Neoantigen-targeted TCR-engineered T cell immunotherapy: current advances and challenges

Abstract

Adoptive cell therapy using T cell receptor-engineered T cells (TCR-T) is a promising approach for cancer therapy with an expectation of no significant side effects. In the human body, mature T cells are armed with an incredible diversity of T cell receptors (TCRs) that theoretically react to the variety of random mutations generated by tumor cells. The outcomes, however, of current clinical trials using TCR-T cell therapies are not very successful especially involving solid tumors. The therapy still faces numerous challenges in the efficient screening of tumor-specific antigens and their cognate TCRs. In this review, we first introduce TCR structure-based antigen recognition and signaling, then describe recent advances in neoantigens and their specific TCR screening technologies, and finally summarize ongoing clinical trials of TCR-T therapies against neoantigens. More importantly, we also present the current challenges of TCR-T cell-based immunotherapies, e.g., the safety of viral vectors, the mismatch of T cell receptor, the impediment of suppressive tumor microenvironment. Finally, we highlight new insights and directions for personalized TCR-T therapy.

Keywords: Neoantigen; Neoantigen-reactive TCRs; TCR-T.

Fig. 1

TCR structure A TCR-CD3 complex and ITAM. Cysteine (S) mediates interchain bridge of disulphide. B The natural TCR complex, CAR and four recombinant TCR complexes

Fig. 2

Neoantigen presentation and regulatory mechanisms in T cell receptor signaling Neoantigen is generated by tumor cell genome mutation, transcribed and translated and cleaved to peptides different from normal self-proteins. Immunogenic neoantigen peptides are bound by MHC molecules (pMHC), and required for recognition by TCR and to initiate immune response. TCR signal is initiated by pMHC recognition of tumor cells or antigen-presenting cells. Then Lck is recruited to TCR-CD3 complex and phosphorylate ITAMs. Zap70 binds to phosphorylated ITAMs and is also phosphorylated itself by Lck. Activated ZAP70 subsequently phosphorylates Lat, which in turn induces the recruitment of adaptor proteins (GRB2, Gads, SLP-76, PLC-γ). Activation of LAT-related effectors results in signal transduction through 3 major signaling pathways. Calmodulin, MAPK and NF-кB signaling pathways. Calmodulin signaling leads to nuclear translocation of NFAT. MAPK signaling leads to actin polymerization and AP-1 activation, a transcription factor of FOS/ JUN complex. NF-кB signaling leads to nuclear translocation of transcription factors of REL and NF-кB

Fig. 3

The workflow of computational prediction and screening pipelines for neoantigens To identify tumor-specific somatic mutations, tumor tissue and normal tissue samples (usually peripheral blood mononuclear cells) are acquired from the patient perform WES/WGS. Additional RNA sequencing provides information on the gene expression of the mutated genes and further confirmation of gene fusion. Peripheral blood cells were used to predict HLA typing performed by RNA-seq or DNA-seq analysis. MHC-peptide binding prediction software predicts peptides presented by MHC. Computational filtering/screening involves three levels: filter 1 is based on RNA expression; filter 2 is based on proteomics mass spectrometry identification; filter 3 is based on database high confidence filtering. By integrating various physical and chemical properties of peptides, computational prediction screening also includes three levels: antigen binding; neoantigen peptide epitope-TCR recognition; immunogenicity calculation to prioritize the predicted peptides and screen out the neoantigens with high confidence that could be recognized by TCR

Fig. 4

Schematic overview and validation of neoantigen and cognate TCR discovery technology. Tumor and/or peripheral blood mononuclear cell (PBMC) derived DNA/RNA are used to perform WES/RNA-seq to identify non-synonymous variants. Through deep learning-based prediction of neoantigen epitopes, select candidate epitopes to synthesize TMG/long peptides. The monocyte-derived APCs should be engineered to promote antigen presentation and T cell activation. Then, immortalized/engineered APCs were loaded with antigen library. When APCs co-cultured with tumor-infiltrating lymphocytes, neoantigen-reactive T cells will be labeled and selected by flow cytometry. The neoantigen-specific TCR are screened by scTCR-seq, and clone candidate TCRs to PBMC derived T cells. Finally, the recognition of neoantigens by T cells is verified by several screening experiments, such as neoepitope tetramers/4-1BB staining, IFN-γ ELISPOT, cytotoxic activity of tumor killing, degranulation. Meanwhile, neoantigen-specific TCRs could be rapid cloned through T cell characterizing by a panel of CXCL13, ENTPD1(CD19) and CD200 etc.