Nucleoprotein (NP) – Specific Antibodies for the Prevention and Treatment of Influenza
- Broad cross-reactive protection against multiple influenza viruses
- For best effect, these therapies should be used in combination with standard vaccines against HA and NA
The 2009 Influenza season clearly demonstrated the need and opportunity for addressing antigenic shift in influenza. Zoonotic influenza A viruses (H1N1 "swine") often express entirely new and unpredictable HA and NA proteins (antigenic shift), to which humans have little or no immunity.
Presented here is a novel mechanism of prevention and treatment of viral infection (e.g.) influenza by using passive immunization with antibodies ("Abs") to one or more conserved viral proteins, including but not limited to conserved influenza viral proteins.
Unlike the external glycoproteins of influenza, internal viral proteins, such as nucleoprotein (NP), are highly conserved between human serotypes and even between human and animal viruses. In fact,there is 95% amino acid identity between the NP in the current H1N1 "swine" influenza virus and the NP in the previously circulating H3N2 human viruses. Thus, it is possible that vaccination with conserved influenza proteins, like NP, might provide broadly cross-reactive protection against multiple influenza viruses.
We tested and observed the following:
- Mice immunized with rNP are resistant to challenge with virulent influenza viruses and exhibit more rapid T cell responses, accelerated viral clearance, reduced morbidity and reduced mortality upon challenge with a normally lethal dose of influenza.
- Protection against challenge infection could be transferred with immune serum, suggesting that antibody was important. Consistent with this conclusion, the transfer of immune serum without antibody resulted in no protection and the immunization of antibody-deficient animals resulted in no protection.
- Mechanistically, the protection provided by transferred antibody requires FcR.
- Protection against challenge infection can be conferred by transferred anti-NP monoclonal antibodies (mAbs).
- Given that essentially all adults have been infected with influenza A virus at some point in their lives, it is not surprising that adult humans have pre-existing circulating antibody titers against NP. However, these antibody titers are low relative to those found in vaccinated mice.
- Anti-NP antibody titers in humans are not boosted in response to TIV vaccine, even though the preparations of TIV that we tested do contain some NP protein.
- Anti-NP antibody secreting cells can be found in human PBL acutely after infection and anti-NP antibody titers in humans are boosted after natural influenza infection, but still not to the levels found after vaccination with rNP in mice.
- Anti-NP antibody titers are also observed in mice after influenza infection, but the titers are lower than in mice immunized with rNP, suggesting that subsequent immunization can boost anti-NP titers even in mice (or humans) previously exposed to influenza virus.
- In fact, we find that anti-NP titers in mice previously infected with influenza can be boosted more than 10-fold by subsequent vaccination with rNP. We also find that in previously infected mice, vaccination with rNP can boost existing immunity, accelerate viral clearance and reduce morbidity and mortality.
Together, these data demonstrate that antibodies against influenza NP are protective against influenza by reducing viral titers, accelerating viral clearance,reducing morbidity and preventing mortality. Monoclonal anti-NP antibodies can be used therapeutically and vaccines that elicit anti-NP antibodies can be used prophylactically, even in individuals who already have pre-existing antibody titers against NP. For best effect these therapies should be used in combination with standard vaccines against HA and NA.
Discoveries at Trudeau Institute have resulted in a novel mechanism of prevention and treatment of viral infection (e.g.) influenza by using passive immunization with antibodies ("Abs") to one or more conserved viral proteins, including but not limited to conserved influenza viral proteins.
Current influenza vaccines elicit Abs to HA and NA envelope proteins. Due to antigenic drift, these vaccines must be reformulated annually to include the envelope proteins predicted to dominate in the following season. By contrast, vaccines that elicit immunity to conserved, often internal viral proteins, such as nucleoprotein ("NP"), provide some protection from multiple subtypes of influenza virus. Trudeau researchers have conducted studies where mice were vaccinated with influenza NP. This approach induced high titers of NP-specific serum Ab, but only poorly detectable NP-specific T cell responses. Nevertheless, this immunization significantly reduced morbidity and viral titers after influenza challenge. Importantly, Ab-deficient mice were not protected by this vaccination strategy. Further, rNP-immune serum could transfer protection to naïve hosts in an Ab-dependent manner. Therefore, Ab to conserved, internal viral proteins, such as NP, provides an unexpected, yet important mechanism of protection against influenza. The results suggest that vaccines designed to elicit optimal heterosubtypic immunity to influenza should promote both Ab and T cell responses to conserved internal proteins.
The Trudeau mouse study results challenge the existing paradigm that T cell responses to conserved epitopes in internal proteins are the exclusive effectors of cross-reactive immunity to heterosubtypic strains of influenza virus and strongly suggest that Abs to these proteins are also an important component of the protective mechanism.
The data developed clearly demonstrate that anti-NP Abs are essential for rNP-derived protection and rNP-immune serum can convey this protection to naïve recipients. Importantly, these results show that a humoral immune response to a single, conserved protein of influenza virus makes a significant contribution to protection.
USPTO Pub. No.: US 2010/0016995 A1 - Immunotherapy to Treat or Prevent Viral Infection (US and PCT Filing Date: September 8, 2008)
Carragher, Damian M., Kaminski, Denis A., Moquin, Amy, Hartson, Louise and Randall, Troy D.: A Novel Role for Non-Neutralizing Antibodies Against Nucleoprotein in Facilitating Resistance to Influenza Virus. The Journal of Immunology, 2008, 181: 4168-4176
Health Research, Inc. on behalf of Trudeau Institute is seeking partners to assist in the further development of this novel discovery into therapeutic products. Partnership opportunities exist in the form of licensing and/or sponsored research with intellectual property rights.
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