# US 11,028,408 B2 · Recombinant influenza viruses and constructs and uses thereof > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 11,028,408 B2 - Document ID: US11028408B2 - Status: Granted - Kind: B2 - Application number: 13/903,891 - Filed: 2013-05-28 - Published: 2014-06-12 - Granted: 2021-06-08 - Earliest claimed: 2012-05-25 - Assignees: University of Maryland, College Park - Applicants: University of Maryland, College Park - Inventors: Daniel R. Perez; Hongjun Chen; Yibin Cai; Lindomar Jose Pena; Matthew Angel - Web page: https://patentgrep.com/patent/US11028408B2 ## CPC - C12N 15/86: Viral vectors - A61K 39/12: Viral antigens - A61K 39/145: Orthomyxoviridae, e.g. influenza virus - C12N 7/00: Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof (preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00) - A61K 2039/525: {Virus} - A61K 2039/543: {intranasal} - A61K 2039/70: {Multivalent vaccine} - C12N 2760/16122: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes - C12N 2760/16134: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein - C12N 2760/16143: viral genome or elements thereof as genetic vector - C12N 2760/16151: Methods of production or purification of viral material - C12N 2760/16162: by genetic engineering - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) - A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D) ## Abstract Disclosed are compositions and methods comprising one or more recombinant influenza viruses. Recombinant influenza viruses with mutated polymerases and/or rearranged genomes are disclosed. Constructs comprising different influenza nucleic acid sequences are also provided. Methods of inducing protecting immunity with the recombinant influenza viruses are disclosed. Also disclosed are methods of plasmid-free production of influenza virus comprising amplicons comprising one or more of influenza genes. ## Claims ### Claim 1 A nucleic acid construct comprising a rearranged influenza type A viral genome segment 2, wherein the rearranged influenza type A viral genome segment 2 comprises: a) a first influenza type A viral genome segment, wherein the first influenza type A viral genome segment is influenza type A viral genome segment 2 which comprises a nucleic acid sequence that encodes PB1 and b) a portion of a second influenza type A viral genome segment, wherein the portion of the second influenza type A viral genome segment comprises the portion of influenza type A viral genome segment 8 comprising a nucleic acid sequence that encodes the influenza type A viral protein, NS2, wherein the NS2 nucleic acid sequence is removed from RNA segment 8 of the genome, wherein the NS2 nucleic acid sequence is downstream of the nucleic acid sequence that encodes PB1, wherein a cleavage site is present between the nucleic acid sequence that encodes PB1 and the NS2 nucleic acid sequence, wherein the cleavage site is a 2A-like-cis-acting hydrolase element (CHYSEL) site, and wherein the nucleic acid sequence that encodes PB1 and the NS2 nucleic acid sequence are co-translatable. ### Claim 2 (depends on 1) The construct of claim 1, wherein the construct further comprises a second rearranged influenza type A viral genome segment comprising a portion of influenza type A viral genome segment 8 comprising at least a portion of a nucleic acid sequence that encodes NS1 and is operably linked to an exogenous sequence. ### Claim 3 (depends on 1) A method of reducing influenza polymerase activity comprising using the construct of claim 1 to produce the influenza by growing the influenza. ### Claim 4 (depends on 3) The method of claim 3, wherein the rearranged genome comprises an NS2 or M2 nucleic acid sequence operably linked to a PB1 nucleic acid sequence. ### Claim 5 (depends on 1) The nucleic acid construct of claim 1, wherein the CHYSEL site is a foot-and-mouth disease virus (FMDV) 2A cleavage site. ### Claim 6 A recombinant type A influenza virus comprising a rearranged influenza type A viral genome segment 2, wherein the rearranged influenza type A viral genome segment 2 comprises: a) a first influenza type A viral genome segment, wherein the first influenza type A viral genome segment is influenza type A viral genome segment 2 which comprises a nucleic acid sequence that encodes PB1 and b) a portion of a second influenza type A viral genome segment, wherein the portion of the second influenza type A viral genome segment comprises the portion of influenza type A viral genome segment 8 comprising a nucleic acid sequence that encodes the influenza type A viral protein, NS2, wherein the NS2 nucleic acid sequence is removed from RNA segment 8 of the genome, wherein the NS2 nucleic acid sequence is downstream of the nucleic acid sequence that encodes PB1, wherein a cleavage site is present between the nucleic acid sequence that encodes PB1 and the NS2 nucleic acid sequence, wherein the cleavage site is a 2A-like-cis-acting hydrolase element (CHYSEL) site, and wherein the nucleic acid sequence that encodes PB1 and the NS2 nucleic acid sequence are co-translatable. ### Claim 7 (depends on 6) The recombinant influenza virus of claim 6, further comprising a rearranged influenza type A viral genome segment comprising an exogenous sequence operably linked to an NS1 sequence. ### Claim 8 (depends on 6) A method of inducing a protective immune response against influenza comprising administering to a subject an effective amount of a composition comprising the recombinant influenza virus of claim 6, wherein the subject generates a protective immune response against influenza. ### Claim 9 (depends on 8) The method of claim 8, wherein the recombinant influenza virus comprises an exogenous sequence. ### Claim 10 (depends on 9) The method of claim 9, wherein the exogenous sequence comprises additional coding and non coding nucleic acid sequences. ### Claim 11 (depends on 6) A vaccine comprising the recombinant influenza virus of claim 6 and a pharmaceutically acceptable carrier. ### Claim 12 (depends on 6) The recombinant type A influenza virus of claim 6, wherein the CHYSEL site is a foot-and-mouth disease virus (FMDV) 2A cleavage site.