scholarly journals Combining Well-Tempered Metadynamics Simulation and SPR Assays to Characterize the Binding Mechanism of the Universal T-Lymphocyte Tetanus Toxin Epitope TT830-843

2021 ◽  
Vol 2021 ◽  
pp. 1-15
Author(s):  
Artur A. M. L. Brandt ◽  
Rodrigo N. Rodrigues-da-Silva ◽  
Josué C. Lima-Junior ◽  
Carlos R. Alves ◽  
Franklin de Souza-Silva
Keyword(s):  
Molecular Dynamics ◽  
Free Energy ◽  
T Cell ◽  
T Cell Activation ◽  
Tetanus Toxin ◽  
Cell Activation ◽  
Cell Epitope ◽  
In Vitro Assays ◽  
Stable Complex ◽  
Free Energy Surface

Peptide TT830-843 from the tetanus toxin is a universal T-cell epitope. It helps in vaccination and induces T-cell activation. However, the fine molecular interaction between this antigen and the major histocompatibility complex (MHC) remains unknown. Molecular analysis of its interaction with murine MHC (H-2) was proposed to explore its immune response efficiency. Molecular dynamics simulations are important mechanisms for understanding the basis of protein-ligand interactions, and metadynamics is a useful technique for enhancing sampling in molecular dynamics. SPR (surface plasmon resonance) assays were used to validate whether the metadynamics results are in accordance with the experimental results. The peptide TT830-843 unbinding process was simulated, and the free energy surface reconstruction revealed a detailed conformational landscape. The simulation described the exiting path as a stepwise mechanism between progressive detachment states. We pointed out how the terminus regions act as anchors for binding and how the detachment mechanism includes the opening of α-helices to permit the peptide’s central region dissociation. The results indicated the peptide/H-2 receptor encounter occurs within a distance lesser than 27.5 Å, and the encounter can evolve to form a stable complex. SPR assays confirmed the complex peptide/H-2 as a thermodynamically stable system, exhibiting enough free energy to interact with TCR on the antigen-presenting cell surface. Therefore, combining in silico and in vitro assays provided significant evidence to support the peptide/H-2 complex formation.

scholarly journals Introduction of Non-natural Amino Acids Into T-Cell Epitopes to Mitigate Peptide-Specific T-Cell Responses

2021 ◽  
Vol 12 ◽  
Author(s):  
Aurélien Azam ◽  
Sergio Mallart ◽  
Stephane Illiano ◽  
Olivier Duclos ◽  
Catherine Prades ◽  
...  
Keyword(s):  
Amino Acids ◽  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Cell Epitope ◽  
T Cell Response ◽  
T Cell Epitopes ◽  
Anchor Residue ◽  
Hla Dr

Non-natural modifications are widely introduced into peptides to improve their therapeutic efficacy, but their impact on immunogenicity remains largely unknown. As the CD4 T-cell response is a key factor in triggering immunogenicity, we investigated the effect of introducing D-amino acids (Daa), amino isobutyric acid (Aib), N-methylation, Cα-methylation, reduced amide, and peptoid bonds into an immunoprevalent T-cell epitope on binding to a set of HLA-DR molecules, recognition, and priming of human T cells. Modifications are differentially accepted at multiple positions, but are all tolerated in the flanking regions. Introduction of Aib and Daa in the binding core had the most deleterious effect on binding to HLA-DR molecules and T-cell activation. Their introduction at the positions close to the P1 anchor residue abolished T-cell priming, suggesting they might be sufficient to dampen peptide immunogenicity. Other modifications led to variable effects on binding to HLA-DR molecules and T-cell reactivity, but none exhibited an increased ability to stimulate T cells. Altogether, non-natural modifications appear generally to diminish binding to HLA-DR molecules and hence T-cell stimulation. These data might guide the design of therapeutic peptides to make them less immunogenic.

scholarly journals Beta-1,3 Oligoglucans Specifically Bind to Immune Receptor CD28 and May Enhance T Cell Activation

2021 ◽  
Vol 22 (6) ◽  
pp. 3124
Author(s):  
Jeffrey Comer ◽  
Molly Bassette ◽  
Riley Burghart ◽  
Mayme Loyd ◽  
Susumu Ishiguro ◽  
...  
Keyword(s):  
Free Energy ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Binding Free Energy ◽  
Receptor Activation ◽  
Free Energy Calculations ◽  
Mrna Levels ◽  
Immunostimulatory Activity ◽  
Immune Receptor

Beta glucans are known to have immunomodulatory effects that mediated by a variety of mechanisms. In this article, we describe experiments and simulations suggesting that beta-1,3 glucans may promote activation of T cells by a previously unknown mechanism. First, we find that treatment of a T lymphoblast cell line with beta-1,3 oligoglucan significantly increases mRNA levels of T cell activation-associated cytokines, especially in the presence of the agonistic anti-CD3 antibody. This immunostimulatory activity was observed in the absence of dectin-1, a known receptor for beta-1,3 glucans. To clarify the molecular mechanism underlying this activity, we performed a series of molecular dynamics simulations and free-energy calculations to explore the interaction of beta-1,3 oligoglucans with potential immune receptors. While the simulations reveal little association between beta-1,3 oligoglucan and the immune receptor CD3, we find that beta-1,3 oligoglucans bind to CD28 near the region identified as the binding site for its natural ligands CD80 and CD86. Using a rigorous absolute binding free-energy technique, we calculate a dissociation constant in the low millimolar range for binding of 8-mer beta-1,3 oligoglucan to this site on CD28. The simulations show this binding to be specific, as no such association is computed for alpha-1,4 oligoglucan. This study suggests that beta-1,3 glucans bind to CD28 and may stimulate T cell activation collaboratively with T cell receptor activation, thereby stimulating immune function.

scholarly journals Melanoma reactive TCR-modified T cells generated without activation retain a less differentiated phenotype and mediate a superior in vivo response

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Siao-Yi Wang ◽  
Tamson V. Moore ◽  
Annika V. Dalheim ◽  
Gina M. Scurti ◽  
Michael I. Nishimura
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Viral Vectors ◽  
Cell Activation ◽  
Adoptive T Cell Therapy ◽  
In Vitro Assays ◽  
T Cell Therapy

AbstractAdoptive T cell therapy with T cell receptor (TCR)-modified T cells has shown promise in treating metastatic melanoma and other malignancies. However, studies are needed to improve the efficacy and durability of responses of TCR-modified T cells. Standard protocols for generating TCR-modified T cells involve activating T cells through CD3 stimulation to allow for the efficient transfer of tumor-reactive receptors with viral vectors. T cell activation results in terminal differentiation and shortening of telomeres, which are likely suboptimal for therapy. In these studies, we demonstrate efficient T cell transduction with the melanoma-reactive TIL1383I TCR through culturing with interleukin 7 (IL-7) in the absence of CD3 activation. The TIL1383I TCR-modified T cells generated following IL-7 culture were enriched with naïve (TN) and memory stem cell populations (TSCM) while maintaining longer telomere lengths. Furthermore, we demonstrated melanoma-reactivity of TIL1383I TCR-modified cells generated following IL-7 culture using in vitro assays and a superior response in an in vivo melanoma model. These results suggest that utilizing IL-7 to generate TCR-modified T cells in the absence of activation is a feasible strategy to improve adoptive T cell therapies for melanoma and other malignancies.

scholarly journals Globally deimmunized lysostaphin evades human immune surveillance and enables highly efficacious repeat dosing

2020 ◽  
Vol 6 (36) ◽  
pp. eabb9011
Author(s):  
Hongliang Zhao ◽  
Seth A. Brooks ◽  
Susan Eszterhas ◽  
Spencer Heim ◽  
Liang Li ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Immune Surveillance ◽  
Cell Epitope ◽  
Wild Type ◽  
Human T Cell ◽  
Novel Variant ◽  
Repeated Dosing ◽  
Infection Study

There is a critical need for novel therapies to treat methicillin-resistant Staphylococcus aureus (MRSA) and other drug-resistant pathogens, and lysins are among the vanguard of innovative antibiotics under development. Unfortunately, lysins’ own microbial origins can elicit detrimental antidrug antibodies (ADAs) that undermine efficacy and threaten patient safety. To create an enhanced anti-MRSA lysin, a novel variant of lysostaphin was engineered by T cell epitope deletion. This “deimmunized” lysostaphin dampened human T cell activation, mitigated ADA responses in human HLA transgenic mice, and enabled safe and efficacious repeated dosing during a 6-week longitudinal infection study. Furthermore, the deimmunized lysostaphin evaded established anti–wild-type immunity, thereby providing significant anti-MRSA protection for animals that were immune experienced to the wild-type enzyme. Last, the enzyme synergized with daptomycin to clear a stringent model of MRSA endocarditis. By mitigating T cell–driven antidrug immunity, deimmunized lysostaphin may enable safe, repeated dosing to treat refractory MRSA infections.

scholarly journals A Multivalent ICAM-1 Binding Nanoparticle which Inhibits ICAM-1 and LFA-1 Interaction Represents a New Tool for the Investigation of Autoimmune-Mediated Dry Eye

2020 ◽  
Vol 21 (8) ◽  
pp. 2758 ◽  
Author(s):  
Pang-Yu Hsueh ◽  
Yaping Ju ◽  
Adrianna Vega ◽  
Maria C. Edman ◽  
J. Andrew MacKay ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Activation ◽  
Dry Eye ◽  
Cell Activation ◽  
Intercellular Adhesion Molecule ◽  
In Vitro Assays ◽  
Lymphocyte Function ◽  
Loss Of Function ◽  
Mixed Cell

The autoimmune disorder, Sjögren’s syndrome (SS), is characterized by lymphocytic infiltration and loss of function of exocrine glands such as the lacrimal gland (LG) and salivary gland. SS-associated changes in the LG are associated with the development of autoimmune-mediated dry eye disease. We have previously reported the accumulation of intercellular adhesion molecule 1 (ICAM-1) in the LG of Non-Obese Diabetic (NOD) mice, a murine model of autoimmune-mediated dry eye in SS, in both LG acinar cells and infiltrating lymphocytes. ICAM-1 initiates T-cell activation and can trigger T-cell migration through binding to lymphocyte function-associated 1 antigen (LFA). To modulate this interaction, this study introduces a new tool, a multivalent biopolymeric nanoparticle assembled from a diblock elastin-like polypeptide (ELP) using the S48I48 (SI) ELP scaffold fused with a mouse ICAM-1 targeting peptide to form IBP-SI. IBP-SI forms a multivalent, monodisperse nanoparticle with a radius of 21.9 nm. Unlike the parent SI, IBP-SI binds mouse ICAM-1 and is internalized by endocytosis into transfected HeLa cells before it accumulates in lysosomes. In vitro assays measuring lymphocyte adhesion to Tumor Necrosis Factor TNF-α-treated bEnd.3 cells, which express high levels of ICAM-1, show that adhesion is inhibited by IBP-SI but not by SI, with IC50 values of 62.7 μM and 81.2 μM, respectively, in two different assay formats. IBP-SI, but not SI, also blocked T-cell proliferation in a mixed lymphocyte reaction by 74% relative to proliferation in an untreated mixed cell reaction. These data suggest that a biopolymeric nanoparticle with affinity for ICAM-1 can disrupt ICAM-1 and LFA interactions in vitro and may have further utility as an in vivo tool or potential therapeutic.

scholarly journals Ex vivo modelling of PD-1/PD-L1 immune checkpoint blockade under acute, chronic, and exhaustion-like conditions of T-cell stimulation

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Alexander Roberts ◽  
Lindsay Bentley ◽  
Tina Tang ◽  
Fay Stewart ◽  
Chiara Pallini ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Immune Checkpoint ◽  
Inflammatory Cytokine ◽  
Cell Activation ◽  
Ex Vivo ◽  
In Vitro Assays ◽  
The Impact

AbstractBlockade of PD-1/PD-L1 interactions is proving an exciting, durable therapeutic modality in a range of cancers whereby T cells are released from checkpoint inhibition to revive their inherent anti-tumour activity. Here we have studied various ways to model ex vivo T cell function in order to compare the impact of the clinically utilised anti-PD-1 antibody, pembrolizumab (Keytruda) on the activation of human T cells: focussing on the release of pro-inflammatory IFNγ and anti-inflammatory IL-10 to assess functionality. Firstly, we investigated the actions of pembrolizumab in an acute model of T-cell activation with either immature or mature allogeneic dendritic cells (DCs); pembrolizumab enhanced IFNγ and IL-10 release from purified CD4+ T-cells in the majority of donors with a bias towards pro-inflammatory cytokine release. Next, we modelled the impact of pembrolizumab in settings of more chronic T-cell activation. In a 7-day antigen-specific response to EBV peptides, the presence of pembrolizumab resulted in a relatively modest increase in both IFNγ and IL-10 release. Where pembrolizumab was assessed against long-term stimulated CD4+ cells that had up-regulated the exhaustion markers TIM-3 and PD-1, there was a highly effective enhancement of the otherwise exhausted response to allogeneic DCs with respect to IFNγ production. By contrast, the restoration of IL-10 production was considerably more limited. Finally, to assess a direct clinical relevance we investigated the consequence of PD-1/PD-L1 blockade in the disease setting of dissociated cells from lung and colon carcinomas responding to allogeneic DCs: here, pembrolizumab once more enhanced IFNγ production from the majority of tumour preparations whereas, again, the increase in IL-10 release was modest at best. In conclusion, we have shown that the contribution of PD-1—revealed by using a canonical blocking antibody to interrupt its interaction with PD-L1—to the production of an exemplar pro- and anti-inflammatory cytokine, respectively, depends in magnitude and ratio on the particular stimulation setting and activation status of the target T cell. We have identified a number of in vitro assays with response profiles that mimic features of dissociated cell populations from primary tumours thereby indicating these represent disease-relevant functional assays for the screening of immune checkpoint inhibitors in current and future development. Such in vitro assays may also support patient stratification of those likely to respond to immuno-oncology therapies in the wider population.

scholarly journals SARS-CoV-2 variant B.1.1.7 caused HLA-A2+ CD8+ T cell epitope mutations for impaired cellular immune response

2021 ◽  
Author(s):  
Chanchan Xiao ◽  
Lipeng Mao ◽  
Zhigang Wang ◽  
Guodong Zhu ◽  
Lijuan Gao ◽  
...  
Keyword(s):  
T Cell ◽  
Immune Responses ◽  
T Cell Activation ◽  
Cell Activation ◽  
Cell Epitope ◽  
T Cell Responses ◽  
Cd8 T Cell ◽  
T Cell Epitopes ◽  
Cell Responses ◽  
Cellular Immune

The rapid spreading of the newly emerged SARS-CoV-2 variant, B.1.1.7, highlighted the requirements to better understand adaptive immune responses to this virus. Since CD8+ T cell responses play an important role in disease resolution and modulation in COVID-19 patients, it is essential to address whether these newly emerged mutations would result in altered immune responses. Here we evaluated the immune properties of the HLA-A2 restricted CD8+ T cell epitopes containing mutations from B.1.1.7, and furthermore performed a comprehensive analysis of the SARS-CoV-2 specific CD8+ T cell responses from COVID-19 convalescent patients recognizing the ancestral Wuhan strain compared to B.1.1.7. First, most of the predicted CD8+ T cell epitopes showed proper binding with HLA-A2, while epitopes from B.1.1.7 had lower binding capability than those from the ancestral strain. In addition, these peptides could effectively induced the activation and cytotoxicity of CD8+ T cells. Our results further showed that at least two site mutations in B.1.1.7 resulted in a decrease in CD8+ T cell activation and a possible immune evasion, namely A1708D mutation in ORF1ab1707-1716 and I2230T mutation in ORF1ab2230-2238. Our current analysis provides information that contributes to the understanding of SARS-CoV-2-specific CD8+ T cell responses elicited by infection of mutated strains.

scholarly journals Experimental Antiglomerular Basement Membrane GN Induced by a Peptide from Actinomyces

2020 ◽  
Vol 31 (6) ◽  
pp. 1282-1295 ◽  
Author(s):  
Qiu-hua Gu ◽  
Megan Huynh ◽  
Yue Shi ◽  
Xiao-yu Jia ◽  
Jie-jian Luo ◽  
...  
Keyword(s):  
T Cell ◽  
Basement Membrane ◽  
Transgenic Mice ◽  
B Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Molecular Mimicry ◽  
Cell Epitope ◽  
Wky Rats ◽  
Bioinformatic Approach

BackgroundAntiglomerular basement membrane (anti-GBM) disease is associated with HLA-DRB1*1501 (the major predisposing genetic factor in the disease), with α3127–148 as a nephritogenic T and B cell epitope. Although the cause of disease remains unclear, the association of infections with anti-GBM disease has been long suspected.MethodsTo investigate whether microbes might activate autoreactive T and B lymphocytes via molecular mimicry in anti-GBM disease, we used bioinformatic tools, including BLAST, SYFPEITHI, and ABCpred, for peptide searching and epitope prediction. We used sera from patients with anti-GBM disease to assess peptides recognized by antibodies, and immunized WKY rats and a humanized mouse model (HLA-DR15 transgenic mice) with each of the peptide candidates to assess pathogenicity.ResultsOn the basis of the critical motif, the bioinformatic approach identified 36 microbial peptides that mimic human α3127–148. Circulating antibodies in sera from patients with anti-GBM recognized nine of them. One peptide, B7, derived from Actinomyces species, induced proteinuria, linear IgG deposition on the GBM, and crescent formation when injected into WKY rats. The antibodies to B7 also targeted human and rat α3127–148. B7 induced T cell activation from human α3127–148-immunized rats. T cell responses to B7 were detected in rats immunized by Actinomyces lysate proteins or recombinant proteins. We confirmed B7’s pathogenicity in HLA-DR15 transgenic mice that developed kidney injury similar to that observed in α3135–145-immunized mice.ConclusionsSera from patients with anti-GBM disease recognized microbial peptides identified through a bioinformatic approach, and a peptide from Actinomyces induced experimental anti-GBM GN by T and B cell crossreactivity. These studies demonstrate that anti-GBM disease may be initiated by immunization with a microbial peptide.

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