Methods of isolating T cells and T cell receptors having antigenic specificity for a cancer-specific mutation from peripheral blood
Inventors
Gros, Alena • Rosenberg, Steven A.
Assignees
US Department of Health and Human Services
Publication Number
US-10544392-B2
Publication Date
2020-01-28
Expiration Date
2036-04-29
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Abstract
Disclosed are methods of isolating T cells and TCRs having antigenic specificity for a mutated amino acid sequence encoded by a cancer-specific mutation. Also disclosed are related methods of preparing a population of cells, populations of cells, TCRs, pharmaceutical compositions, and methods of treating or preventing cancer.
Core Innovation
The invention provides methods of isolating T cells and T cell receptors (TCRs) having antigenic specificity for mutated amino acid sequences encoded by cancer-specific mutations, by obtaining these T cells from a patient's peripheral blood mononuclear cells (PBMCs). The methods involve selecting T cells that express programmed cell death 1 (PD-1), enriching these cells, identifying cancer-specific mutations in genes of the patient's cancer cells that encode mutated amino acid sequences, inducing autologous antigen presenting cells (APCs) to present these mutated amino acid sequences, co-culturing the PD-1+ T cells with these APCs, and selecting T cells with antigenic specificity for the mutated amino acid sequences presented in the context of the patient's major histocompatibility complex (MHC) molecules.
A related method comprises isolating nucleotide sequences encoding TCRs from the enriched PD-1+ T cells, introducing these sequences into further PBMC populations to obtain cells expressing the TCRs, co-culturing these cells with autologous APCs presenting the mutated amino acid sequences, and selecting antigen-specific T cells. Additionally, isolated TCRs, polypeptides, proteins, nucleic acids, recombinant expression vectors, host cells, populations of cells, pharmaceutical compositions, and methods of treating or preventing cancer using these components are disclosed.
The problem being addressed arises from difficulties in identifying and isolating T cells and TCRs that specifically recognize cancer antigens, which hinders the successful use of adoptive cell therapy (ACT) for cancer. Existing methods may require invasive techniques like surgery or biopsy to obtain tumor-infiltrating lymphocytes, which are not always feasible. There is a need for improved methods to obtain cancer-reactive T cells and TCRs from more accessible sources such as peripheral blood, which generally contains infrequent cancer antigen-reactive T cells.
Claims Coverage
The patent claims include 16 claims with multiple inventive features focusing on isolated nucleic acids encoding specific TCRs or antigen-binding portions, recombinant vectors, host cells, populations of cells, and pharmaceutical compositions comprising these elements.
Isolated nucleic acids encoding specific TCRs or antigen-binding portions
Nucleic acids comprising nucleotide sequences encoding TCRs or antigen-binding portions that include amino acid sequences of SEQ ID NOs: 5-10 or 13-18, including variants comprising amino acid sequences SEQ ID NOs: 11-12 or 19-20, and optionally including substituted constant regions (SEQ ID NOs: 45 and 46) with specific residue options.
Isolated nucleic acids encoding polypeptides with defined TCR sequences
Nucleic acids encoding polypeptides comprising amino acid sequences of SEQ ID NOs: 5-10 or 13-18, optionally comprising sequences SEQ ID NOs: 11-12 or 19-20, and further including substituted constant regions as described, or sequences SEQ ID NOs: 51-52 or 53-54.
Isolated nucleic acids encoding proteins comprising paired TCR chains
Nucleic acids encoding proteins with a first polypeptide chain comprising SEQ ID NOs: 5-7 or 13-15 and a second polypeptide chain comprising SEQ ID NOs: 8-10 or 16-18, optionally including variable regions SEQ ID NOs: 11-12 or 19-20, and optionally including substituted constant regions SEQ ID NOs: 45 and 46 or full-length chains SEQ ID NOs: 51-52 or 53-54.
Recombinant expression vectors, host cells, and cell populations
Vectors comprising the nucleic acids encoding said TCRs, host cells containing these vectors, and populations of such host cells expressing the TCRs.
Pharmaceutical compositions
Pharmaceutical compositions comprising populations of host cells expressing the nucleic acid-encoded TCRs and a pharmaceutically acceptable carrier.
The patent claims center on isolated nucleic acids encoding specified TCRs or their antigen-binding portions, recombinant vectors and host cells expressing these TCRs, and related pharmaceutical compositions, thus covering compositions and methods for producing TCR-expressing cells for targeted cancer immunotherapy.
Stated Advantages
Obtaining cancer antigen-reactive T cells from peripheral blood, which is more accessible and abundant compared to tumor tissue.
Avoiding invasive techniques like surgery or biopsy required to obtain tumor-infiltrating lymphocytes.
Effectively and efficiently identifying and enriching infrequent cancer antigen-reactive T cells from peripheral blood.
Enabling ACT administration to patients without available tumors for TIL harvest.
Reducing the cost of ACT, making it available to more patients.
Rapidly assessing a large number of mutations restricted by all MHC molecules of the patient simultaneously to identify the full repertoire of mutation-reactive T cells.
Distinguishing immunogenic cancer mutations from silent or non-immunogenic mutations to identify targetable mutated amino acid sequences.
Providing personalized T cells and TCRs specific to patient-unique cancer mutations for cancer treatment or prevention.
Avoiding technical biases and labor-intensive steps associated with traditional cancer antigen identification methods.
Minimizing or eliminating destruction of normal cells while targeting cancer cells, thereby reducing toxicity.
Potential success in treating cancers resistant to chemotherapy, surgery, or radiation.
Documented Applications
Isolating T cells and TCRs having antigenic specificity for mutated amino acid sequences encoded by cancer-specific mutations from peripheral blood.
Preparing populations of cells expressing isolated TCRs or antigen-binding portions thereof for adoptive cell therapy.
Producing recombinant expression vectors and host cells expressing mutation-specific TCRs.
Formulating pharmaceutical compositions comprising TCR-expressing host cells or related materials for cancer treatment or prevention.
Methods of treating or preventing cancer by administering the pharmaceutical compositions or cell populations expressing mutation-specific TCRs.
Screening and identifying mutated neo-epitopes recognized by TCRs or T cells to personalize cancer immunotherapy.
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