# US 11,111,310 B2 · Chimeric polypeptides and methods of altering the membrane localization of the same

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

- Number: US 11,111,310 B2
- Document ID: US11111310B2
- Status: Granted
- Kind: B2
- Application number: 15/938,600
- Filed: 2018-03-28
- Published: 2018-11-08
- Granted: 2021-09-07
- Earliest claimed: 2017-03-28
- Assignees: Cell Design Labs, Inc.
- Applicants: Cell Design Labs, Inc.
- Inventors: Stephen Santoro; Scott Coyle; Levi Rupp; Peter Emtage
- Web page: https://patentgrep.com/patent/US11111310B2

## CPC

- C07K 16/30: from tumour cells
- A61K 38/177: {Receptors; Cell surface antigens; Cell surface determinants}
- A61K 39/00: Medicinal preparations containing antigens or antibodies (materials for immunoassay G01N33/53)
- A61K 39/001112: {CD19 or B4}
- A61K 39/39558: {against tumor tissues, cells, antigens}
- 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 40/4211: {CD19 or B4}
- A61K 40/4215: {Receptors for tumor necrosis factors [TNF], e.g. lymphotoxin receptor [LTR], CD30}
- A61K 48/005: {characterised by an aspect of the 'active' part of the composition delivered, i.e. the nucleic acid delivered}
- A61P 35/00: Antineoplastic agents
- C07K 14/7051: {T-cell receptor (TcR)-CD3 complex}
- C07K 14/70517: {CD8}
- C07K 14/70521: {CD28, CD152}
- C07K 14/70578: {NGF-receptor/TNF-receptor superfamily, e.g. CD27, CD30, CD40, CD95 (NGF-receptor C07K14/71, TNF-receptor C07K14/7151)}
- C07K 14/7153: {for colony-stimulating factors [CSF]}
- C07K 14/721: {Steroid/thyroid hormone superfamily, e.g. GR, EcR, androgen receptor, oestrogen receptor}
- C07K 16/2878: {against the NGF-receptor/TNF-receptor superfamily, e.g. CD27, CD30, CD40, CD95}
- C07K 19/00: Hybrid peptides {, i.e. peptides covalently bound to nucleic acids, or non-covalently bound protein-protein complexes}
- C12N 5/0637: {Immunosuppressive T lymphocytes, e.g. regulatory T cells or Treg}
- C12N 5/0646: {Natural killers cells [NK], NKT cells}
- C12N 15/62: DNA sequences coding for fusion proteins
- C12N 15/63: Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- A61K 2039/5156: {expressing foreign proteins}
- A61K 2039/5158: {Antigen-pulsed cells, e.g. T-cells}
- A61K 2239/11: Antigen recognition domain
- A61K 2239/22: Intracellular domain
- A61K 2239/23: On/off switch
- C07K 2319/02: containing a signal sequence
- C07K 2319/03: containing a transmembrane segment
- C07K 2319/035: containing a signal for targeting to the external surface of a cell, e.g. to the outer membrane of Gram negative bacteria, GPI- anchored eukaryote proteins
- C12N 5/0636: {T lymphocytes}
- 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

Methods and compositions are provided for reversibly localizing proteins to the exterior part of the cell surface. Compositions provided herein can include nucleic acids that encode polypeptides of interest and the ligand binding domain (LBD) of a nuclear hormone receptor. Medical applications are provided, including controlling the toxicity of CAR T cells.

## Claims

### Claim 1

A single-chain chimeric polypeptide, wherein the single-chain chimeric polypeptide comprises:
a transmembrane domain; and
a hormone receptor ligand binding domain,
wherein:
the transmembrane domain and the hormone receptor ligand binding domain directly abut each other or are separated by 1 to 500 amino acids;
the transmembrane domain and the hormone receptor ligand binding domain are not both present in the same endogenous single-chain polypeptide in a mammal; and
wherein the hormone receptor ligand binding domain is a ligand binding domain of a steroid receptor.

### Claim 2 (depends on 1)

A nucleic acid comprising a nucleotide sequence encoding the single-chain chimeric polypeptide of any claim 1.

### Claim 3 (depends on 2)

A vector comprising the nucleic acid of claim 2.

### Claim 4 (depends on 3)

An isolated cell comprising the vector of claim 3.

### Claim 5 (depends on 1)

The single-chain chimeric polypeptide of claim 1, wherein the transmembrane domain is a domain from: a chain of a T cell receptor, β chain of a T cell receptor, ζ chain of a T cell receptor, CD28, CD3ε, CD3δ, CD3γ, CD33, CD37, CD64, CD80, CD45, CD4, CD5, CD8a, CD9, CD16, CD22, CD86, CD134, CD137, or CD154.

### Claim 6 (depends on 1)

The single-chain chimeric polypeptide of claim 1, wherein the hormone receptor ligand binding domain is a ligand binding domain from an estrogen receptor, a progesterone receptor, or an androgen receptor.

### Claim 7 (depends on 6)

The single-chain chimeric polypeptide of claim 6, wherein the hormone receptor ligand binding domain is a ligand binding domain from an estrogen receptor.

### Claim 8 (depends on 1)

The single-chain chimeric polypeptide of claim 1, wherein the transmembrane domain and the hormone receptor ligand binding domain directly abut each other.

### Claim 9 (depends on 1)

The single-chain chimeric polypeptide of claim 1, wherein the transmembrane domain and the hormone receptor ligand binding domain are separated by 1 to 500 amino acids.

### Claim 10 (depends on 9)

The single-chain chimeric polypeptide of claim 9, wherein the transmembrane domain and the hormone receptor ligand binding domain are separated by 1 to 250 amino acids.

### Claim 11 (depends on 10)

The single-chain chimeric polypeptide of claim 10, wherein the transmembrane domain and the hormone receptor ligand binding domain are separated by 1 to 50 amino acids.

### Claim 12

A single-chain chimeric polypeptide, wherein the single-chain chimeric polypeptide comprises:
an extracellular hormone receptor ligand binding domain; and
a transmembrane domain,
wherein:
the extracellular hormone receptor ligand binding domain and the transmembrane domain directly abut each other or are separated by 1 to 700 amino acids;
the transmembrane domain and the extracellular hormone receptor ligand binding domain are not both present in a single endogenous single-chain polypeptide in a mammal; and
wherein the extracellular hormone receptor ligand binding domain is a ligand binding domain of a steroid receptor.

### Claim 13 (depends on 12)

A nucleic acid comprising a nucleotide sequence encoding the single-chain chimeric polypeptide of claim 12.

### Claim 14 (depends on 13)

A vector comprising the nucleic acid of claim 13.

### Claim 15 (depends on 14)

An isolated cell comprising the vector of claim 14.

### Claim 16 (depends on 12)

The single-chain chimeric polypeptide of claim 12, wherein the transmembrane domain is a domain from: a chain of a T cell receptor, β chain of a T cell receptor, ζ chain of a T cell receptor, CD28, CD3ε, CD3δ, CD3γ, CD33, CD37, CD64, CD80, CD45, CD4, CD5, CD8α, CD9, CD16, CD22, CD86, CD134, CD137, or CD154.

### Claim 17 (depends on 12)

The single-chain chimeric polypeptide of claim 12, wherein the extracellular hormone receptor ligand binding domain is a ligand binding domain from an estrogen receptor, a progesterone receptor, or an androgen receptor.

### Claim 18 (depends on 17)

The single-chain chimeric polypeptide of claim 17, wherein the extracellular hormone receptor ligand binding domain is a ligand binding domain from an estrogen receptor.

### Claim 19 (depends on 12)

The single-chain chimeric polypeptide of claim 12, wherein the extracellular hormone receptor ligand binding domain is a ligand binding domain from an estrogen receptor.

### Claim 20 (depends on 19)

The single-chain chimeric polypeptide of claim 19, wherein the extracellular hormone receptor ligand binding domain is a ligand binding domain from a human estrogen receptor.

### Claim 21 (depends on 20)

The single-chain chimeric polypeptide of claim 20, wherein the ligand binding domain of the human estrogen receptor has a wildtype sequence, except that the ligand binding domain comprises one or both of a substitution at amino acid position 521 and a substitution at amino acid position 537, each numbered relative to the full-length wildtype sequence of human hormone receptor.

### Claim 22 (depends on 12)

The single-chain chimeric polypeptide of claim 12, wherein the extracellular hormone receptor ligand binding domain and the transmembrane domain directly abut each other.

### Claim 23 (depends on 22)

The single-chain chimeric polypeptide of claim 22, wherein the extracellular hormone receptor ligand binding domain and the transmembrane domain are separated by 1 to 700 amino acids.

### Claim 24 (depends on 23)

The single-chain chimeric polypeptide of claim 23, wherein the extracellular hormone receptor ligand binding domain and the transmembrane domain are separated by 1 to 500 amino acids.

### Claim 25 (depends on 24)

The single-chain chimeric polypeptide of claim 24, wherein the extracellular hormone receptor ligand binding domain and the transmembrane domain are separated by 1 to 250 amino acids.

### Claim 26 (depends on 25)

The single-chain chimeric polypeptide of claim 25, wherein the extracellular hormone receptor ligand binding domain and the transmembrane domain are separated by 1 to 100 amino acids.

### Claim 27 (depends on 26)

The single-chain chimeric polypeptide of claim 26, wherein the extracellular hormone receptor ligand binding domain and the transmembrane domain are separated by 1 to 50 amino acids.

### Claim 28 (depends on 27)

The single-chain chimeric polypeptide of claim 27, wherein the extracellular hormone receptor ligand binding domain and the transmembrane domain are separated by 1 to 25 amino acids.

### Claim 29 (depends on 12)

The single-chain chimeric polypeptide of claim 12, wherein, when going in the N-terminal to C-terminal direction, the single-chain chimeric polypeptide comprises an extracellular antigen-binding domain, the extracellular hormone receptor ligand binding domain, and the transmembrane domain.

### Claim 30 (depends on 12)

The single-chain chimeric polypeptide of claim 12, wherein, when going in the C-terminal to N-terminal direction, the single-chain chimeric polypeptide comprises an extracellular antigen-binding domain, the extracellular hormone receptor ligand binding domain, and the transmembrane domain.

### Claim 31 (depends on 12)

The single-chain chimeric polypeptide of claim 12, wherein, when going in the N-terminal to C-terminal direction, the single-chain chimeric polypeptide comprises the extracellular hormone receptor ligand binding domain, an extracellular antigen-binding domain, and the transmembrane domain.

### Claim 32 (depends on 12)

The single-chain chimeric polypeptide of claim 12, wherein, when going in the C-terminal to N-terminal direction, the single-chain chimeric polypeptide comprises the extracellular hormone receptor ligand binding domain, an extracellular antigen-binding domain, and the transmembrane domain.