New England Journal of Medicine - NEJM
Vaccines against Avian Influenza ? A Race against Time
Gregory A. Poland, M.D.
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</td> <td align="center" valign="top"><table border="0" cellpadding="0" cellspacing="0"> <tbody><tr><th align="center" nowrap="nowrap" valign="top">Volume 354:1411-1413</th> <td align="center" nowrap="nowrap"></td> <th align="center" nowrap="nowrap" valign="top">March 30, 2006</th> <td align="center" nowrap="nowrap"></td> <th align="center" nowrap="nowrap" valign="top">Number 13</th></tr></tbody></table>http://content.nejm.org/cgi/content/full/354/13/1411
</td></tr></tbody></table>
Vaccines against Avian Influenza ? A Race against Time
Gregory A. Poland, M.D.
<table border="0" cellpadding="0" cellspacing="0" height="49" width="640"><tbody><tr><td nowrap="nowrap" valign="top">
</td> <td align="center" valign="top"><table border="0" cellpadding="0" cellspacing="0"> <tbody><tr><th align="center" nowrap="nowrap" valign="top">Volume 354:1411-1413</th> <td align="center" nowrap="nowrap"></td> <th align="center" nowrap="nowrap" valign="top">March 30, 2006</th> <td align="center" nowrap="nowrap"></td> <th align="center" nowrap="nowrap" valign="top">Number 13</th></tr></tbody></table>http://content.nejm.org/cgi/content/full/354/13/1411
</td></tr></tbody></table>
Avian influenza A (H5N1) virus poses an important pandemic threat.<sup> </sup>A study by the Congressional Budget Office estimates that the<sup> </sup>consequences of a severe pandemic could, in the United States,<sup> </sup>include 200 million people infected, 90 million clinically ill,<sup> </sup>and 2 million dead.<sup>1</sup> The study estimates that 30 percent of<sup> </sup>all workers would become ill and 2.5 percent would die, with<sup> </sup>30 percent of workers missing a mean of three weeks of work<sup> </sup>? resulting in a decrease in the gross domestic product<sup> </sup>of 5 percent. Furthermore, 18 million to 45 million people would<sup> </sup>require outpatient care, and economic costs would total approximately<sup> </sup>$675 billion. As of March 10, 2006, the World Health Organization<sup> </sup>(WHO) had reported 176 confirmed human cases of influenza A<sup> </sup>(H5N1) across seven countries, with 97 deaths (a 55 percent<sup> </sup>mortality rate for identified cases).<sup>2</sup>
<sup> </sup> A major worry is that influenza A (H5N1) continues to evolve<sup> </sup>into antigenically distinct clades ? infecting mammalian<sup> </sup>hosts other than humans, expanding its ecologic niche of domestic<sup> </sup>fowl to include wild migratory birds, and causing outbreaks<sup> </sup>among birds in more than 30 countries. The virus is close to<sup> </sup>meeting the criteria for a pandemic virus ? one that is<sup> </sup>new, can cause human illness, and can be transmitted from human<sup> </sup>to human<sup>3</sup><sup>,</sup><sup>4</sup><sup>,</sup><sup>5</sup> ? and the world is currently in phase three<sup> </sup>of the six WHO phases of alert for pandemic influenza (higher<sup> </sup>numbers represent greater seriousness). Influenza A (H5N1) is<sup> </sup>not yet pandemic because of a single factor: the inefficiency<sup> </sup>of human-to-human transmission. Once such transmission is efficient<sup> </sup>and sustained, even assuming that the current mortality rate<sup> </sup>of approximately 50 percent decreases, we will be in the midst<sup> </sup>of a serious pandemic.
<sup> </sup>
For this reason, maintaining the public health requires attempts<sup> </sup>to mitigate, avert, and treat infection with influenza A (H5N1)<sup> </sup>virus, and the key to meeting these goals is the development,<sup> </sup>testing, licensing, manufacturing, and stockpiling of vaccines.<sup> </sup>Safe and effective vaccines are likely to be the single most<sup> </sup>important public health tool for decreasing the morbidity, mortality,<sup> </sup>and economic effects of pandemic influenza ? particularly<sup> </sup>in view of the reported resistance of influenza A (H5N1) to<sup> </sup>antiviral agents.<sup>6</sup>
<sup>
</sup><sup> </sup>
Thus, the data reported in this issue of the Journal by Treanor<sup> </sup>et al.<sup>7</sup> from their multicenter randomized, double-blind, placebo-controlled<sup> </sup>clinical trial of a subvirion influenza A (H5N1) vaccine are<sup> </sup>important and informative. Enrolled in the study were 451 healthy<sup> </sup>adults 18 to 64 years of age who received two doses of the vaccine<sup> </sup>without adjuvant, each of which contained 90, 45, 15, or 7.5<sup> </sup>?g of hemagglutinin antigen, or placebo. The vaccine was<sup> </sup>produced from a human isolate (A/Vietnam/1203/2004 [H5N1]) of<sup> </sup>a virulent clade 1 influenza A (H5N1) virus with the use of<sup> </sup>a plasmid rescue system, with only the hemagglutinin and neuraminidase<sup> </sup>genes expressed. The rest of the genes were derived from an<sup> </sup>avirulent egg-adapted influenza A/PR/8/34 strain. The hemagglutinin<sup> </sup>gene was further modified to replace six basic amino acids associated<sup> </sup>with high pathogenicity in birds at the cleavage site between<sup> </sup>hemagglutinin 1 and hemagglutinin 2. Immunogenicity was assessed<sup> </sup>by microneutralization and hemagglutination-inhibition assays<sup> </sup>with the use of the vaccine virus, although a subgroup of samples<sup> </sup>were tested with the use of the wild-type influenza A/Vietnam/1203/2004<sup> </sup>(H5N1) virus.
<sup> </sup>
The results of the vaccine in the study by Treanor et al. (referred<sup> </sup>to here as the "1203 vaccine") give pause. Although the 1203<sup> </sup>vaccine was safe, with an unremarkable adverse-event profile,<sup> </sup>its immunogenicity was poor to moderate at best. In fact, in<sup> </sup>only one group did more than 50 percent of the subjects reach<sup> </sup>the immunogenicity threshold (defined a priori) of an antibody<sup> </sup>titer of 1:40 or greater (typically thought of as seroprotective)<sup> </sup>? the subjects who received two doses of 90 ?g each<sup> </sup>28 days apart ? a total dose 12 times that of seasonal<sup> </sup>influenza vaccines. Notably, the current worldwide manufacturing<sup> </sup>capacity for influenza vaccine is estimated at only 900 million<sup> </sup>doses (at the dose level of 15 ?g). The requirement of<sup> </sup>two doses of 90 ?g per person means that only 75 million<sup> </sup>persons (1.25 percent of the world's population) could be fully<sup> </sup>immunized, and of those, only half would achieve seroprotection.<sup> </sup>Thus, vaccines must contain much less influenza hemagglutinin<sup> </sup>to be widely useful as a global public health measure.
<sup> </sup>
And there are some additional provisos. An antibody titer of<sup> </sup>1:40 does not guarantee protection from infection. People with<sup> </sup>lower titers show protection against influenza, and people with<sup> </sup>higher titers can have symptomatic infection. Moreover, the<sup> </sup>assumption that a titer of 1:40 is seroprotective is based on<sup> </sup>circulating strains of seasonal influenza.
Whether the same<sup> </sup>will prove to be true for new influenza viruses in people whose<sup> </sup>immune systems have not been primed is unknown. However, even<sup> </sup>moderate levels of seroprotection could be useful for the public<sup> </sup>health by preventing or decreasing transmissibility, severe<sup> </sup>symptoms, complications, or death.
<sup> </sup>
An important issue is whether the 1203 vaccine offers cross-protection<sup> </sup>against other H5N1 strains of influenza A. A lethal human infection<sup> </sup>with an antigenically distinct influenza A (H5N1) strain is<sup> </sup>discussed elsewhere in this issue of the Journal.<sup>8</sup> From an immunologic<sup> </sup>standpoint, it is probable that more than one H5N1 vaccine will<sup> </sup>be needed. We know that the Indonesian clade 2 influenza A (H5N1)<sup> </sup>viruses are antigenically distinct from the clade 1 viruses<sup> </sup>from which the 1203 vaccine was developed. Preliminary evidence<sup> </sup>from serologic studies of laboratory-confirmed cases of influenza<sup> </sup>A (H5N1) infection also suggests that cross-protection between<sup> </sup>these two influenza A (H5N1) clades may be limited (Katz J:<sup> </sup>personal communication).
Therefore, further studies are warranted<sup> </sup>to establish the level of cross-neutralizing antibody against<sup> </sup>heterologous influenza A (H5N1) viruses, such as those in clade<sup> </sup>2, that is generated by vaccination with the 1203 vaccine. Such<sup> </sup>cross-neutralization is of great importance, because at the<sup> </sup>current time, the 1203 vaccine is being stockpiled for use in<sup> </sup>the event of an influenza A (H5N1) pandemic. In any case, one<sup> </sup>candidate for a clade 2 vaccine is now available, and others<sup> </sup>are being developed by the WHO Influenza Network.
<sup> </sup>
Additional factors for which data are needed include differences<sup> </sup>in vaccine-induced immunity according to age, sex, immune status,<sup> </sup>and ethnic group. Some of these data could be derived from the<sup> </sup>results of Treanor et al. on further analysis. Age may be particularly<sup> </sup>important; those who have died in past pandemics and from influenza<sup> </sup>A (H5N1) infection are disproportionately children, adolescents,<sup> </sup>and young adults.
<sup> </sup>
Studies of different dose levels of vaccines administered with<sup> </sup>MF59 (a licensed adjuvant in Europe), aluminum hydroxide, or<sup> </sup>other adjuvants are urgently needed. We know from previous work<sup> </sup>that new hemagglutinin proteins (including H5) in people who<sup> </sup>have not been primed are poorly immunogenic.<sup>9</sup><sup>,</sup><sup>10</sup> In recognition<sup> </sup>of this fact, the Department of Health and Human Services and<sup> </sup>the National Institutes of Health have funded studies of more<sup> </sup>than 30 candidate vaccines. Early results from some of these<sup> </sup>trials should be available in the next 6 to 12 months. Previous<sup> </sup>studies of a new influenza A (H5N3) vaccine administered with<sup> </sup>MF59 adjuvant showed that vaccine administered without adjuvant<sup> </sup>was poorly immunogenic but that vaccine administered with MF59<sup> </sup>adjuvant in two doses, each as low as 7.5 ?g, was highly<sup> </sup>immunogenic and resulted in cross-neutralizing antibodies against<sup> </sup>influenza A (H5N1).<sup>11</sup><sup>,</sup><sup>12</sup> Studies of an influenza A (H2N2) vaccine<sup> </sup>administered with alum adjuvant had similar results:
hemagglutination-inhibition<sup> </sup>titers increased significantly at doses as low as 1.9 ?g.<sup>9</sup><sup> </sup>
The immediate development and testing of such antigen-sparing<sup> </sup>vaccines administered with adjuvant are imperative both to improve<sup> </sup>immunogenicity and to increase the number of doses available<sup> </sup>(if lower doses are effective). In addition, live attenuated<sup> </sup>cold-adapted influenza vaccines are safe, are immunogenic, and<sup> </sup>have the relevant advantage of cross-protection against heterologous<sup> </sup>influenza strains ? suggesting a promising avenue to the<sup> </sup>development of pandemic vaccines. A contract for the development<sup> </sup>of such vaccines has been awarded to MedImmune. Other approaches<sup> </sup>to vaccine development involve DNA, adenovirus vectors,<sup>13</sup> and<sup> </sup>cell-culture manufacturing techniques to increase the speed<sup> </sup>and capacity of vaccine production. These approaches are promising,<sup> </sup>particularly since reverse-genetics reassortant vaccine candidates<sup> </sup>can be generated within weeks.<sup>14</sup>
<sup>
</sup><sup> </sup>
Thirty years ago, the United States attempted to respond to<sup> </sup>the threat of pandemic influenza with a vaccine approach. Now,<sup> </sup>armed with a greater understanding of the science, we have the<sup> </sup>capacity and the responsibility to embark on multiple, parallel<sup> </sup>avenues of vaccine development. In addition, we need efficient,<sup> </sup>rapid, high-yield, low-cost manufacturing innovations; the rapid<sup> </sup>generation of candidate vaccines for other, potentially pandemic<sup> </sup>influenza viruses (including emerging clade-2 influenza A [H5N1]<sup> </sup>viruses); and the rapid movement of those vaccines into clinical<sup> </sup>trials. In turn, this effort will require creativity along the<sup> </sup>entire pipeline: in the development and manufacture of candidate<sup> </sup>vaccines; the synchronization among countries of regulatory<sup> </sup>approaches; the resolution of issues concerning liability and<sup> </sup>intellectual property; ensuring the efficiency of clinical trials;<sup> </sup>and the use of methods to stockpile and rapidly deploy these<sup> </sup>vaccines. To do otherwise, with the pandemic clock ticking,<sup> </sup>could prove to be too little, too late.
<sup> </sup>
Dr. Poland reports serving as the chair of a data monitoring<sup> </sup>and safety board for an investigational trial of an influenza<sup> </sup>peptide vaccine being conducted by Merck Research Laboratories.<sup> </sup>No other potential conflict of interest relevant to this article<sup> </sup>was reported.<sup> </sup>
Source Information
From the Mayo Vaccine Research Group, the Program in Translational Immunovirology and Biodefense, and the Department of Internal Medicine, Mayo Clinic College of Medicine, Rochester, Minn.
References
<sup> </sup> A major worry is that influenza A (H5N1) continues to evolve<sup> </sup>into antigenically distinct clades ? infecting mammalian<sup> </sup>hosts other than humans, expanding its ecologic niche of domestic<sup> </sup>fowl to include wild migratory birds, and causing outbreaks<sup> </sup>among birds in more than 30 countries. The virus is close to<sup> </sup>meeting the criteria for a pandemic virus ? one that is<sup> </sup>new, can cause human illness, and can be transmitted from human<sup> </sup>to human<sup>3</sup><sup>,</sup><sup>4</sup><sup>,</sup><sup>5</sup> ? and the world is currently in phase three<sup> </sup>of the six WHO phases of alert for pandemic influenza (higher<sup> </sup>numbers represent greater seriousness). Influenza A (H5N1) is<sup> </sup>not yet pandemic because of a single factor: the inefficiency<sup> </sup>of human-to-human transmission. Once such transmission is efficient<sup> </sup>and sustained, even assuming that the current mortality rate<sup> </sup>of approximately 50 percent decreases, we will be in the midst<sup> </sup>of a serious pandemic.
<sup> </sup>
For this reason, maintaining the public health requires attempts<sup> </sup>to mitigate, avert, and treat infection with influenza A (H5N1)<sup> </sup>virus, and the key to meeting these goals is the development,<sup> </sup>testing, licensing, manufacturing, and stockpiling of vaccines.<sup> </sup>Safe and effective vaccines are likely to be the single most<sup> </sup>important public health tool for decreasing the morbidity, mortality,<sup> </sup>and economic effects of pandemic influenza ? particularly<sup> </sup>in view of the reported resistance of influenza A (H5N1) to<sup> </sup>antiviral agents.<sup>6</sup>
<sup>
</sup><sup> </sup>
Thus, the data reported in this issue of the Journal by Treanor<sup> </sup>et al.<sup>7</sup> from their multicenter randomized, double-blind, placebo-controlled<sup> </sup>clinical trial of a subvirion influenza A (H5N1) vaccine are<sup> </sup>important and informative. Enrolled in the study were 451 healthy<sup> </sup>adults 18 to 64 years of age who received two doses of the vaccine<sup> </sup>without adjuvant, each of which contained 90, 45, 15, or 7.5<sup> </sup>?g of hemagglutinin antigen, or placebo. The vaccine was<sup> </sup>produced from a human isolate (A/Vietnam/1203/2004 [H5N1]) of<sup> </sup>a virulent clade 1 influenza A (H5N1) virus with the use of<sup> </sup>a plasmid rescue system, with only the hemagglutinin and neuraminidase<sup> </sup>genes expressed. The rest of the genes were derived from an<sup> </sup>avirulent egg-adapted influenza A/PR/8/34 strain. The hemagglutinin<sup> </sup>gene was further modified to replace six basic amino acids associated<sup> </sup>with high pathogenicity in birds at the cleavage site between<sup> </sup>hemagglutinin 1 and hemagglutinin 2. Immunogenicity was assessed<sup> </sup>by microneutralization and hemagglutination-inhibition assays<sup> </sup>with the use of the vaccine virus, although a subgroup of samples<sup> </sup>were tested with the use of the wild-type influenza A/Vietnam/1203/2004<sup> </sup>(H5N1) virus.
<sup> </sup>
The results of the vaccine in the study by Treanor et al. (referred<sup> </sup>to here as the "1203 vaccine") give pause. Although the 1203<sup> </sup>vaccine was safe, with an unremarkable adverse-event profile,<sup> </sup>its immunogenicity was poor to moderate at best. In fact, in<sup> </sup>only one group did more than 50 percent of the subjects reach<sup> </sup>the immunogenicity threshold (defined a priori) of an antibody<sup> </sup>titer of 1:40 or greater (typically thought of as seroprotective)<sup> </sup>? the subjects who received two doses of 90 ?g each<sup> </sup>28 days apart ? a total dose 12 times that of seasonal<sup> </sup>influenza vaccines. Notably, the current worldwide manufacturing<sup> </sup>capacity for influenza vaccine is estimated at only 900 million<sup> </sup>doses (at the dose level of 15 ?g). The requirement of<sup> </sup>two doses of 90 ?g per person means that only 75 million<sup> </sup>persons (1.25 percent of the world's population) could be fully<sup> </sup>immunized, and of those, only half would achieve seroprotection.<sup> </sup>Thus, vaccines must contain much less influenza hemagglutinin<sup> </sup>to be widely useful as a global public health measure.
<sup> </sup>
And there are some additional provisos. An antibody titer of<sup> </sup>1:40 does not guarantee protection from infection. People with<sup> </sup>lower titers show protection against influenza, and people with<sup> </sup>higher titers can have symptomatic infection. Moreover, the<sup> </sup>assumption that a titer of 1:40 is seroprotective is based on<sup> </sup>circulating strains of seasonal influenza.
Whether the same<sup> </sup>will prove to be true for new influenza viruses in people whose<sup> </sup>immune systems have not been primed is unknown. However, even<sup> </sup>moderate levels of seroprotection could be useful for the public<sup> </sup>health by preventing or decreasing transmissibility, severe<sup> </sup>symptoms, complications, or death.
<sup> </sup>
An important issue is whether the 1203 vaccine offers cross-protection<sup> </sup>against other H5N1 strains of influenza A. A lethal human infection<sup> </sup>with an antigenically distinct influenza A (H5N1) strain is<sup> </sup>discussed elsewhere in this issue of the Journal.<sup>8</sup> From an immunologic<sup> </sup>standpoint, it is probable that more than one H5N1 vaccine will<sup> </sup>be needed. We know that the Indonesian clade 2 influenza A (H5N1)<sup> </sup>viruses are antigenically distinct from the clade 1 viruses<sup> </sup>from which the 1203 vaccine was developed. Preliminary evidence<sup> </sup>from serologic studies of laboratory-confirmed cases of influenza<sup> </sup>A (H5N1) infection also suggests that cross-protection between<sup> </sup>these two influenza A (H5N1) clades may be limited (Katz J:<sup> </sup>personal communication).
Therefore, further studies are warranted<sup> </sup>to establish the level of cross-neutralizing antibody against<sup> </sup>heterologous influenza A (H5N1) viruses, such as those in clade<sup> </sup>2, that is generated by vaccination with the 1203 vaccine. Such<sup> </sup>cross-neutralization is of great importance, because at the<sup> </sup>current time, the 1203 vaccine is being stockpiled for use in<sup> </sup>the event of an influenza A (H5N1) pandemic. In any case, one<sup> </sup>candidate for a clade 2 vaccine is now available, and others<sup> </sup>are being developed by the WHO Influenza Network.
<sup> </sup>
Additional factors for which data are needed include differences<sup> </sup>in vaccine-induced immunity according to age, sex, immune status,<sup> </sup>and ethnic group. Some of these data could be derived from the<sup> </sup>results of Treanor et al. on further analysis. Age may be particularly<sup> </sup>important; those who have died in past pandemics and from influenza<sup> </sup>A (H5N1) infection are disproportionately children, adolescents,<sup> </sup>and young adults.
<sup> </sup>
Studies of different dose levels of vaccines administered with<sup> </sup>MF59 (a licensed adjuvant in Europe), aluminum hydroxide, or<sup> </sup>other adjuvants are urgently needed. We know from previous work<sup> </sup>that new hemagglutinin proteins (including H5) in people who<sup> </sup>have not been primed are poorly immunogenic.<sup>9</sup><sup>,</sup><sup>10</sup> In recognition<sup> </sup>of this fact, the Department of Health and Human Services and<sup> </sup>the National Institutes of Health have funded studies of more<sup> </sup>than 30 candidate vaccines. Early results from some of these<sup> </sup>trials should be available in the next 6 to 12 months. Previous<sup> </sup>studies of a new influenza A (H5N3) vaccine administered with<sup> </sup>MF59 adjuvant showed that vaccine administered without adjuvant<sup> </sup>was poorly immunogenic but that vaccine administered with MF59<sup> </sup>adjuvant in two doses, each as low as 7.5 ?g, was highly<sup> </sup>immunogenic and resulted in cross-neutralizing antibodies against<sup> </sup>influenza A (H5N1).<sup>11</sup><sup>,</sup><sup>12</sup> Studies of an influenza A (H2N2) vaccine<sup> </sup>administered with alum adjuvant had similar results:
hemagglutination-inhibition<sup> </sup>titers increased significantly at doses as low as 1.9 ?g.<sup>9</sup><sup> </sup>
The immediate development and testing of such antigen-sparing<sup> </sup>vaccines administered with adjuvant are imperative both to improve<sup> </sup>immunogenicity and to increase the number of doses available<sup> </sup>(if lower doses are effective). In addition, live attenuated<sup> </sup>cold-adapted influenza vaccines are safe, are immunogenic, and<sup> </sup>have the relevant advantage of cross-protection against heterologous<sup> </sup>influenza strains ? suggesting a promising avenue to the<sup> </sup>development of pandemic vaccines. A contract for the development<sup> </sup>of such vaccines has been awarded to MedImmune. Other approaches<sup> </sup>to vaccine development involve DNA, adenovirus vectors,<sup>13</sup> and<sup> </sup>cell-culture manufacturing techniques to increase the speed<sup> </sup>and capacity of vaccine production. These approaches are promising,<sup> </sup>particularly since reverse-genetics reassortant vaccine candidates<sup> </sup>can be generated within weeks.<sup>14</sup>
<sup>
</sup><sup> </sup>
Thirty years ago, the United States attempted to respond to<sup> </sup>the threat of pandemic influenza with a vaccine approach. Now,<sup> </sup>armed with a greater understanding of the science, we have the<sup> </sup>capacity and the responsibility to embark on multiple, parallel<sup> </sup>avenues of vaccine development. In addition, we need efficient,<sup> </sup>rapid, high-yield, low-cost manufacturing innovations; the rapid<sup> </sup>generation of candidate vaccines for other, potentially pandemic<sup> </sup>influenza viruses (including emerging clade-2 influenza A [H5N1]<sup> </sup>viruses); and the rapid movement of those vaccines into clinical<sup> </sup>trials. In turn, this effort will require creativity along the<sup> </sup>entire pipeline: in the development and manufacture of candidate<sup> </sup>vaccines; the synchronization among countries of regulatory<sup> </sup>approaches; the resolution of issues concerning liability and<sup> </sup>intellectual property; ensuring the efficiency of clinical trials;<sup> </sup>and the use of methods to stockpile and rapidly deploy these<sup> </sup>vaccines. To do otherwise, with the pandemic clock ticking,<sup> </sup>could prove to be too little, too late.
<sup> </sup>
Dr. Poland reports serving as the chair of a data monitoring<sup> </sup>and safety board for an investigational trial of an influenza<sup> </sup>peptide vaccine being conducted by Merck Research Laboratories.<sup> </sup>No other potential conflict of interest relevant to this article<sup> </sup>was reported.<sup> </sup>
Source Information
From the Mayo Vaccine Research Group, the Program in Translational Immunovirology and Biodefense, and the Department of Internal Medicine, Mayo Clinic College of Medicine, Rochester, Minn.
References
- <!-- null -->
- Congressional Budget Office. A potential influenza pandemic: possible macroeconomic effects and policy issues. December 8, 2005. (Accessed March 10, 2006, at http://www.dhhs.state.nh.us/DHHS/CDC...bo-economy.htm.)<!-- HIGHWIRE ID="354:13:1411:1" --> <!-- /HIGHWIRE --><!-- null -->
- World Health Organization. Avian influenza. (Accessed March 10, 2006, at http://www.who.int/csr/disease/avian_influenza/en/.)<!-- HIGHWIRE ID="354:13:1411:2" --><!-- /HIGHWIRE --><!-- null -->
- Buxton Bridges C, Katz JM, Seto WH, et al. Risk of influenza A (H5N1) infection among health care workers exposed to patients with influenza A (H5N1), Hong Kong. J Infect Dis 2000;181:344-348.<!-- HIGHWIRE ID="354:13:1411:3" --> [CrossRef][ISI][Medline]<!-- /HIGHWIRE --><!-- null -->
- Katz JM, Lim W, Bridges CB, et al. Antibody response in individuals infected with avian influenza A (H5N1) viruses and detection of anti-H5 antibody among household and social contacts. J Infect Dis 1999;180:1763-1770.<!-- HIGHWIRE ID="354:13:1411:4" --> [CrossRef][ISI][Medline]<!-- /HIGHWIRE --><!-- null -->
- Ungchusak K, Auewarakul P, Dowell SF, et al. Probable person-to-person transmission of avian influenza A (H5N1). N Engl J Med 2005;352:333-340.<!-- HIGHWIRE ID="354:13:1411:5" --> <nobr>[Abstract/Full Text]</nobr><!-- /HIGHWIRE --><!-- null -->
- de Jong MD, Thanh TT, Khanh TH, et al. Oseltamivir resistance during treatment of influenza A (H5N1) infection. N Engl J Med 2005;353:2667-2672.<!-- HIGHWIRE ID="354:13:1411:6" --> <nobr>[Abstract/Full Text]</nobr><!-- /HIGHWIRE --><!-- null -->
- Treanor JJ, Campbell JD, Zangwill KM, Rowe T, Wolff M. Safety and immunogenicity of an inactivated subvirion influenza A (H5N1) vaccine. N Engl J Med 2006;354:1343-1351.<!-- HIGHWIRE ID="354:13:1411:7" --> <nobr>[Abstract/Full Text]</nobr><!-- /HIGHWIRE --><!-- null -->
- Shu Y, Yu H, Li D. Lethal avian influenza A (H5N1) infection in a pregnant woman in Anhui Province, China. N Engl J Med 2006;354:1421-1422.<!-- HIGHWIRE ID="354:13:1411:8" --> <nobr>[Full Text]</nobr><!-- /HIGHWIRE --><!-- null -->
- Hehme N, Engelmann H, Kuenzel W, Neumeier E, Saenger R. Immunogenicity of a monovalent, aluminum-adjuvanted influenza whole virus vaccine for pandemic use. Virus Res 2004;103:163-171.<!-- HIGHWIRE ID="354:13:1411:9" --> [CrossRef][ISI][Medline]<!-- /HIGHWIRE --><!-- null -->
- Hehme N, Engelmann H, Kuenzel W, Neumeier E, Saenger R. Pandemic preparedness: lessons learnt from H2N2 and H9N2 candidate vaccines. Med Microbiol Immunol (Berl) 2002;191:203-208.<!-- HIGHWIRE ID="354:13:1411:10" --> [CrossRef][ISI][Medline]<!-- /HIGHWIRE --><!-- null -->
- Nicholson KG, Colegate AE, Podda A, et al. Safety and antigenicity of non-adjuvanted and MF59-adjuvanted influenza A/Duck/Singapore/97 (H5N3) vaccine: a randomised trial of two potential vaccines against H5N1 influenza. Lancet 2001;357:1937-1943.<!-- HIGHWIRE ID="354:13:1411:11" --> [CrossRef][ISI][Medline]<!-- /HIGHWIRE --><!-- null -->
- Stephenson I, Nicholson KG, Colegate A, et al. Boosting immunity to influenza H5N1 with MF59-adjuvanted H5N3 A/Duck/Singapore/97 vaccine in a primed human population. Vaccine 2003;21:1687-1693.<!-- HIGHWIRE ID="354:13:1411:12" --> [CrossRef][ISI][Medline]<!-- /HIGHWIRE --><!-- null -->
- Hoelscher MA, Garg S, Bangari DS, et al. Development of adenoviral-vector-based pandemic influenza vaccine against antigenically distinct human H5N1 strains in mice. Lancet 2006;367:475-481.<!-- HIGHWIRE ID="354:13:1411:13" --> [CrossRef][ISI][Medline]<!-- /HIGHWIRE --><!-- null -->
- Wood JM, Robertson JS. From lethal virus to life-saving vaccine: developing inactivated vaccines for pandemic influenza. Nat Rev Microbiol 2004;2:842-847.<!-- HIGHWIRE ID="354:13:1411:14" --> [CrossRef][ISI][Medline]<!-- /HIGHWIRE -->
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