# US 2017/0283497 A1 · Ror1 (ntrkr1) specific chimeric antigen receptors for cancer immunotherapy

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- Number: US 2017/0283497 A1
- Document ID: US20170283497A1
- Status: Published application
- Kind: A1
- Application number: 15/329,932
- Filed: 2015-07-29
- Published: 2017-10-05
- Earliest claimed: 2014-07-29
- Assignees: Cellectis
- Applicants: Cellectis
- Inventors: Cècile SCHIFFER-MANNIOUI
- Web page: https://patentgrep.com/patent/US20170283497A1

## CPC

- C07K 16/2803: {against the immunoglobulin superfamily}
- 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 40/4251: {Kinases, e.g. Raf or Src}
- A61K 48/00: Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
- A61P 35/00: Antineoplastic agents
- A61P 37/00: Drugs for immunological or allergic disorders
- A61P 37/04: Immunostimulants
- C07K 14/7051: {T-cell receptor (TcR)-CD3 complex}
- C07K 14/70517: {CD8}
- C07K 16/30: from tumour cells
- C12N 5/0636: {T lymphocytes}
- C12N 5/10: Cells modified by introduction of foreign genetic material
- C12N 15/62: DNA sequences coding for fusion proteins
- A61K 2039/505: {comprising antibodies}
- A61K 2039/572: {cytotoxic response}
- C07K 19/00: Hybrid peptides {, i.e. peptides covalently bound to nucleic acids, or non-covalently bound protein-protein complexes}
- 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 2319/02: containing a signal sequence
- C07K 2319/03: containing a transmembrane segment
- C07K 2319/33: fusions for targeting to specific cell types, e.g. tissue specific targeting, targeting of a bacterial subspecies
- C12N 2510/00: Genetically modified cells
- C12N 2740/15043: viral genome or elements thereof as genetic vector
- 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
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

The present invention relates to Chimeric Antigen Receptors (CAR) that are recombinant chimeric proteins able to redirect immune cell specificity and reactivity toward selected membrane antigens, and more particularly in which extracellular ligand binding is a scFV derived from a ROR1 monoclonal antibody, conferring specific immunity against ROR1 positive cells. The engineered immune cells endowed with such CARs are particularly suited for treating lymphomas and leukemia, and for solid tumors such as breast, colon, lung, and kidney tumors

## Claims

### Claim at position 1

1-54. (canceled)

### Claim 55

A ROR1 (NTRKR1) specific chimeric antigen receptor (CAR) having one of the polypeptide structure selected from V3, V5 and V1, said structure comprising an extra cellular ligand binding-domain comprising VH and VL from a monoclonal anti-ROR1 antibody, a hinge, a CD8α transmembrane domain and a cytoplasmic domain including a CD3 zeta signaling domain and a co-stimulatory domain from 4-1BB.

### Claim 56 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein the CD8α transmembrane domain shares at least 80% sequence identity with SEQ ID NO.6.

### Claim 57 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein the hinge is a CD8α hinge, a IgG1 hinge or a FcγRIIIα hinge.

### Claim 58 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein the hinge shares at least 80% sequence identity, for the structures V3, V5 and V1, respectively with SEQ ID NO. 4 (CD8α), SEQ ID NO. 5 (IgG1) and SEQ ID NO. 3 (FcγRIIIα).

### Claim 59 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein the polypeptide structure V3 comprises a CD8α hinge with at least 80% sequence identity to the amino acid sequence of SEQ ID NO.4 and a CD8α transmembrane domain with at least 80% sequence identity to the amino acid sequence of SEQ ID NO:6.

### Claim 60 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein the polypeptide structure V5 comprises an IgG1 hinge with at least 80% sequence identity to SEQ ID NO. 5 and a CD8α transmembrane domain with at least 80% sequence identity to SEQ ID NO.6.

### Claim 61 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein the polypeptide structure V1 comprises a FcγRIIIα hinge with at least 80% sequence identity to SEQ ID NO.3 and a CD8α transmembrane domain with at least 80% sequence identity to SEQ ID NO:6

### Claim 62 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein said extra cellular ligand binding-domain comprises:
a variable heavy VH chain comprising CDR regions from a mouse monoclonal antibody H10 of SEQ ID NO. 54 (CDR-H1), SEQ ID NO:55 (CDR-H2) and SEQ ID NO:56 (CDR-H3), and;
a variable light VL chain comprising CDR regions from a mouse monoclonal antibody H10 of SEQ ID NO. 59 (CDR-L1), SEQ ID NO:60 (CDR-L2) and SEQ ID NO:61 (CDR-L3).

### Claim 63 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein said extra cellular ligand binding-domain comprises:
a variable heavy VH chain comprising CDR regions from a mouse monoclonal antibody D10 of SEQ ID NO. 28 (CDR-H1), SEQ ID NO.29 (CDR-H2) and SEQ ID NO.30 (CDR-H3), and;
a variable light VL chain comprising CDR regions from a mouse monoclonal antibody D10 of SEQ ID NO. 33 (CDR-L1), SEQ ID NO:34 (CDR-L2) and SEQ ID NO:35 (CDR-L3).

### Claim 64 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein said extra cellular ligand binding-domain comprises VH and VL chains with at least 80% sequence identity to SEQ ID NO:53 (H10-VH) and SEQ ID NO:58 (H10-VL), respectively.

### Claim 65 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein said extra cellular ligand binding-domain comprises VH and VL chains with at least 80% sequence identity to SEQ ID NO:27 (D10-VH) and SEQ ID NO:32 (D10-VL), respectively.

### Claim 66 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein said extra cellular ligand binding-domain comprises humanized VH and VL chains from a H10 or D10 antibody.

### Claim 67 (depends on 66)

The ROR1 specific CAR according to claim 66, wherein said
VH chain comprises a polypeptide encoded by SEQ ID NO: 57, and
VL chain comprises a polypeptide encoded by SEQ ID NO: 62.

### Claim 68 (depends on 66)

The ROR1 specific CAR according to claim 66, wherein said
VH chain comprises a polypeptide encoded by SEQ ID NO: 31, and
VL chain comprises a polypeptide encoded by SEQ ID NO: 36.

### Claim 69 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein said CAR shares at least 80% sequence identity with SEQ ID NO:117 (H10v3-CAR sequence).

### Claim 70 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein said CAR shares at least 80% sequence identity with SEQ ID NO:93 (D10v3-CAR sequence).

### Claim 71 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein said CAR shares at least 80% sequence identity with SEQ ID NO:95 (D10v5-CAR sequence).

### Claim 72 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein said co-stimulatory domain from 4-1BB shares at least 80% sequence identity with SEQ ID NO.8.

### Claim 73 (depends on 55)

The ROR1 specific CAR according to claim 55, wherein said CD3 zeta signaling domain shares at least 80% sequence identity with SEQ ID NO. 9.

### Claim 74 (depends on 55)

The ROR1 specific CAR according to claim 55, further comprising a signal peptide.

### Claim 75 (depends on 55)

A polynucleotide encoding the CAR according to claim 55.

### Claim 76 (depends on 75)

An expression vector comprising the polynucleotide of claim 75.

### Claim 77 (depends on 55)

An engineered immune cell expressing the ROR1 specific CAR according to claim 55 at a surface of the immune cell.

### Claim 78 (depends on 77)

The engineered immune cell according to claim 77, wherein the immune cell is derived from an inflammatory T-lymphocyte, a cytotoxic T-lymphocyte, a regulatory T-lymphocyte or a helper T-lymphocyte.

### Claim 79 (depends on 77)

A method of treating a disease or condition in a subject comprising administering to the subject a therapeutically effective amount of the engineered immune cell according to claim 77.

### Claim 80 (depends on 77)

The method according to claim 77, wherein the disease or condition is a pre-malignant or malignant cancer characterized by ROR1-expressing cells.

### Claim 81 (depends on 79)

The method according to claim 79, wherein the disease or condition is a haematological cancer, such as leukemia.

### Claim 82 (depends on 81)

The method according to claim 81, wherein the haematological cancer is chronic lymphocytic leukemia (CLL), the Small Lymphocytic Lymphoma (SLL), acute myelogenous leukemia, chronic myelogenous leukemia, myelodysplastic syndrome, Mantle Cell Lymphoma (MCL), or Acute Lymphoblastic Leukemia (ALL) with a t(1;19) chromosome translocation.

### Claim 83 (depends on 79)

The method according to claim 79, wherein the disease or condition is a solid tumor.

### Claim 84 (depends on 77)

The engineered immune cell according to claim 77, wherein said cell comprises a suppressed T-Cell Receptor.

### Claim 85 (depends on 77)

The engineered immune cell according to claim 77, wherein said immune cell is mutated to confer resistance to at least one immune suppressive or chemotherapy drug.