Structure-Based, Rational Design of T Cell Receptors

Abstract

Adoptive cell transfer using engineered T cells is emerging as a promising treatment for metastatic melanoma. Such an approach allows one to introduce T cell receptor (TCR) modifications that, while maintaining the specificity for the targeted antigen, can enhance the binding and kinetic parameters for the interaction with peptides (p) bound to major histocompatibility complexes (MHC). Using the well-characterized 2C TCR/SIYR/H-2K(b) structure as a model system, we demonstrated that a binding free energy decomposition based on the MM-GBSA approach provides a detailed and reliable description of the TCR/pMHC interactions at the structural and thermodynamic levels. Starting from this result, we developed a new structure-based approach, to rationally design new TCR sequences, and applied it to the BC1 TCR targeting the HLA-A2 restricted NY-ESO-1157–165 cancer-testis epitope. Fifty-four percent of the designed sequence replacements exhibited improved pMHC binding as compared to the native TCR, with up to 150-fold increase in affinity, while preserving specificity. Genetically engineered CD8+ T cells expressing these modified TCRs showed an improved functional activity compared to those expressing BC1 TCR. We measured maximum levels of activities for TCRs within the upper limit of natural affinity, KD = ∼1 − 5 μM. Beyond the affinity threshold at KD < 1 μM we observed an attenuation in cellular function, in line with the “half-life” model of T cell activation. Our computer-aided protein-engineering approach requires the 3D-structure of the TCR-pMHC complex of interest, which can be obtained from X-ray crystallography. We have also developed a homology modeling-based approach, TCRep 3D, to obtain accurate structural models of any TCR-pMHC complexes when experimental data is not available. Since the accuracy of the models depends on the prediction of the TCR orientation over pMHC, we have complemented the approach with a simplified rigid method to predict this orientation and successfully assessed it using all non-redundant TCR-pMHC crystal structures available. These methods potentially extend the use of our TCR engineering method to entire TCR repertoires for which no X-ray structure is available. We have also performed a steered molecular dynamics study of the unbinding of the TCR-pMHC complex to get a better understanding of how TCRs interact with pMHCs. This entire rational TCR design pipeline is now being used to produce rationally optimized TCRs for adoptive cell therapies of stage IV melanoma.

Immune Response to Cancer Therapy: Mounting an Effective Antitumor Response and Mechanisms of Resistance

“…Antigens (Ag), typically foreign substances of environmental, viral, or bacterial origin, products of somatically altered proteins, or debris from dying (apoptotic) cells are processed and presented by major histocompatibility complex (MHC) on antigen presenting cells, including (but not limited to) dendritic cells, macrophages, and B cells. CD8 + T cells utilize T cell receptors (TCRs) to recognize MHC-presented peptides and subsequently mount an antigen-specific cytolytic attack [8, 9]. In particular, Ag-TCR engagement ultimately leads to activation and proliferation of CD8 + T cells that play a crucial role in autoimmunity, response to pathogens, and tumor suppression [10–13].…”

“…Genetic rearrangement of TCRs during T cell development enables the recognition of a broad spectrum of processed Ag in the adult. Individual T cells vary with respect to precise structure of TCRs, and this variation endows each T cell with a different specificity for a distinct foreign entity [8]. There is growing evidence that solid tumors contain T cells specific to tumor Ags, but have either been rendered tolerant or inhibited from killing tumor cells by expression of suppressive cytokines such as interleukin (IL)-10 [14] and arginase [15], expression of ligands for immune checkpoints molecules [16], downregulation of MHC class I on tumor cells [17], or other immunosuppressive factors expressed within the TME [18].…”

Identification of Rare High-Avidity, Tumor-Reactive CD8+ T Cells by Monomeric TCR–Ligand Off-Rates Measurements on Living Cells

“…1A). We first evaluated this novel approach by direct visualization of the dissociation process on individual human CD8 þ SUP-T1 cells expressing TCRs of increasing affinities for the HLA-A Ã 0201-restricted tumor epitope NY-ESO-1 157-165 (18), using a flow cytometer generating simultaneous high-resolution microscopy imaging (ImageStream X MarkII; Fig. 1B).…”

A generalized framework for computational design and mutational scanning of T-cell receptor binding interfaces

“…Structure-guided design has been used to generate a small number of high affinity TCRs, as well as manipulate binding specificity (Haidar et al, 2009;Malecek et al, 2014;Pierce et al, 2014;Zoete et al, 2013). However, although the TCR-pMHC structural database has grown significantly in recent years, wide-scale application of structure-guided TCR design is hindered by several complexities.…”

“…Not only can interactions be manipulated in a way that more appropriately addresses peptide specificity, affinity increments can in principle be more tightly controlled. Towards these goals, structureguided design has been used to modify a small number of TCRs (Haidar et al, 2009;Malecek et al, 2014;Pierce et al, 2014;Zoete et al, 2013). Recently, we used structure-guided design to engineer variants of the DMF5 TCR, which has been used clinically in immunotherapy for melanoma and continues to serve as a model TCR for improving cancer immunotherapy (Johnson et al, 2009).…”