Structure-Based, Rational Design of T Cell Receptors.

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, K D = ∼1 - 5 μM. Beyond the affinity threshold at K D < 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.

Modelling the consequences of a reduction in alcohol consumption among patients with alcohol dependence based on real-life observational data.

BACKGROUND: Most available pharmacotherapies for alcohol-dependent patients target abstinence; however, reduced alcohol consumption may be a more realistic goal. Using randomized clinical trial (RCT) data, a previous microsimulation model evaluated the clinical relevance of reduced consumption in terms of avoided alcohol-attributable events. Using real-life observational data, the current analysis aimed to adapt the model and confirm previous findings about the clinical relevance of reduced alcohol consumption. METHODS: Based on the prospective observational CONTROL study, evaluating daily alcohol consumption among alcohol-dependent patients, the model predicted the probability of drinking any alcohol during a given day. Predicted daily alcohol consumption was simulated in a hypothetical sample of 200,000 patients observed over a year. Individual total alcohol consumption (TAC) and number of heavy drinking days (HDD) were derived. Using published risk equations, probabilities of alcohol-attributable adverse health events (e.g., hospitalizations or death) corresponding to simulated consumptions were computed, and aggregated for categories of patients defined by HDDs and TAC (expressed per 100,000 patient-years). Sensitivity analyses tested model robustness. RESULTS: Shifting from >220 HDDs per year to 120-140 HDDs and shifting from 36,000-39,000 g TAC per year (120-130 g/day) to 15,000-18,000 g TAC per year (50-60 g/day) impacted substantially on the incidence of events (14,588 and 6148 events avoided per 100,000 patient-years, respectively). Results were robust to sensitivity analyses. CONCLUSIONS: This study corroborates the previous microsimulation modeling approach and, using real-life data, confirms RCT-based findings that reduced alcohol consumption is a relevant objective for consideration in alcohol dependence management to improve public health.