# US 12,758,582 B2 · Anti-CD33 antibodies 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 12,758,582 B2 - Document ID: US12758582B2 - Status: Granted - Kind: B2 - Application number: 17/995,089 - Filed: 2021-03-31 - Published: 2023-06-22 - Granted: 2026-10-13 - Earliest claimed: 2020-03-31 - Assignees: Fred Hutchinson Cancer Center - Applicants: Fred Hutchinson Cancer Center - Inventors: Roland B. Walter; George S. Laszlo - Web page: https://patentgrep.com/patent/US12758582B2 ## CPC - A61K 35/17: Lymphocytes; B-cells; T-cells; Natural killer cells; Interferon-activated or cytokine-activated lymphocytes (when activated by a specific antigen A61K39/00) - A61P 35/02: specific for leukemia - C07K 16/2803: {against the immunoglobulin superfamily} - 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) - A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS - 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 A suite of novel anti-CD33 antibodies is described. The provided antibodies are pan-binders, binding the C2-set Ig-like domain in the presence or absence of the V-set Ig-like domain of CD33; are C2-set specific binders, binding the C2-set Ig-like domain only in the absence of the V-set Ig-like domain of CD33; or are V-set binders, binding the V-set Ig-like domain of CD33. The antibodies provide novel therapeutic and diagnostic tools against CD33-related disorders, such as acute myeloid leukemia (AML). ## Claims ### Claim 1 A CD33-binding antibody or antigen biding fragment thereof comprising a binding domain comprising a six-member complementarity determining region (CDR) set having a CDR light chain (L) 1, a CDRL2, a CDRL3, a CDR heavy chain (H) 1, a CDRH2, a CDRH3, wherein: (i) the CDRL1 has the sequence of SEQ ID NO: 20, the CDRL2 has the sequence of SEQ ID NO: 15, the CDRL3 has the sequence of SEQ ID NO: 16, the CDRH1 has the sequence of SEQ ID NO: 21, the CDRH2 has the sequence of SEQ ID NO: 22, and the CDRH3 has the sequence of SEQ ID NO: 23, according to North; (ii) the CDRL1 has the sequence of SEQ ID NO: 253, the CDRL2 has the sequence YTS, the CDRL3 has the sequence of SEQ ID NO: 16, the CDRH1 has the sequence of SEQ ID NO: 254, the CDRH2 has the sequence of SEQ ID NO: 255, and the CDRH3 has the sequence of SEQ ID NO: 23, according to IMGT; (iii) the CDRL1 has the sequence of SEQ ID NO: 20, the CDRL2 has the sequence of SEQ ID NO: 187, the CDRL3 has the sequence of SEQ ID NO: 16, the CDRH1 has the sequence of SEQ ID NO: 291, the CDRH2 has the sequence of SEQ ID NO: 292, and the CDRH3 has the sequence of SEQ ID NO: 293, according to Kabat; or (iv) the CDRL1 has the sequence of SEQ ID NO: 20, the CDRL2 has the sequence of SEQ ID NO: 187, the CDRL3 has the sequence of SEQ ID NO: 16, the CDRH1 has the sequence of SEQ ID NO: 325, the CDRH2 has the sequence of SEQ ID NO: 326, and the CDRH3 has the sequence of SEQ ID NO: 293, according to Chothia; and wherein the CD33-binding antibody or antigen binding fragment thereof is part of an anti-CD33 immunotoxin, an anti-CD33 antibody-drug conjugate, an antibody-fluorophore conjugate, an anti-CD33 antibody-radioisotope conjugate, or an anti-CD33 multispecific antibody. ### Claim 2 (depends on 1) The CD33-binding antibody or antigen binding fragment thereof of claim 1, comprising a variable light chain having the sequence of SEQ ID NO: 40 or a sequence having at least 90% sequence identity to the sequence of SEQ ID NO: 40, and a variable heavy chain having the sequence of SEQ ID NO: 41 or a sequence having at least 90% sequence identity to the sequence of SEQ ID NO: 41. ### Claim 3 (depends on 1) The CD33-binding antibody or antigen binding fragment thereof of claim 1, wherein the antigen binding fragment is a single chain variable fragment (scFv) in a VH-VL orientation or a VL-VH orientation. ### Claim 4 (depends on 3) The CD33-binding antibody or antigen binding fragment thereof of claim 3, wherein the scFv has the sequence of as set forth in SEQ ID NO. 154 or SED ID NO: 155. ### Claim 5 (depends on 1) The CD33-binding antibody or antigen binding fragment thereof of claim 1, wherein the antibody or antigen binding fragment thereof is part of an anti-CD33 immunotoxin comprising holotoxin, hemotoxin, abrin, bouganin, Bryodin 1, diphtheria toxin (DT), gelonin, mistletoe lectin, modeccin, pokeweed antiviral protein (PAP), Pseudomonas exotoxin (PE), ricin, or saporin. ### Claim 6 (depends on 1) The CD33-binding antibody or antigen fragment thereof of claim 1, where the anti-CD33 antibody-drug conjugate comprises a drug comprising: monomethyl auristatin E (MMAE), vedotin, dolastatin, auristatin, calicheamicin, pyrrolobenzodiazepine (PBD), nemorubicin, PNU-159682, anthracycline, duocarmycin, vinca alkaloid, taxane, trichothecene, CC1065, camptothecin, elinafide, taxol, cytochalasin B, gramicidin D, ethidium bromide, emetine, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicin, doxorubicin, daunorubicin, dihydroxy anthracenedione, mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, and/or propranolol. ### Claim 7 (depends on 1) The CD33-binding antibody or antigen binding fragment thereof of claim 1, wherein the anti-CD33 antibody-radioisotope conjugate comprises a radioisotope comprising: 225Ac, 228Ac, 111Ag, 124Am, 72As, 74As, 211At, 209At, 194Au, 128Ba, 7Be, 206Bi, 245Bk, 246Bk, 76Br, 11C, 47Ca, 254Cf, 242Cm, 51Cr, 67Cu, 153Dy, 157Dy, 159Dy, 165Dy, 166Dy, 171Er, 250Es, 254Es, 147Eu, 157Eu, 52Fe, 59Fe, 251Fm, 252Fm, 253Fm, 66Ga, 72Ga, 146Gd, 153Gd, 68Ge, 170Hf, 171Hf, 193Hg, 193mHg, 160mHo, 130I, 131I, 135I, 114mln, 185Ir, 42K, 43K, 76Kr, 79Kr, 81mKr, 132La, 262Lr, 169Lu, 174mLu, 176mLu, 257Mld, 260Md, 28Mg, 52Mn, 90Mo, 24Na, 95Nb, 138Nd, 57Ni, 66Ni, 234Np, 15O, 182Os, 189mOs, 191Os, 32P, 201Pb, 101Pd, 143Pr, 191Pt, 243Pu, 225Ra, 81Rb, 188Re, 105Rh, 211Rn, 103Ru, 35S, 44Sc, 72Se, 153Sm, 125Sn, 91Sr, 173Ta, 154Tb, 127Te, 234Th, 45Ti, 166Tm, 230U, 237U, 240U, 48V, 178V, 181W, 188WV, 125Xe, 127Xe, 133Xe, 133mXe, 135Xe, 85mY, 86Y, 90Y, 93Y, 169Yb, 175Yb, 65Zn, 71mZn, 86Zr, 95Zr, and/or 97Zr. ### Claim 8 (depends on 1) The CD33-binding antibody or antigen binding fragment thereof of claim 1, wherein the anti-CD33 multispecific antibody is a bispecific antibody, a trispecific antibody, or a tetraspecific antibody. ### Claim 9 (depends on 8) The CD33-binding antibody or antigen binding fragment thereof of claim 8, wherein the anti-CD33 multispecific antibody comprises a binding domain that activates an immune cell. ### Claim 10 (depends on 9) The CD33-binding antibody or antigen binding fragment thereof of claim 9, wherein the immune cell is a T-cell, natural killer (NK) cell, or a macrophage. ### Claim 11 (depends on 10) The CD33-binding antibody or antigen binding fragment thereof of claim 10, wherein the T-cell is a CD3 T-cell, a CD4 T-cell, a CD8 T-cell, a central memory T-cell, an effector memory T-cell, and/or a naïve T-cell. ### Claim 12 (depends on 1) The CD33-binding antibody or antigen binding fragment thereof of claim 1, further comprising a Gly-Ser linker comprising a formula (GlyxSery)n, wherein x and y are independently an integer from 0 to 10 provided that x and y are not both 0 and wherein n is an integer of 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. ### Claim 13 (depends on 12) The CD33-binding antibody or antigen binding fragment thereof of claim 12, wherein the Gly-Ser linker comprises SEQ ID NO: 100, SEQ ID NO: 102, SEQ ID NO: 103, SEQ ID NO: 104, SEQ ID NO: 105, SEQ ID NO: 106, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, (Gly2Ser)1, SEQ ID NO: 111, SEQ ID NO: 112, or SEQ ID NO: 113. ### Claim 14 (depends on 1) A composition comprising a CD33-binding antibody or antigen binding fragment thereof of claim 1 and a pharmaceutically acceptable carrier. ### Claim 15 (depends on 14) A method of treating a CD33-related disorder in a subject in need thereof, comprising administering a therapeutically effective amount of the composition of claim 14 to the subject, thereby treating the CD33-related disorder in the subject in need thereof. ### Claim 16 (depends on 15) The method of claim 15, wherein the CD33-related disorder comprises acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL), chronic myelogenous leukemia (CML), chronic myelomonocytic leukemia (CMML), mast cell leukemia, myelodysplastic syndrome (MDS), B-cell acute lymphoblastic leukemia (B-ALL), T-cell acute lymphoblastic leukemia (T-ALL), or megakaryocytic leukemia. ### Claim 17 (depends on 15) The method of claim 15, further comprising determining whether the subject expresses or lacks a V-set domain of CD33, and if the subject expresses the V-set domain of CD33, selecting a combination therapy comprising: a composition comprising: a first binding domain comprising a first six-member complementarity determining region (CDR) set having a first CDR light chain (L) 1, a first CDRL2, a first CDRL3, a first CDR heavy chain (H) 1, a first CDRH2, and a first CDRH3, wherein the first CDRL1 has the sequence of SEQ ID NO: 20, the first CDRL2 has the sequence of SEQ ID NO: 15, the first CDRL3 has the sequence SEQ ID NO: 16, the first CDRH1 has the sequence of SEQ ID NO: 21, the first CDRH2 has the sequence of SEQ ID NO: 22, and the first CDRH3 has the sequence of SEQ ID NO: 23, according to North, and a second binding domain comprising a second six-member CDR set having a CDRL1, a CDRL2, a CDRL3, a CDRH1, a CDRH2, and a CDRH3 wherein, the second CDRL1 has the sequence of SEQ ID NO: 30, the second CDRL2 has the sequence of SEQ ID NO: 208, the second CDRL3 has the sequence SEQ ID NO: 32, the second CDRH1 has the sequence of SEQ ID NO: 209, the second CDRH2 has the sequence of SEQ ID NO: 210, and the second CDRH3 has the sequence of SEQ ID NO: 211, according to North; or the second CDRL1 has the sequence of SEQ ID NO: 174, the second CDRL2 has the sequence of SEQ ID NO: 225, the second CDRL3 has the sequence SEQ ID NO: 176, the second CDRH1 has the sequence of SEQ ID NO: 226, the second CDRH2 has the sequence of SEQ ID NO: 227, and the second CDRH3 has the sequence of SEQ ID NO: 228, according to North; or if the subject does not express the V-set domain of CD33, selecting a therapy comprising: a composition comprising a first binging domain comprising a first six-member CDR set having a first CDRL1, a first CDRL2, a first CDRL3, a first CDRH1, a first CDRH2, and a first CDRH3, wherein the first CDRL1 has the sequence of SEQ ID NO: 20, the first CDRL2 has the sequence of SEQ ID NO: 15, the first CDRL3 has the sequence SEQ ID NO: 16, the first CDRH1 has the sequence of SEQ ID NO: 21, the first CDRH2 has the sequence of SEQ ID NO: 22, and the first CDRH3 has the sequence of SEQ ID NO: 23, according to North; and a second binding domain comprising a second six-member CDR set having a second CDRL1, a second CDRL2, a second CDRL3, a second CDRH1, a second CDRH2, and a second CDRH3 wherein, the second CDRL1 has the sequence of SEQ ID NO: 180, the second CDRL2 has the sequence of SEQ ID NO: 229, the second CDRL3 has the sequence SEQ ID NO: 182, the second CDRH1 has the sequence of SEQ ID NO: 230, the second CDRH2 has the sequence of SEQ ID NO: 231, and the second CDRH3 has the sequence of SEQ ID NO: 232, according to North; the second CDRL1 has the sequence of SEQ ID NO: 198, the second CDRL2 has the sequence of SEQ ID NO: 229, the second CDRL3 has the sequence SEQ ID NO: 182, the second CDRH1 has the sequence of SEQ ID NO: 240, the second CDRH2 has the sequence of SEQ ID NO: 241, and the second CDRH3 has the sequence of SEQ ID NO: 242, according to North; the second CDRL1 has the sequence of SEQ ID NO: 202, the second CDRL2 has the sequence of SEQ ID NO: 243, the second CDRL3 has the sequence SEQ ID NO: 204, the second CDRH1 has the sequence of SEQ ID NO: 244, the second CDRH2 has the sequence of SEQ ID NO: 245, and the second CDRH3 has the sequence of SEQ ID NO: 246, according to North; the second CDRL1 has the sequence of SEQ ID NO: 186, the second CDRL2 has the sequence of SEQ ID NO: 15, the second CDRL3 has the sequence SEQ ID NO: 188, the second CDRH1 has the sequence of SEQ ID NO: 233, the second CDRH2 has the sequence of SEQ ID NO: 234, and the second CDRH3 has the sequence of SEQ ID NO: 235, according to North; or the second CDRL1 has the sequence of SEQ ID NO: 192, the second CDRL2 has the sequence of SEQ ID NO: 236, the second CDRL3 has the sequence SEQ ID NO: 194, the second CDRH1 has the sequence of SEQ ID NO: 237, the second CDRH2 has the sequence of SEQ ID NO: 238, and the second CDRH3 has the sequence of SEQ ID NO: 239, according to North. ### Claim 18 A kit comprising a composition comprising a first binding domain comprising a first six-member complementarity determining region (CDR) set having a first CDRL1, a first CDRL2, a first CDRL3, a first CDRH1, a first CDRH2, and a first CDRH3, wherein the first CDRL1 has the sequence of SEQ ID NO: 20, the first CDRL2 has the sequence of SEQ ID NO: 15, the first CDRL3 has the sequence SEQ ID NO: 16, the first CDRH1 has the sequence of SEQ ID NO: 21, the first CDRH2 has the sequence of SEQ ID NO: 22, and the first CDRH3 has the sequence of SEQ ID NO: 23, according to North; and a second binding domain comprising a second six-member CDR set having a second CDRL1, a second CDRL2, a second CDRL3, a second CDRH1, a second CDRH2, and a second CDRH3 wherein: the second CDRL1 has the sequence of SEQ ID NO: 180, the second CDRL2 has the sequence of SEQ ID NO: 229, the second CDRL3 has the sequence SEQ ID NO: 182, the second CDRH1 has the sequence of SEQ ID NO: 230, the second CDRH2 has the sequence of SEQ ID NO: 231, and the second CDRH3 has the sequence of SEQ ID NO: 232, according to North; the second CDRL1 has the sequence of SEQ ID NO: 198, the second CDRL2 has the sequence of SEQ ID NO: 229, the second CDRL3 has the sequence SEQ ID NO: 182, the second CDRH1 has the sequence of SEQ ID NO: 240, the second CDRH2 has the sequence of SEQ ID NO: 241, and the second CDRH3 has the sequence of SEQ ID NO: 242, according to North; the second CDRL1 has the sequence of SEQ ID NO: 202, the second CDRL2 has the sequence of SEQ ID NO: 243, the second CDRL3 has the sequence SEQ ID NO: 204, the second CDRH1 has the sequence of SEQ ID NO: 244, the second CDRH2 has the sequence of SEQ ID NO: 245, and the second CDRH3 has the sequence of SEQ ID NO: 246, according to North; the second CDRL1 has the sequence of SEQ ID NO: 186, the second CDRL2 has the sequence of SEQ ID NO: 15, the second CDRL3 has the sequence SEQ ID NO: 188, the second CDRH1 has the sequence of SEQ ID NO: 233, the second CDRH2 has the sequence of SEQ ID NO: 234, and the second CDRH3 has the sequence of SEQ ID NO: 235, according to North; or the second CDRL1 has the sequence of SEQ ID NO: 192, the second CDRL2 has the sequence of SEQ ID NO: 236, the second CDRL3 has the sequence SEQ ID NO: 194, the second CDRH1 has the sequence of SEQ ID NO: 237, the second CDRH2 has the sequence of SEQ ID NO: 238, and the second CDRH3 has the sequence of SEQ ID NO: 239, according to North.