# US 10,766,960 B2 · Heterodimerized polypeptide > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 10,766,960 B2 - Document ID: US10766960B2 - Status: Granted - Kind: B2 - Application number: 14/654,895 - Filed: 2013-12-26 - Published: 2015-12-03 - Granted: 2020-09-08 - Earliest claimed: 2012-12-27 - Assignees: Chugai Seiyaku Kabushiki Kaisha - Applicants: Chugai Seiyaku Kabushiki Kaisha - Inventors: Tomoyuki Igawa; Futa Mimoto; Hitoshi Katada; Hirotake Shiraiwa - Web page: https://patentgrep.com/patent/US10766960B2 ## CPC - C07K 16/283: {against Fc-receptors, e.g. CD16, CD32, CD64 (CD23 C07K16/2851)} - A61P 29/00: Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID] - A61P 35/00: Antineoplastic agents - A61P 37/00: Drugs for immunological or allergic disorders - A61P 37/02: Immunomodulators - C07K 16/00: Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies - C07K 16/303: {Liver or Pancreas} - C07K 16/46: Hybrid immunoglobulins (hybrids of an immunoglobulin with a peptide not being an immunoglobulin C07K19/00) - C07K 2317/31: multispecific - C07K 2317/52: Constant or Fc region; Isotype - C07K 2317/524: CH2 domain - C07K 2317/72: Increased effector function due to an Fc-modification - C07K 2317/732: Antibody-dependent cellular cytotoxicity [ADCC] - C07K 2319/33: fusions for targeting to specific cell types, e.g. tissue specific targeting, targeting of a bacterial subspecies - 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) - A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS ## Abstract The present inventors produced a heterodimerized polypeptide having an Fc region formed from two polypeptides with different amino acid sequences (a first polypeptide and a second polypeptide), and succeeded in producing a heterodimerized polypeptide containing an Fc region with improved function compared to that of a homodimer in which the Fc region is composed of only the first polypeptide or the second polypeptide by conventional technology. ## Claims ### Claim 1 A heterodimer comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises an IgG Fc region comprising a set of amino acids selected from the group consisting of sets (a) to (aj) below (all positions by EU numbering): (a) amino acid E at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; amino acid D at position 327; (b) amino acid S at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (c) amino acid L at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (d) amino acid L at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (e) amino acid S at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (f) amino acid F at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (g) amino acid E at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (h) amino acid F at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (i) amino acid V at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (j) amino acid D at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (k) amino acid Q at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (l) amino acid I at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (m) amino acid M at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (n) amino acid T at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (o) amino acid A at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (p) amino acid G at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (q) amino acid H at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (r) amino acid V at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (s) amino acid D at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (t) amino acid Q at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (u) amino acid I at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (v) amino acid M at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (w) amino acid T at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (x) amino acid A at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (y) amino acid G at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (z) amino acid H at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid M at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (aa) amino acid F at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid I at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (ab) amino acid E at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid I at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (ac) amino acid D at position 234; amino acid Q at position 235; amino acid W at position 236; amino acid I at position 239; amino acid D at position 268; amino acid E at position 270; and amino acid A at position 298; (ad) amino acid V at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid I at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (ae) amino acid I at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid I at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (af) amino acid L at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid I at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (ag) amino acid E at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid I at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (ah) amino acid D at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid I at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; (ai) amino acid F at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid I at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; and (aj) amino acid T at position 234; amino acid Y at position 235; amino acid W at position 236; amino acid I at position 239; amino acid D at position 268; amino acid A at position 298; and amino acid D at position 327; and the second polypeptide comprises an IgG Fc region that is different from the Fc region of the first polypeptide and comprises a set of amino acids selected from the group consisting of sets (ak) to (aq) (all positions by EU numbering): (ak) amino acid E at position 270; amino acid D at position 326; amino acid K at position 330; and amino acid E at position 334; (al) amino acid E at position 270; amino acid D at position 326; amino acid M at position 330; and amino acid E at position 334; (am) amino acid E at position 270; amino acid D at position 326; amino acid A at position 330; and amino acid E at position 334; (an) amino acid E at position 270; amino acid D at position 326; amino acid F at position 330; and amino acid E at position 334; (ao) amino acid E at position 270; amino acid D at position 326; amino acid I at position 330; and amino acid E at position 334; (ap) amino acid E at position 270; amino acid D at position 326; amino acid Y at position 330; and amino acid E at position 334; and (aq) amino acid E at position 270; amino acid D at position 326; amino acid H at position 330; and amino acid E at position 334. ### Claim 2 (depends on 1) The heterodimer of claim 1, wherein the amino acid sequence of the first polypeptide differs from the amino acid sequence of the second polypeptide at one or more additional positions selected from the group consisting of positions 137, 138, 139, 147, 192, 193, 196, 198, 199, 203, 214, 231, 233, 242, 263, 272, 274, 278, 288, 290, 316, 317, 320, 324, 335, 337, 338, 340, 341, 358, 360, 362, 364, 383, 384, 385, 386, 387, 390, 397, and 422 (all positions by EU numbering), wherein the difference at each such additional position increases the difference between isoelectric points of the first polypeptide and the second polypeptide. ### Claim 3 (depends on 2) The heterodimer of claim 2, wherein the one or more additional positions comprise mutation(s) in either the first or second polypeptide at EU numbering position(s) selected from the group consisting of positions 196, 199, 231, 233, 242, 263, 272, 316, 358, 364, 383, 387, and 397, and further comprise mutation(s) in the other polypeptide at EU numbering position(s) selected from the group consisting of positions 137, 138, 139, 147, 192, 193, 198, 199, 203, 214, 274, 278, 288, 290, 316, 317, 320, 324, 335, 337, 338, 340, 341, 358, 360, 362, 383, 384, 385, 386, 390, and 422, with all of said mutations being mutations relative a naturally occurring IgG Fc region amino acid sequence. ### Claim 4 (depends on 1) The heterodimer of claim 1, wherein the heterodimer is an antigen-binding molecule. ### Claim 5 (depends on 4) The heterodimer of claim 4, wherein the antigen-binding molecule is an antibody, a bispecific antibody, or an Fc fusion molecule. ### Claim 6 (depends on 1) A pharmaceutical composition comprising the heterodimer of claim 1 and a pharmaceutically acceptable carrier. ### Claim 7 (depends on 1) The heterodimer of claim 1, wherein, in the first polypeptide, the amino acid at position 234 (EU numbering) is E, D, T, or L, and the amino acid at position 327 (EU numbering) is D. ### Claim 8 (depends on 1) The heterodimer of claim 1, wherein, in the first polypeptide, the amino acid at position 234 (EU numbering) is L, F, E, or D, and the amino acid at position 270 (EU numbering) is E. ### Claim 9 (depends on 1) The heterodimer of claim 1, wherein, in the first polypeptide, the amino acid at position 234 (EU numbering) is V, I, T, M, or L, and the amino acid at position 327 (EU numbering) is D. ### Claim 10 (depends on 1) The heterodimer of claim 1, wherein, in the first polypeptide, the amino acid at position 234 (EU numbering) is V, E, D, T, I, L, or F, and the amino acid at position 327 (EU numbering) is D; and in the second polypeptide, the amino acid at position 330 (EU numbering) is A or K. ### Claim 11 (depends on 1) The heterodimer of claim 1, wherein, in the first polypeptide, the amino acid at position 234 (EU numbering) is F, E, D, S, or L, and the amino acid at position 270 (EU numbering) is E; and in the second polypeptide, the amino acid at position 330 (EU numbering) is A, F, or K. ### Claim 12 (depends on 1) The heterodimer of claim 1, wherein, when compared to a naturally occurring IgG Fc region, the IgG Fc regions of the first and second polypeptides together exhibit an alteration of function selected from the group consisting of: enhanced binding to an Fcγ receptor, decreased binding to an Fcγ receptor, and increased selectivity of binding to one Fcγ receptor over another. ### Claim 13 (depends on 12) The heterodimer of claim 12, wherein each Fcγ receptor is selected from the group consisting of FcγRIa, FcγRIIa R, FcγRIIa H, FcγRIIb, FcγRIIIaF, and FcγRIIIaV. ### Claim 14 (depends on 12) The heterodimer of claim 12, wherein the alteration of function is increased selectivity of binding to one Fcγ receptor over another. ### Claim 15 (depends on 14) The heterodimer of claim 14, wherein the alteration of function is increased selectivity of binding to at least one activating Fcγ receptor over an inhibitory Fcγ receptor, or vice versa; wherein the at least one activating Fcγ receptor is selected from the group consisting of FcγRIa, FcγRIIa R, FcγRIIa H, FcγRIIIaF, and FcγRIIIaV; and wherein the inhibitory Fcγ receptor is FcγRIIb. ### Claim 16 (depends on 15) The heterodimer of claim 15, wherein binding to the at least one activating Fcγ receptor is selectively enhanced compared to binding to the inhibitory Fcγ receptor. ### Claim 17 (depends on 1) The heterodimer of claim 1, wherein the heterodimer is any one of (A) to (D) below: (A) the first polypeptide comprises a set of amino acids selected from the group consisting of sets (a) to (h), (r) to (z), and (ad) to (aj); and the second polypeptide comprises the set of amino acids (ak); (B) the first polypeptide comprises a set of amino acids selected from the group consisting of sets (f), (g), (j), (aa), (ab), and (ac); and the second polypeptide comprises the set of amino acids (an); (C) the first polypeptide comprises a set of amino acids selected from the group consisting of sets (c), (e) to (g), and (i) to (q); and the second polypeptide comprises the set of amino acids (al); or (D) the first polypeptide comprises a set of amino acids selected from the group consisting of sets (c), (e) to (h), and (aa) to (ac); and the second polypeptide comprises the set of amino acids (am).