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Functional influenza virus like particles (VLP)

A technology for influenza virus and influenza virus infection, applied in the direction of viruses, viral peptides, antiviral agents, etc., can solve the problems of weak antigenicity, low copy number, and inability to block virus attachment

Active Publication Date: 2008-12-17
NOVAVAX
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, although both the full-length M2 protein and M2-HBcAg VLPs induce detectable antibodies and protection in mice, it is unlikely that future influenza vaccines will be based solely on the M2 protein because of the high number of copies of the M2 protein present per virion Low, weak antigenicity, does not elicit antibodies that bind free influenza virions, and does not block viral attachment to cellular receptors (i.e., neutralizes the virus)

Method used

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  • Functional influenza virus like particles (VLP)
  • Functional influenza virus like particles (VLP)
  • Functional influenza virus like particles (VLP)

Examples

Experimental program
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Embodiment 1

[0200] Materials and methods

[0201] Avian influenza A / Hong Kong / 1073 / 99 (H9N2) virus HA, NA and M1 genes were expressed in Spodoptera frugiperda cells (Sf-9S cell line; ATCCPTA-4047) using the baculovirus bacmid vector expression system. The HA, NA, and M1 genes were synthesized by reverse transcription and polymerase chain reaction (PCR) using RNA isolated from avian influenza A / Hong Kong / 1073 / 99 (H9N2) virus ( figure 1 , 2 and 3). Reverse transcription and PCR used oligonucleotide primers specific for the HA, NA and M1 genes of the avian influenza A / Hong Kong / 1073 / 99 (H9N2) virus (Table 1). The cDNA copies of these genes were first cloned into the bacterial subcloning vector pCR2.1TOPO. From the resulting three pCR2.1TOPO-based plasmids, the HA, NA and M1 genes were inserted downstream of the AcMNPV polyhedrin promoter in the baculovirus transfer vector pFastBac1 (InVitrogen), resulting in three pFastBac1-based plasmids: pHA, pNA and pM1, which express these influenza vi...

Embodiment 2

[0220] RT-PCR Cloning of Avian Influenza A / Hong Kong / 1073 / 99 Virus Gene

[0221] It is an object of the present invention to provide synthetic nucleic acid sequences capable of directing the production of recombinant influenza virus proteins. Such synthetic nucleic acid sequences were obtained using reverse transcription and polymerase chain reaction (PCR) methods using native viral genomic RNA isolated from influenza virus. For the purposes of this application, nucleic acid sequence refers to RNA, DNA, cDNA or any synthetic variant thereof which encodes the protein in question.

[0222]Avian Influenza A / Hong Kong / 1073 / 99 (H9N2) virus was provided by Dr. K. Subbarao (Centers for Disease Control, Atlanta, Ga., USA). Under CDC's level 3 biosafety (B SL3) precautionary conditions, viral genomic RNA was isolated by acid phenol RNA extraction using Trizol LS reagent (Invitrogen, Carlsbad, Calif. USA). cDNA molecules of these viral RNAs were obtained by reverse transcription using...

Embodiment 3

[0224] RT-PCR Cloning of Human Influenza A / Sydney / 5 / 94(H3N2) Virus Gene

[0225] Influenza A / Sydney / 5 / 94 (H3N2) virus was provided by Dr. M. Massare (Novavax, Inc., Rockville, Md.). Viral genomic RNA was isolated by acid phenol RNA extraction using Trizol LS reagent (Invitrogen) at Novavax.Inc under BSL2 containment conditions. cDNA molecules of these viral RNAs were obtained by reverse transcription and PCR using oligonucleotide primers specific for the HA, NA, M1, M2 and NP proteins (Table 1). The PCR fragment was cloned between the EcoRI sites in the bacterial subcloning vector pCR2.1TOPO to obtain five recombinant plasmids containing HA, NA, M1, M2 and NP cDNA clones.

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Abstract

The present invention discloses and claims virus like particles (VLPs) that express and / or contains seasonal influenza virus proteins, avian influenza virus proteins and / or influenza virus proteins from viruses with pandemic potential. The invention includes vector constructs comprising said proteins, cells comprising said constructs, formulations and vaccines comprising VLPs of the inventions. The invention also includes methods of making and administrating VLPs to vertebrates, including methods of inducing substantial immunity to either seasonal and avian influenza, or at least one symptom thereof.

Description

[0001] This application claims priority to: Provisional Application 60 / 727,513 filed October 18, 2005; Provisional Application 60 / 780,847 filed May 10, 2006; Provisional Application 60 / 800,006 filed May 15, 2006 ; Provisional Application 60 / 831,196, filed July 17, 2006; Provisional Application 60 / 832,116, filed July 21, 2006; and Provisional Application 60 / 845,495, filed September 19, 2006; All content is for all purposes. Background of the invention [0002] Influenza viruses are members of the family Orthomyxoviridae (for review see Murphy and Webster, 1996). There are three subtypes of influenza viruses, called A, B, and C. Influenza virions contain a segmented negative-sense RNA genome. Influenza virions include the following proteins: hemagglutinin (HA), neuraminidase (NA), matrix (M1), proton ion channel protein (M2), nucleoprotein (NP), polymerase basic protein 1 (PB1 ), polymerase basic protein 2 (PB2), polymerase acidic protein (PA), and nonstructural protein 2 (NS...

Claims

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Application Information

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IPC IPC(8): A61K39/00
CPCA61K39/145A61K2039/5258C12N7/00C12N2760/16123C12N2760/16134A61K2039/55505A61K2039/55555A61K39/12A61P31/14A61P31/16A61P37/04A61P43/00C07K14/11
Inventor 盖尔·史密斯里克·布赖特彼得·普什科张金友库塔布·马穆德
Owner NOVAVAX
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