Exploring the pre-immune landscape of antigen-specific T cells.

BACKGROUND: Adaptive immune responses to newly encountered pathogens depend on the mobilization of antigen-specific clonotypes from a vastly diverse pool of naive T cells. Using recent advances in immune repertoire sequencing technologies, models of the immune receptor rearrangement process, and a database of annotated T cell receptor (TCR) sequences with known specificities, we explored the baseline frequencies of T cells specific for defined human leukocyte antigen (HLA) class I-restricted epitopes in healthy individuals. METHODS: We used a database of TCR sequences with known antigen specificities and a probabilistic TCR rearrangement model to estimate the baseline frequencies of TCRs specific to distinct antigens epitopespecificT-cells. We verified our estimates using a publicly available collection of TCR repertoires from healthy individuals. We also interrogated a database of immunogenic and non-immunogenic peptides is used to link baseline T-cell frequencies with epitope immunogenicity. RESULTS: Our findings revealed a high degree of variability in the prevalence of T cells specific for different antigens that could be explained by the physicochemical properties of the corresponding HLA class I-bound peptides. The occurrence of certain rearrangements was influenced by ancestry and HLA class I restriction, and umbilical cord blood samples contained higher frequencies of common pathogen-specific TCRs. We also identified a quantitative link between specific T cell frequencies and the immunogenicity of cognate epitopes presented by defined HLA class I molecules. CONCLUSIONS: Our results suggest that the population frequencies of specific T cells are strikingly non-uniform across epitopes that are known to elicit immune responses. This inference leads to a new definition of epitope immunogenicity based on specific TCR frequencies, which can be estimated with a high degree of accuracy in silico, thereby providing a novel framework to integrate computational and experimental genomics with basic and translational research efforts in the field of T cell immunology.

Self-Delivering RNAi Targeting PD-1 Improves Tumor-Specific T Cell Functionality for Adoptive Cell Therapy of Malignant Melanoma.

Adoptive cell therapy (ACT) is becoming a prominent alternative therapeutic treatment for cancer patients relapsing on traditional therapies. In parallel, antibodies targeting immune checkpoint molecules, such as cytotoxic-T-lymphocyte-associated antigen 4 (CTLA-4) and cell death protein 1 pathway (PD-1), are rapidly being approved for multiple cancer types, including as first line therapy for PD-L1-expressing non-small-cell lung cancer. The combination of ACT and checkpoint blockade could substantially boost the efficacy of ACT. In this study, we generated a novel self-delivering small interfering RNA (siRNA) (sdRNA) that knocked down PD-1 expression on healthy donor T cells as well as patient-derived tumor-infiltrating lymphocytes (TIL). We have developed an alternative chemical modification of RNA backbone for improved stability and increased efficacy. Our results show that T cells treated with sdRNA specific for PD-1 had increased interferon γ (IFN-γ) secreting capacity and that this modality of gene expression interference could be utilized in our rapid expansion protocol for production of TIL for therapy. TIL expanded in the presence of PD-1-specific sdRNA performed with increased functionality against autologous tumor as compared to control TIL. This method of introducing RNAi into T cells to modify the expression of proteins could easily be adopted into any ACT protocol and will lead to the exploration of new combination therapies.

VDJdb: a curated database of T-cell receptor sequences with known antigen specificity.

The ability to decode antigen specificities encapsulated in the sequences of rearranged T-cell receptor (TCR) genes is critical for our understanding of the adaptive immune system and promises significant advances in the field of translational medicine. Recent developments in high-throughput sequencing methods (immune repertoire sequencing technology, or RepSeq) and single-cell RNA sequencing technology have allowed us to obtain huge numbers of TCR sequences from donor samples and link them to T-cell phenotypes. However, our ability to annotate these TCR sequences still lags behind, owing to the enormous diversity of the TCR repertoire and the scarcity of available data on T-cell specificities. In this paper, we present VDJdb, a database that stores and aggregates the results of published T-cell specificity assays and provides a universal platform that couples antigen specificities with TCR sequences. We demonstrate that VDJdb is a versatile instrument for the annotation of TCR repertoire data, enabling a concatenated view of antigen-specific TCR sequence motifs. VDJdb can be accessed at https://vdjdb.cdr3.net and https://github.com/antigenomics/vdjdb-db.

Explaining the eventual transient saturation of climate-carbon cycle feedback.

BACKGROUND: Coupled climate-carbon cycle simulations generally show that climate feedbacks amplify the buildup of CO2 under respective anthropogenic emission. The effect of climate-carbon cycle feedback is characterised by the feedback gain: the relative increase in CO2 increment as compared to uncoupled simulations. According to the results of the recent Coupled Climate-Carbon Cycle Model Intercomparison Project (C4MIP), the gain is expected to increase during the 21st century. This conclusion is not supported by the climate model developed at the A.M. Obukhov Institute of Atmospheric Physics at the Russian Academy of Sciences (IAP RAS CM). The latter model shows an eventual transient saturation of the feedback gain. This saturation is manifested in a change of climate-carbon cycle feedback gain which grows initially, attains a maximum, and then decreases, eventually tending to unity. RESULTS: Numerical experiments with the IAP RAS CM as well as an analysis of the conceptual framework demonstrate that this eventual transient saturation results from the fact that transient climate sensitivity decreases with time. CONCLUSION: One may conclude that the eventual transient saturation of the climate-carbon cycle feedback is a fundamental property of the coupled climate-carbon system that manifests itself on a relevant time scale.

Recognition of alpha-amino acid derivatives by N,N'-dibenzylated S,S-(+)-tetrandrine.

Complexation of free and N-acetylated alpha-amino acid anions (Gly, Ala, Phe) and some structurally related guests by a dicationic cyclophane-type N,N'-dibenzylated chiral derivative of a bisisoquinoline macrocyclic alkaloid S,S-(+)-tetrandrine (DBT) has been studied by (1)H-NMR titrations in D(2)O. In contrast to other macrocyclic hosts like cyclodextrins and calixarenes, DBT shows highest affinity and large enantioselectivity (K(S)/K(R) >/=10) toward smaller N-acetylalanine and binds larger phenylalanine derivatives more weakly and non-selectively. With 1,2,3,4-tetrahydroisoquinoline-3-carboxylate, a rigid analog of phenylalanine, binding again becomes enantioselective with K(S)/K(R)= 3.8. The binding specificity of DBT is rationalized on the basis of molecular mechanics calculations.

Ketones as building blocks for dynamic combinatorial libraries: highly active neuraminidase inhibitors generated via selection pressure of the biological target.

New and potent inhibitors of neuraminidase, a key enzyme in the influenza virus activity, have been discovered in dynamic combinatorial libraries based on ketones and amines as building blocks. Selective synthesis of a number of inhibitors among multiple theoretically possible combinations of building blocks is driven by the presence of the target enzyme.

Target-induced formation of neuraminidase inhibitors from in vitro virtual combinatorial libraries.

Neuraminidase, a key enzyme responsible for influenza virus propagation, has been used as a template for selective synthesis of small subsets of its own inhibitors from theoretically highly diverse dynamic combinatorial libraries. We show that the library building blocks, aldehydes and amines, form significant amounts of the library components resulting from their coupling by reductive amination only in the presence of the enzyme. The target amplifies the best hits at least 120-fold. The dynamic libraries synthesized and screened in such an in vitro virtual mode form the components that possess high inhibitory activity, as confirmed by enzyme assays with independently synthesized individual compounds.

Synthesis and mass spectrometry studies of branched oxime ether libraries. Mapping the substitution motif via linker stability and fragmentation pattern.

The oxime ether chemistry has recently been used as a convenient approach to preparing potentially highly diverse combinatorial libraries. The synthetically easiest way to form the libraries is convergent, i.e., via reaction of a branched scaffold containing two or more aminooxy linker groups, with a variety of carbonyl substituents. We show here that such reactions between aldehydes and ketones of different structure with the scaffolds containing different types of aminooxy groups can lead to the formation of virtually all expected components in the model mixtures 1-3 formed from three scaffolds (7-9) and eight substituents (R(1)-R(8)). One important problem with the branched libraries is that the libraries formed from the more complex scaffolds, such as 11, contain multiple regioisomers. The results of extensive analysis of a variety of library components by mass spectrometry presented here show that the differences in the MS-MS fragmentation energies for different linkers yield regiochemical information essential for identification of individual library components.