# US 10,308,930 B2 · Methods to accelerate antibody diversification > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 10,308,930 B2 - Document ID: US10308930B2 - Status: Granted - Kind: B2 - Application number: 15/514,052 - Filed: 2015-11-05 - Published: 2018-06-28 - Granted: 2019-06-04 - Earliest claimed: 2014-11-06 - Assignees: Albert Einstein College of Medicine - Applicants: Albert Einstein College of Medicine, Inc. - Inventors: Xiaohua Wang; Matthew Scharff - Web page: https://patentgrep.com/patent/US10308930B2 ## CPC - C12N 15/1024: {In vivo mutagenesis using high mutation rate "mutator" host strains by inserting genetic material, e.g. encoding an error prone polymerase, disrupting a gene for mismatch repair} - C07K 14/005: from viruses - C07K 16/00: Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies - C12N 9/78: acting on carbon to nitrogen bonds other than peptide bonds (3.5) - C12N 15/113: Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; {Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing (when used in plants C12N15/8218)} - C12N 15/86: Viral vectors - C07K 2317/14: Specific host cells or culture conditions, e.g. components, pH or temperature - C07K 2319/80: containing a DNA binding domain, e.g. Lacl or Tet-repressor - C12N 2310/14: interfering nucleic acids [NA] - C12N 2310/531: Stem-loop; Hairpin - C12N 2710/10043: viral genome or elements thereof as genetic vector - C12N 2740/15043: viral genome or elements thereof as genetic vector - 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) ## Abstract Methods for increasing somatic hypermutation of antibodies, especially in vitro expression of an isolated retroviral protein in an isolated mammalian immunoglobulin-producing cell, are provided. ## Claims ### Claim 1 A method of increasing somatic hypermutation of an immunoglobulin loci comprising effecting expression of a Tat protein in (i) an isolated mammalian immunoglobulin-producing cell, or (ii) an isolated immunoglobulin-producing cell of a cell line derived from a mammal, in an amount effective to increase somatic hypermutation of an immunoglobulin loci. ### Claim 2 (depends on 1) The method of claim 1, wherein expression is effected in an isolated mammalian immunoglobulin-producing B cell. ### Claim 3 (depends on 1) The method of claim 1, wherein expression is effected in the isolated immunoglobulin-producing cell of a cell line derived from a mammal. ### Claim 4 (depends on 3) The method of claim 3, wherein the isolated immunoglobulin-producing cell of a cell line derived from a mammal is a cell that does not constitutively express immunoglobulins, but which has been genetically engineered to express immunoglobulins. ### Claim 5 (depends on 3) The method of claim 3, wherein the isolated immunoglobulin-producing cell of a cell line derived from a mammal is a Ramos cell line. ### Claim 6 (depends on 1) The method of claim 1, wherein the isolated immunoglobulin-producing cell of a cell line derived from a mammal is a hybridoma. ### Claim 7 (depends on 1) The method of claim 1, wherein the cell secretes immunoglobulins. ### Claim 8 (depends on 1) The method of claim 1, wherein the cell expresses immunoglobulins on a cell surface thereof. ### Claim 9 (depends on 1) The method of claim 1, wherein the expression of the isolated exogenous protein is of a retroviral protein in the cell is effected by contacting the cell with a vector encoding the isolated retroviral protein. ### Claim 10 (depends on 1) The method of claim 1, wherein the isolated retroviral protein is an HIV-2 Tat protein. ### Claim 11 (depends on 9) The method of claim 9, wherein the vector is a lentiviral vector. ### Claim 12 (depends on 9) The method of claim 9, wherein the vector is an adenoviral vector. ### Claim 13 (depends on 1) The method of claim 1, wherein the expression of the HIV Tat protein in the cell is effected by contacting the cell with a pCDNA-tat eukaryotic expression vector encoding the exogenous HIV Tat protein. ### Claim 14 (depends on 1) The method of claim 1, further comprising recovering one or more immunoglobulins produced by the cell of (i) or (ii) subsequent to effecting expression of the isolated exogenous protein therein. ### Claim 15 (depends on 14) The method of claim 14, further comprising screening the one or more immunoglobulins for activity against a predetermined antigen. ### Claim 16 (depends on 1) The method of claim 1, further comprising reducing expression of Spt5 in the cell of (i) by contacting the cell with a RNAi nucleic acid directed to Spt5. ### Claim 17 (depends on 16) The method of claim 16, wherein the RNAi nucleic acid directed to Spt5 is a siRNA or an shRNA. ### Claim 18 (depends on 17) The method of claim 17, wherein the RNAi nucleic acid is an shRNA delivered by a vector.