# US 2020/0399343 A1 · Improved t cell compositions and methods

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- Number: US 2020/0399343 A1
- Document ID: US20200399343A1
- Status: Published application
- Kind: A1
- Application number: 16/604,690
- Filed: 2018-04-18
- Published: 2020-12-24
- Earliest claimed: 2017-04-19
- Assignees: Allogene Therapeutics, Inc.
- Applicants: Allogene Therapeutics, Inc.
- Inventors: Javier Fernando CHAPARRO RIGGERS; Thomas John VAN BLARCOM; Bijan Andre BOLDAJPOUR
- Web page: https://patentgrep.com/patent/US20200399343A1

## CPC

- C07K 14/7051: {T-cell receptor (TcR)-CD3 complex}
- A61K 40/11: T-cells, e.g. tumour infiltrating lymphocytes [TIL] or regulatory T [Treg] cells; Lymphokine-activated killer [LAK] cells
- A61K 40/22: Immunosuppressive or immunotolerising
- A61K 40/31: Chimeric antigen receptors [CAR]
- A61K 40/418: {Antigens related to induction of tolerance to non-self}
- A61P 37/06: Immunosuppressants, e.g. drugs for graft rejection
- C07K 14/005: from viruses
- C07K 16/2803: {against the immunoglobulin superfamily}
- C07K 16/2878: {against the NGF-receptor/TNF-receptor superfamily, e.g. CD27, CD30, CD40, CD95}
- C12N 5/0636: {T lymphocytes}
- C12N 5/0646: {Natural killers cells [NK], NKT cells}
- C12N 7/00: Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof (preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00)
- A61K 40/50: characterised by the use of allogeneic cells
- A61K 2035/124: {the cells being hematopoietic, bone marrow derived or blood cells}
- 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 2501/50: Cell markers; Cell surface determinants
- C12N 2501/59: Lectins
- C12N 2510/00: Genetically modified cells
- C12N 2710/16122: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2740/16043: 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 provides compositions and methods that downregulate major histocompatibility class I molecule cell surface expression, and uses of such compositions and methods for improving the functional activities of isolated T cells (e.g., gene-modified antigen-specific T cells, such as chimeric antigen receptor T (CAR-T) cells). In particular, the present invention provides methods and compositions for bolstering the therapeutic efficacy of CAR-T cells.

## Claims

### Claim 1

An isolated T cell comprising
(i) a viral protein which decreases cell surface expression level of major histocompatibility complex (MHC) Class I relative to cell surface expression level of MHC Class I of an isolated T cell that does not comprise the viral protein, and
(ii) a chimeric antigen receptor (CAR) comprising an extracellular ligand-binding domain, a transmembrane domain, and an intracellular signaling domain.

### Claim 2 (depends on 1)

The isolated T cell of claim 1, wherein the viral protein is a cytomegalovirus (CMV) protein, adenovirus protein, herpesvirus protein, or human immunodeficiency virus protein.

### Claim 3 (depends on 1, 2)

The isolated T cell of claim 1 or 2, wherein the viral protein is ICP47, K3, K5, E19, US3, US6, US2, U21, Nef, US10, or U21.

### Claim 4 (depends on 1, 2, 3)

The isolated T cell of any one of claims 1 to 3, wherein the viral protein does not significantly decrease cell surface expression level of the CAR relative to cell surface expression level of the CAR of an isolated T cell that comprises the CAR but does not comprise the viral protein.

### Claim 5 (depends on 1, 2, 3, 4)

The isolated T cell of any one of claims 1 to 4, wherein the isolated T cell further comprises an NK cell antagonist.

### Claim 6 (depends on 5)

The isolated T cell of claim 5, wherein the NK cell antagonist is an anti-NK cell inhibitory receptor agonist antibody or an anti-NK cell activating receptor antagonist antibody.

### Claim 7 (depends on 6)

The isolated T cell of claim 6, wherein the anti-NK cell inhibitory receptor antibody comprises a single-chain variable fragment (scFv).

### Claim 8 (depends on 6, 7)

The isolated T cell of claim 6 or 7, wherein the anti-NK cell inhibitory receptor antibody specifically binds to a killer cell immunoglobulin-like receptor (KIR), a CD94-NKG2A/C/E heterodimer, a 2B4 (CD244) receptor, or a Killer cell lectin-like receptor G1 (KLRG1) receptor.

### Claim 9 (depends on 8)

The isolated T cell of claim 8, wherein the KIR is KIR2DL1, KIR2DL2, KIR2DL3, KIR3DL1, KIR3DL2, KIR3DL3, KIR2DL5A, KIR2DL5B, or KIR2DL4.

### Claim 10 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9)

The isolated T cell of any one of claims 1 to 9, wherein the isolated T cell exhibits improved in vivo persistence relative to in vivo persistence of a second isolated T cell, wherein the second isolated T cell comprises all components of the isolated T cell except it does not comprise the viral protein.

### Claim 11 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10)

The isolated T cell of any one of claims 1 to 10, wherein the isolated T cell elicits no or a reduced graft-versus-host disease (GVHD) response in a histoincompatible recipient as compared to the GVHD response elicited by a second isolated T cell, wherein the second isolated T cell comprises all components of the isolated T cell except it does not comprise the viral protein.

### Claim 12 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11)

A CAR-T cell population comprising: a plurality of the isolated T cell of any one of claims 1 to 11.

### Claim 13 (depends on 12)

The CAR-T cell population of claim 12, wherein cell surface expression level of MHC Class I is decreased by at least about 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% relative to cell surface expression level of MHC Class I on T cells that do not comprise the viral protein.

### Claim 14 (depends on 13)

The CAR-T cell population of claim 13, wherein the cell surface expression level of MHC Class I is measured by flow cytometry.

### Claim 15

A method of generating an isolated T cell, wherein the method comprises modifying a T-cell expressing a CAR to express a viral protein, wherein the CAR comprises an extracellular ligand-binding domain, a transmembrane domain, and an intracellular signaling domain.

### Claim 16 (depends on 14, 15)

The method of claim 14 or 15, further comprising modifying the T cell to express an anti-NK cell antagonist.

### Claim 17 (depends on 14, 15, 16)

The method of any one of claims 14 to 16, wherein the viral protein is stably introduced into the cell.

### Claim 18 (depends on 14, 15, 16, 17)

The method of any one of claims 14 to 17, wherein a polynucleotide that encodes the viral protein is introduced to the cell by a transposon/transposase system, a viral-based gene transfer system, or electroporation.

### Claim 19 (depends on 14, 15, 16, 17, 18)

The method of any one of claims 14 to 18, wherein a polynucleotide that encodes the chimeric antigen receptor is introduced to the cell by a transposon/transposase system, electroporation or a viral-based gene transfer system.

### Claim 20 (depends on 19)

The method of claim 19, wherein the viral-based gene transfer system comprises recombinant retrovirus or lentivirus.

### Claim 21 (depends on 16, 17, 18, 19, 20)

The method of any one of claims 16 to 20, wherein a polynucleotide that encodes the NK cell antagonist is introduced to the cell by a transposon/transposase system, by viral-based gene transfer system or by electroporation.

### Claim 22 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11)

A pharmaceutical composition comprising the isolated T cell of any one of claims 1 to 11, for use in treating a disorder.

### Claim 23 (depends on 22)

The pharmaceutical composition of claim 22, wherein the disorder is cancer, autoimmune disease, or infection.

### Claim 24 (depends on 22, 23)

The pharmaceutical composition of claim 22 or 23, wherein the cells are to be provided more than once.

### Claim 25 (depends on 24)

The pharmaceutical composition of claim 24, wherein the cells are to be provided to the subject at least about 1, 2, 3, 4, 5, 6, 7, or more days apart.

### Claim 26 (depends on 25)

The pharmaceutical composition of claim 25, wherein the disorder is a viral disease, a bacterial disease, a cancer, an inflammatory disease, an immune disease, or an aging-associated disease.

### Claim 27 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14)

A method for treating a disorder in a subject, comprising administering the isolated T cell of any one of claims 1 to 14 to the subject.

### Claim 28 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14)

A method of reducing GVHD in a subject, comprising administering the isolated T cell of any one of claims 1 to 14 to the subject.

### Claim 29 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14)

A method of improving persistence in subject, comprising administering the isolated T cell of any one of claims 1 to 14 to the subject.

### Claim 30 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14)

A method of lengthening the time of a durable response in a subject, comprising administering the isolated T cell of any one of claims 1 to 14 to the subject.

### Claim 31 (depends on 27, 28, 29, 30)

The method of any one of claims 27 to 30, wherein the cells are provided to the subject more than once.

### Claim 32 (depends on 27, 28, 29, 30, 31)

The method of any one of claims 27 to 31, wherein the subject has been previously treated with a therapeutic agent prior to administration of the isolated T cell.

### Claim 33 (depends on 32)

The method of claim 32, wherein the therapeutic agent is an antibody or chemotherapeutic agent.

### Claim 34 (depends on 27, 28, 29, 30, 31, 32, 33)

The method of any one of claims 27 to 33, further comprising administering an NK cell antagonist.

### Claim 35 (depends on 34)

The method of claim 34, wherein the NK cell antagonist is an anti-NK cell inhibitory receptor antibody.

### Claim 36 (depends on 35)

The method of claim 35, wherein the anti-NK cell inhibitory receptor antibody is an anti-KIR antibody.

### Claim 37 (depends on 27, 28, 29, 30, 31, 32, 33, 34, 35, 36)

The method of any one of claims 27 to 36, wherein the disorder is a viral disease, a bacterial disease, a cancer, an inflammatory disease, an immune disease, or an aging-associated disease.

### Claim 38 (depends on 37)

The method according to claim 37, wherein the cancer is a hematological malignancy or a solid cancer.

### Claim 39 (depends on 38)

The method according to claim 38, wherein the hematological malignancy is selected from acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic myelogenous leukemia (CML), chronic eosinophilic leukemia (CEL), myelodysplasia syndrome (MDS), non-Hodgkin's lymphoma (NHL), or multiple myeloma (MM).

### Claim 40 (depends on 38)

The method according to claim 38, wherein the solid cancer is selected from biliary cancer, bladder cancer, bone and soft tissue carcinoma, brain tumor, breast cancer, cervical cancer, colon cancer, colorectal adenocarcinoma, colorectal cancer, desmoid tumor, embryonal cancer, endometrial cancer, esophageal cancer, gastric cancer, gastric adenocarcinoma, glioblastoma multiforme, gynecological tumor, head and neck squamous cell carcinoma, hepatic cancer, lung cancer, malignant melanoma, osteosarcoma, ovarian cancer, pancreatic cancer, pancreatic ductal adenocarcinoma, primary astrocytic tumor, primary thyroid cancer, prostate cancer, renal cancer, renal cell carcinoma, rhabdomyosarcoma, skin cancer, soft tissue sarcoma, testicular germ-cell tumor, urothelial cancer, uterine sarcoma, or uterine cancer.

### Claim 41 (depends on 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40)

The method of any one of claims 27 to 40, further comprising administering to the subject one or more additional therapeutic agent(s).

### Claim 42 (depends on 41)

The method of claim 41, wherein the additional therapeutic agent is an antibody or chemotherapeutic agent.

### Claim 43

A polynucleotide encoding (i) a viral protein which decreases cell surface expression level of a major histocompatibility complex (MHC) Class I molecule relative to cell surface expression level of MHC Class I molecule of an isolated T cell that does not comprise the viral protein and (ii) a chimeric antigen receptor (CAR).

### Claim 44 (depends on 43)

A vector comprising the polynucleotide of claim 43.

### Claim 45 (depends on 44)

The vector of claim 44, wherein the vector is a viral vector.