# US 10,118,965 B2 · Compositions and methods related to anti-EGFR antibody drug conjugates

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- Number: US 10,118,965 B2
- Document ID: US10118965B2
- Status: Granted
- Kind: B2
- Application number: 15/276,231
- Filed: 2016-09-26
- Published: 2017-03-30
- Granted: 2018-11-06
- Earliest claimed: 2015-09-25
- Assignees: LegoChem Biosciences, Inc.
- Applicants: LegoChem Biosciences, Inc.
- Inventors: Yong Zu Kim; Yun Hee Park; Hyo Jung Choi; Ji Eun Jung; Jeong Hee Ryu; Ho Young Song; Jeiwook Chae; Chul-Woong Chung
- Web page: https://patentgrep.com/patent/US10118965B2

## CPC

- C07K 16/2863: {against receptors for growth factors, growth regulators}
- A61K 47/6817: {Toxins}
- A61K 47/6849: {the antibody targeting a receptor, a cell surface antigen or a cell surface determinant}
- A61K 47/6889: {Conjugates wherein the antibody being the modifying agent and wherein the linker, binder or spacer confers particular properties to the conjugates, e.g. peptidic enzyme-labile linkers or acid-labile linkers, providing for an acid-labile immuno conjugate wherein the drug may be released from its antibody conjugated part in an acidic, e.g. tumoural or environment}
- 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)

## Abstract

In some aspects, the invention relates to an antibody-drug conjugate, comprising an anti-epidermal growth factor receptor (“EGFR”) antibody; a linker; and an active agent. The antibody-drug conjugate may comprise a self-immolative group. The linker may comprise an O-substituted oxime, e.g., wherein the oxygen atom of the oxime is substituted with a group that covalently links the oxime to the drug; and the carbon atom of the oxime is substituted with a group that covalently links the oxime to the antibody.

## Claims

### Claim 1

An antibody drug conjugate, having the structure of Formula I:
wherein:
G is a glucuronic acid moiety or
wherein R3 is hydrogen or a carboxyl protecting group; and each R4 is independently hydrogen or a hydroxyl protecting group;
A represents an anti-EGFR antibody;
B represents an active agent;
W is —C(O)—, —C(O)NR′—, —C(O)O—, —S(O)2NR′—, —P(O)R″NR′—, —S(O)NR′—, or —PO2NR′—, in each case where the C(O), S, or P is directly bound to the phenyl ring, and R′ and R″ are each independently hydrogen, (C1-C8)alkyl, (C3-C8)cycloalkyl, (C1-C8)alkoxy, (C1-C8)alkylthio, mono- or di-(C1-C8)alkylamino, (C3-C20)heteroaryl, or (C6-C20)aryl;
each Z independently represents hydrogen, (C1-C8)alkyl, halogen, cyano, or nitro;
n is an integer from 1 to 3;
L comprises an alkylene chain of 3 to 50 atoms that covalently links A to W, wherein the chain further comprises an oxime and either:
(a) the oxygen atom of the oxime is substituted with a group that covalently links the oxime to W; and the carbon atom of the oxime is substituted with a group that covalently links the oxime to A; or
(b) the carbon atom of the oxime is substituted with a group that covalently links the oxime to W; and the oxygen atom of the oxime is substituted with a group that covalently links the oxime to A;
R1 and R2 are each independently hydrogen, (C1-C8)alkyl, or (C3-C8)cycloalkyl, or R1 and R2 taken together with the carbon atom to which they are attached form a (C3-C8)cycloalkyl ring; and
the moieties connecting A and B, taken together, form the linker.

### Claim 2 (depends on 1)

The antibody-drug conjugate of claim 1, wherein W represents —C(O)NR′—, wherein C(O) is bonded to the phenyl ring and NR′ is bonded to L.

### Claim 3 (depends on 1)

The antibody-drug conjugate of claim 1, wherein the linker comprises at least one isoprenyl unit, represented by

### Claim 4 (depends on 3)

The antibody-drug conjugate of claim 3, wherein the isoprenyl unit covalently links the oxime to the antibody.

### Claim 5 (depends on 4)

The antibody-drug conjugate of claim 4, wherein the linker comprises:

### Claim 6 (depends on 1)

The antibody-drug conjugate of claim 1, wherein the linker comprises at least one polyethylene glycol unit, represented by

### Claim 7 (depends on 6)

The antibody-drug conjugate of claim 6, wherein the linker comprises an oxime, and the at least one polyethylene glycol unit covalently links the oxime to the active agent.

### Claim 8 (depends on 2)

The antibody-drug conjugate of claim 2, wherein the linker comprises a connection unit represented by —(CH2)r(V(CH2)p)q—, wherein:
r is an integer of 1 to 10;
p is an integer of 0 to 10;
q is an integer of 1 to 10;
V is a single bond, —O—, —S—, —NR21—, —C(O)NR22—, —NR23C(O)—, —NR24SO2—, or —SO2NR25—; and
R21 to R25 are each independently hydrogen, (C1-C6)alkyl, (C1-C6)alkyl(C6-C20)aryl, or (C1-C6)alkyl(C3-C20)heteroaryl.

### Claim 9 (depends on 1)

The antibody-drug conjugate of claim 1, wherein the linker comprises a connection unit represented by (CH2CH2X)w—, wherein:
X represents —O—, (C1-C8)alkylene, or —NR21—;
R21 represents hydrogen, (C1-C6)alkyl, (C1-C6)alkyl(C6-C20)aryl, or (C1-C6)alkyl(C3-C20)heteroaryl; and
w is an integer of 1 to 10.

### Claim 10 (depends on 1)

The antibody-drug conjugate of claim 1, wherein the linker comprises a linking moiety formed by a 1,3-dipolar cycloaddition reaction, hetero-Diels-Alder reaction, nucleophilic substitution reaction, non-aldol type carbonyl reaction, addition to carbon-carbon multiple bond, oxidation reaction, or click reaction.

### Claim 11 (depends on 10)

The antibody-drug conjugate of claim 10, wherein the linking moiety is represented by any one of Formulas A, B, C, or D:
wherein:
L1 is a single bond or alkylene having 1 to 30 carbon atoms;
R11 is hydrogen or alkyl having 1 to 10 carbon atoms; and
L2 is alkylene having 1 to 30 carbon atoms.

### Claim 12 (depends on 11)

The antibody-drug conjugate of claim 11, wherein:
the linker comprises
V is a single bond, —O—, —S—, —NR21—, —C(O)NR22—, —NR23C(O)—, —NR24SO2—, or —SO2NR25—;
R21 to R25 are each independently hydrogen, (C1-C6)alkyl, (C1-C6)alkyl(C6-C20)aryl, or (C1-C6)alkyl(C3-C20)heteroaryl;
r is an integer of 1 to 10;
p is an integer of 0 to 10;
q is an integer of 1 to 10; and
L1 is a single bond.

### Claim 13 (depends on 1)

The antibody-drug conjugate of claim 1, comprising the structure of Formula III:
wherein A represents the antibody; and B represents the active agent.

### Claim 14 (depends on 1)

The antibody-drug conjugate of claim 1, wherein the linker is covalently bound to the antibody by a thioether bond, and the thioether bond comprises a sulfur atom of a cysteine of the antibody.

### Claim 15 (depends on 1)

The antibody-drug conjugate of claim 1, wherein the antibody is a monoclonal antibody, polyclonal antibody, antibody fragment, Fab, Fab′, Fab′-SH, F(ab′)2, Fv, single chain Fv (“scFv”), diabody, linear antibody, bispecific antibody, multispecific antibody, chimeric antibody, humanized antibody, human antibody, or fusion protein comprising the antigen-binding portion of an antibody.

### Claim 16 (depends on 1)

The antibody-drug conjugate of claim 1, wherein the antibody can specifically bind an extracellular epitope on EGFR.

### Claim 17 (depends on 1)

The antibody-drug conjugate of claim 1, wherein the active agent is a chemotherapeutic agent or a toxin.

### Claim 18 (depends on 1)

The antibody-drug conjugate of claim 1, wherein the active agent is selected from:
(a) erlotinib, bortezomib, fulvestrant, sutent, letrozole, imatinib mesylate, PTK787/ZK 222584, oxaliplatin, 5-fluorouracil, leucovorin, rapamycin, lapatinib, lonafarnib, sorafenib, gefitinib, AG1478, AG1571, thiotepa, cyclophosphamide, busulfan, improsulfan, piposulfan, benzodopa, carboquone, meturedopa, uredopa, ethylenimine, altretamine, triethylenemelamine, trietylenephosphoramide, triethiylenethiophosphoramide, trimethylolomelamine, bullatacin, bullatacinone, camptothecin, topotecan, bryostatin, callystatin, CC-1065, adozelesin, carzelesin, bizelesin, cryptophycin 1, cryptophycin 8,dolastatin, duocarmycin, KW-2189, CB1-TM1, eleutherobin, pancratistatin, sarcodictyin, spongistatin, chlorambucil, chlornaphazine, cholophosphamide, estramustine, ifosfamide, mechlorethamine, melphalan, novembichin, phenesterine, prednimustine, trofosfamide, uracil mustard, carmustine, chlorozotocin, fotemustine, lomustine, nimustine, ranimnustine, calicheamicin, calicheamicin gamma 1, calicheamicin omega 1, dynemicin, dynemicin A, clodronate, esperamicin, neocarzinostatin chromophore, aclacinomysins, actinomycin, antrmycin, azaserine, bleomycins, cactinomycin, carabicin, carninomycin, carzinophilin, chromomycins, dactinomycin, daunorubicin, detorubucin, 6-diazo-5-oxo-L-norleucine, doxorubicin, morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubucin, liposomal doxorubicin, deoxydoxorubicin, epirubicin, esorubicin, marcellomycin, mitomycin C, mycophenolic acid, nogalamycin, olivomycins, peplomycin, potfiromycin, puromycin, quelamycin, rodorubicin, streptomigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorubicin, 5-fluorouracil, denopterin, methotrexate, pteropterin, trimetrexate, fludarabine, 6-mercaptopurine, thiamiprine, thiguanine, ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine, calusterone, dromostanolone propionate, epitiostanol, mepitiostane, testolactone, aminoglutethimide, mitotane, trilostane, folinic acid, aceglatone, aldophosphamide glycoside, aminolevulinic acid, eniluracil, amsacrine, bestrabucil, bisantrene, edatraxate, defofamine, demecolcine, diaziquone, elfornithine, elliptinium acetate, etoglucid, gallium nitrate, hydroxyurea, lentinan, lonidainine, maytansine, ansamitocins, mitoguazone, mitoxantrone, mopidanmol, nitraerine, pentostatin, phenamet, pirarubicin, losoxantrone, 2-ethylhydrazide, procarbazine, polysaccharide-k, razoxane, rhizoxin, sizofiran, spirogermanium, tenuazonic acid, triaziquone, 2,2′,2″-trichlorotriethylamine, T-2 toxin, verracurin A, roridin A, and anguidine, urethane, vindesine, dacarbazine, mannomustine, mitobronitol, mitolactol, pipobroman, gacytosine, arabinoside, cyclophosphamide, thiotepa, paclitaxel, albumin-engineered nanoparticle formulation of paclitaxel, doxetaxel, chlorambucil, gemcitabine, 6-thioguanine, mercaptopurine, cisplatin, carboplatin, vinblastine, platinum, etoposide, ifosfamide, mitoxantrone, vincristine, vinorelbine, novantrone, teniposide, edatrexate, daunomycin, aminopterin, xeloda, ibandronate, CPT-11, topoisomerase inhibitor RFS 2000, difluoromethylornithine, retinoic acid, capecitabine, or pharmaceutically acceptable salts, solvates or acids of any of the foregoing;
(b) monokine, a lymphokine, a polypeptide hormone, parathyroid hormone, thyroxine, relaxin, prorelaxin, a glycoprotein hormone, follicle stimulating hormone, thyroid stimulating hormone, luteinizing hormone, hepatic growth factor fibroblast growth factor, prolactin, placental lactogen, tumor necrosis factor-α, tumor necrosis factor-β, mullerian-inhibiting substance, mouse gonadotropin-associated peptide, inhibin, activin, vascular endothelial growth factor, thrombopoietin, erythropoietin, an osteoinductive factor, an interferon, interferon-α, interferon-β, interferon-γ, a colony stimulating factor (“CSF”), macrophage-CSF, granulocyte-macrophage-CSF, granulocyte-CSF, an interleukin (“IL”), IL-1, IL-la, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, a tumor necrosis factor, TNF-α, TNF-β, a polypeptide factor, LIF, kit ligand, or a combination of any of the foregoing;
(c) diphtheria toxin, botulium toxin, tetanus toxin, dysentery toxin, cholera toxin, amanitin, α-amanitin, pyrrolobenzodiazepine, tetrodotoxin, brevetoxin, ciguatoxin, ricin, AM toxin, auristatin, tubulysin, geldanamycin, maytansinoid, calicheamycin, daunomycin, doxorubicin, methotrexate, vindesine, SG2285, dolastatin, auristatin, cryptophycin, camptothecin, rhizoxin, CC-1065, duocarmycin, an enediyne antibiotic, esperamicin, epothilone, a toxoid, or a combination of any of the foregoing;
(d) an affinity ligand, wherein the affinity ligand is a substrate, an inhibitor, a stimulating agent, a neurotransmitter, a radioisotope, or a combination of any of the foregoing;
(e) a radioactive label, 32P, 35S, a fluorescent dye, an electron dense reagent, an enzyme, biotin, streptavidin, dioxigenin, a hapten, an immunogenic protein, a nucleic acid molecule with a sequence complementary to a target, or a combination of any of the foregoing;
(f) an immunomodulatory compound, an anti-cancer agent, an anti-viral agent, an anti-bacterial agent, an anti-fungal agent, and an anti-parasitic agent, or a combination of any of the foregoing;
(g) tamoxifen, raloxifene, droloxifene, 4-hydroxytamoxifen, trioxifene, keoxifene, LY117018, onapristone, or toremifene;
(h) 4(5)-imidazoles, aminoglutethimide, megestrol acetate, exemestane, letrozole, or anastrozole;
(i) flutamide, nilutamide, bicalutamide, leuprolide, goserelin, or troxacitabine;
(j) an aromatase inhibitor;
(k) a protein kinase inhibitor;
(l) a lipid kinase inhibitor;
(m) an antisense oligonucleotide;
(n) a ribozyme;
(o) a vaccine; and
(p) an anti-angiogenic agent.

### Claim 19 (depends on 1)

The antibody-drug conjugate of claim 1, wherein the active agent is amanitin, auristatin, calicheamicin, camptothecin, cryptophycin, daunomycin, dolastatin, doxorubicin, duocarmycin, epothilone, esperamicin, geldanamycin, maytansinoid, methotrexate, monomethyl auristatin E (“MMAE”), monomethyl auristatin F (“MMAF”), pyrrolobenzodiazepine, rhizoxin, SG2285, tubulysin, vindesine, or toxoid.

### Claim 20 (depends on 1)

The antibody-drug conjugate of claim 1, wherein B is:
wherein y is an integer of 1 to 10.

### Claim 21 (depends on 1)

A pharmaceutical composition comprising the antibody-drug conjugate of claim 1.

### Claim 22 (depends on 21)

A method of treating cancer in a subject, comprising administering the pharmaceutical composition of claim 21 to the subject.

### Claim 23 (depends on 8)

The antibody-drug conjugate of claim 8, wherein the linker comprises at least one isoprenyl unit, represented by

### Claim 24 (depends on 9)

The antibody-drug conjugate of claim 9, wherein the linker comprises at least one isoprenyl unit, represented by

### Claim 25 (depends on 11)

The antibody-drug conjugate of claim 11, wherein the linker comprises at least one isoprenyl unit, represented by

### Claim 26 (depends on 1)

The antibody-drug conjugate of claim 1, wherein L satisfies at least one of the following (i) to (iii):
(i) the alkylene includes at least one unsaturated bond,
(ii) the alkylene includes at least one heteroarylene,
(iii) the alkylene is substituted with one or more alkyls having 1 to 20 carbon atoms.