# US 2021/0277067 A1 · Modified cytomegalovirus proteins and stabilized complexes > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 2021/0277067 A1 - Document ID: US20210277067A1 - Status: Published application - Kind: A1 - Application number: 17/223,207 - Filed: 2021-04-06 - Published: 2021-09-09 - Earliest claimed: 2018-10-17 - Assignees: Glaxosmithkline Biologicals SA - Applicants: Glaxosmithkline Biologicals SA - Inventors: Enrico MALITO; Matthew James BOTTOMLEY; Andrea CARFI; Sumana CHANDRAMOULI; Kate LUISI - Web page: https://patentgrep.com/patent/US20210277067A1 ## CPC - A61K 39/12: Viral antigens - C07K 14/045: Cytomegalovirus - A61K 39/245: Herpetoviridae, e.g. herpes simplex virus - C07K 14/005: from viruses - A61K 38/00: Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00) - C12N 2710/16122: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes - C12N 2710/16134: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein - 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) - 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) - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) ## Abstract Described are mutant human cytomegalovirus (HCMV) pentamer complex polypeptides, methods of making them, and their use in HCMV protein complexes and compositions. In particular, the use of the modified HCMV polypeptides to stabilize HCMV complexes or unmask a pentamer epitope is described. ## Claims ### Claim 1 A complex comprising one or more mutant Human Cytomegalovirus (HCMV) polypeptides, wherein the one or more mutant HCMV polypeptides are: (a) a pUL128 polypeptide, or complex-forming fragment thereof, that has cysteine (C) at residue 142, numbered with respect to SEQ ID NO: 13; a gL polypeptide, or complex-forming fragment thereof, that has a cavity filling mutant at residue 140 and has a cysteine (C) at residue 150, numbered with respect to SEQ ID NO: 7; and a pUL130 polypeptide, or complex-forming fragment thereof, that has a cysteine (C) at residue 64 and a cysteine (C) at residue 95, numbered with respect to SEQ ID NO: 17; (b) a pUL128 polypeptide, or complex-forming fragment thereof, that has cysteine (C) at residue 142, numbered with respect to SEQ ID NO: 13; a gL polypeptide, or complex-forming fragment thereof, that has a cysteine (C) at residue 150, numbered with respect to SEQ ID NO: 7; and a pUL130 polypeptide, or complex-forming fragment thereof, that has a cysteine (C) at residue 64 and a cysteine (C) at residue 95, numbered with respect to SEQ ID NO: 17; (c) a pUL128 polypeptide, or complex-forming fragment thereof, that has cysteine (C) at residue 142, numbered with respect to SEQ ID NO: 13; a gL polypeptide, or complex-forming fragment thereof, that has a cavity filling mutant at residue 145 and has a cysteine (C) at residue 150, numbered with respect to SEQ ID NO: 7; and a pUL130 polypeptide, or complex-forming fragment thereof, that has a cysteine (C) at residue 64 and a cysteine (C) at residue 95, numbered with respect to SEQ ID NO: 17; (d) a pUL128 polypeptide, or complex-forming fragment thereof, that has cysteine (C) at residue 142, numbered with respect to SEQ ID NO: 13; a gL polypeptide, or complex-forming fragment thereof, that has a cavity filling mutant at residue 224 and has a cysteine (C) at residue 150, numbered with respect to SEQ ID NO: 7; and a pUL130 polypeptide, or complex-forming fragment thereof, that has a cysteine (C) at residue 64 and a cysteine (C) at residue 95, numbered with respect to SEQ ID NO: 17; (e) a pUL128 polypeptide, or complex-forming fragment thereof, that has cysteine (C) at residue 142, numbered with respect to SEQ ID NO: 13; a gL polypeptide, or complex-forming fragment thereof, that has a cavity filling mutant at residue 224, numbered with respect to SEQ ID NO: 7; and a pUL130 polypeptide, or complex-forming fragment thereof, that has a cysteine (C) at residue 95, numbered with respect to SEQ ID NO: 17; (f) a pUL128 polypeptide, or complex-forming fragment thereof, that has a cavity filling mutant at residue 77 and has a cavity filling mutant at residue 103, numbered with respect to SEQ ID NO: 13; and a gL polypeptide, or complex-forming fragment thereof, that has a cavity filling mutant at residue 140 and has a cavity filling mutant at residue 145, numbered with respect to SEQ ID NO: 7; (g) a pUL128 polypeptide, or complex-forming fragment thereof, that has a cavity filling mutant at residue 77 and has a cavity filling mutant at residue 103, numbered with respect to SEQ ID NO: 13; and a gL polypeptide, or complex-forming fragment thereof, that has a cavity filling mutant at residue 140, has a cavity filling mutant at residue 145, and has a cavity filling mutant at residue 224, numbered with respect to SEQ ID NO: 7; (h) a gL polypeptide, or complex-forming fragment thereof, that has a cavity filling mutant at residue 140, numbered with respect to SEQ ID NO: 7; and a pUL131 polypeptide, or complex-forming fragment thereof, that has a repacking hydrophobic mutant at residue 52 and has a repacking hydrophobic mutant at residue 67, numbered with respect to SEQ ID NO: 21; (i) a pUL13 I polypeptide, or complex-forming fragment thereof, that has a cavity filling mutant at residue 86, numbered with respect to SEQ ID NO: 21; or (j) a pUL128 polypeptide, or complex-forming fragment thereof, that has a cysteine (C) at residue 83, numbered with respect to SEQ ID NO: 13; and a pUL130 polypeptide, or complex-forming fragment thereof, that has a cysteine (C) at residue 167, numbered with respect to SEQ ID NO: 17. ### Claim 2 (depends on 1) The complex of claim 1, wherein the cavity filling mutant at residue 140, numbered with respect to SEQ ID NO: 7, is tryptophan (W), phenylalanine (F), tyrosine (Y), valine (V), isoleucine (I), or leucine (L). ### Claim 3 (depends on 1) The complex of claim 1, wherein the cavity filling mutant at residue 145, numbered with respect to SEQ ID NO: 7, is tryptophan (W), phenylalanine (F), tyrosine (Y), valine (V), isoleucine (I), or leucine (L). ### Claim 4 (depends on 1) The complex of claim 1, wherein the cavity filling mutant at residue 224, numbered with respect to SEQ ID NO: 7, is tryptophan (W), phenylalanine (F), tyrosine (Y), valine (V), isoleucine (I), or leucine (L). ### Claim 5 (depends on 1) The complex of claim 1, wherein the cavity filling mutant at residue 77, numbered with respect to SEQ ID NO: 13, is tryptophan (W), phenylalanine (F), tyrosine (Y), valine (V), isoleucine (I), or leucine (L). ### Claim 6 (depends on 1) The complex of claim 1, wherein the cavity filling mutant at residue 103, numbered with respect to SEQ ID NO: 13, is tryptophan (W), phenylalanine (F), tyrosine (Y), valine (V), isoleucine (I), or leucine (L). ### Claim 7 (depends on 1) The complex of claim 1, wherein the repacking hydrophobic mutant at residue 52, numbered with respect to SEQ ID NO: 21, is tryptophan (W), phenylalanine (F), methionine (M), cysteine (C), alanine (A), leucine (L), isoleucine (I), valine (V) or proline (P). ### Claim 8 (depends on 1) The complex of claim 1, wherein the repacking hydrophobic mutant at residue 67, numbered with respect to SEQ ID NO: 21, is tryptophan (W), phenylalanine (F), methionine (M), cysteine (C), alanine (A), leucine (L), isoleucine (I), valine (V) or proline (P). ### Claim 9 (depends on 1) The complex of claim 1, wherein the cavity filling mutant at residue 86, numbered with respect to SEQ ID NO: 21, is tryptophan (W), phenylalanine (F), tyrosine (Y), valine (V), isoleucine (I), or leucine (L). ### Claim 10 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9) The complex of any of claims 1-9 wherein the complex has an increased thermostability as compared to a control complex in the same conditions. ### Claim 11 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10) The complex of any of claims 1-10, wherein the complex is an HCMV pentamer complex comprising one or more mutant HCMV polypeptides. ### Claim 12 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11) An immunogenic composition comprising the complex of any of claims 1-11 and, optionally, a non-antigen component. ### Claim 13 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11) An isolated nucleic acid molecule comprising one or more operably linked polynucleotide sequences that encode the complex of any of claims 1-11. ### Claim 14 (depends on 13) An expression vector comprising the isolated nucleic acid molecule of claim 13. ### Claim 15 (depends on 13, 14) A host cell comprising the isolated nucleic acid molecule of claim 13 or the expression vector of claim 14. ### Claim 16 (depends on 15) A method of making a complex, comprising cultivating the host cell of claim 15. ### Claim 17 (depends on 12) A method of inducing an immune response against Human Cytomegalovirus (HCMV), comprising administering to a subject an immunologically effective amount of the immunogenic composition of claim 12. ### Claim 18 (depends on 1) A method of inhibiting Human Cytomegalovirus (HCMV) entry into a cell, comprising contacting the cell with a complex according to claim 1.