# US 12,698,329 B2 · T cell recruiting polypeptides capable of binding CD123 and TCR α/β

> Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt

- Number: US 12,698,329 B2
- Document ID: US12698329B2
- Status: Granted
- Kind: B2
- Application number: 18/486,205
- Filed: 2023-10-13
- Published: 2024-06-20
- Granted: 2026-08-04
- Earliest claimed: 2016-11-16
- Assignees: Ablynx N.V.
- Applicants: Ablynx N.V.
- Inventors: Diane Van Hoorick; Annelies Roobrouck; Catelijne Stortelers; João Vieira; Edward McGowan
- Web page: https://patentgrep.com/patent/US12698329B2

## CPC

- C07K 16/2809: {against the T-cell receptor (TcR)-CD3 complex}
- C07K 16/3061: {Blood cells}
- A61K 39/0011: {Cancer antigens}
- A61K 39/001119: {Receptors for interleukins [IL]}
- A61P 11/06: Antiasthmatics
- A61P 17/02: for treating wounds, ulcers, burns, scars, keloids, or the like
- A61P 19/02: for joint disorders, e.g. arthritis, arthrosis
- A61P 25/00: Drugs for disorders of the nervous system
- 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 35/02: specific for leukemia
- A61P 37/02: Immunomodulators
- A61P 37/04: Immunostimulants
- A61P 37/08: Antiallergic agents (antiasthmatic agents A61P11/06; ophthalmic antiallergics A61P27/14)
- A61P 43/00: Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
- C07K 16/2866: {against receptors for cytokines, lymphokines, interferons}
- C07K 16/468: {Immunoglobulins having two or more different antigen binding sites, e.g. multifunctional antibodies}
- A61K 45/06: Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
- A61K 2039/5158: {Antigen-pulsed cells, e.g. T-cells}
- A61K 2039/572: {cytotoxic response}
- C07K 2317/22: from camelids, e.g. camel, llama or dromedary
- C07K 2317/24: containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
- C07K 2317/31: multispecific
- C07K 2317/565: Complementarity determining region [CDR]
- C07K 2317/569: Single domain, e.g. dAb, sdAb, VHH, VNAR or nanobody®
- C07K 2317/622: Single chain antibody (scFv)
- C07K 2317/94: Stability, e.g. half-life, pH, temperature or enzyme-resistance
- 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

## Abstract

Polypeptides are provided that bind CD123 on a target cell and the constant domain of TCR on a T cell. The polypeptides can be used in methods for treatment of CD123 associated cancers or inflammatory conditions.

## Claims

### Claim 1

A nucleic acid encoding a polypeptide comprising a first immunoglobulin single variable domain (ISV) and a second ISV, wherein the first ISV specifically binds TCR and essentially consists of 4 framework regions (FR1 to FR4, respectively) and 3 complementarity determining regions (CDR1 to CDR3, respectively), in which:
i) CDR1 is chosen from the group consisting of:
a) SEQ ID NO: 181; and
b) amino acid sequences that have 3, 2 or 1 amino acid(s) difference with the amino acid sequence of SEQ ID NO: 181, wherein
at position 2 the D has been changed into A, S, E, or G;
at position 4 the H has been changed into Y;
at position 5 the K has been changed into L;
at position 6 the I has been changed into L;
at position 8 the F has been changed into I or V; and/or
at position 10 the G has been changed into S; and
ii) CDR2 is chosen from the group consisting of:
c) SEQ ID NO: 192; and
d) amino acid sequences that have 3, 2 or 1 amino acid(s) difference with the amino acid sequence of SEQ ID NO: 192, wherein
at position 1 the D has been changed into T or R;
at position 3 the S has been changed into T or A;
at position 5 the G has been changed into S or A;
at position 7 the Q has been changed into D, E, T, A or V;
at position 8 the T has been changed into A or V; and/or
at position 9 the D has been changed into A, Q, N, V or S;
and
iii) CDR3 is chosen from the group consisting of:
e) SEQ ID NO: 218; and
f) amino acid sequences that have 3, 2 or 1 amino acid(s) difference with the amino acid sequence of SEQ ID NO: 218, wherein
at position 1 the F has been changed into Y, L or G;
at position 4 the I has been changed into L;
at position 5 the Y has been changed into W; and/or
at position 8 the D has been changed into N or S;
and wherein the second ISV specifically binds CD123 and essentially consists of 4 framework regions (FR1 to FR4, respectively) and 3 complementarity determining regions (CDR1 to CDR3, respectively), in which:
i) CDR1 is chosen from the group consisting of:
a) SEQ ID NO: 11; and
b) amino acid sequences that have 3, 2 or 1 amino acid(s) difference with the amino acid sequence of SEQ ID NO: 11, wherein
at position 3 the T has been changed into S or P;
at position 6 the I has been changed into S;
at position 7 the N has been changed into D; and/or
at position 8 the D has been changed into V or A;
and
ii) CDR2 is SEQ ID NO: 17;
and
iii) CDR3 is chosen from the group consisting of:
c) SEQ ID NO: 21; and
d) amino acid sequences that have 1 amino acid(s) difference with the amino acid sequence of SEQ ID NO: 21, wherein at position 3 the P has been changed into A.

### Claim 2 (depends on 1)

The nucleic acid according to claim 1, which is in the form of a genetic construct.

### Claim 3 (depends on 1)

An expression vector comprising the nucleic acid according to claim 1.

### Claim 4 (depends on 1)

A host or host cell comprising the nucleic acid according to claim 1.

### Claim 5 (depends on 1)

A method for the production of a polypeptide, said method at least comprising the steps of:
a) expressing, in a suitable host cell or in another suitable expression system, the nucleic acid according to claim 1; optionally followed by
b) isolating and/or purifying the polypeptide.

### Claim 6 (depends on 1)

A composition comprising the nucleic acid according to claim 1.

### Claim 7 (depends on 6)

The composition according to claim 6, which is a pharmaceutical composition.

### Claim 8 (depends on 7)

The composition according to claim 7, which further comprises at least one pharmaceutically acceptable carrier, diluent or excipient and/or adjuvant.

### Claim 9 (depends on 1)

A kit comprising the nucleic acid according to claim 1.

### Claim 10 (depends on 1)

The nucleic acid of claim 1, wherein the first ISV essentially consists of 4 framework regions (FR1 to FR4, respectively) and 3 complementarity determining regions (CDR1 to CDR3, respectively), in which CDR1 is SEQ ID NO: 181, CDR2 is SEQ ID NO: 192, and CDR3 is SEQ ID NO: 218.

### Claim 11 (depends on 1)

The nucleic acid of claim 1, wherein the first ISV is chosen from the group consisting of SEQ ID NOs: 42 and 78-180 and an amino acid sequence having a sequence identity of more than 90% with one of SEQ ID NOs: 42 and 78-180.

### Claim 12 (depends on 1)

The nucleic acid of claim 1, comprising a third ISV, wherein the third ISV specifically binds CD123 and essentially consists of 4 framework regions (FR1 to FR4, respectively) and 3 complementarity determining regions (CDR1 to CDR3, respectively), in which:
i) CDR1 is SEQ ID NO: 16;
and
ii) CDR2 is chosen from the group consisting of:
a) SEQ ID NO: 18; and
b) amino acid sequences that have 3, 2 or 1 amino acid(s) difference with the amino acid sequence of SEQ ID NO: 18, wherein
at position 3 the Y has been changed into W;
at position 6 the N has been changed into S; and/or
at position 10 the Q has been changed into E;
and
iii) CDR3 is chosen from the group consisting of:
c) SEQ ID NO: 23; and
d) amino acid sequences that have 2 or 1 amino acid(s) difference with the amino acid sequence of SEQ ID NO: 23, wherein
at position 4 the E has been changed into R; and/or
at position 5 the T has been changed into D or Y.

### Claim 13 (depends on 1)

The nucleic acid of claim 1, wherein the second ISV essentially consists of 4 framework regions (FR1 to FR4, respectively) and 3 complementarity determining regions (CDR1 to CDR3, respectively), in which CDR1 is SEQ ID NO: 11, CDR2 is SEQ ID NO: 17, and CDR3 is SEQ ID NO: 21.

### Claim 14 (depends on 1)

The nucleic acid of claim 1, wherein the second ISV is chosen from the group consisting of SEQ ID NOs: 1-6 and an amino acid sequence having a sequence identity of more than 90% with one of SEQ ID NOs: 1-6.

### Claim 15 (depends on 1)

The nucleic acid of claim 1, further comprising a third ISV, wherein the third ISV specifically binds CD123, wherein the second ISV binds to an epitope on CD123 that is different from the epitope on CD123 bound by the third ISV.

### Claim 16 (depends on 15)

The nucleic acid of claim 15, wherein the second ISV is chosen from the group consisting of SEQ ID NOs: 1-6 and an amino acid sequence having more than 90% identity with one of SEQ ID NOs: 1-6, and wherein the third ISV is chosen from the group consisting of SEQ ID NOs: 7-10 and an amino acid sequence having more than 90% identity with one of SEQ ID NOs: 7-10.

### Claim 17 (depends on 1)

The nucleic acid of claim 1, wherein each of the first ISV and the second ISV essentially consist of a single domain antibody, a dAb, a Nanobody, a VHH, a humanized VHH, a camelized VH or a VHH which has been obtained by affinity maturation.

### Claim 18 (depends on 1)

The nucleic acid of claim 1, wherein the polypeptide is chosen from the group consisting of SEQ ID NOs: 47, 49, 52, 53, 55, 56 and 58-61 and an amino acid sequence having a sequence identity of more than 90% with one of SEQ ID NOs: 47, 49, 52, 53, 55, 56 and 58-61.

### Claim 19

A nucleic acid encoding a polypeptide comprising a first immunoglobulin single variable domain (ISV) and a second ISV, wherein the first ISV specifically binds TCR and essentially consists of 4 framework regions (FR1 to FR4, respectively) and 3 complementarity determining regions (CDR1 to CDR3, respectively), in which:
i) CDR1 is chosen from the group consisting of:
a) SEQ ID NO: 181; and
b) amino acid sequences that have 1 amino acid difference with the amino acid sequence of SEQ ID NO: 181;
and
ii) CDR2 is chosen from the group consisting of:
c) SEQ ID NO: 192; and
d) amino acid sequences that have 1 amino acid difference with the amino acid sequence of SEQ ID NO: 192;
and
iii) CDR3 is chosen from the group consisting of:
e) SEQ ID NO: 218; and
f) amino acid sequences that have 1 amino acid difference with the amino acid sequence of SEQ ID NO: 218;
and wherein the second ISV specifically binds CD123 and essentially consists of 4 framework regions (FR1 to FR4, respectively) and 3 complementarity determining regions (CDR1 to CDR3, respectively), in which:
i) CDR1 is chosen from the group consisting of:
a) SEQ ID NO: 11; and
b) amino acid sequences that have 1 amino acid difference with the amino acid sequence of SEQ ID NO: 11;
and
ii) CDR2 is chosen from the group consisting of:
c) SEQ ID NO: 17; and
d) amino acid sequences that have 1 amino acid difference with the amino acid sequence of SEQ ID NO: 17;
and
iii) CDR3 is chosen from the group consisting of:
e) SEQ ID NO: 21; and
f) amino acid sequences that have 1 amino acid difference with the amino acid sequence of SEQ ID NO: 21.