Nucleic acid molecules encoding ferritin-hemagglutinin fusion proteins
Inventors
Nabel, Gary J. • Kanekiyo, Masaru • Wei, Chih-jen • MCTAMNEY, Patrick M. • YASSINE, Hadi M. • BOYINGTON, Jeffrey C.
Assignees
US Department of Health and Human Services
Publication Number
US-10137190-B2
Publication Date
2018-11-27
Expiration Date
2032-09-24
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Abstract
Novel vaccines are provided that elicit broadly neutralizing anti-influenza antibodies. Some vaccines comprise nanoparticles that display hemagglutinin trimers from influenza virus on their surface. The nanoparticles comprise fusion proteins comprising a monomeric subunit of ferritin joined to at least a portion of an influenza hemagglutinin protein. Some portions comprise the ectodomain while some portions are limited to the stem region. The fusion proteins self-assemble to form the hemagglutinin-displaying nanoparticles. Some vaccines comprise only the stem region of an influenza hemagglutinin protein joined to a trimerization domain. Such vaccines can be used to vaccinate an individual against infection by heterologous influenza viruses and influenza virus that are antigenically divergent from the virus from which the nanoparticle hemagglutinin protein was obtained. Also provided are fusion proteins and nucleic acid molecules encoding such proteins.
Core Innovation
The invention provides novel influenza vaccines comprising nanoparticles that display hemagglutinin trimers from influenza virus on their surface. These nanoparticles are composed of fusion proteins that include a monomeric subunit of ferritin joined to at least a portion of an influenza hemagglutinin protein, such as the ectodomain or the stem region. The fusion proteins self-assemble to form the hemagglutinin-displaying nanoparticles, which can effectively elicit broadly neutralizing influenza antibodies.
The problem addressed by the invention is the limitation of current influenza vaccines, which are generally strain-specific, produced via time-consuming processes in embryonated eggs, and present manufacturing constraints. These vaccines typically induce neutralizing antibodies targeted at variable antigenic sites in the HA globular head, limiting protection against antigenically divergent strains. There is a need for vaccines that induce broadly neutralizing antibodies with potent and broad protection against heterologous and antigenically divergent influenza viruses.
The invention solves this need by employing ferritin-based nanoparticles that display influenza hemagglutinin proteins in their native trimeric form, using fusion proteins linking ferritin subunits to HA portions. This approach allows for easy manufacturing, potent immunogenicity, and induction of broadly neutralizing antibodies against conserved HA regions such as the stem. The nanoparticles formed have an octahedral cage structure of 24 subunits, which provides a multivalent display enhancing immune responses compared to monovalent forms. The vaccines can comprise monovalent or multivalent nanoparticles incorporating HA proteins from diverse influenza strains thereby conferring broad protection.
Claims Coverage
The patent claims 17 claims including a main independent claim directed to a nucleic acid molecule encoding a fusion protein. The inventive features focus on the structural composition of the fusion protein, its functional capabilities, and methods of production.
Nucleic acid encoding fusion protein with self-assembling ferritin subunit
A nucleic acid molecule encoding a fusion protein comprising a monomeric ferritin subunit protein joined to an influenza hemagglutinin protein, wherein the ferritin subunit comprises a domain enabling the fusion protein to self-assemble into nanoparticles.
Selection of ferritin subunit from diverse ferritin sources
The monomeric ferritin subunit protein can be selected from bacterial, plant, algal, insect, fungal, or mammalian ferritins, including a preferred Helicobacter pylori ferritin monomeric subunit.
Ferritin subunit comprising specific amino acid regions or sequences
The ferritin monomeric subunit can comprise a region corresponding to amino acids 5-167 of SEQ ID NO:2, or amino acid sequences selected from SEQ ID NO:2 and SEQ ID NO:5, including sequences with high sequence identity (80-99%).
Hemagglutinin protein region specificity and sequence identity
The hemagglutinin protein portion can comprise at least 25 amino acids from influenza virus hemagglutinin of various specified strains or regions such as ectodomain, stem region, or sequences corresponding to SEQ ID NOs:8, 11, 14, 17, 20, 23, 26, 29, 32, 35, and 38 with high sequence identity (80-99%).
Immune elicitation capabilities of hemagglutinin protein
The hemagglutinin protein is capable of eliciting an immune response against specified influenza hemagglutinin proteins or sequences, including eliciting neutralizing antibodies against influenza virus.
Fusion protein amino acid sequences
The fusion protein comprises amino acid sequences selected from SEQ ID NO:41, 44, 47, 50, 53, 56, 59, 62, 65, and 68 or from SEQ ID NOs:101, 104, 107, 110, 113, 116, 119, 122, 125, and 128, sequences encoding stem regions or stabilized constructs.
Composition of hemagglutinin protein domains in fusion protein
The hemagglutinin protein may include domains capable of trimerization, the stem region, ectodomain, or defined regions from distal to second helical coiled coil to proximal to transmembrane domain, and specific influenza strain sequences as listed in claims.
Functional linkage to promoter and recombinant expression
The nucleic acid molecule can be functionally linked to a promoter and used in recombinant cells or viruses for expression of fusion proteins.
Methods to produce nanoparticles and vaccines
Methods include introducing nucleic acid molecules encoding the fusion protein into cells, allowing expression and self-assembly into nanoparticles, and purifying these for vaccine production.
The claims collectively cover nucleic acid molecules encoding fusion proteins comprising monomeric ferritin subunits joined to influenza hemagglutinin proteins that self-assemble into nanoparticles displaying hemagglutinin trimers. They cover variations in ferritin source, hemagglutinin sequences and domains, functional aspects of the fusion proteins, methods of producing the nanoparticles, and vaccine compositions, emphasizing broad influenza immune response elicitation and nanoparticle formation.
Stated Advantages
The vaccines are easily manufactured with improved potency and elicit broadly neutralizing anti-influenza antibodies.
The multivalent display of hemagglutinin on nanoparticles induces stronger B cell responses than monovalent forms and can elicit T-cell independent antibody responses.
The vaccines provide broad protection against heterologous and antigenically divergent influenza virus strains, including evolving seasonal and pandemic strains.
Ferritin-based nanoparticles mimic physiological trimeric viral spikes with rigid and symmetrical display allowing optimal immune system access to broadly neutralizing epitopes.
Preexisting immunity to ferritin does not diminish hemagglutinin-specific antibody responses, minimizing concerns of autoreactivity.
Documented Applications
Vaccines comprising HA-ferritin fusion protein nanoparticles for immunizing individuals against influenza virus infection.
Use of nanoparticles displaying hemagglutinin trimers to elicit immune responses, including broadly neutralizing antibodies to diverse influenza strains.
Methods of vaccinating subjects with monovalent or multivalent influenza vaccines comprising ferritin-based nanoparticles.
Boosting immunity with heterologous vaccine compositions comprising stabilized stem region HA-ferritin nanoparticles.
Production methods for vaccines involving recombinant expression of fusion proteins and nanoparticle purification.
Use in veterinary and human medicine to protect against infection by homologous, heterologous, and antigenically divergent influenza viruses.
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