Viral & host immunomodulators in improved Fowlpox virus recombinant vector vaccines for use in poultry against highly pathogenic Avian Influenza H5N1

直到1997年，禽流感病毒，甚至是在鸡和火鸡中造成高死亡率的高致病性毒株（HPAI），被认为对人类构成的直接威胁很小，需要首先将遗传信息（或重组）与已经适应人类的流感病毒混合，这被认为最有可能发生在猪身上。当18名在香港与家禽直接接触的人直接感染了一种病毒，导致6人死亡时，这种评估发生了变化。该病毒携带独特的表面标记H5和N1。此后，H5N1病毒在东南亚广泛传播，到达欧洲和非洲，并继续罕见地感染人类（约200例，死亡率持续高至51%）以及动物园动物和猫。显然，它可以通过感染候鸟而广泛传播，并继续进化和多样化，有可能通过突变或重组直接发展成人类大流行毒株。正在采取措施加强对可能出现的人类大流行病毒（可能保留也可能不保留H5和N1标记）的保护，但人们认识到，控制家禽和鸟类中的病毒对于减少出现这种大流行毒株的风险非常重要。在高致病性禽流感集中爆发的情况下，英国和欧盟的标准做法是控制和屠宰。然而，面对20世纪90年代中期在墨西哥广泛传播的高致病性禽流感（H5N2），疫苗接种被引入并部分成功地根除了高致病性禽流感（HPAI），但未能根除低致病性毒株（LPAI）。这一运动是一个里程碑，因为除了使用基于与流行毒株相同或“同源”类型的灭活流感病毒的传统疫苗外，还首次将编码流感病毒H5部分的转基因禽痘病毒用作所谓的“重组疫苗”。面对目前的H5N1流行，一些国家已经实施了疫苗接种规划。在中国，已经使用了三种类型的疫苗：(i)传统的、不匹配的或“异源的”（H5N2）灭活流感疫苗，（ii） H5N1的灭活低致病性禽流感衍生物，通过基因改造改造成更安全的骨架，以及（iii）表达H5和N1的重组禽痘病毒。在法国，由于重组禽痘病毒和转基因H5N1病毒尚未获得许可，因此使用了一种传统的异源灭活疫苗（在墨西哥使用H5N2作为同源疫苗）。鉴于H5N1毒株的进化速度，在疫苗中使用较旧的H5毒株，特别是在墨西哥使用的H5N2毒株，引起了一些关注。目前的疫苗保护接种过疫苗的禽类，但并不总能防止它们受到感染，四分之一的受感染禽类成为携带者。由于病毒的进化，疫苗变得不那么有效，人们担心疫苗无法阻止鸟与鸟之间的传播，这将导致病毒变异增加的恶性循环。传统的和重组的鸡痘病毒疫苗可诱导良好的保护性抗体反应，但在诱导“细胞反应”方面不太好，这种“细胞反应”是使机体清除受感染细胞所必需的。在小鼠身上的工作表明，预期更好的细胞反应不是针对H5和N1，而是来自流感病毒的内部和非结构蛋白，因此我们将基于这些蛋白以及H5和N1制造重组鸡痘病毒。我们可能还需要调整抗体和细胞反应之间的平衡。这可以通过将鸡免疫调节剂加入重组鸡痘病毒疫苗中来实现，我们将在实验情况下进行尝试。然而，我们认为最好通过改变鸡痘病毒自身的一些免疫调节因子来实现这种调整；我们已经证明这种方法可以对抗一种不同的家禽病毒病。

Until 1997, avian influenza viruses, even the highly pathogenic strains (HPAI) that cause high mortality in chickens and turkeys, were thought to pose little direct threat to humans, needing first to mix genetic information (or recombine), thought most likely to occur in pigs, with influenza viruses already adapted to humans. This assessment changed when 18 people in direct contact with poultry in Hong Kong were directly infected with a virus, killing six. The virus carried the distinctive surface markers H5 and N1. The H5N1 virus has since spread widely throughout SE Asia, reaching Europe and Africa, and continues to infect humans infrequently (about 200 cases with a continued high mortality rate of 51%) as well as zoo animals and cats. It can clearly spread widely by infection of migratory birds and continues to evolve and diversify, threatening to develop into a human pandemic strain, either directly by mutation or by recombination. Measures are being taken to improve protection against emergence of a possible human pandemic virus (which may or may not retain the H5 and N1 markers) but it is recognised that control of the virus in poultry and birds is important to reducing the risk of the emergence of such a pandemic strain. Standard practise in the UK and EU in the event of focal outbreaks of HPAI is containment and slaughter. However, in the face of widespread HPAI (H5N2) in Mexico in the mid-1990's, vaccination was introduced and was partially successful, in so far as it eradicated HPAI, but it failed to eradicate low pathogenicity strains (LPAI). This campaign was a landmark in that as well as using conventional vaccines based on killed influenza viruses of the same or 'homologous' type as the epidemic strain, a genetically modified Fowlpox virus coding for the H5 part of the influenza virus was used for the first time as a so called 'recombinant vaccine'. In the face of the current H5N1 epidemic, several countries have introduced vaccination programmes. In China, three types of vaccine have been used: (i) conventional, mismatched or 'heterologous' (H5N2) killed influenza vaccine, (ii) a killed, LPAI derivative of H5N1, engineered into a safer backbone by genetic modification, and (iii) a recombinant Fowlpox virus expressing both H5 and N1. In France, as the recombinant Fowlpox virus and the genetically modified H5N1 have not been licensed, a conventional, heterologous, killed vaccine (the H5N2 used as a homologous vaccine in Mexico) has been used. With the speed of evolution of the H5N1 strain, the use of older versions of H5 in vaccines, especially that from the H5N2 strain used in Mexico, causes some concern. Current vaccines protect vaccinated birds but do not always prevent them become infected and a quarter of those infected birds become carriers. As vaccines becomes less effective due to evolution of the virus, there are fears that the vaccine's inability to stop bird-to-bird transmission will contribute to a vicious circle of increased virus variation. The conventional and recombinant Fowlpox virus vaccines induce good, protective antibody responses but are not so good at inducing the 'cellular responses' necessary to allow the body to clear itself of infected cells. Work in mice has shown that better cellular responses would be expected not against H5 and N1 but from the internal and non-structural proteins of influenza virus so we will make recombinant Fowlpox viruses based on these proteins as well as on H5 and N1. We may also need to adjust the balance of antibody versus cellular responses. This could be done by incorporating chicken immune regulators into the recombinant Fowlpox virus vaccines, and we will try this in the experimental situation. However we think it would be preferable to achieve this adjustment by changing some of the Fowlpox virus's own immune regulators; we already have demonstrated this approach can work against a different virus disease of poultry.