# US 4,870,023 A · Recombinant baculovirus occlusion bodies in vaccines and biological insecticides > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 4,870,023 A - Document ID: US4870023A - Status: Granted - Kind: A - Application number: 07/153,736 - Filed: 1988-02-08 - Granted: 1989-09-26 - Earliest claimed: 1987-03-16 - Assignees: American Biogenetic Sciences, Inc. - Inventors: Malcolm J. Fraser; Elliot D. Rosen; Victoria A. Ploplis - Web page: https://patentgrep.com/patent/US4870023A ## CPC - C12N 15/86: Viral vectors - C07K 14/005: from viruses - C12N 15/62: DNA sequences coding for fusion proteins - A61K 39/00: Medicinal preparations containing antigens or antibodies (materials for immunoassay G01N33/53) - C07K 2319/00: Fusion polypeptide - C12N 2710/14122: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes - C12N 2710/14143: viral genome or elements thereof as genetic vector - C12N 2760/16122: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes - Y02A 50/30: Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change - Y10S 530/82: Proteins from microorganisms - Y10S 530/826: Viruses - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) - C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00) - 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) - Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE - Y10S: TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS ## Abstract The present invention is directed to recombinant baculoviruses which encode fusion polyhedrin proteins capable of forming occlusion bodies containing foreign peptides. The recombinant baculoviruses of the invention are formed by insertion into or replacement of regions of the polyhedrin gene that are not essential for occlusion body formation, with foreign DNA fragments by recombinant DNA techniques. The recombinant occlusion bodies produced in accordance with the present invention have uses in vaccine formulations, immunoassays, immobilized enzyme reactions, as biological insecticides, and as expression vectors. ## Claims ### Claim 1 A recombinant occlusion body comprising repeating subunits in which each subunit comprises a polyhedrin fusion protein comprising a portion of the polyhedrin protein which participates in crystallization, fused to a foreign amino acid sequence. ### Claim 2 (depends on 1) The recombinant occlusion body according to claim 1 in which the foreign amino acid sequence is related to an epitope of a pathogenic microorganism. ### Claim 3 (depends on 2) The recombinant occlusion body according to claim 2 in which the pathogenic microorganism comprises a virus. ### Claim 4 (depends on 3) The recombinant occlusion body according to claim 3 in which the epitope comprises influenza hemagglutinin. ### Claim 5 (depends on 4) The recombinant occlusion body according to claim 4 in which the epitope comprises amino acids 98-106 of influenza hemagglutinin. ### Claim 6 (depends on 3) The recombinant occlusion body according to claim 3 in which the virus comprises Hepatitis A virus. ### Claim 7 (depends on 2) The recombinant occlusion body according to claim 2 in which the foreign amino acid sequence is exposed on the surface of the occlusion body. ### Claim 8 (depends on 1) The recombinant occlusion body according to claim 1 in which the foreign amino acid sequence comprises an antigenic determinant of a foreign protein. ### Claim 9 (depends on 1) The recombinant occlusion body according to claim 1 in which the foreign amino acid sequence replaces all or a portion of the amino terminus of the polyhedrin protein amino acid sequence. ### Claim 10 (depends on 1) The recombinant occlusion body according to claim 1 in which the foreign amino acid sequence replaces all or a portion of a region homologous to the amino acid sequence substantially as depicted in FIG. 1 from amino acid residue number 37 to 49. ### Claim 11 (depends on 5) The recombinant occlusion body according to claim 5 in which the foreign amino acid sequence replaces all or a portion of a region homologous to the amino acid sequence substantially as depicted in FIG. 1 from amino acid residue number 37 to 49. ### Claim 12 (depends on 1) The recombinant occlusion body according to claim 1 in which the foreign amino acid sequence is inserted after amino acid residue number 1 of the Autographa polyhedrin sequence substantially depicted in FIG. 2. ### Claim 13 (depends on 1) The recombinant occlusion body according to claim 1 in which the foreign amino acid sequence replaces amino acid residue number 43 of the Autographa polyhedrin sequence substantially depicted in FIG. 2. ### Claim 14 (depends on 1) The recombinant occlusion body according to claim 1 in which the foreign amino acid sequence replaces amino acid residue number 50 of the Autographa polyhedrin sequence substantially depicted in FIG. 2. ### Claim 15 A polyhedrin fusion protein which is capable of crystallizing with other polyhedrin proteins to form recombinant occlusion bodies, comprising: a portion of the polyhedrin protein which participates in crystallization fused to a foreign amino acid sequence. ### Claim 16 (depends on 15) The recombinant polyhedrin protein of claim 15 in which the foreign amino acid sequence comprises an epitope of a pathogenic microorganism. ### Claim 17 (depends on 16) The recombinant polyhedrin protein of claim 16 in which the pathogenic microorganism comprises a virus. ### Claim 18 (depends on 17) The recombinant polyhedrin protein of claim 17 in which the pathogenic microorganism comprises influenza virus. ### Claim 19 (depends on 18) The recombinant polyhedrin protein of claim 18 in which the foreign amino acid sequence comprises amino acids 98-106 of the influenza hemagglutinin. ### Claim 20 (depends on 17) The recombinant polyhedrin protein of claim 17 in which the pathogenic microorganism comprises Hepatitis A virus. ### Claim 21 (depends on 15) The recombinant polyhedrin protein of claim 15 in which the second amnio acid sequence replaces all or a portion of the amino terminus of the polyhedrin protein. ### Claim 22 (depends on 15) The recombinant polyhedrin protein of claim 15 in which the second amino acid sequence replaces all or a portion of a region homologous to the amino acid sequence substantially as depicted in FIG. 1 from amino acid residue number 37 to 49. ### Claim 23 (depends on 15) The recombinant polyhedrin protein of claim 15 in which the second amino acid sequence comprises an antigenic determinant of a foreign protein. ### Claim 24 (depends on 15) The recombinant polyhedrin protein of claim 15 in which the foreign amino acid sequence is inserted after amino acid residue number 1 of the Autographa polyhedrin sequence substantially as depicted in FIG. 2. ### Claim 25 (depends on 15) The recombinant polyhedrin protein of claim 15 in which the foreign amino acid sequence replaces amino acid residue number 43 of the Autographa polyhedrin sequence substantially as depicted in FIG. 2. ### Claim 26 (depends on 15) The recombinant polyhedrin protein of claim 15 in which the foreign amino acid sequence replaces amino acid residue number 50 of the Autographa polyhedrin sequence substantially as depicted in FIG. 2. ### Claim 27 A recombinant virus which directs the expression of polyhedrin fusion proteins that crystallize to form recombinant occlusion bodies, comprising: (a) a polyhedrin promoter; and (b) a nucleotide sequence encoding a polyhedrin fusion protein comprising (i) a first nucleotide sequence encoding a portion of the polyhedrin structural protein that participates in crystallization and (ii) a second nucleotide sequence encoding a foreign protein, in which the first and second nucleotide sequences are in the same translational reading frame uninterrupted by translation termination signals; and in which the nucleotide sequence encoding the polyhedrin fusion protein is under the control of the polyhedrin promoter so that polyhedrin fusion proteins which crystallize to form recombinant occlusion bodies are produced in a suitable host infected with recombinant virus. ### Claim 28 (depends on 27) The recombinant virus according to claim 27 comprising a baculovirus. ### Claim 29 (depends on 28) The recombinant virus according to claim 28 comprising a nuclear polyhedrosis virus. ### Claim 30 (depends on 29) The recombinant virus according to claim 29 comprising Autographa californica nuclear polyhedrosis virus. ### Claim 31 (depends on 29) The recombinant virus according to claim 29 comprising Heliothis zea nuclear polyhedrosis virus. ### Claim 32 (depends on 28) The recombinant virus according to claim 28 comprising a granulosis virus. ### Claim 33 (depends on 27) The recombinant virus according to claim 27 in which the foreign peptide comprises an epitope of a pathogenic microorganism. ### Claim 34 (depends on 33) The recombinant virus according to claim 33 in which the pathogenic microorganism comprises a virus. ### Claim 35 (depends on 34) The recombinant virus according to claim 34 in which the epitope is related to an epitope of influenza hemagglutinin. ### Claim 36 (depends on 27) The recombinant virus according to claim 27 in which the nucleotide sequence encoding the foreign peptide replaces all or part of the polyhedrin gene that encodes the amino terminus of the polyhedrin protein. ### Claim 37 (depends on 27) The recombinant virus according to claim 27 in which the nucleotide sequence encoding the foreign peptide replaces all or part of a region of the polyhedrin gene that is homologous to the nucleotide sequence substantially as depicted in FIG. 1 from nucleotide number 142 to 180. ### Claim 38 (depends on 27) The recombinant virus according to claim 27 in which the nucleotide sequence encoding the foreign peptide is inserted after amino acid number 1 of the Autographa polyhedrin gene substantially a depicted in FIG. 2. ### Claim 39 (depends on 27) The recombinant virus according to claim 27 in which the nucleotide sequence encoding the foreign peptide replaces the part of the Autographa polyhedrin gene that encodes amino acid residue number 43 substantially as depicted in FIG. 2. ### Claim 40 (depends on 27) The recombinant virus according to claim 27 in which the nucleotide sequence encoding the foreign peptide replaces the part of the Autographa polyhedrin gene that encodes amino acid residue number 50 substantially as depicted in FIG. 2. ### Claim 41 A transfer vector encoding a polyhedrin fusion protein that crystallizes to form recombinant occlusion bodies comprising: (a) a first nucleotide sequence encoding a portion of the polyhedrin structural protein that participates in crystallization; and (b) a second nucleotide sequence encoding a foreign peptide in which the first and second nucleotide sequences are in the same translational reading frame uninterrupted by translation termination signals; and (c) baculovirus flanking sequences surrounding the first and second nucleotide sequence so that recombination with baculovirus can occur in vivo. ### Claim 42 (depends on 41) The transfer vector according to claim 41, in which the nucleotide sequence encoding the foreign peptide replaces all or part of the polyhedrin gene that encodes the amino terminus of the polyhedrin protein. ### Claim 43 (depends on 41) The transfer vector according to claim 41, in which the nucleotide sequence encoding the foreign peptide replaces all or part of a region of the polyhedrin gene that is homologous to the nucleotide sequence substantially as depicted in FIG. 1 from nucleotide number 142 to 180. ### Claim 44 (depends on 41) The transfer vector according to claim 41 in which the nucleotide sequence encoding the foreign peptide is inserted after the coding sequence for amino acid number 1 of the Autographa polyhedrin gene substantially as depicted in FIG. 2. ### Claim 45 (depends on 41) The transfer vector according to claim 41 in which the nucleotide sequence encoding the foreign peptide replaces the part of the Autographa polyhedrin gene that encodes amino acid residue number 43 substantially as depicted in FIG. 2. ### Claim 46 (depends on 41) The transfer vector according to claim 41 in which the nucleotide sequence encoding the foreign peptide replaces the part of the Autographa polyhedrin gene that encodes amino acid residue number 50 substantially as depicted in FIG. 2. ### Claim 47 (depends on 41) The transfer vector according to claim 41, comprising pAV15InHem-43, substantially as deposited with the NRRL and assigned accession number B18308. ### Claim 48 (depends on 41) The transfer vector according to claim 41, comprising pAV15InHem-50, substantially as deposited with the NRRL and assigned accession number B18309. ### Claim 49 (depends on 41) The transfer vector according to claim 41, comprising pAV17bInHem-1, substantially as deposited with the NRRL and assigned accession number B18310. ### Claim 50 (depends on 41) The transfer vector according to claim 41, comprising pAV17bInHem-2, substantially as deposited with the NRRL and assigned accession number B18311. ### Claim 51 A recombinant vector, pBRX13, substantially as deposited with the NRRL and assigned accession number B18312.