US 10,590,182 B2Granted

Binding-triggered transcriptional switches and methods of use thereof

AssigneeThe Regents of the University of California

InventorsWendell A. Lim, Leonardo Morsut, Kole T. Roybal, Justin Thomas Farlow, Satoshi Toda

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Abstract

The present disclosure provides binding-triggered transcriptional switch polypeptides, nucleic acids comprising nucleotide sequences encoding the binding-triggered transcriptional switch polypeptides, and host cells genetically modified with the nucleic acids. The present disclosure also provides chimeric Notch receptor polypeptides, nucleic acids comprising nucleotide sequences encoding the chimeric Notch receptor polypeptides, and host cells transduced and/or genetically modified with the nucleic acids. The present disclosure provides transgenic organisms comprising a nucleic acid encoding a binding triggered transcriptional switch polypeptide and/or a chimeric Notch receptor polypeptide of the present disclosure. Binding triggered transcriptional switch polypeptides and chimeric Notch receptor polypeptides of the present disclosure are useful in a variety of applications, which are also provided.

Claims18

Dependent claims

A method of locally modulating an activity of a cell, the method comprising:

expressing in the cell a synthetic Notch receptor comprising, in N-terminal to C-terminal order:
an extracellular domain comprising a first member of a specific binding pair that is heterologous to the Notch receptor;
a Notch receptor regulatory region comprising Lin-12 Notch repeats A-C, heterodimerization domains HD-N and HD-C, a binding-induced proteolytic cleavage site, and a transmembrane domain;
and
an intracellular domain heterologous to the Notch receptor;
and
contacting the cell with a second member of the specific binding pair, wherein binding of the first member of the specific binding pair to the second member of the specific binding pair induces cleavage of the binding-induced proteolytic cleavage site to activate the intracellular domain, thereby producing an activated intracellular domain, wherein the activated intracellular domain modulates an activity of the cell selected from the group consisting of:
expression of a gene product of the cell, proliferation of the cell, apoptosis of the cell, non-apoptotic death of the cell, differentiation of the cell, dedifferentiation of the cell, migration of the cell, secretion of a molecule from the cell and cellular adhesion of the cell.

5 more elements

A method of modulating an activity of a cell, the method comprising:

contacting the cell with a second member of a first specific binding pair and a second member of a second specific binding pair, wherein the cell expresses:
i) a first synthetic Notch receptor comprising, in N-terminal to C-terminal order:
an extracellular domain comprising a first member of the first specific binding pair that is heterologous to the Notch receptor;
a Notch receptor regulatory region comprising Lin-12 Notch repeats A-C, heterodimerization domains HD-N and HD-C, a binding-induced proteolytic cleavage site, and a transmembrane domain;
and
an intracellular domain heterologous to the Notch receptor;
and
ii) at least a second synthetic Notch receptor comprising, in N-terminal to C-terminal order:
an extracellular domain comprising the first member of the second specific binding pair that is heterologous to the Notch receptor;
a Notch receptor regulatory region comprising Lin-12 Notch repeats A-C,
heterodimerization domains HD-N and HD-C, a binding-induced proteolytic cleavage site, and a transmembrane domain;
and
an intracellular domain heterologous to the Notch receptor;
wherein the intracellular domain of the first synthetic Notch receptor provides a first effector function and the intracellular domain of the second synthetic Notch receptor provides a second effector function that is different from the first effector function when binding of the first and second members of the first and second specific binding pairs induces cleavage of the binding-induced proteolytic cleavages sites to activate the first and second intracellular domains.

11 more elements

A method of modulating an activity of a cell, the method comprising:

contacting the cell with a second member of a first specific binding pair, wherein the cell expresses:
i) a first synthetic Notch receptor comprising, in N-terminal to C-terminal order:
an extracellular domain comprising a first member of the first specific binding pair that is heterologous to the Notch receptor;
a Notch receptor regulatory region comprising Lin-12 Notch repeats A-C, heterodimerization domains HD-N and HD-C, a binding-induced proteolytic cleavage site, and a transmembrane domain;
and
an intracellular domain heterologous to the Notch receptor;
and
ii) at least a second synthetic Notch receptor comprising, in N-terminal to C-terminal order:
an extracellular domain comprising the first member of a second specific binding pair that is heterologous to the Notch receptor;
a Notch receptor regulatory region comprising Lin-12 Notch repeats A-C, heterodimerization domains HD-N and HD-C, a binding-induced proteolytic cleavage site, and a transmembrane domain;
and
an intracellular domain heterologous to the Notch receptor;
wherein the second synthetic Notch receptor is expressed from a nucleotide sequence operably linked to a transcriptional control element that is activated or repressed by the intracellular domain of the first synthetic Notch receptor.

10 more elements

Dependencies are inferred from the claim text.

Facts

Application: 15/543,220Filed 23 Feb 2016 · us provisional application
Priority: 24 Feb 2015
Published: US 2018/0208636 A126 Jul 2018
Granted: 17 Mar 2020
Term ends: Not estimatedThe source records don’t support an estimate.

Classification33 codes

C07K 14/705
From animals; from humans: receptors; Cell surface antigens; Cell surface determinants
A61K 40/11
Characterised by the cell type used: t-cells, e.g. tumour infiltrating lymphocytes [TIL] or regulatory T [Treg] cells; Lymphokine-activated killer…
A61K 40/31
Characterised by the recombinant expression of specific molecules in the cells of the immune system: chimeric antigen receptors [CAR]
A61K 40/32
Characterised by the recombinant expression of specific molecules in the cells of the immune system: t-cell receptors [TCR]

Family

Application62/120,256Us provisional application · filed 24 Feb 2015
Application62/257,153Us provisional application · filed 18 Nov 2015
Application62/269,758Us provisional application · filed 18 Dec 2015
US 2018/0208636 A1Pre-grant publication · published 26 Jul 2018
US 10,590,182 B2This patent

From the US filing history. Foreign family members aren’t listed.