Hemagglutinin Stalk- and Neuraminidase-Specific Monoclonal Antibodies Protect against Lethal H10N8 Influenza Virus Infection in Mice

Abstract

Between November 2013 and February 2014, China reported three human cases of H10N8 influenza virus infection in the Jiangxi province, two of which were fatal. Using hybridoma technology, we isolated a panel of H10-and N8-directed monoclonal antibodies (MAbs) and further characterized the binding reactivity of these antibodies (via enzyme-linked immunosorbent assay) to a range of purified virus and recombinant protein substrates. The H10-directed MAbs displayed functional hemagglutination inhibition (HI) and neutralization activity, and the N8-directed antibodies displayed functional neuraminidase inhibition (NI) activity against H10N8. Surprisingly, the HI-reactive H10 antibodies, as well as a previously generated, group 2 hemagglutinin (HA) stalk-reactive antibody, demonstrated NI activity against H10N8 and an H10N7 strain; this phenomenon was absent when virus was treated with detergent, suggesting the anti-HA antibodies inhibited neuraminidase enzymatic activity through steric hindrance. We tested the prophylactic efficacy of one representative H10-reactive, N8-reactive, and group 2 HA stalk-reactive antibody in vivo using a BALB/c challenge model. All three antibodies were protective at a high dose (5 mg/kg). At a low dose (0.5 mg/kg), only the anti-N8 antibody prevented weight loss. Together, these data suggest that antibody targets other than the globular head domain of the HA may be efficacious in preventing influenza virus-induced morbidity and mortality. IMPORTANCEAvian H10N8 and H10N7 viruses have recently crossed the species barrier, causing morbidity and mortality in humans and other mammals. Although these reports are likely isolated incidents, it is possible that more cases may emerge in future winter seasons, similar to H7N9. Furthermore, regular transmission of avian influenza viruses to humans increases the risk of adaptive mutations and reassortment events, which may result in a novel virus with pandemic potential. Currently, no specific therapeutics or vaccines are available against the H10N8 influenza virus subtype. We generated a panel of H10-and N8-reactive MAbs. Although these antibodies may practically be developed into therapeutic agents, characterizing the protective potential of MAbs that have targets other than the HA globular head domain will provide insight into novel antibody-mediated mechanisms of protection and help to better understand correlates of protection for influenza A virus infection. R ecently, avian influenza A viruses of the H10 subtype have been reported to infect seals and humans and have generated concern over their pandemic potential. Three human cases of H10N8 virus have been reported in China so far, two of which were fatal (1-3). Furthermore, an avian H10N7 strain was found to be the etiological agent responsible for the massive die-off harbor seals in the Baltic Sea, an epidemic that killed more than 10% of the local seal population (4-6). The receptor binding profile of H10 viruses is currently debated (7-12), but the subtype has been proven to cause ...

An influenza A virus (H7N9) anti-neuraminidase monoclonal antibody with prophylactic and therapeutic activity in vivo

“…Here we show that the anti-NA mAb 3c10-3 protects mice from lethal challenge with A(H7N9) virus, and reduces morbidity and mortality when used therapeutically in already-infected mice. Although a number of anti-NA mAbs with protective efficacy in vivo have been described recently (Doyle et al, 2013; Jiang et al, 2013; Shoji et al, 2011; Wan et al, 2013; Wohlbold et al, 2016), this is the first report of an anti-N9 mAb that has therapeutic activity.…”

“…However, it is well known that anti-NA antibodies also can confer some protection (Monto and Kendal, 1973; Murphy et al, 1972; Schild, 1969), although the level of NA in split vaccines vary and currently are not standardized. The growing number of reports detailing the protective efficacy of anti-NA mAb in vivo (Doyle et al, 2013; Jiang et al, 2013; Shoji et al, 2011; Wan et al, 2013; Wohlbold et al, 2016), further support a re-emphasis on including anti-NA antibody responses in the analysis of protection against influenza infection and provides additional support for standardizing the amount of NA contained in each vaccine dose.…”

Epitope specificity plays a critical role in regulating antibody-dependent cell-mediated cytotoxicity against influenza A virus

“…Although present in substantially lower quantities than HA-binding antibodies, NAbinding antibodies can inhibit NA activity, which has been shown to confer protection from infection in vivo (28)(29)(30). Certain HAbinding antibodies have been shown to exhibit NA-inhibiting (NAI) activity through steric hindrance of the NA enzymatic site (28). Therefore, we examined whether NA-binding antibodies by themselves could induce ADCC or influence ADCC induced by HA stalk-binding bnAbs.…”

“…In addition to HA, a substantial antibody response is also generated against the second major IAV surface glycoprotein, NA. Although present in substantially lower quantities than HA-binding antibodies, NAbinding antibodies can inhibit NA activity, which has been shown to confer protection from infection in vivo (28)(29)(30). Certain HAbinding antibodies have been shown to exhibit NA-inhibiting (NAI) activity through steric hindrance of the NA enzymatic site (28).…”

An influenza A virus (H7N9) anti-neuraminidase monoclonal antibody protects mice from morbidity without interfering with the development of protective immunity to subsequent homologous challenge

“…Six of these mAbs are directed against various epitopes contained within the HA surface protein while one targets the matrix protein 2 (M2). In addition to the HA and M2, anti-neuraminidase (NA) mAb have also proven to be protective in the mouse model but, to our knowledge, have yet to enter clinical development (Doyle et al, 2013; Jiang et al, 2015; Shoji et al, 2011; Wan et al, 2013; Wohlbold et al, 2016; DiLillo et al, 2016). One potential concern regarding mAb as antivirals include the possibility that their use may drive the evolution of viral escape mutations which could be resistant not only to the mAb but to natural or vaccine-induced immunity.…”