# US 2020/0399365 A1 · Site-specific antibody-drug conjugates

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- Number: US 2020/0399365 A1
- Document ID: US20200399365A1
- Status: Published application
- Kind: A1
- Application number: 16/684,222
- Filed: 2019-11-14
- Published: 2020-12-24
- Earliest claimed: 2015-04-15
- Assignees: MedImmune, LLC; Adc Therapeutics SA
- Applicants: Medimmune Limited; Adc Therapeutics S.A.
- Inventors: Patricius Hendrikus Cornelis VAN BERKEL; Philip Wilson HOWARD
- Web page: https://patentgrep.com/patent/US20200399365A1

## CPC

- C07K 16/2803: {against the immunoglobulin superfamily}
- A61K 47/68035: {the drug being a pyrrolobenzodiazepine}
- A61K 47/6867: {the tumour determinant being from a cell of a blood cancer}
- A61P 35/02: specific for leukemia
- C07D 487/04: Ortho-condensed systems
- C07K 2317/24: containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
- C07K 2317/53: Hinge
- Y02P 20/55: Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups
- 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)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C07D: HETEROCYCLIC COMPOUNDS (macromolecular compounds C08)
- Y02P: CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS

## Abstract

Site-specific antibody-drug conjugates are described, in particular conjugates comprising pyrrolobenzodiazepines (PBDs) having a labile protecting group in the form of a linker. The site of conjugation, along with modification of the antibody moiety, allows for improved safety and efficacy of the ADC.

## Claims

### Claim at position 1

-143. (canceled)

### Claim 144

A conjugate of formula L-(DL)p, where DL is of formula I or II:
wherein:
L is an antibody (Ab) which binds CD22, and which comprises:
heavy chains comprising an amino acid substitution of each of HC226 and HC229 according to the EU index as set forth in Kabat,
light chains each having an amino acid substitution of the interchain cysteine residue κLC214 or λLC213 according to the EU index as set forth in Kabat, and
heavy chains each retaining the unsubstituted interchain cysteine HC220 according to the EU index as set forth in Kabat;
when there is a double bond present between C2′ and C3′, R12 is selected from the group consisting of:
(ia) C5-10 aryl group, optionally substituted by one or more substituents selected from the group comprising: halo, nitro, cyano, ether, carboxy, ester, C1-7 alkyl, C3-7 heterocyclyl and bis-oxy-C1-3 alkylene;
(ib) C1-5 saturated aliphatic alkyl;
(ic) C3-6 saturated cycloalkyl;
(id)
wherein each of R21, R22 and R23 are independently selected from H, C1-3 saturated alkyl, C2-3 alkenyl, C2-3 alkynyl and cyclopropyl, where the total number of carbon atoms in the R12 group is no more than 5;
(ie)
wherein one of R25a and R25b is H and the other is selected from: phenyl, which phenyl is optionally substituted by a group selected from halo, methyl, methoxy; pyridyl; and thiophenyl; and
(if)
where R24 is selected from: H; C1-3 saturated alkyl; C2-3 alkenyl; C2-3 alkynyl; cyclopropyl; phenyl, which phenyl is optionally substituted by a group selected from halo, methyl, methoxy; pyridyl; and thiophenyl;
when there is a single bond present between C2′ and C3′,
R12 is
where R26a and R26b are independently selected from H, F, C1-4 saturated alkyl, C2-3 alkenyl, which alkyl and alkenyl groups are optionally substituted by a group selected from C1-4 alkyl amido and C1-4 alkyl ester; or, when one of R26a and R26b is H, the other is selected from nitrile and a C1-4 alkyl ester;
R6 and R9 are independently selected from H, R, OH, OR, SH, SR, NH2, NHR, NRR′, nitro, Me3Sn and halo;
where R and R′ are independently selected from optionally substituted C1-12 alkyl, C3-20 heterocyclyl and C5-20 aryl groups;
R7 is selected from H, R, OH, OR, SH, SR, NH2, NHR, NHRR′, nitro, Me3Sn and halo;
R″ is a C3-12 alkylene group, which chain may be interrupted by one or more heteroatoms, e.g. O, S, NRN2 (where RN2 is H or C1-4 alkyl), and/or aromatic rings, e.g. benzene or pyridine;
Y and Y′ are selected from O, S, or NH;
R6′, R7′, R9′ are selected from the same groups as R6, R7 and R9 respectively;
[Formula I]
RL1′ is a linker for connection to the antibody (Ab);
R11a is selected from OH, ORA, where RA is C1-4 alkyl, and SOzM, where z is 2 or 3 and M is a monovalent pharmaceutically acceptable cation;
R20 and R21 either together form a double bond between the nitrogen and carbon atoms to which they are bound or;
R20 is selected from H and RC, where RC is a capping group;
R21 is selected from OH, ORA and SOzM;
when there is a double bond present between C2 and C3, R2 is selected from the group consisting of:
(ia) C5-10 aryl group, optionally substituted by one or more substituents selected from the group comprising: halo, nitro, cyano, ether, carboxy, ester, C1-7 alkyl, C3-7 heterocyclyl and bis-oxy-C1-3 alkylene;
(ib) C1-5 saturated aliphatic alkyl;
(ic) C3-6 saturated cycloalkyl;
(id)
wherein each of R11, R12 and R13 are independently selected from H, C1-3 saturated alkyl, C2-3 alkenyl, C2-3 alkynyl and cyclopropyl, where the total number of carbon atoms in the R2 group is no more than 5;
(ie)
wherein one of R15a and R1b is H and the other is selected from: phenyl, which phenyl is optionally substituted by a group selected from halo, methyl, methoxy; pyridyl; and thiophenyl; and
(if)
where R14 is selected from: H; C1-3 saturated alkyl; C2-3 alkenyl; C2-3 alkynyl; cyclopropyl; phenyl, which phenyl is optionally substituted by a group selected from halo, methyl, methoxy; pyridyl; and thiophenyl;
when there is a single bond present between C2 and C3,
R2 is
where R16a and R16b are independently selected from H, F, C1-4 saturated alkyl, C2-3 alkenyl, which alkyl and alkenyl groups are optionally substituted by a group selected from C1-4 alkyl amido and C1-4 alkyl ester; or, when one of R16a and R16b is H, the other is selected from nitrile and a C1-4 alkyl ester;
[Formula I]
R22 is of formula IIIa, formula IIIb or formula IIIc:
(a)
where A is a C5-7 aryl group, and either
(i) Q1 is a single bond, and Q2 is selected from a single bond and —Z—(CH2)n—, where Z is selected from a single bond, O, S and NH and n is from 1 to 3; or
(ii) Q1 is —CH═CH—, and Q2 is a single bond;
(b)
where;
RC1, RC2 and RC3 are independently selected from H and unsubstituted C1-2 alkyl;
(c)
where Q is selected from O—RL2′, S—RL2′, and NRN—RL2′, and RN is selected from H, methyl and ethyl
X is selected from the group comprising: O—RL2′, CO2—RL2′, CO—RL2′, NH—C(═O)—RL2′, NHNH—RL2′, CONHNH—RL2′,
NRNRL2′, wherein RN is selected from the group comprising H and C1-4 alkyl;
RL2′ is a linker for connection to the antibody (Ab);
R10 and R11 either together form a double bond between the nitrogen and carbon atoms to which they are bound or;
R10 is H and R11 is selected from OH, ORA and SOzM;
R30 and R31 either together form a double bond between the nitrogen and carbon atoms to which they are bound or;
R30 is H and R31 is selected from OH, ORA and SOzM;
[Formula I and II]
wherein the conjugation of the drug moiety to the antibody is at HC220 according to the EU index as set forth in Kabat.

### Claim 145 (depends on 144)

The conjugate of claim 144, wherein R7 is a C1-4 alkyloxy group.

### Claim 146 (depends on 144)

The conjugate of claim 144, wherein Y is O and R″ is C3-7 alkylene.

### Claim 147 (depends on 144)

The conjugate of claim 144, wherein R6 and R9 are H.

### Claim 148 (depends on 144)

The conjugate of claim 144, wherein there is a double bond between C2′ and C3′, and R12 is:
(a) a C5-7 aryl group, which may bear one to three substituent groups selected from methoxy, ethoxy, fluoro, chloro, cyano, bis-oxy-methylene, methyl-piperazinyl, morpholino and methyl-thiophenyl; or
(b) methyl, ethyl or propyl; or
(c) cyclopropyl; or
(d) a group of formula:
wherein the total number of carbon atoms in the R12 group is no more than 4; Or
(e) the group:
or
(f) a group of formula:
wherein R24 is selected from H and methyl.

### Claim 149 (depends on 144)

The conjugate of claim 144, wherein there is a single bond between C2′ and C3′, R12 is
and:
(a) R26a and R26b are both H; or
(b) R26a and R26b are both methyl; or
(c) one of R26a and R26b is H, and the other is selected from C1-4 saturated alkyl, C2-3 alkenyl, which alkyl and alkenyl groups are optionally substituted.
[Formula I]

### Claim 150 (depends on 144)

The conjugate of claim 144, wherein there is a double bond between C2 and C3, and R2 is:
(a) a C5-7 aryl group, which may bear one to three substituent groups selected from methoxy, ethoxy, fluoro, chloro, cyano, bis-oxy-methylene, methyl-piperazinyl, morpholino and methyl-thiophenyl; or
(b) methyl, ethyl or propyl; or
(c) cyclopropyl; or
(d) a group of formula:
wherein the total number of carbon atoms in the R2 group is no more than 4; or
(e) the group:
or
(f) a group of formula:
wherein R14 is selected from H and methyl.

### Claim 151 (depends on 144)

The conjugate of claim 144, wherein there is a single bond between C2 and C3, R2 is
and:
(a) R16a and R16b are both H; or
(b) R16a and R16b are both methyl; or
(c) one of R16a and R16b is H, and the other is selected from C1-4 saturated alkyl, C2-3 alkenyl, which alkyl and alkenyl groups are optionally substituted.

### Claim 152 (depends on 144)

The conjugate of claim 144, wherein R20 is RC, wherein RC is a group:
where the asterisk indicates the point of attachment to the N10 position, G2 is a terminating group, L3 is a covalent bond or a cleavable linker L1, L2 is a covalent bond or together with OC(═O) forms a self-immolative linker.
[Formula II]

### Claim 153 (depends on 144)

The conjugate of claim 144, wherein:
(a) R22 is of formula iiia, A is phenyl, Q1 is a single bond, Q2 is a single bond; or
(b) R22 is of formula iiib, and RC1, RC2 and RC3 are all H; and X is NH—RL2′.

### Claim 154 (depends on 144)

The conjugate of claim 144, wherein R6′, R7′, R9′, and Y′ are the same as R6, R7, R9, and Y.

### Claim 155 (depends on 144)

The conjugate of claim 144, wherein L-RL1′ or L-RL2′ is a group:
where the asterisk indicates the point of attachment to the PBD, Ab is the antibody, L1 is a cleavable linker, A is a connecting group connecting L1 to the antibody, L2 is a covalent bond or together with —OC(═O)— forms a self-immolative linker.

### Claim 156 (depends on 155)

The conjugate of claim 155, wherein L1 comprises a dipeptide and the group —X1—X2— in dipeptide, —NH—X1—X2—CO—, is selected from:
-Phe-Lys-,
-Val-Ala-,
-Val-Lys-,
-Ala-Lys-,
-Val-Cit-.

### Claim 157 (depends on 156)

The conjugate according to claim 156, wherein C(═O)O and L2 together form the group:
where the asterisk indicates the point of attachment to the PBD, the wavy line indicates the point of attachment to the linker L1, Y is NH, O, C(═O)NH or C(═O)O, and n is 0 to 3.

### Claim 158 (depends on 144)

The conjugate of claim 144 of formula

### Claim 159 (depends on 144)

The conjugate of claim 144, wherein the antibody comprises:
a heavy chain comprising the amino acid sequence of SEQ ID NO.110, or fragment thereof, wherein each of the cysteines at positions 109 and 112 in SEQ ID NO: 110, if present, is substituted by an amino acid that is not cysteine;
a light chain comprising,
(i) the amino acid sequence of SEQ ID NO. 150, or fragment thereof, wherein the cysteine at position 105, if present, is substituted by an amino acid that is not cysteine; or
(ii) the amino acid sequence of SEQ ID NO. 160, or fragment thereof, wherein the cysteine at position 102, if present, is substituted by an amino acid that is not cysteine;
wherein the drug moiety is conjugated to the cysteine at position 103 of SEQ ID NO.110.

### Claim 160 (depends on 144)

The conjugate of claim 144, wherein the antibody comprises:
a heavy chain comprising the amino acid sequence of SEQ ID NO.113 or SEQ ID NO.114, and
a light chain comprising the amino acid sequence of SEQ ID NO.151, SEQ ID NO.152, SEQ ID NO.153, SEQ ID NO.161, SEQ ID NO.162, or SEQ ID NO.163;
wherein the drug moiety is conjugated to the cysteine at position 103 of SEQ ID NO.113.

### Claim 161 (depends on 144)

The conjugate of claim 144, wherein the antibody comprises a heavy chain having a substitution of the amino acid at position 234 in the EU index set forth in Kabat and/or a substitution of the residue at position 235 in the EU index set forth in Kabat, optionally wherein the substituted amino acids are replaced by alanine, glycine, valine, or isoleucine.

### Claim 162 (depends on 144)

The conjugate of claim 144, wherein the antibody comprises a VH domain having the amino acid sequence of SEQ ID NO. 1.

### Claim 163 (depends on 162)

The conjugate of claim 162, wherein the antibody further comprises a VL domain having the amino acid sequence of SEQ ID NO. 2.

### Claim 164 (depends on 144)

The conjugate of claim 144, wherein the antibody in an intact antibody, optionally wherein the antibody is humanised, deimmunised or resurfaced.

### Claim 165 (depends on 144)

The conjugate of claim 144, wherein the drug loading (p) of drugs (D) to antibody (Ab) is 2 or 4.

### Claim 166 (depends on 144)

The conjugate of claim 144, for use in therapy.

### Claim 167 (depends on 144)

The conjugate of claim 144, for use in the treatment of a proliferative disease in a subject, optionally wherein the disease is cancer.

### Claim 168 (depends on 144)

A pharmaceutical composition comprising the conjugate of claim 144, and a pharmaceutically acceptable diluent, carrier or excipient, optionally further comprising a therapeutically effective amount of a chemotherapeutic agent.