their use in a pandemic situation should be discussed. A recombinant haemagglutinin vaccine manufactured by a baculovirus expression system in insect cells has been

submitted to the US FDA for licensure (Cox and Hollister, 2009; FDA, 2009). Other recombinant approaches include VLPs (virus like particles) produced in a baculovirus system (Kang et al., 2009), a haemagglutinin–flagellin fusion protein produced in an E. coli system (Treanor et al., 2010), and haemagglutinin produced in a plant based system (Chichester et al., 2009). It has been proposed that large quantities of these recombinant vaccines could be manufactured much faster than conventional egg-based vaccine and thus would have great value in the event of a pandemic (Executive Office of the President, 2010). However, in order for this goal to be realised, the necessary manufacturing infrastructure would have to be built, recognising that it may only be needed infrequently, and a regulatory pathway developed to demonstrate that such vaccines are safe and efficacious. The ultimate goal must of course be a ‘universal’ vaccine that will protect against all subtypes of influenza (Du et al., 2010). If such a vaccine existed, it could become routine to vaccinate most of the world’s population to give them protection to both seasonal and pandemic influenza, and there would no longer be a need for pandemic planning and response. Conserved epitopes or antigens that are common to all or to wide groups of HA subtypes, such as the M2e antigen (Fiers et al., 2009; Turley et al., 2011) or T cell epitopes on the nucleoprotein and matrix proteins (Berthoud et al., 2011) are being utilised in novel vaccines, and results in small animal models have been encouraging, but these approaches await examination of their effectiveness in humans, both in trial situations, and in a real emergency. Approaching the problem from a different angle, broadly neutralising antibodies have been identified that bind to the stalk region or other highly conserved epitopes of the haemagglutinin (Corti et al., 2011; Ekiert et al., 2011). Such antibodies have potential use as therapeutic agents, although their use may be limited to life-threatening situations as they are likely to be highly expensive to manufacture. However, such research has opened the door to the development of broadly protective vaccines based upon the epitopes recognised by these newly discovered antibodies (Wang et al., 2010). For the foreseeable future, then, we will be reliant on mounting an effective vaccine response in response to an emergency, and this is dependent on numerous factors – the candidate vaccine virus, vaccine manufacturers, manufacturing staff, egg supply, licensing procedures, vaccination devices (syringes, needles, labels, packaging), distribution, health practitioners. This review has focused on many of the practical hurdles that need to be surmounted to deliver large amounts of safe and effective pandemic vaccine to the general public. In spite of this, the existing system works, but it is less than ideal in many ways, and there needs to be a continued focus on improvement. This will require close collaboration between influenza and vaccine experts, manufacturers, regulators and public health authorities around the world. Fortunately this community has worked together very well in the past to build the current system and will no do doubt do so again in the future. References Aoki, F.Y., 1998. Amantadine and rimantadine. In: Nicholson, K.G., Webster, R.G., Hay, A.J. (Eds.), Textbook of Influenza. Blackwell Science, Oxford, pp. 457–476. Banzhoff, A., Pellegrini, M., Del Giudice, G., Fragapane, E., Groth, N., Podda, A., 2008. MF59-adjuvanted vaccines for seasonal and pandemic influenza prophylaxis. Influenza and Other Respiratory Viruses 2 (6), 243–249. Berthoud, T.K., Hamill, M., Lillie, P.J., Hwenda, L., Collins, K.A., Ewer, K.J., Milicic, A., Poyntz, H.C., Lambe, T., Fletcher, H.A., Hill, A.V., Gilbert, S.C., 2011. Potent CD8+ T-cell immunogenicity in humans of a novel heterosubtypic influenza A vaccine, MVA-NP+M1. Clinical Infectious Diseases 52 (1), 1–7. Bucher, D., 2011. High Yield Reassortants for the Flu Vaccine: The Darwinian http://www.who.int/vaccine research/diseases/influenza/18 feb Approach, 2011 Doris Bucher ws.pdf (accessed 15.08.11). Cabinet Office, 2010. National Risk Register of Civil Emergencies, 2010 edition, pp. 7–10. http://www.cabinetoffice.gov.uk/sites/default/files/resources/ nationalriskregister-2010.pdf (accessed 15.08.11).

J.S. Robertson, S.C. Inglis / Virus Research 162 (2011) 39–46 Carter, N.J., Plosker, G.L., 2008. Prepandemic influenza vaccine H5N1 (split virion, inactivated, adjuvanted) [Prepandrix]: a review of its use as an active immunization against influenza A subtype H5N1 virus. BioDrugs 22 (5), 279–292. Chan, M., 2009. World now at the Start of 2009 Influenza Pandemic, http://www.who.int/mediacentre/news/statements/2009/h1n1 pandemic phase6 20090611/en/index.html (accessed 15.08.11). Chan, P.K., 2002. Outbreak of avian influenza A(H5N1) virus infection in Hong Kong in 1997. Clinical Infectious Diseases 34 (Suppl. 2), S58–S64. Chen, G.L., Subbarao, K., 2009. Live attenuated vaccines for pandemic influenza. Current Topics in Microbiology and Immunology 333, 109–132. Chichester, J.A., Haaheim, L.R., Yusibov, V., 2009. Using plant cells as influenza vaccine substrates. Expert Review of Vaccines 8 (4), 493–498. CIDRAP, 2008. US has Enough H5N1 Vaccine for 13 Million People, http://www.cidrap.umn.edu/cidrap/content/influenza/panflu/news/ mar2008panupdate.html (accessed 15.08.11). Coelingh, K., 2011. LAIV for Increasing Pandemic Vaccine Supply, http://www.who.int/influenza vaccines plan/resources/coelingh.pdf (accessed 15.08.11). Corti, D., Voss, J., Gamblin, S.J., Codoni, G., Macagno, A., Jarrossay, D., Vachieri, S.G., Pinna, D., Minola, A., Vanzetta, F., Silacci, C., Fernandez-Rodriguez, B.M., Agatic, G., Bianchi, S., Giacchetto-Sasselli, I., Calder, L., Sallusto, F., Collins, P., Haire, L.F., Temperton, N., Langedijk, J.P., Skehel, J.J., Lanzavecchia, A., 2011. A neutralizing antibody selected from plasma cells that binds to group 1 and group 2 influenza A hemagglutinins. Science 333 (6044), 850–856. Cox, M.M., Hollister, J.R., 2009. FluBlok, a next generation influenza vaccine manufactured in insect cells. Biologicals 37 (3), 182–189. Du, L., Zhou, Y., Jiang, S., 2010. Research and development of universal influenza vaccines. Microbes and Infection 12 (4), 280–286. Ekiert, D.C., Friesen, R.H., Bhabha, G., Kwaks, T., Jongeneelen, M., Yu, W., Ophorst, C., Cox, F., Korse, H.J., Brandenburg, B., Vogels, R., Brakenhoff, J.P., Kompier, R., Koldijk, M.H., Cornelissen, L.A., Poon, L.L., Peiris, M., Koudstaal, W., Wilson, I.A., Goudsmit, J., 2011. A highly conserved neutralizing epitope on group 2 influenza A viruses. Science 333 (6044), 843–850. EMA, 2008. European Medicines Agency. Guideline on Dossier Structure and Content for Pandemic Influenza Vaccine Marketing Authorisation Application (Revision), http://www.ema.europa.eu/docs/en GB/document library/Scientific guideline/ 2009/09/WC500003869.pdf (accessed 15.08.11). EMA, 2009a. European Medicines Agency. European Medicines Agency Recommends Authorisation for two Vaccines for Influenza Pandemic (H1N1) 2009, http://www.ema.europa.eu/ema/index.jsp?curl=pages/news and events/news/ 2009/11/news detail 000038.jsp&murl=menus/news and events/news and events.jsp&mid=WC0b01ac058004d5c1&jsenabled=true (accessed 15.08.11). EMA, 2009b. European Medicines Agency. European Medicines Agency Recommends Authorisation of Additional Vaccine for Influenza Pandemic (H1N1) 2009, http://www.ema.europa.eu/ema/index.jsp?curl=pages/news and events/news/ 2009/11/news detail 000013.jsp&murl=menus/news and events/news and events.jsp&mid=WC0b01ac058004d5c1&jsenabled=true (accessed 15.08.11). EMA, 2011a. European Medicines Agency. Prepandrix, http://www.ema.europa.eu/ ema/index.jsp?curl=pages/medicines/human/medicines/000822/human med 000986.jsp&murl=menus/medicines/medicines.jsp&mid= WC0b01ac058001d124 (accessed 15.08.11). EMA, 2011b. European Medicines Agency. Medicines Authorised During Pandemic, http://www.ema.europa.eu/ema/index.jsp?curl=pages/special topics/general/ general content 000251.jsp&murl=menus/special topics/special topics. jsp&mid=WC0b01ac05801db932 (accessed 15.08.11). EMA, 2011c. European Medicines Agency. Focetria, http://www.ema.europa.eu/ ema/index.jsp?curl=pages/medicines/human/medicines/000710/human med 000796.jsp&murl=menus/medicines/medicines.jsp&mid= WC0b01ac058001d124 (accessed 16.09.11). EMA, 2011d. European Medicines Agency. European Medicines Agency Recommends Restricting use of Pandemrix, http://www.ema.europa.eu/ docs/en GB/document library/Press release/2011/07/WC500109182.pdf (accessed 16.09.11). Executive Office of the President, 2010. Report to the President on Reengineering the Influenza Vaccine Production Enterprise to Meet the Challenges of Pandemic Influenza. Chapter V. Recombinant Influenza Vaccines, http://www.whitehouse.gov/sites/default/files/microsites/ostp/PCASTInfluenza-Vaccinology-Report.pdf (accessed 15.08.11). FDA, 2009. FDA Briefing Document. Influenza Vaccine, FluBlok® , http://www.fda. gov/downloads/advisorycommittees/committeesmeetingmaterials/ bloodvaccinesandotherbiologics/vaccinesandrelatedbiologicalproducts advisorycommittee/ucm190835.pdf (accessed 15.08.11). Ferguson, N.M., Cummings, D.A., Cauchemez, S., Fraser, C., Riley, S., Meeyai, A., Iamsirithaworn, S., Burke, D.S., 2005. Strategies for containing an emerging influenza pandemic in Southeast Asia. Nature 437, 209–214. Fiers, W., De Filette, M., El Bakkouri, K., Schepens, B., Roose, K., Schotsaert, M., Birkett, A., Saelens, X., 2009. M2e-based universal influenza A vaccine. Vaccine 27 (45), 6280–6283. Fodor, E., Devenish, L., Engelhardt, O.G., Palese, P., Brownlee, G.G., Garcia-Sastre, A., 1999. Rescue of influenza A virus from recombinant DNA. Journal of Virology 73, 9679–9682. Friede, M., Palkonyay, L., Alfonso, C., Pervikov, Y., Torelli, G., Wood, D., Kieny, M.P., 2011. WHO initiative to increase global and equitable access to influenza vaccine in the event of a pandemic: supporting developing country production capacity through technology transfer. Vaccine 29 (Suppl. 1), A2–A7.

45

Furminger, I.G.S., 1998. Vaccine production. In: Nicholson, K.G., Webster, R.G., Hay, A.J. (Eds.), Textbook of Influenza. Blackwell Science, Oxford, pp. 324–332. GAP-II, 2011. Second Consultation on the Global Action Plan for Influenza Vaccines, http://www.who.int/influenza vaccines plan/en/ (accessed 15.08.11). Garten, R.J., Davis, C.T., Russell, C.A., Shu, B., Lindstrom, S., Balish, A., Sessions, W.M., Xu, X., Skepner, E., Deyde, V., Okomo-Adhiambo, M., Gubareva, L., Barnes, J., Smith, C.B., Emery, S.L., Hillman, M.J., Rivailler, P., Smagala, J., de Graaf, M., Burke, D.F., Fouchier, R.A., Pappas, C., Alpuche-Aranda, C.M., López-Gatell, H., Olivera, H., López, I., Myers, C.A., Faix, D., Blair, P.J., Yu, C., Keene, K.M., Dotson Jr., P.D., Boxrud, D., Sambol, A.R., Abid, S.H., St George, K., Bannerman, T., Moore, A.L., Stringer, D.J., Blevins, P., Demmler-Harrison, G.J., Ginsberg, M., Kriner, P., Waterman, S., Smole, S., Guevara, H.F., Belongia, E.A., Clark, P.A., Beatrice, S.T., Donis, R., Katz, J., Finelli, L., Bridges, C.B., Shaw, M., Jernigan, D.B., Uyeki, T.M., Smith, D.J., Klimov, A.I., Cox, N.J., 2009. Antigenic and genetic characteristics of swine-origin 2009 A(H1N1) influenza viruses circulating in humans. Science 325, 197–201. Gerdil, C., 2003. The annual production cycle for influenza vaccine. Vaccine 21, 1776–1779. GISRS, 2011. Global Influenza Surveillance and Response System, http://www.who.int/influenza/gisrs laboratory/en/ (accessed 15.08.11). Han, F., Lin, L., Warby, S.C., Faraco, J., Li, J., Dong, S.X., An, P., Zhao, L., Wang, L.H., Li, Q.Y., Yan, H., Gao, Z.C., Yuan, Y., Strohl, K.P., Mignot, E., 2011. Narcolepsy onset is seasonal and increased following the 2009 H1N1 pandemic in China. Annals of Neurology 70 (3), 410–417. Hardelid, P., Fleming, D.M., McMenamin, J., Andrews, N., Robertson, C., Sebastianpillai, P., Ellis, J., Carman, W., Wreghitt, T., Watson, J.M., Pebody, R.G., 2011. Effectiveness of pandemic and seasonal influenza vaccine in preventing pandemic influenza A(H1N1)2009 infection in England and Scotland 2009–2010. Euro Surveillance 16 (2), pii: 19763. Harvey, R., Guilfoyle, K.A., Roseby, S., Robertson, J.S., Engelhardt, O.G., 2011. Improved antigen yield in pandemic H1N1 (2009) candidate vaccine viruses with chimeric haemagglutinin molecules. Journal of Virology 85, 6086–6090. Harvey, R., Nicolson, C., Johnson, R.E., Guilefoyle, K.A., Major, D.L., Robertson, J.S., Engelhardt, O.G., 2010. Improved haemagglutinin antigen content in H5N1 candidate vaccine viruses with chimeric haemagglutinin molecules. Vaccine 28, 8008–8014. HPA, 2008. Health Protection Agency (UK). Vaccines and Anti-viral http://www.hpa.org.uk/Topics/InfectiousDiseases/InfectionsAZ/ Drugs, AvianInfluenza/GeneralInformation/FrequentlyAskedQuestions/ influaVaccinesandantiviraldrugs/ (accessed 15.08.11). HPA, 2010. Health Protection Agency (UK). Epidemiological Report of Pandemic (H1N1) 2009 in the UK, http://www.hpa.org.uk/web/HPAwebFile/ HPAweb C/1284475321350 (accessed 15.08.11). Hurt, A.C., Holien, J.K., Parker, M.W., Barr, I.G., 2009. Oseltamivir resistance and the H274Y neuraminidase mutation in seasonal, pandemic and highly pathogenic influenza viruses. Drugs 69 (18), 2523–2531. IFPMA, 2011. Industry Perspective: Strategies and Costs Associated with Increasing Seasonal Influenza Vaccine use from High to Low Income Countries Since 2006, http://www.who.int/influenza vaccines plan/resources/watson1.pdf (accessed 15.08.11). Kang, S.M., Pushko, P., Bright, R.A., Smith, G., Compans, R.W., 2009. Influenza virus-like particles as pandemic vaccines. Current Topics in Microbiology and Immunology 333, 269–289. Kieny, M.-P., 2011. Review of Global Activities and Achievements for GAP Since 2006, http://www.who.int/influenza vaccines plan/resources/mpk.pdf (accessed 15.08.11). Kilbourne, E.D., 1969. Future influenza vaccines and the use of genetic recombinants. Bulletin of the World Health Organization 41, 643–645. Kilbourne, E.D., Schulman, J.L., Schild, G.C., Schloer, G., Swanson, J., Bucher, D., 1971. Correlated studies of a recombinant influenza virus vaccine. I. Derivation and characterization of the virus and vaccine. Journal of Infectious Diseases 124, 449–462. Lemaitre, M., Carrat, F., 2010. Comparative age distribution of influenza morbidity and mortality during seasonal influenza epidemics and the 2009 H1N1 pandemic. BMC Infectious Diseases 10, 162. Leroux-Roels, I., Bernhard, R., Gérard, P., Dramé, M., Hanon, E., Leroux-Roels, G., 2008. Broad Clade 2 cross-reactive immunity induced by an adjuvanted clade 1 rH5N1 pandemic influenza vaccine. PLoS One 3 (2), e1665. Maassab, H.F., Bryant, M.L., 1999. The development of live attenuated coldadapted influenza virus vaccine for humans. Reviews in Medical Virology 9 (4), 237–244. McNicholl, I.R., McNicholl, J.J., 2001. Neuraminidase inhibitors: zanamivir and oseltamivir. Annals of Pharmacotherapy 35 (1), 57–70. MPA, 2011. Medical Products Agency (Sweden). A Swedish Registry Based Cohort Study Provides Strengthened Evidence of an Association Between Vaccination with Pandemrix and Narcolepsy in Children and Adolescents, http://www.lakemedelsverket.se/english/All-news/NYHETER-2011/ASwedish-registry-based-cohort-study-provides-strengthened-evidenceof-an-association-between-vaccination-with-Pandemrix-and-narcolepsy-inchildren-and-adolescents-/ (accessed 16.09.11). Neumann, G., Kawaoka, Y., 2011. The first influenza pandemic of the new millennium. Influenza and Other Respiratory Viruses 5, 157–166. Nicolson, C., Major, D., Wood, J.M., Robertson, J.S., 2005. Generation of influenza vaccine viruses on Vero cells by reverse genetics: an H5N1 candidate vaccine strain produced under a quality system. Vaccine 23, 2943–2952.

46

J.S. Robertson, S.C. Inglis / Virus Research 162 (2011) 39–46

NIHW, 2011. National Institute for Health and Welfare (Finland). Association Between Pandemrix and Narcolepsy Confirmed Among Finnish Children and Adolescents, http://www.thl.fi/en US/web/en/pressrelease?id= 26352 (accessed 16.09.11). Nishiura, H., 2010. The virulence of pandemic influenza A (H1N1) 2009: an epidemiological perspective on the case–fatality ratio. Expert Review of Respiratory Medicine 4, 329–338. Nguyen-Van-Tam, J.S., Openshaw, P.J., Hashim, A., Gadd, E.M., Lim, W.S., Semple, M.G., Read, R.C., Taylor, B.L., Brett, S.J., McMenamin, J., Enstone, J.E., Armstrong, C., Nicholson, K.G., 2010. Risk factors for hospitalisation and poor outcome with pandemic A/H1N1 influenza: United Kingdom first wave (May–September 2009). Thorax 65, 645–651. OIE, 2005. Chapter 2.7.12. Avian Influenza, http://web.oie.int/fr/normes/mmanual/ A 00037.htm (accessed 15.08.11). OFFLU, 2011. OIE/FAO Network of Expertise on Animal Influenza, http://www.offlu.net/index.html (accessed 15.08.11). Pan, K., Deem, M.W., 2011. Quantifying selection and diversity in viruses by entropy methods, with application to the haemagglutinin of H3N2 influenza. Journal of the Royal Society, Interface [Epub ahead of print]. Potter, C.W., 1998. Chronicle of influenza pandemics. In: Nicholson, K.G., Webster, R.G., Hay, A.J. (Eds.), Textbook of Influenza. Blackwell Science, Oxford, pp. 3–18. Rappuoli, R., 2003. Cell-culture-based vaccine production: technological options. Engineering and Vaccine Production for an Influenza Pandemic 36 (3), http://www.nae.edu/Publications/Bridge/EngineeringandVaccineProduction foranInfluenzaPandemic/Cell-Culture-BasedVaccineProductionTechnological Options.aspx (accessed 15.08.11). Robertson, J.S., Engelhardt, O.G., 2010. Developing vaccines to combat pandemic influenza. Viruses 2, 532–546. Robertson, J.S., 2011. Egg- and Cell-derived Influenza Vaccines: Advantages and Drawbacks, http://www.who.int/vaccine research/diseases/ influenza/18 feb 2011 Jim Robertson ws.pdf (accessed 15.08.11). Robertson, J.S., Nicolson, C., Harvey, R., Johnson, R., Major, D., Guilfoyle, K., Roseby, S., Newman, R., Collin, R., Wallis, C., Engelhardt, O.G., Wood, J.M., Le, J., Manojkumar, R., Pokorny, B.A., Silverman, J., Devis, R., Bucher, D., Verity, E., Agius, C., Camuglia, S., Ong, C., Rockman, S., Curtis, A., Schoofs, P., Zoueva, O., Xie, H., Li, X., Lin, Z., Ye, Z., Chen, L.M., O’Neill, E., Balish, A., Lipatov, A.S., Guo, Z., Isakova, I., Davis, C.T., Rivailler, P., Gustin, K.M., Belser, J.A., Maines, T.R., Tumpey, T.M., Xu, X., Katz, J.M., Klimov, A., Cox, N.J., Donis, R.O., 2011. The development of vaccine viruses against pandemic A(H1N1) influenza. Vaccine 29, 1836–1843. Rott, R., 1992. The pathogenic determinant of influenza virus. Veterinary Microbiology 33, 303–310. Schubert, C., 2009. Swine flu agitates the adjuvant debate. Nature Medicine 15, 986–987. Sinovac, 2011. Panflu® – Pandemic Influenza Vaccine (Inactivated, Adjuvanted), http://www.sinovac.com/?optionid=460 (accessed 15.08.11). Subbarao, K., Chen, H., Swayne, D., Mingay, L., Fodor, E., Brownlee, G., Xu, X., Lu, X., Katz, J., Cox, N., Matsuoka, Y., 2003. Evaluation of a genetically modified reassortant H5N1 influenza A virus vaccine candidate generated by plasmid-based reverse genetics. Virology 305, 192–200. Taubenberger, J.K., Morens, D.M., 2006. 1918 Influenza: the mother of all pandemics. Emerging Infectious Diseases 12, 15–22. Treanor, J.J., Taylor, D.N., Tussey, L., Hay, C., Nolan, C., Fitzgerald, T., Liu, G., Kavita, U., Song, L., Dark, I., Shaw, A., 2010. Safety and immunogenicity of a recombinant hemagglutinin influenza–flagellin fusion vaccine (VAX125) in healthy young adults. Vaccine 28 (52), 8268–8274. Turley, C.B., Rupp, R.E., Johnson, C., Taylor, D.N., Wolfson, J., Tussey, L., Kavita, U., Stanberry, L., Shaw, A., 2011. Safety and immunogenicity of a recombinant M2e-flagellin influenza vaccine (STF2.4xM2e) in healthy adults. Vaccine 29 (32), 5145–5152. UPI, 2009. First Human H1N1 Vaccine Trials Begin, http://www.upi.com/ Science News/2009/07/22/First-human-H1N1-vaccine-trials-begin/UPI22641248274043/ (accessed 15.08.11). Valenciano, M., Kissling, E., Cohen, J.M., Oroszi, B., Barret, A.S., Rizzo, C., Nunes, B., Pitigoi, D., Larrauri Cámara, A., Mosnier, A., Horvath, J.K., O’Donnell, J., Bella, A., Guiomar, R., Lupulescu, E., Savulescu, C., Ciancio, B.C., Kramarz, P., Moren, A., 2011. Estimates of pandemic influenza vaccine effectiveness in Europe, 2009–2010: results of influenza monitoring vaccine effectiveness in Europe (IMOVE) multicentre case–control study. PLoS Medicine 8 (1), e1000388. Wang, T.T., Tan, G.S., Hai, R., Pica, N., Ngai, L., Ekiert, D.C., Wilson, I.A., García-Sastre, A., Moran, T.M., Palese, P., 2010. Vaccination with a synthetic peptide from the influenza virus hemagglutinin provides protection against distinct viral subtypes. Proceedings of the National Academy of Sciences of the United States of America 107 (44), 18979–18984.

Ward, P., Small, I., Smith, J., Suter, P., Dutkowski, R., 2005. Oseltamivir (Tamifluw) and its potential for use in the event of an influenza pandemic. Journal of Antimicrobial Chemotherapy 55 (Suppl. S1), i5–i21. Webby, R.J., Perez, D.R., Coleman, J.S., Guan, Y., Knight, J.H., Govorkova, E.A., McClain-Moss, L.R., Peiris, J.S., Rehg, J.E., Tuomanen, E.I., Webster, R.G., 2004. Responsiveness to a pandemic alert: use of reverse genetics for rapid development of influenza vaccines. Lancet 363, 1099–1103. WER, 2009. Pandemic influenza A (H1N1) 2009 virus vaccine – conclusions and recommendations from the October 2009 meeting of the immunization Strategic Advisory Group of Experts. Weekly Epidemiological Record 84 (49), 505–508. WER, 2011. Antigenic and genetic characteristics of influenza A(H5N1) and influenza A(H9N2) viruses for development of candidate vaccines viruses for pandemic preparedness. Weekly Epidemiological Record 86 (11), 93–100. 2011. 64th World Health Assembly, Resolution 64, WHA, http://apps.who.int/gb/ebwha/pdf files/WHA64/A64 8-en.pdf (accessed 15.08.11). WHO, 2005. Annex 5. WHO Biosafety Risk Assessment and Guidelines for the Production and Quality Control of Human Influenza Pandemic Vachttp://www.who.int/biologicals/publications/ECBS%202005%20 cines, Annex%205%20Influenza.pdf (accessed 15.08.11). 2007. Global Stockpile of H5N1 Vaccine ‘feasible’, WHO, http://www.who.int/mediacentre/news/releases/2007/pr21/en/ index.html (accessed 15.08.11). WHO, 2009a. Pandemic Influenza Vaccine Manufacturing Process and Timeline, http://www.who.int/csr/disease/swineflu/notes/h1n1 vaccine 20090806/en/ index.html (accessed 15.08.11). WHO, 2009b. Availability of a Candidate Reassortant Vaccine Virus for the Novel Influenza A (H1N1) Vaccine Development. 27 May 2009. IDCDCRG15, http://www.who.int/csr/resources/publications/swineflu/2009 05 27IDCDCRG15a.pdf (accessed 15.08.11). WHO, 2009c. Availability of a Candidate Reassortant Vaccine Virus for the Novel Influenza A (H1N1) Vaccine Development. 27 May 2009. NIBRG121, http://www.who.int/csr/resources/publications/swineflu/2009 05 27 NIBRG121a.pdf (accessed 15.08.11). WHO, 2009d. Availability of a Candidate Reassortant Vaccine Virus for the Novel Influenza A (H1N1) Vaccine Development. 27 May 2009. X-179A, http://www.who.int/csr/resources/publications/swineflu/2009 05 27 X179Aa.pdf (accessed 15.08.11). WHO, 2009e. Availability of a Candidate Reassortant Vaccine Virus for the Novel Influenza A (H1N1) Vaccine Development. 4 June 2009. IVR-153, http://www.who.int/csr/resources/publications/swineflu/ivr153 20090608 en.pdf (accessed 15.08.11). WHO, 2009f. Availability of a New Candidate Reassortant Vaccine Virus for Pandemic (H1N1) 2009 Virus Vaccine Development. 6 August 2009. NIBRG-121xp, http://www.who.int/csr/disease/swineflu/guidance/vaccines/candidates/ cp122 2009 0608 availability of new cr vaccine virus nibrg-121-final.pdf (accessed 15.08.11). WHO, 2009g. Availability of two New Candidate Reassortant Vaccine Viruses for Pandemic (H1N1) 2009 Virus Vaccine Development. 14 September 2009. X-181 and X-181A, http://www.who.int/csr/resources/publications/ swineflu/x 181 x 181a.pdf (accessed 15.08.11). 2011a. Influenza Pandemic Threat: Current Situation, WHO, http://www.who.int/csr/disease/avian influenza/pandemic/en/index.html (accessed 15.08.11). 2011b. Recommendations for Influenza Vaccines, WHO, http://www.who.int/csr/disease/influenza/vaccinerecommendations/en/ (accessed 15.08.11). WHO, 2011c. H5N1 Avian Influenza: Timeline of Major Events, http://www.who. (accessed int/csr/disease/avian influenza/H5N1 avian influenza update.pdf 15.08.11). WHO, 2011d. Cumulative Number of Confirmed Human Cases of Avian Influenza A(H5N1) Reported to WHO, http://www.who.int/csr/disease/avian influenza/country/cases table 2011 08 09/en/index.html (accessed 15.08.11). Wikipedia, 2011. Global Spread of H5N1, http://en.wikipedia.org/wiki/Global spread of H5N1 (accessed 15.08.11). Williams, M.S., 1993. Single-radial-immunodiffusion as an in vitro potency assay for human inactivated viral vaccines. Veterinary Microbiology 37 (3–4), 253–262. Wong, S.S., Yuen, K.Y., 2006. Avian influenza virus infections in humans. Chest 129 (1), 156–168. Wood, J.W., Williams, M.S., 1998. History of inactivated influenza vaccines. In: Nicholson, K.G., Webster, R.G., Hay, A.J. (Eds.), Textbook of Influenza. Blackwell Science, Oxford, pp. 317–323.