A Replicating Modified Vaccinia Tiantan Strain Expressing an Avian-Derived Influenza H5N1 Hemagglutinin Induce Broadly Neutralizing Antibodies and Cross-Clade Protective Immunity in Mice

Xiao, Haixia; Liu, Li; Zhu, Qingyu; Tan, Zhiwu; Yu, Wenbo; Tang, Xian; Zhan, Dawei; Du, Yanhua; Wang, Haibo; Liu, Di; Li, Zhixin; Yuen, Kwok-Yung; Ho, David D.; Gao, George F.; Chen, Zhiwei

In this study, we investigated the breath and efficacy of two MVTT based vaccines expressing H5 derived from A/Bar-headed Goose/Qinghai/1/2005 or A/Anhui/1/2005 against homologous and heterologous human H5N1 influenza viruses. HPAIV has been considered a serious public health threat with pandemic potential. If an HPAIV pandemic occurs, it is estimated to claim the lives of over a hundred million people [29, 30] . However, currently licensed vaccines are only effective to a small number of specific strains. The difficulty to predict the pandemic strain together with long and slow process of vaccine production make the efficient development of a vaccine with broad cross-protection an important step for pandemic preparedness. Here, we describe a Modified Vaccinia Virus TianTan strain (MVTT) based H5N1 vaccine, expressing the H5 gene of a goose-derived Qinghai strain A/Bar-headed Goose/Qinghai/1/2005, which can induce broadly neutralizing antibodies and cross-clade protective immunity in mice. Currently, human vaccines have mainly focused on clade 1 strains, among which VNM/1203/04-based vaccines are the most extensively studied. In contrast, both A/BhG/QH/1/05 and A/ Anhui/1/2005, although recommended by the WHO (World Health Organization) as one of the vaccine targeted strains, have yet to be comprehensively studied for their potential as a human vaccine candidate. In the past few years, viruses that are closely related to A/BhG/QH/1/05 have been found to have spread rapidly to Europe, India and Africa via migratory birds, and were likely the causative agents for several human infections throughout Europe and China, including our recently identified A/Xinjiang/ 1/06 [31] [32] [33] . In this study, we characterized the immunogenicity of MVTT HA-QH and MVTT HA-AH , and the antibody response induced against multiple H5N1 influenza viruses representing clades 1 (VN1203 and VN1194), 2.1 (ID05 and ID04), 2.2 (Qinghai and Turkey), 2.3.2 (Chinese duck E319) and 2.3.4 (Fujian and Anhui) (Fig. 1) using the pseudoviral NT assay. We found that Nabs induced by MVTT HA-QH neutralized pseudoviruses from all major clades tested. The MVTT HA-QH -induced serum inhibited viruses from clade 2.2 most optimally, followed by clades 2.1, 2.3.2, 2.3.4 and 1, suggesting that sequence variation in the antigenic sites probably accounted for the reduced NT titers of divergent H5N1 strains (Table 1) . Moreover, we characterized a MVTT HA-QH -induced antibody response against the human A/ Xinjiang/1/06 strain (Fig. 1) , a virus recently identified from a Chinese patient in Xinjiang province that is northwest to the Qinghai province. Despite sequence variations in the antigenic sites, probably due to adaptive infections in humans (Table 1) , the HA gene of A/Xinjiang/1/06 remains closely related to A/BhG/ QH/1/05 (Fig. 1) . It is, therefore, not surprising to see that MVTT HA-QH induced neutralizing and HI antibody responses that reacted equally well against both A/BhG/QH/1/05 and HA XJ /WSN (Table 3 ). In particular, the A86V and A156T mutations in A/Xinjiang/1/06 did not seem to confer resistance to the NT and HI antibodies induced by MVTT HA-QH (Table 1) . Furthermore, we determined the cross-reactivity of MVTT HA-QHinduced NT and HI antibodies against another human virus, the clade 1 A/VN/1194/04. Interestingly, although ten major antigenic mutations were identified between A/BhG/QH/1/05 and A/VN/1194/04 (Table 1) , which accounted for titer drops in HI (,8-fold) an