US 2022/0017864 A1 · METHODS OF IDENTIFYING IMMUNOMODULATORY GENES

Abstract

Disclosed herein are methods for identifying immunomodulatory genes. In some embodiments, the method comprises of screening a candidate gene comprising: a) expressing an exogenous cellular receptor, or a functional portion thereof, in a plurality of immune cells; b) introducing into said plurality of immune cells: i. a guiding polynucleic acid, or a nucleic acid encoding said guiding polynucleic acid, wherein said guiding polynucleic acid targets said candidate gene; and ii. an exogenous nuclease, or a nucleic acid encoding said exogenous nuclease; thereby generating a plurality of engineered immune cells comprising a genomic disruption in said candidate gene; c) contacting said plurality of engineered immune cells with a plurality of cells expressing a cognate antigen of said exogenous cellular receptor or a functional portion thereof, thereby performing an in vitro assay; and d) determining a readout of said in vitro assay.

Claims92

Dependent claims

A method of screening a plurality of single candidate genes, said method comprising:

a. expressing an exogenous cellular receptor, or a functional fragment thereof, in a plurality of separate populations of immune cells, wherein each population comprises a plurality of immune cells;
b. introducing into each of said separate populations of immune cells a CRISPR system that comprises:
i. a guide nucleic acid that binds a portion of a single candidate gene, wherein said single candidate gene is different for each of said separate populations of immune cells;
and
ii. an exogenous nuclease, or a nucleic acid encoding said exogenous nuclease;
thereby generating a plurality of separate populations of engineered immune cells that comprise a genomic disruption in said single candidate gene, wherein said genomic disruption that suppresses expression of said single candidate gene;
c. performing an in vitro assay that comprises contacting said plurality of engineered immune cells with a plurality of cells expressing a cognate antigen of said exogenous cellular receptor or said functional fragment thereof in vitro;
and
d. obtaining a readout from said in vitro assay, to thereby determine an effect of said genomic disruption that suppresses expression of said single candidate gene on said plurality of separate populations of engineered immune cells.

6 more elements

Dependencies are inferred from the claim text.

Methods of identifying immunomodulatory genes

Life of this patent

Methods of identifying immunomodulatory genes

Life of this patent

Abstract

Claims92