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KR-102964264-B1 - Antigenic respiratory syncytial virus polypeptide

KR102964264B1KR 102964264 B1KR102964264 B1KR 102964264B1KR-102964264-B1

Abstract

The present disclosure relates to an antigenic RSV polypeptide for use in inducing antibodies against respiratory syncytial virus (RSV). An antigenic polypeptide comprising an RSV polypeptide and a ferritin protein is also disclosed.

Inventors

  • 나벨, 그레이 제이.
  • 웨이, 취-젠
  • 스완슨, 커트
  • 달, 프레딥
  • 다라니프라가다, 램
  • 베지브, 매그너스

Assignees

  • 사노피

Dates

Publication Date
20260513
Application Date
20190402
Priority Date
20180403

Claims (20)

  1. It is an antigenic RSV polypeptide containing the respiratory syncytial virus (RSV) F polypeptide, and Here, an N-linked glycan attaches to the asparagine of the RSV polypeptide, blocking the epitope of the RSV polypeptide shared between pre-fusion RSV F and post-fusion RSV F, and RSV F polypeptide comprises the amino acid sequence of amino acids 1 to 478 of SEQ ID NO. 17, and Asparagine is that which corresponds to positions 288, 308, or 467 of sequence number 17, Antigenic RSV polypeptide.
  2. It is an antigenic RSV polypeptide containing RSV F polypeptide, and Here, an N-linked glycan attaches to the asparagine of the RSV polypeptide, blocking the epitope of the RSV polypeptide shared between pre-fusion RSV F and post-fusion RSV F, and RSV F polypeptide comprises the sequence of amino acids 1 to 478 of SEQ ID NO. 23, and Asparagine is that which corresponds to positions 288, 308, or 467 of sequence number 23, Antigenic RSV polypeptide.
  3. In paragraph 1 or 2, Antigenic RSV polypeptide containing pre-fusion RSV F.
  4. In paragraph 1 or 2, Antigenic RSV polypeptide recognized by a pre-fusion RSV F-specific antibody selected from D25 or AM14.
  5. In paragraph 3, Antigenic RSV polypeptide, wherein pre-fusion RSV F contains an epitope not found in post-fusion RSV F, which is a site 0 epitope, wherein the site 0 epitope contains amino acid residues corresponding to amino acids 62–69 and 196–209 of SEQ ID NO. 26.
  6. In paragraph 1 or 2, Antigenic RSV polypeptide containing RSV F after fusion.
  7. In paragraph 1 or 2, Antigenic RSV polypeptide containing ferritin protein.
  8. In Paragraph 7, Ferritin is an antigenic RSV polypeptide containing a mutation that replaces a surface-exposed amino acid with cysteine.
  9. In paragraph 1 or 2, The antigenic RSV polypeptide contains ferritin protein, and Ferritin contains mutations that replace surface-exposed amino acids with cysteine, and Ferritin comprises one or more of the E12C, S26C, S72C, A75C, K79C, S100C, and S111C mutations of Helicobacter pylori (H. pylori) ferritin , or one or more mutations in non-Helicobacter pylori (H. pylori) ferritin corresponding to the E12C, S26C, S72C, A75C, K79C, S100C, and/or S111C mutations of Helicobacter pylori (H. pylori) ferritin as determined by paired or structural alignment , Antigenic RSV polypeptide.
  10. In paragraph 1 or 2, The antigenic RSV polypeptide contains ferritin protein, and The antigenic RSV polypeptide comprises one or more immunostimulatory moietyes linked to ferritin via surface-exposed amino acids, and One or more immunostimulatory moietyes comprise Toll-like receptor (TLR) agonists, pathogen-associated molecular pattern (PAMP) and/or interferon gene stimulator (STING) agonists, Antigenic RSV polypeptide.
  11. In paragraph 10, an antigenic RSV polypeptide in which the surface-exposed amino acid is cysteine generated from a mutation.
  12. In paragraph 1 or 2, The antigenic RSV polypeptide contains ferritin protein, and Ferritin is a mutation that replaces surface-exposed asparagine with a non-asparagine amino acid, Antigenic RSV polypeptide.
  13. An antigenic RSV polypeptide in which, in paragraph 12, asparagine is at position 19 of Helicobacter pylori (H. pylori) ferritin, or at a non- Helicobacter pylori (H. pylori) ferritin position corresponding to position 19 of Helicobacter pylori (H. pylori) ferritin when determined by pairing or structural alignment.
  14. In paragraph 1 or 2, The antigenic RSV polypeptide contains ferritin protein, and Ferritin is an antigenic RSV polypeptide containing a mutation that replaces internal cysteine with serine, alanine, glycine, threonine, or asparagine.
  15. An antigenic RSV polypeptide according to claim 14, wherein the internal cysteine is at position 31 of Helicobacter pylori (H. pylori) ferritin, or at a non-H. pylori (H. pylori) ferritin position corresponding to position 31 of Helicobacter pylori (H. pylori) ferritin when determined by pairing or structural alignment.
  16. In paragraph 1 or 2, The antigenic RSV polypeptide contains ferritin protein, and An antigenic RSV polypeptide comprising an epitope not found in post-fusion RSV F, wherein the site 0 epitope comprises amino acid residues 62–69 and 196–209 of SEQ ID NO. 26.
  17. In paragraph 1 or 2, An antigenic RSV polypeptide in which an amino acid residue of the RSV F polypeptide at a position corresponding to position 328 of sequence number 26 is substituted with asparagine.
  18. In paragraph 1 or 2, An antigenic RSV polypeptide in which an amino acid residue of the RSV F polypeptide at a position corresponding to position 348 of sequence number 26 is substituted with asparagine.
  19. In paragraph 1 or 2, An antigenic RSV polypeptide in which an amino acid residue of the RSV F polypeptide at a position corresponding to position 507 of sequence number 26 is substituted with asparagine.
  20. In paragraph 1 or 2, An antigenic RSV polypeptide in which the amino acid residue of the RSV F polypeptide at the position corresponding to lysine 498 of sequence number 26 is substituted with leucine.

Description

Antigenic respiratory syncytial virus polypeptide This application claims the benefit of U.S. Provisional Application No. 62/652,199 filed on April 3, 2018, the entirety of which is incorporated herein by reference. The present application comprises a list of sequences submitted electronically in ASCII format, the entirety of which is incorporated herein by reference. The said ASCII copy created on March 27, 2019, is named 2019-03-27_01121-0031-00PCT_SL_ST25.txt and has a size of 187,354 bytes. Despite significant success in the field of vaccines, new breakthroughs are needed to protect humans from many life-threatening infectious diseases. Many currently licensed vaccines rely on decade-old technology to produce attenuated or inactivated dead pathogens, which have inherent safety issues and often stimulate only short-term, weak immune responses requiring multiple administrations. While advancements in genetic and biochemical engineering have enabled the development of therapies for challenging disease targets, these applications in the field of vaccines have not been fully realized. Now, recombinant protein technology allows for the design of optimal antigens. Furthermore, nanoparticles have increasingly demonstrated their potential for optimal antigen presentation and targeted drug delivery. Nanoparticles attached to multiple antigens have been shown to possess increased binding affinity due to the polyvalent labeling of their molecular cargoes and the ability to penetrate biological barriers more efficiently due to their nanoscale size. Helicobacter pylori ( H. pylori ) ferritin nanoparticles fused to influenza virus hemagglutinin (HA) protein improved antigen stability and increased immunogenicity in a mouse influenza model (see reference [Kanekiyo et al., Nature 499:102~106 (2013)]). This fusion protein self-assembled into octahedral-symmetric nanoparticles and provided eight trimeric HA spikes to deliver a potent immune response in various preclinical models when used with an adjuvant. Respiratory syncytial virus (RSV) is a major cause of severe respiratory disease in infants and the elderly. Despite decades of research, the need for an unmet vaccine remains. Although the need for a vaccine is clear, the development of an RSV vaccine was hampered in the 1960s when clinical trials using formalin-inactivated RSV virus caused more severe disease in infants following RSV infection. See [Hurwitz (2011) Expert Rev Vaccines 10(10): 1415~1433.]. More recently, clinical programs using the post-fusion form of the RSV F antigen have failed to elicit sufficient efficacy in adults. See [Faloon et al. (2017) JID 216:1362~1370]. However, RSV F antigen stabilized in the pre-fusion form can elicit a superior neutralization response compared to the post-fusion antigen that failed in clinical trials. Here, a series of novel polypeptides, nanoparticles, compositions, methods, and uses associated with RSV polypeptides are presented. A novel RSV F polypeptide was generated comprising a polypeptide in which the epitope of the RSV polypeptide shared between pre-fusion RSV F and post-fusion RSV F is blocked by an N-glycan, for example, at a glycosylation site added by a mutation. Additionally, antigenic polypeptides and nanoparticles comprising these novel RSV polypeptides and ferritin were generated. An antigenic ferritin polypeptide comprising RSV G polypeptide was also generated. Furthermore, a self-adjuvant antigenic polypeptide comprising RSV polypeptide and ferritin was developed, wherein an immunostimulatory moiety, such as an adjuvant, was chemically attached directly to the antigenic polypeptide. Direct conjugation of an immunostimulatory moiety to an antigenic polypeptide enables targeted co-delivery of the immunostimulatory moiety and the RSV polypeptide within a single macromolecule, which can significantly reduce the potential for systemic toxicity associated with conventional vaccines that contain the antigen and the immunostimulatory molecule, such as the adjuvant, as separate molecules. The co-delivery of the immunostimulatory moiety together with the RSV polypeptide within a macromolecule and their multi-presentation can also reduce the total dose required to induce protection, thereby lowering manufacturing burden and costs. Figures 1a–1d show exemplary RSV Pre-F-NP polypeptide structures. (Figure 1a) A linear diagram listing residue numbers corresponding to the N-terminus of each segment. The numbering follows sequence number 26. Domains 1–3 are labeled DI, DII, and DIII, respectively, and heptad repeat region A (HRA) and heptad repeat region B (HRB) are also labeled. C-terminal ferritin is labeled (ferritin nanoparticle). The F1 and F2 fragments of the RSV F moiety are labeled under the carton. When the peptide 27 fragment (p27) fusion peptide (FP) and the purine cleavage site (purine site) are deleted and replaced with a flexible linker to form a single-strand F construct, the region between the F1 and F2