Nucleic acid molecules encoding ferritin-hemagglutinin fusion proteins

Patent metadata

Field	Value
Publication number	US-10137190-B2
Application number	US-201615244321-A
Country	US
Kind code	B2
Filing date	Aug 23, 2016
Priority date	Sep 23, 2011
Publication date	Nov 27, 2018
Grant date	Nov 27, 2018

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.

First claim

What is claimed is:

1. A nucleic acid molecule comprising a nucleotide sequence encoding a fusion protein comprising a monomeric ferritin subunit protein joined to an influenza hemagglutinin protein, wherein the monomeric ferritin subunit comprises a domain that allows the fusion protein to self-assemble into nanoparticles.

2. The nucleic acid molecule of claim 1 , wherein the monomeric ferritin subunit protein is selected from the group consisting of a bacterial ferritin, a plant ferritin, an algal ferritin, an insect ferritin, a fungal ferritin and a mammalian ferritin.

3. The nucleic acid molecule of claim 1 , wherein the monomeric subunit is a monomeric subunit of a Helicobacter pylori ferritin protein.

4. The nucleic acid molecule of claim 1 , wherein the monomeric ferritin subunit protein
comprises a region corresponding to amino acids 5-167 of SEQ ID NO:2.

5. The nucleic acid molecule of claim 1 , wherein the monomeric subunit comprises an amino acid sequence selected from the group consisting of SEQ ID NO:2 and SEQ ID NO:5.

6. The nucleic acid molecule of claim 1 , wherein the hemagglutinin protein comprises a region selected from the group consisting of:
a) a region comprising an amino acid sequence comprising at least 25 amino acids from a hemagglutinin protein from an influenza virus selected from the group consisting of A/New Caledonia/20/1999 (1999 NC, H1), A/California/04/2009 (2009 CA, H1), A/Singapore/1/1957 (1957 Sing, H2), A/Hong Kong/1/1968 (1968 HK, H3), A/Brisbane/10/2007 (2007 Bris, H3), A/Indonesia/05/2005 (2005 Indo, H5), B/Florida/4/2006 (2006 Flo, B), A/Perth/16/2009 (2009 Per, H3), A/Brisbane/59/2007 (2007 Bris, H1), and, B/Brisbane/60/2008 (2008 Bris, B);
b) a region corresponding to amino acids 1-519 of SEQ ID NO:8;
c) a region comprising amino acids 1-519 of SEQ ID NO:8; and
d) a region comprising an amino acid sequence at least about 80% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:8, SEQ ID NO:11, SEQ ID NO:14, SEQ ID NO:17, SEQ ID NO:20, SEQ ID NO:23, SEQ ID NO:26, SEQ ID NO:29, SEQ ID NO:32, SEQ ID NO:35, and SEQ ID NO:38.

7. The nucleic acid molecule of claim 1 , wherein the hemagglutinin protein is capable of eliciting an immune response to a protein comprising an amino acid sequence selected from the group consisting of SEQ ID NO:8, SEQ ID NO:11, SEQ ID NO:14, SEQ ID NO:17, SEQ ID NO:20, SEQ ID NO:23, SEQ ID NO:26, SEQ ID NO:29, SEQ ID NO:32, SEQ ID NO:35, and SEQ ID NO:38.

8. The nucleic acid molecule of claim 1 , wherein the fusion protein comprises an amino acid sequence selected from the group consisting of SEQ ID NO:41, SEQ ID NO:44, SEQ ID NO:47, SEQ ID NO:50, SEQ ID NO:53, SEQ ID NO:56, SEQ ID NO:59, SEQ ID NO:62, SEQ ID NO:65, and SEQ ID NO:68.

9. The nucleic acid molecule of claim 1 , wherein the influenza hemagglutinin protein is selected from the group consisting of:
a) an influenza hemagglutinin protein comprising a region selected from the group consisting of a region capable of allowing trimerization of the hemagglutinin protein, a stem region, an ectodomain, and a region comprising the amino acid sequence from the amino acid residue immediately distal to the last amino acid of the second helical coiled coil to the amino acid residue proximal to the first amino acid of the transmembrane domain;
b) an influenza hemagglutinin protein comprising the stem region from an influenza virus selected from the group consisting of A/New Caledonia/20/1999 (1999 NC, H1), A/California/04/2009 (2009 CA, H1), A/Singapore/1/1957 (1957 Sing, H2), A/Hong Kong/1/1968 (1968 HK, H3), A/Brisbane/10/2007 (2007 Bris, H3), A/Indonesia/05/2005 (2005 Indo, H5), B/Florida/4/2006 (2006 Flo, B), A/Perth/16/2009 (2009 Per, H3), A/Brisbane/59/2007 (2007 Bris, H1), and, B/Brisbane/60/2008 (2008 Bris, B);
c) an influenza hemagglutinin protein comprising a region corresponding to amino acids 1-519 of SEQ ID NO:8;
d) an influenza hemagglutinin protein comprising an amino acid sequence at least about 80% identical to SEQ ID NO:71, SEQ ID NO:74, SEQ ID NO:77, SEQ ID NO:80, SEQ ID NO:83, SEQ ID NO:86, SEQ ID NO:89, SEQ ID NO:92, SEQ ID NO:95, or SEQ ID NO:98; and,
e) an influenza hemagglutinin protein comprising an amino acid sequence selected from the group consisting of amino acids 1-519 of SEQ ID NO:8 and SEQ ID NO:11.

10. The nucleic acid molecule of claim 1 , wherein the fusion protein comprises a sequence selected from the group consisting of SEQ ID NO:101, SEQ ID NO:104 SEQ ID NO:107 SEQ ID NO:110 SEQ ID NO:113 SEQ ID NO:116 SEQ ID NO:119 SEQ ID NO:122 SEQ ID NO:125 and SEQ ID NO:128.

11. The nucleic acid molecule of claim 1 , wherein the nucleotide sequence is functionally linked to a promoter.

12. The nucleic acid molecule of claim 1 , wherein the portion of the nucleotide sequence encoding the influenza hemagglutinin protein comprises a sequence at least 80% identical to a sequence selected from the group consisting of SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:16, SEQ ID NO:19, SEQ ID NO:22, SEQ ID NO:25, SEQ ID NO:28, SEQ ID NO:31, SEQ ID NO:34, SEQ ID NO:37, SEQ ID NO:70, SEQ ID NO:73, SEQ ID NO:76, SEQ ID NO:79, SEQ ID NO:82, SEQ ID NO:85, SEQ ID NO:88, SEQ ID NO:91, SEQ ID NO:94, and SEQ ID NO:97.

13. The nucleic acid molecule of claim 1 , wherein the nucleotide sequence comprises a sequence selected from the group consisting of SEQ ID NO:40, SEQ ID NO:43, SEQ ID NO:46, SEQ ID NO:49, SEQ ID NO:52, SEQ ID NO:55, SEQ ID NO:58, SEQ ID NO:61, SEQ ID NO:64, SEQ ID NO:67, SEQ ID NO:100, SEQ ID NO:103, SEQ ID NO:106, SEQ ID NO:109, SEQ ID NO:112, SEQ ID NO:115, SEQ ID NO:118, SEQ ID NO:121, SEQ ID NO:124, and SEQ ID NO:127.

14. A method of producing a nanoparticle, comprising:
a) introducing into a cell a nucleic acid molecule comprising a nucleotide sequence encoding a fusion protein, wherein the fusion protein comprises a monomeric ferritin subunit joined to an influenza hemagglutinin protein, and wherein the monomeric ferritin subunit protein comprises a domain that allows the fusion protein to self-assemble into nanoparticles; and, b) purifying the nanoparticles from the cell.

15. The method of claim 14 , wherein the monomeric ferritin subunit protein is selected from the group consisting of a bacterial ferritin, a plant ferritin, an algal ferritin, an insect ferritin, a fungal ferritin and a mammalian ferritin.

16. The method of claim 14 , wherein the monomeric subunit is a monomeric subunit of a Helicobacter pylori ferritin protein.

17. A method to produce a vaccine against influenza virus, the method comprising:
a) introducing into a cell a nucleic acid molecule comprising a nucleotide sequence encoding a fusion protein, wherein the fusion protein comprises a monomeric ferritin subunit joined to an influenza hemagglutinin protein, and wherein the monomeric ferritin subunit protein comprises a domain that allows the fusion protein to self-assemble into nanoparticles; and,
b) purifying the nanoparticles from the cell to produce the vaccine.

Assignees

• Us Health

Inventors

• Nabel Gary J
• Kanekiyo Masaru
• Wei Chih-Jen
• Mctamney Patrick M
• Yassine Hadi M
• Boyington Jeffrey C

Classifications

• A61P37/04

  Immunostimulants · CPC title

• C12N2760/16134

  Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein · CPC title

• C07K14/005

  from viruses · CPC title

• C07K2319/40

  containing a tag for immunodetection, or an epitope for immunisation · CPC title

• A61K2039/70

  Multivalent vaccine · CPC title