Method of detecting new immunogenic T cell epitopes and isolating new antigen-specific T cell receptors by means of an MHC cell library

Patent metadata

Field	Value
Publication number	US-12215313-B2
Application number	US-202117313761-A
Country	US
Kind code	B2
Filing date	May 6, 2021
Priority date	Mar 16, 2015
Publication date	Feb 4, 2025
Grant date	Feb 4, 2025

Abstract

The present invention relates to the field of immunotherapy, in particular, to adoptive T cell therapy, T cell receptor (TCR) gene therapy and vaccination. The invention provides a method for preparing a nucleic acid encoding the TCR alpha chain construct (TRA) and TCR beta chain construct (TRB) of a TCR construct specific for an epitope from an antigen presented on major histocompatibility complex (MHC), comprising contacting T cells isolated from a donor with a library of artificial antigen presenting cells (APC) comprising cells expressing all MHC I or MHC II alleles present in the donor, preferably, in K562 cells. The TCR construct can be expressed in a T cell, which is useful for adoptive T cell therapy, e.g., of cancer, viral infections or autoimmune diseases. The invention further provides a method for identifying the epitope recognized by said TCR. Immunogenic epitopes recognized by said TCRs can be used to develop vaccine formulations to induce antigen-specific T cell immunity in patients. The invention further provides pairs of two TCR constructs and respective immunogenic epitopes obtained by the method of the invention, wherein the epitopes are from human papillomavirus (HPV) 16 (also designated alphapapillomavirus 9) oncoprotein E5 and human cytomegalovirus (CMV) protein pp65.

First claim

The invention claimed is:

1. A method for preparing a nucleic acid encoding the TRA and TRB of a TCR construct specific for an immunodominant epitope from a defined antigen presented on a MHC, comprising
(a) stimulating T cells isolated from a donor with professional antigen presenting cells presenting epitopes of said defined antigen, to enrich antigen-specific T cells; and
(b) contacting said T cells with a library of cells, wherein each cell expresses a single MHC allele, wherein the library comprises cells expressing all MHC II alleles present in the donor, and wherein the cells of said library present epitopes of said defined antigen; wherein T cells expressing TCR specific for said epitopes become activated T cells; and
(c) selecting T cells activated by said contact in step b, preferably, based on an activation marker expressed by said activated T cells; and
(d) isolating the nucleic acids encoding the TCR alpha and TCR beta chains of the TCR of said activated T cells selected in step c.

2. The method of claim 1 , wherein the library of cells comprises MHC II-expressing K562 cells.

3. The method of claim 1 , wherein the method further comprises optimizing the sequence of the nucleic acid of step d.

4. A method for preparing a T cell expressing a TCR construct specific for an epitope from a defined antigen presented on a MHC, comprising carrying out the method of claim 1 , and expressing said nucleic acids encoding the TRA and TRB in a T cell.

5. A method for identifying an epitope capable of being presented by a MHC in a defined antigen, comprising carrying out steps (a)-(d) of claim 1 and identifying the epitope capable of activating T cells transfected with nucleic acids encoding the isolated TRA and TRB constituting the TCR construct, wherein the epitope is optionally prepared in peptide or nucleic acid form.

6. The method of claim 3 , further comprising optimizing codon usage of the TRA and TRB.

7. The method of claim 3 , further comprising combining human variable regions with murine constant regions or minimal murine constant regions.

8. The method of claim 1 , wherein the professional antigen presenting cells express CD74 and HLA-DM.

Assignees

• Max Delbrueck Centrum Fuer Molekulare Medizin Helmholtz Gemeinschaft
• Helmholtz Zentrum Muenchen Deutsches Forschungszentrum Gesundheit & Umwelt Gmbh
• Zentrum Muenchen Deutsches Forschungszentrum Fuer Gesundheit Und Umwelt Gmbh

Inventors

• Lorenz Felix
• Uckert Wolfgang
• Ellinger Christian
• Schendel Dolores

Classifications

• C12N2710/16134

  Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein · CPC title

• C12N2710/16122

  New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes · CPC title

• C12N2710/20034

  Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein · CPC title

• C12N2710/20022

  New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes · CPC title

• C07K14/005

  from viruses · CPC title