# US 11,299,546 B2 · FLT3-specific chimeric antigen receptors and methods using same

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- Number: US 11,299,546 B2
- Document ID: US11299546B2
- Status: Granted
- Kind: B2
- Application number: 16/304,552
- Filed: 2017-05-26
- Published: 2019-07-25
- Granted: 2022-04-12
- Earliest claimed: 2016-05-27
- Assignees: The United States of America, as represented by the Secretary, Department of Health and Human Services
- Applicants: The United States of America, as represented by the Secretary, Department of Health and Human Services
- Inventors: Christopher D. Chien; Terry J. Fry
- Web page: https://patentgrep.com/patent/US11299546B2

## CPC

- C07K 16/2863: {against receptors for growth factors, growth regulators}
- A61K 38/00: Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00)
- A61K 39/395: Antibodies (agglutinins A61K38/36 {; as drug carriers A61K47/50}); Immunoglobulins; Immune serum, e.g. antilymphocytic serum
- A61K 40/11: T-cells, e.g. tumour infiltrating lymphocytes [TIL] or regulatory T [Treg] cells; Lymphokine-activated killer [LAK] cells
- A61K 40/31: Chimeric antigen receptors [CAR]
- A61K 40/421: {Immunoglobulin superfamily}
- A61P 35/00: Antineoplastic agents
- C07K 16/2809: {against the T-cell receptor (TcR)-CD3 complex}
- C07K 16/2815: {against CD8}
- G01N 33/57505: of the blood, e.g. leukaemia
- G01N 33/5759: {involving compounds localised on the membrane of tumour or cancer cells}
- A61K 2039/505: {comprising antibodies}
- A61K 2239/31: characterized by the route of administration
- A61K 2239/38: characterised by the dose, timing or administration schedule
- A61K 2239/48: Blood cells, e.g. leukemia or lymphoma
- A61K 2300/00: Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
- C07K 2317/24: containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
- C07K 2317/53: Hinge
- C07K 2317/622: Single chain antibody (scFv)
- C07K 2317/64: comprising a combination of variable region and constant region components
- C07K 2319/03: containing a transmembrane segment
- G01N 2333/715: for cytokines; for lymphokines; for interferons
- 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
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)

## Abstract

An embodiment of the invention provides a chimeric antigen receptor (C AR) comprising an antigen binding domain specific for FLT3, a transmembrane domain, and an intracellular T cell signaling domain. Nucleic acids, recombinant expression vectors, host cells, populations of cells, antibodies, or antigen binding portions thereof, and pharmaceutical compositions relating to the CARs are disclosed. Methods of detecting the presence of a proliferative disorder, e.g., cancer, in a mammal and methods of treating or preventing a proliferative disorder, e.g., cancer, in a mammal are also disclosed.

## Claims

### Claim 1

A chimeric antigen receptor (CAR) comprising an antigen binding domain specific for FLT3, a transmembrane domain, and an intracellular T cell signaling domain, wherein the antigen binding domain comprises a single chain variable fragment (scFv) comprising from N-terminus to C-terminus (a) the heavy chain variable region CDR1, CDR2, and CDR3 sequences of SEQ ID NOS: 6, 8, and 10 and (b) the light chain variable region CDR1, CDR2, and CDR3 sequences of SEQ ID NOS: 14, 16, and 18.

### Claim 2 (depends on 1)

The CAR of claim 1, wherein the antigen binding domain comprises the light chain variable region comprising the sequences of SEQ ID NOS: 13-19.

### Claim 3 (depends on 1)

The CAR of claim 1, wherein the antigen binding domain comprises the heavy chain variable region comprising the sequences of SEQ ID NOS: 5-11.

### Claim 4 (depends on 1)

The CAR of claim 1, wherein the antigen binding domain comprises the linker sequence of SEQ ID NO: 12.

### Claim 5 (depends on 1)

The CAR of claim 1, wherein the antigen binding domain comprises SEQ ID NOS: 5-19.

### Claim 6 (depends on 1)

The CAR of claim 1, wherein the transmembrane domain comprises CD8 amino acid sequence comprising the CD8α hinge sequence of SEQ ID NO: 25 and the transmembrane domain of sequence SEQ ID NO: 26.

### Claim 7 (depends on 1)

The CAR of claim 1, wherein the intracellular T cell signaling domain comprises the 4-1BB amino acid sequence of SEQ ID NO: 27.

### Claim 8 (depends on 1)

The CAR of claim 1, wherein the intracellular T cell signaling domain comprises the CD3 zeta amino acid sequence of SEQ ID NO: 28.

### Claim 9 (depends on 1)

The CAR of claim 1, wherein the CAR further comprises the spacer comprising SEQ ID NOS: 21-24.

### Claim 10 (depends on 1)

The CAR of claim 1, wherein the CAR comprises any one of the sequence of SEQ ID NOS: 1, 2, 29, or 30.

### Claim 11 (depends on 1)

A nucleic acid comprising a nucleotide sequence encoding the CAR of claim 1.

### Claim 12 (depends on 11)

A recombinant expression vector comprising the nucleic acid of claim 11.

### Claim 13 (depends on 12)

An isolated host cell comprising the recombinant expression vector of claim 12.

### Claim 14 (depends on 13)

A population of cells comprising at least one host cell of claim 13.

### Claim 15 (depends on 1)

A pharmaceutical composition comprising the CAR of claim 1, and a pharmaceutically acceptable carrier.

### Claim 16 (depends on 1)

A method of detecting the presence of cancer, comprising:
(a) contacting a sample comprising one or more cells with the CAR of claim 1, thereby forming a complex, and
(b) detecting the complex, wherein detection of the complex is indicative of the presence of cancer.

### Claim 17 (depends on 16)

The method of claim 16, wherein the cancer is pre-B cell precursor acute lymphoblastic leukemia or acute myeloid leukemia.

### Claim 18 (depends on 13)

A method of treating or preventing cancer in a mammal, the method comprising administering to the mammal the host cell of claim 13 or a population thereof in an effective amount to treat or prevent cancer in the mammal, wherein the host cell is a T cell or non-T cell with an immune-effector function, and wherein the cancer is FLT3+.

### Claim 19 (depends on 18)

The method of claim 18, wherein the cancer is pre-B cell precursor acute lymphoblastic leukemia or acute myeloid leukemia.

### Claim 20 (depends on 16)

The method of claim 16, wherein the method further comprises measuring FLT3 expression levels in a biological sample from a mammal; and
determining if the FLT3 expression levels of the biological sample are increased compared to a sample from a control mammal without a proliferative disorder.