scholarly journals Human T cells employ conserved AU-rich elements to fine-tune IFN-γ production

2019 ◽  
Author(s):  
Julian J. Freen-van Heeren ◽  
Branka Popović ◽  
Aurélie Guislain ◽  
Monika C. Wolkers
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Responses ◽  
Activated T Cells ◽  
Cell Responses ◽  
Human T Cells ◽  
Engineered T Cells ◽  
Ifn Γ ◽  
Post Transcriptional Regulation ◽  
Fine Tune

ABSTRACTLong-lasting CD8+ T cell responses are critical in combatting infections and tumors. The pro-inflammatory cytokine IFN-γ is a key effector molecule herein. We recently showed that in murine T cells, the production of IFN-γ is tightly regulated through AU-rich elements (AREs) that are located in the 3’ Untranslated Region (UTR). Loss of AREs resulted in prolonged cytokine production in activated T cells and boosted anti-tumoral T cell responses. Here, we investigated whether these findings can be translated to primary human T cells. Utilizing CRISPR-Cas9 technology, we deleted the ARE region from the IFNG 3’UTR in peripheral blood-derived human T cells. Loss of AREs stabilized the IFNG mRNA in T cells and supported a higher proportion of sustained IFN-γ protein-producing T cells. Importantly, this was also true for tumor antigen-specific T cells. MART-1 TCR engineered T cells that were gene-edited for ARE-deletion showed increased percentages of IFN-γ producing MART-1-specific ARE-Del T cells in response to MART-1 expressing tumor cells. Combined, our study reveals that ARE-mediated post-transcriptional regulation is highly conserved between murine and human T cells. Furthermore, generating antigen-specific ARE-Del T cells is feasible, a feature that could potentially be exploited for therapeutical purposes.

scholarly journals Interaction of Both SH2 Domains of SHP-2 with a PD-1 Homodimer Is Required for PD-1-Mediated Inhibition of T Cell Responses

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 859-859 ◽  
Author(s):  
Nikolaos Patsoukis ◽  
Asia Council ◽  
Anders Berg ◽  
Kankana Bardhan ◽  
Jessica D Weaver ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
T Cell Responses ◽  
Sh2 Domain ◽  
Activated T Cells ◽  
Sh2 Domains ◽  
Cell Responses ◽  
Human T Cells

Abstract Programmed death-1 (PD-1) is a checkpoint receptor expressed on activated T-cells. PD-1 has a key role in maintenance of peripheral tolerance but also restrains anti-viral and anti-tumor immunity. Although PD-1 blockade leads to durable clinical responses in a significant fraction of patients, the majority of patients have only transient responses, emphasizing the need for better understanding of the mechanism of PD-1-mediated T cell inhibition. PD-1 consists of a single N-terminal IgV-like domain, a 20 amino acid stalk separating the IgV domain from the plasma membrane, a transmembrane domain, and a cytoplasmic tail containing two tyrosine-based structural motifs, an immunoreceptor tyrosine-based inhibitory motif (ITIM) and an immunoreceptor tyrosine-based switch motif (ITSM). SHP-2 tyrosine phosphatase interacts with the ITSM and has a critical role in PD-1-mediated inhibition but the precise mechanism is poorly understood. We sought to determine how PD-1: SHP-2 interaction leads to inhibition of T-cell responses. SHP-2 contains two SH2 domains, a phosphatase (PTP) domain and a C-terminus tail (C-tail), forming a structure of N-SH2-C-SH2-PTP-C-tail. We generated five GST-fusion proteins in which GST was fused with either SHP-2 full length, N-SH2, C-SH2 C-SH2-PTP (lacking the N-terminus SH2 domain), or PTP. Pull-down assays using lysates from human T cells revealed that PD-1 interacted with GST-SHP-2 fusion protein only after TCR/CD3-mediated activation with simultaneous PD-1 ligation, and the interaction of PD-1 with SHP-2 was mediated via the SH2 domains of SHP-2. The SH2 domains of SHP-2 have a crucial and distinct role in regulating SHP-2 PTPase activity. In the absence of a tyrosine-phosphorylated ligand, N-SH2 binds the PTP domain leading to an auto-inhibitory closed conformation that blocks the PTP site. Phosphorylation of Y542 in the SHP-2 C-tail leads to an intramolecular interaction of Y542 with the N-SH2 domain that relieves N-SH2 binding to the PTP domain and thereby reverses basal inhibition of the PTPase. Phosphorylation of Y580 in the SHP-2 C-tail relieves the auto-inhibitory closed conformation by interaction with the C-SH2 domain. Subsequent high affinity intermolecular interaction of the N-SH2 with a phosphorylated protein partner completely disrupts its PTP recognition surface, reversing the auto-inhibitory conformation and activating the PTPase activity, whereas the C-SH2 domain contributes to binding energy and specificity. We found that in activated T cells, PD-1-associated SHP-2 was phosphorylated in the tyrosines of the C-tail. To determine whether PD-1 selectively interacts with a specific SH2 domain of SHP-2, we mutagenized the functional sites of N-SH2 and C-SH2 domains at arginines 32 and 138, respectively, to alanine (R32A and R138A) and transfected COS cells with cDNA of SHP-2 wild type or each SH2 mutant together with PD-1 and TCR proximal kinase Fyn, which is required for PD-1 phosphorylation. Immunoprecipitation and immunoblot showed that mutagenesis of either SH2 domain abrogated interaction of SHP-2 with PD-1, indicating that both SH2 domains of SHP-2 are involved in the interaction with PD-1. Surprisingly, each SH2 domain of SHP-2 interacted with tyrosine phosphorylated ITSM of PD-1, as determined by immunoblot with a phopho-PD-1 antibody specific for the phosphorylated tyrosine residue Y245 and by disruption of both N-SH2:PD-1 and C-SH2:PD-1 interaction by mutation of PD-1 ITSM tyrosine residue Y245. These results indicate that SHP-2 brings together two tyrosine phosphorylated PD-1 molecules by interaction with N-SH2 and C-SH2 domains. To determine the functional implications of PD-1 homodimerization, we cultured human T cells in the presence of a soluble dimeric PD-L1 or a monomeric PD-L1. Although dimeric PD-L1 inhibited T cell proliferation and IFN-g production, monomeric PD-L1 had the opposite effect. Our results reveal a previously unidentified mechanism of PD-1: SHP-2 interaction and have implications for the development of PD-1-binding compounds to selectively suppress T cell responses by dimerizing PD-1 or to enhance T cell activation by disrupting PD-1 homodimerization. Our findings open new avenues for the development of selective PD-1-binding compounds in order to augment T cell responses for the induction of antitumor immunity or to suppress aberrant T cell activation in autoimmunity and graft versus host disease. Disclosures No relevant conflicts of interest to declare.

scholarly journals Batf3-Dependent Dendritic Cells Promote Optimal CD8 T Cell Responses against Respiratory Poxvirus Infection

2018 ◽  
Vol 92 (16) ◽  
Author(s):  
Pritesh Desai ◽  
Vikas Tahiliani ◽  
Georges Abboud ◽  
Jessica Stanfield ◽  
Shahram Salek-Ardakani
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Vaccinia Virus ◽  
Respiratory Infection ◽  
Host Resistance ◽  
T Cell Responses ◽  
Cell Responses ◽  
Ifn Γ ◽  
The Impact

ABSTRACTRespiratory infection with vaccinia virus (VacV) elicits robust CD8+T cell responses that play an important role in host resistance. In the lung, VacV encounters multiple tissue-resident antigen-presenting cell (APC) populations, but which cell plays a dominant role in priming of virus-specific CD8+effector T cell responses remains poorly defined. We used Batf3−/−mice to investigate the impact of CD103+and CD8α+dendritic cell (DC) deficiency on anti-VacV CD8+T cell responses. We found that Batf3−/−mice were more susceptible to VacV infection, exhibiting profound weight loss, which correlated with impaired accumulation of gamma interferon (IFN-γ)-producing CD8+T cells in the lungs. This was largely due to defective priming since early in the response, antigen-specific CD8+T cells in the draining lymph nodes of Batf3−/−mice expressed significantly reduced levels of Ki67, CD25, and T-bet. These results underscore a specific role for Batf3-dependent DCs in regulating priming and expansion of effector CD8+T cells necessary for host resistance against acute respiratory VacV infection.IMPORTANCEDuring respiratory infection with vaccinia virus (VacV), a member ofPoxviridaefamily, CD8+T cells play important role in resolving the primary infection. Effector CD8+T cells clear the virus by accumulating in the infected lungs in large numbers and secreting molecules such as IFN-γ that kill virally infected cells. However, precise cell types that regulate the generation of effector CD8+T cells in the lungs are not well defined. Dendritic cells (DCs) are a heterogeneous population of immune cells that are recognized as key initiators and regulators of T-cell-mediated immunity. In this study, we reveal that a specific subset of DCs that are dependent on the transcription factor Batf3 for their development regulate the magnitude of CD8+T cell effector responses in the lungs, thereby providing protection during pulmonary VacV infection.

scholarly journals Peripheral T-Cell Reactivity to Heat Shock Protein 70 and Its Cofactor GrpE from Tropheryma whipplei Is Reduced in Patients with Classical Whipple's Disease

2017 ◽  
Vol 85 (8) ◽  
Author(s):  
Lucia Trotta ◽  
Kathleen Weigt ◽  
Katina Schinnerling ◽  
Anika Geelhaar-Karsch ◽  
Gerrit Oelkers ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Heat Shock ◽  
Heat Shock Protein ◽  
Heat Shock Protein 70 ◽  
T Cell Responses ◽  
Tropheryma Whipplei ◽  
Content Type ◽  
Cell Responses ◽  
Ifn Γ

ABSTRACT Classical Whipple's disease (CWD) is characterized by the lack of specific Th1 response toward Tropheryma whipplei in genetically predisposed individuals. The cofactor GrpE of heat shock protein 70 (Hsp70) from T. whipplei was previously identified as a B-cell antigen. We tested the capacity of Hsp70 and GrpE to elicit specific proinflammatory T-cell responses. Peripheral mononuclear cells from CWD patients and healthy donors were stimulated with T. whipplei lysate or recombinant GrpE or Hsp70 before levels of CD40L, CD69, perforin, granzyme B, CD107a, and gamma interferon (IFN-γ) were determined in T cells by flow cytometry. Upon stimulation with total bacterial lysate or recombinant GrpE or Hsp70 of T. whipplei, the proportions of activated effector CD4+ T cells, determined as CD40L+ IFN-γ+, were significantly lower in patients with CWD than in healthy controls; CD8+ T cells of untreated CWD patients revealed an enhanced activation toward unspecific stimulation and T. whipplei-specific degranulation, although CD69+ IFN-γ+ CD8+ T cells were reduced upon stimulation with T. whipplei lysate and recombinant T. whipplei-derived proteins. Hsp70 and its cofactor GrpE are immunogenic in healthy individuals, eliciting effective responses against T. whipplei to control bacterial spreading. The lack of specific T-cell responses against these T. whipplei-derived proteins may contribute to the pathogenesis of CWD.

scholarly journals Recruitment of highly functional SARS-CoV-2-specific CD8+ T cell receptors mediating cytotoxicity of virus-infected target cells in non-severe COVID-19

2021 ◽  
Author(s):  
Karolin I. Wagner ◽  
Laura M. Mateyka ◽  
Sebastian Jarosch ◽  
Vincent Grass ◽  
Simone Weber ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Responses ◽  
Hla Class I ◽  
Cd8 T Cell ◽  
Target Cells ◽  
Cell Responses ◽  
Cell Immunity ◽  
Engineered T Cells

T cell immunity is crucial for the control of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections and has been widely characterized on a quantitative level. In contrast, the quality of such T cell responses has been poorly investigated, in particular in the case of CD8+ T cells. Here, we explored the quality of SARS-CoV-2-specific CD8+ T cell responses in individuals who recovered from mild symptomatic infections, through which protective immunity should develop, by functional characterization of their T cell receptor (TCR) repertoire. CD8+ T cell responses specific for SARS-CoV-2-derived epitopes were low in frequency but could be detected robustly early as well as late - up to twelve months - after infection. A pool of immunodominant epitopes, which accurately identified previous SARSCoV- 2 infections, was used to isolate TCRs specific for epitopes restricted by common HLA class I molecules. TCR-engineered T cells showed heterogeneous functional avidity and cytotoxicity towards virus-infected target cells. High TCR functionality correlated with gene signatures of T cell function and activation that, remarkably, could be retrieved for each epitope:HLA combination and patient analyzed. Overall, our data demonstrate that highly functional HLA class I TCRs are recruited and maintained upon mild SARS-CoV-2 infection. Such validated epitopes and TCRs could become valuable tools for the development of diagnostic tests determining the quality of SARS-CoV-2-specific CD8+ T cell immunity, and thereby investigating correlates of protection, as well as to restore functional immunity through therapeutic transfer of TCR-engineered T cells.

scholarly journals Modulation of let-7 miRNAs controls the differentiation of effector CD8 T cells

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Alexandria C Wells ◽  
Keith A Daniels ◽  
Constance C Angelou ◽  
Eric Fagerberg ◽  
Amy S Burnside ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Cytotoxic T Lymphocytes ◽  
Cd8 T Cells ◽  
T Cell Responses ◽  
Cd8 T Cell ◽  
Cell Receptor ◽  
Activated T Cells ◽  
Cell Responses

The differentiation of naive CD8 T cells into effector cytotoxic T lymphocytes upon antigen stimulation is necessary for successful antiviral, and antitumor immune responses. Here, using a mouse model, we describe a dual role for the let-7 microRNAs in the regulation of CD8 T cell responses, where maintenance of the naive phenotype in CD8 T cells requires high levels of let-7 expression, while generation of cytotoxic T lymphocytes depends upon T cell receptor-mediated let-7 downregulation. Decrease of let-7 expression in activated T cells enhances clonal expansion and the acquisition of effector function through derepression of the let-7 targets, including Myc and Eomesodermin. Ultimately, we have identified a novel let-7-mediated mechanism, which acts as a molecular brake controlling the magnitude of CD8 T cell responses.

Potential Tolerizing Capacity of Human Dendritic Cells Featuring High Levels of Expression and Activity of the Tryptophan Metabolizing Enzyme Indoleamine 2,3 Dioxygenase.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 623-623
Author(s):  
Andreas Heitger ◽  
Birgit Juergens ◽  
Ursula Hainz ◽  
Dietmar Fuchs
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
T Cell Responses ◽  
Cell Fraction ◽  
Cell Responses ◽  
Indoleamine 2,3 Dioxygenase ◽  
Ifn Γ ◽  
Human Dendritic Cells ◽  
Expression And Activity

Abstract An enhanced tryptophan metabolism mediated by the enzymatic activity of indoleamine 2,3 dioxygenase (IDO) has recently been demonstrated to profoundly affect T cell responses. By the present study we explored whether human dendritic cells (DCs) displaying high IDO expression and activity, down-regulate allogeneic T cell responses. A comparison of lipopolysaccaride (LPS), interferon-γ (IFN-γ), and CD40L as DC maturation agents showed that most abundant IDO expression and activity in DCs was observed when immature DCs were exposed to a combination of LPS and IFN-γ for 48 hours. This time period of maturation was associated with the development of a mature DC phenotype. In contrast, semi-mature DCs, i.e. DCs matured for 4 hours only, were IDO negative. In co-cultures with allogeneic T cells both types of DCs began to metabolize tryptophan, as determined by decreasing concentrations of tryptophan and increasing concentrations of kynurenines in cell culture supernatants, but mature IDO positive DCs did so at a faster rate (complete consumption of tryptophan within 16 hours of co-culture) than semi-mature DCs. A comparison of the allo-stimulatory capacity of semi-mature IDO negative DCs and mature IDO positive DCs showed that at a high DC/T cell ratio (1:1) IDO positive DCs had a significantly reduced capacity to stimulate allogeneic T cells (median 63% reduction, n=5). The reduction of the allogeneic T cell response induced by IDO positive DCs was reversed upon the addition of the IDO inhibitor methylhydantoin-tryptophan to the co-cultures, suggesting an IDO dependent mechanism. Furthermore, allogeneic T cells exposed to IDO positive DCs had an increased rate of apoptosis in the activated cell fraction and after 8 days of co-culture contained a cell fraction (~30%) displaying a CD4+CD25+highFOXP3+ phenotype. These latter cells, when enriched by fluorescent activated cell sorting (FACS), were able to suppress the proliferative response of naive T cells to anti-CD3 mediated stimulation, which indicates the generation of a regulatory T cell population by IDO positive DCs. Together, these findings suggest that the amount of IDO expression and activity by DCs is one feature to govern the type of response of stimulated T cells. Human DCs can be induced to display high levels of IDO expression and activity and, thereby, acquire the ability to effectivley modulate allogeneic T cell responses towards tolerance by eliminating allo-reactive T cells through apoptosis and augmentation of their regulatory rather than their effector potential. Our current approaches address whether this property can be employed to use DCs for the generation of allo-antigen specific tolerance in the setting of hematopoietic cell transplantation.

Ex-Vivo Characterization of Polyclonal Memory CD8+ T-Cell Responses to PRAME-Specific Peptides in Patients with Acute Leukemia and Chronic Myeloid Leukemia

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2910-2910
Author(s):  
Katayoun Rezvani ◽  
Agnes S. M. Yong ◽  
Abdul Tawab ◽  
Behnam Jafarpour ◽  
Rhoda Eniafe ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Myeloid Leukemia ◽  
Ex Vivo ◽  
T Cell Responses ◽  
Cd8 T Cell ◽  
High Avidity ◽  
Cell Responses ◽  
Ifn Γ

Abstract PRAME (Preferentially expressed antigen of melanoma) is aberrantly expressed in hematological malignancies and may be a useful target for immunotherapy in leukemia. We studied CD8+ T-cell responses to four HLA-A*0201-restricted PRAME-derived epitopes (PRA100, PRA142, PRA300, PRA425) in HLA-A*0201-positive patients with acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic myeloid leukemia (CML) and healthy donors, using PRA300/HLA-A*0201 tetramer staining, intracellular cytokine (IC) assay and ex-vivo and cultured ELISPOT analysis. CD8+ T-cells recognizing PRAME peptides were detected directly ex-vivo in 4/10 ALL, 6/10 AML, 3/10 CML patients and 3/10 donors. The frequency of PRAME-specific CD8+ T-cells was greater in patients with AML, CML and ALL than in healthy controls. All peptides were immunogenic in patients, whilst PRA300 was the only immunogenic peptide in donors. High PRAME expression in patient peripheral blood mononuclear cells was associated with responses to two or more PRAME epitopes (4/7 vs. 0/23 in individuals with low PRAME expression, P = 0.001), suggesting a PRAME-driven T-cell response. In 2 patients studied PRA300/HLA-A*0201+ CD8+T-cells were found to be a mixture of effector and central memory phenotypes. To determine the functional avidity of the PRAME T-cell response, the response of CD8+ T-cells to stimulation with 2 concentrations of peptide was measured by IC-IFN-γ staining. High-avidity CD8+ T-cells were defined as those capable of producing IFN-γ in response to the lower concentration of peptide (0.1μM), while low-avidity CD8+ T-cells were those that only produced IFN-γ in response to the higher concentration of peptide (10 μM). Both high and low-avidity CD8+ T-cell responses could be detected for all peptides tested (median 1.05, 0.90, 0.52, 0.40 high/lowavidity ratios for PRA100, PRA142, PRA300 and PRA425 respectively). In patients with high PRAME expression (>0.001 PRAME/ABL) low-avidity CD8+ T-cell responses to PRAME peptides were more prominent than high-avidity responses, suggesting selective deletion of high-avidity T-cells. In contrast, in some patients with levels <0.001 PRAME/ABL, we could detect the presence of high-avidity CD8+ T-cell responses to PRAME. PRAME-specific CD8+ T-cells were further characterized by IC staining for IL-2, IL-4 and IL-10 production and CD107a mobilization (as a marker of cytotoxicity). Following stimulation with the relevant PRAME peptide, there was no significant production of IL-2, IL-4 or IL-10, suggesting a Tc1 effector response but no significant CD107a mobilization was detected despite significant CD107a mobilization in the same patient in response to CMVpp65495. This finding suggests that patients with leukemia have a selective functional impairment of PRAME-specific CD8+ T-cells, consistent with PRAME-specific T cell exhaustion. However, PRAME-specific T-cells were readily expanded in the presence of cytokines in short-term cultures in-vitro to produce IFN-γ, suggesting that it may be possible to improve the functional capacity of PRAME-specific T-cells for therapeutic purposes. These results provide evidence for spontaneous T-cell reactivity against multiple epitopes of PRAME in ALL, AML and CML and support the usefulness of PRAME as a target for immunotherapy in leukemia. The predominance of low-avidity PRAME-specific CD8+ T-cells suggests that achievement of a state of minimal residual disease may be required prior to peptide vaccination to augment T-cell immune surveillance.

The Response to Repeated Vaccination with PR1 and WT1 Peptides Admixed in Montanide Adjuvant Is Transient and Fails to Induce Sustained Anti-Leukemia Responses in Patients with Myeloid Malignancies.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4096-4096
Author(s):  
Katayoun Rezvani ◽  
Agnes S. M. Yong ◽  
Stephan Mielke ◽  
Behnam Jafarpour ◽  
Bipin N. Savani ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cell Response ◽  
T Cell Responses ◽  
Cd8 T Cell ◽  
T Cell Response ◽  
Gm Csf ◽  
Cell Responses ◽  
Ifn Γ

Abstract Abstract 4096 Poster Board III-1031 We previously demonstrated the immunogenicity of a combined vaccine approach employing two leukemia-associated antigenic peptides, PR1 and WT1 (Rezvani Blood 2008). Eight patients with myeloid malignancies received one subcutaneous 0.3 mg and 0.5 mg dose each of PR1 and WT1 vaccines in Montanide adjuvant, with 100 μg of granulocyte-macrophage colony-stimulating factor (GM-CSF). CD8+ T-cell responses against PR1 or WT1 were detected in all patients as early as 1 week post-vaccination. However, responses were only sustained for 3-4 weeks. The emergence of PR1 or WT1-specific CD8+ T-cells was associated with a significant but transient reduction in minimal residual disease (MRD) as assessed by WT1 expression, suggesting a vaccine-induced anti-leukemia response. Conversely, loss of response was associated with reappearance of WT1 transcripts. We hypothesized that maintenance of sustained or at least repetitive responses may require frequent boost injections. We therefore initiated a phase 2 study of repeated vaccination with PR1 and WT1 peptides in patients with myeloid malignancies. Five patients with acute myeloid leukemia (AML) and 2 patients with myelodysplastic syndrome (MDS) were recruited to receive 6 injections at 2 week intervals of PR1 and WT1 in Montanide adjuvant, with GM-CSF as previously described. Six of 7 patients completed 6 courses of vaccination and follow-up as per protocol, to monitor toxicity and immunological responses. Responses to PR1 or WT1 vaccine were detected in all patients after only 1 dose of vaccine. However, additional boosting did not further increase the frequency of PR1 or WT1-specific CD8+ T-cell response. In 4/6 patients the vaccine-induced T-cell response was lost after the fourth dose and in all patients after the sixth dose of vaccine. To determine the functional avidity of the vaccine-induced CD8+ T-cell response, the response of CD8+ T-cells to stimulation with 2 concentrations of PR1 and WT1 peptides (0.1 and 10 μM) was measured by IC-IFN-γ staining. Vaccination led to preferential expansion of low avidity PR1 and WT1 specific CD8+ T-cell responses. Three patients (patients 4, 6 and 7) returned 3 months following the 6th dose of PR1 and WT1 peptide injections to receive a booster vaccine. Prior to vaccination we could not detect the presence of PR1 and WT1 specific CD8+ T-cells by direct ex-vivo tetramer and IC-IFN-γ assay or with 1-week cultured IFN-γ ELISPOT assay, suggesting that vaccination with PR1 and WT1 peptides in Montanide adjuvant does not induce memory CD8+ T-cell responses. This observation is in keeping with recent work in a murine model where the injection of minimal MHC class I binding peptides derived from self-antigens mixed with IFA adjuvant resulted in a transient effector CD8+ T cell response with subsequent deletion of these T cells and failure to induce CD8+ T cell memory (Bijker J Immunol 2007). This observation can be partly explained by the slow release of vaccine peptides from the IFA depot without systemic danger signals, leading to presentation of antigen in non-inflammatory lymph nodes by non-professional antigen presenting cells (APCs). An alternative explanation for the transient vaccine-induced immune response may be the lack of CD4+ T cell help. In summary these data support the immunogenicity of PR1 and WT1 peptide vaccines. However new approaches will be needed to induce long-term memory responses against leukemia antigens. To avoid tolerance induction we plan to eliminate Montanide adjuvant and use GM-CSF alone. Supported by observations that the in vivo survival of CD8+ T-effector cells against viral antigens are improved by CD4+ helper cells, we are currently attempting to induce long-lasting CD8+ T-cell responses to antigen by inducing CD8+ and CD4+ T-cell responses against class I and II epitopes of WT1 and PR1. Disclosures: No relevant conflicts of interest to declare.

scholarly journals The Early Kinetics of Cytomegalovirus-Specific CD8+ T-Cell Responses Are Not Affected by Antigen Load or the Absence of Perforin or Gamma Interferon

2008 ◽  
Vol 82 (10) ◽  
pp. 4931-4937 ◽  
Author(s):  
Daniel M. Andrews ◽  
Christopher E. Andoniou ◽  
Peter Fleming ◽  
Mark J. Smyth ◽  
Mariapia A. Degli-Esposti
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Expansion ◽  
T Cell Responses ◽  
Gamma Interferon ◽  
Murine Cytomegalovirus ◽  
Cell Responses ◽  
Ifn Γ ◽  
T Cell Expansion ◽  
Kinetics Of

ABSTRACT Both innate and adaptive immune responses participate in the control of murine cytomegalovirus (mCMV) infection. In some mouse strains, like BALB/c, the control of infection relies on the activities of CD8+ T cells. mCMV-specific CD8+ T-cell responses are unusual in that, even after mCMV has been controlled in the periphery, the numbers of circulating virus-specific CD8+ T cells remain high compared to those observed in other viral infections. To better understand the generation and maintenance of mCMV-specific CD8+ T-cell responses, we evaluated how antigen load and effector molecules, such as perforin (Prf) and gamma interferon (IFN-γ), influence these responses during acute infection in vivo. Viral burden affected the magnitude, but not the early kinetics, of antigen-specific CD8+ T-cell responses. Similarly, the magnitude of virus-specific CD8+ T-cell expansion was affected by Prf and IFN-γ, but contraction of antigen-specific responses occurred normally in both Prf- and IFN-γ-deficient mice. These data indicate that control of mCMV-specific CD8+ T-cell expansion and contraction is more complex than anticipated and, despite the role of Prf or IFN-γ in controlling viral replication, a full program of T-cell expansion and contraction can occur in their absence.

scholarly journals Asian Elephant T Cell Responses to Elephant Endotheliotropic Herpesvirus

2017 ◽  
Vol 92 (6) ◽  
Author(s):  
Angela Fuery ◽  
Ann M. Leen ◽  
Rongsheng Peng ◽  
Matthew C. Wong ◽  
Hao Liu ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Activity ◽  
T Cell Responses ◽  
T Cell Response ◽  
Asian Elephant ◽  
Hemorrhagic Disease ◽  
Asian Elephants ◽  
Cell Responses ◽  
Ifn Γ

ABSTRACTElephant endotheliotropic herpesvirus (EEHV) can cause lethal hemorrhagic disease in juvenile Asian elephants, an endangered species. One hypothesis to explain this vulnerability of some juvenile elephants is that they fail to mount an effective T cell response to the virus. To our knowledge, there have been no studies of Asian elephant T cell responses to EEHV. To address this deficiency, we validated the gamma interferon (IFN-γ) enzyme-linked immunospot assay for tracking antigen-directed T cell activity by monitoring rabies-specific responses in vaccinated elephants. In addition, we generated monoclonal antibodies to Asian elephant CD4 and CD8 to facilitate phenotypic T cell profiling. Using these tools, we screened healthy elephants with a history of EEHV infection for reactivity against nine EEHV proteins whose counterparts in other herpesviruses are known to induce T cell responses in their natural hosts. We identified glycoprotein B (gB) and the putative regulatory protein E40 as the most immunogenic T cell targets (IFN-γ responses in five of seven elephants), followed by the major capsid protein (IFN-γ responses in three of seven elephants). We also observed that IFN-γ responses were largely from CD4+T cells. We detected no activity against the predicted major immediate early (E44) and large tegument (E34) proteins, both immunodominant T cell targets in humans latently infected with cytomegalovirus. These studies identified EEHV-specific T cells in Asian elephants for the first time, lending insight into the T cell priming that might be required to protect against EEHV disease, and will guide the design of effective vaccine strategies.IMPORTANCEEndangered Asian elephants are facing many threats, including lethal hemorrhagic disease from elephant endotheliotropic herpesvirus (EEHV). EEHV usually establishes chronic, benign infections in mature Asian elephants but can be lethal to juvenile elephants in captivity and the wild. It is the leading cause of death in captive Asian elephants in North America and Europe. Despite the availability of sensitive tests and protocols for treating EEHV-associated illness, these measures are not always effective. The best line of defense would be a preventative vaccine. We interrogated normal healthy elephants previously infected with EEHV for T cell responses to nine EEHV proteins predicted to induce cellular immune responses. Three proteins elicited IFN-γ responses, suggesting their potential usefulness as vaccine candidates. Our work is the first to describe T cell responses to a member of the proposed fourth subfamily of mammalian herpesviruses, theDeltaherpesvirinae, within a host species in the clade Afrotheria. An EEHV vaccine would greatly contribute to the health care of Asian and African elephants that are also susceptible to this disease.

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