Engineering chimeric human and mouse major histocompatibility complex (MHC) class I tetramers for the production of T-cell receptor (TCR) mimic antibodies

Abstract

Therapeutic monoclonal antibodies targeting cell surface or secreted antigens are among the most effective classes of novel immunotherapies. However, the majority of human proteins and established cancer biomarkers are intracellular. Peptides derived from these intracellular proteins are presented on the cell surface by major histocompatibility complex class I (MHC-I) and can be targeted by a novel class of T-cell receptor mimic (TCRm) antibodies that recognise similar epitopes to T-cell receptors. Humoural immune responses to MHC-I tetramers rarely generate TCRm antibodies and many antibodies recognise the α3 domain of MHC-I and β2 microglobulin (β2m) that are not directly involved in presenting the target peptide. Here we describe the production of functional chimeric human-murine HLA-A2-H2Dd tetramers and modifications that increase their bacterial expression and refolding efficiency. These chimeric tetramers were successfully used to generate TCRm antibodies against two epitopes derived from wild type tumour suppressor p53 (RMPEAAPPV and GLAPPQHLIRV) that have been used in vaccination studies. Immunisation with chimeric tetramers yielded no antibodies recognising the human α3 domain and β2m and generated TCRm antibodies capable of specifically recognising the target peptide/MHC-I complex in fully human tetramers and on the cell surface of peptide pulsed T2 cells. Chimeric tetramers represent novel immunogens for TCRm antibody production and may also improve the yield of tetramers for groups using these reagents to monitor CD8 T-cell immune responses in HLA-A2 transgenic mouse models of immunotherapy.

Comprehensive mutagenesis identifies the peptide repertoire of a p53 T-cell receptor mimic antibody that displays no toxicity in mice transgenic for human HLA-A*0201

“…BSN transcripts were most abundant in NCI-H1930, OCI-Ly3, SUDHL1 (all non-Hodgkin’s lymphoma) and AU65 (breast cancer). Notably, despite being HLA-A2+, NCI-H1930 and SUDHL1 did not bind T1-116C [ 8 ]. UBR3 transcripts were more abundant in cancer cell lines than in normal tissues.…”

“…We have previously reported the production of a TCRm antibody, T1-116C, which targets a wild type p53 peptide (RMPEAAPPV; p53RMP) that is presented by HLA-A*0201 [ 8 ]. Targeting the p53 tumour suppressor is appealing as p53 is widely dysregulated, p53-derived peptides are more abundantly presented by MHC-I on cancer cells and therefore p53-targeting TCRm could have broad therapeutic applications [ 9 ].…”

pMHC Structural Comparisons as a Pivotal Element to Detect and Validate T-Cell Targets for Vaccine Development and Immunotherapy—A New Methodological Proposal

“…Also here, tetramer technology can be applied to give a picture of the real T-cell repertoire in each specific cancer type or even for individual cancers, as well as to select the best immunogenic peptides (the predicted targets that effectively stimulate a T-cell response) to establish a personalized vaccine design to be applied in distinct tumor types. Based on an initial analysis of DNA/RNA sequencing from the individual tumor cells, followed by the inference and modelling of putative immunogenic peptidome and analysis of the structural impact caused by changes in amino acid sequence variants, the synthesis of these peptides (or putative targets) using tetramer technology makes it possible to identify tumor-specific T cells, providing a fingerprint for each tumor type [42]. through the sliding window (In the figure, α, β, δ, and the last nonamer (γ) are represented), which make it possible to model the complete cellular peptidome in a prevalent MHC allele of interest (e.g., HLA-A*02:01).…”

TCR-like antibodies in cancer immunotherapy

“…These include TCR-like antibodies targeting peptide/MHC complexes derived from tumor protein 53 (TP53) [36], macrophage migration inhibitory factor (MIF) [40], proteinase 3 (PR1) [41], and WT1 [15, 44]. In addition to ADCC and CDC effects, the mouse hybridoma-derived TCR-like antibodies can also be utilized therapeutically to detect the expression of peptide/MHC complexes on the tumor cell surface as phage library-derived Fab antibodies [30, 32, 33, 37, 49, 51].…”