scholarly journals Antigen-Responsive CD4+ T Cells from C3H Mice Chronically Infected with Leishmania amazonensis Are Impaired in the Transition to an Effector Phenotype

2006 ◽  
Vol 74 (3) ◽  
pp. 1547-1554 ◽  
Author(s):  
Amanda E. Ramer ◽  
Yannick F. Vanloubbeeck ◽  
Douglas E. Jones
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Population ◽  
T Cell Response ◽  
Leishmania Amazonensis ◽  
Effector Memory ◽  
Intrinsic Defect ◽  
Antigen Stimulation ◽  
Ifn Γ

ABSTRACT C3HeB/FeJ mice challenged with Leishmania major develop a polarized Th1 response and subsequently heal, whereas Leishmania amazonensis challenge leads to chronic lesions with high parasite loads at 10 weeks postinfection. In this study, a comparison of draining lymph node cells from L. amazonensis- and L. major-infected mice at 10 weeks postinfection showed equivalent percentages of effector/memory phenotype CD44hi CD4+ T cells producing interleukin-2 (IL-2) and proliferating after antigen stimulation. However, these cells isolated from L. amazonensis-infected mice were not skewed toward either a Th1 or Th2 phenotype in vivo, as evidenced by their unbiased Th1/Th2 transcription factor mRNA profile. In vivo antigen stimulation with added IL-12 failed to enhance gamma interferon (IFN-γ) production of CD4+ T cells from L. amazonensis-infected mice. Antigen stimulation of CD4+ T cells from L. amazonensis-infected mice in vitro in the presence of IL-12 resulted in production of only 10 to 15% of the IFN-γ produced by T cells from L. major-infected mice under identical conditions. These results suggest that the CD4+ T-cell response during chronic L. amazonensis infection is limited during the transition from an early activated CD4+ T-cell population to an effector cell population and demonstrate that these T cells have an intrinsic defect beyond the presence or absence of IL-12 during antigen stimulation.

scholarly journals Short-Lived Effector CD8 T Cells Induced by Genetically Attenuated Malaria Parasite Vaccination Express CD11c

2013 ◽  
Vol 81 (11) ◽  
pp. 4171-4181 ◽  
Author(s):  
Laura A. Cooney ◽  
Megha Gupta ◽  
Sunil Thomas ◽  
Sebastian Mikolajczak ◽  
Kimberly Y. Choi ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Inflammatory Cytokines ◽  
Plasmodium Yoelii ◽  
T Cell Response ◽  
Parasite Development ◽  
Effector Memory ◽  
Cell Phenotype ◽  
Effector Cells ◽  
Ifn Γ

ABSTRACTVaccination with a single dose of genetically attenuated malaria parasites can induce sterile protection against sporozoite challenge in the rodentPlasmodium yoeliimodel. Protection is dependent on CD8+T cells, involves perforin and gamma interferon (IFN-γ), and is correlated with the expansion of effector memory CD8+T cells in the liver. Here, we have further characterized vaccine-induced changes in the CD8+T cell phenotype and demonstrated significant upregulation of CD11c on CD3+CD8b+T cells in the liver, spleen, and peripheral blood. CD11c+CD8+T cells are predominantly CD11ahiCD44hiCD62L−, indicative of antigen-experienced effector cells. Followingin vitrorestimulation with malaria-infected hepatocytes, CD11c+CD8+T cells expressed inflammatory cytokines and cytotoxicity markers, including IFN-γ, tumor necrosis factor alpha (TNF-α), interleukin-2 (IL-2), perforin, and CD107a. CD11c−CD8+T cells, on the other hand, expressed negligible amounts of all inflammatory cytokines and cytotoxicity markers tested, indicating that CD11c marks multifunctional effector CD8+T cells. Coculture of CD11c+, but not CD11c−, CD8+T cells with sporozoite-infected primary hepatocytes significantly inhibited liver-stage parasite development. Tetramer staining for the immunodominant circumsporozoite protein (CSP)-specific CD8+T cell epitope demonstrated that approximately two-thirds of CSP-specific cells expressed CD11c at the peak of the CD11c+CD8+T cell response, but CD11c expression was lost as the CD8+T cells entered the memory phase. Further analyses showed that CD11c+CD8+T cells are primarily KLRG1+CD127−terminal effectors, whereas all KLRG1−CD127+memory precursor effector cells are CD11c−CD8+T cells. Together, these results suggest that CD11c marks a subset of highly inflammatory, short-lived, antigen-specific effector cells, which may play an important role in eliminating infected hepatocytes.

scholarly journals Enhanced CAR-T activity against established tumors by polarizing human T cells to secrete interleukin-9

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Lintao Liu ◽  
Enguang Bi ◽  
Xingzhe Ma ◽  
Wei Xiong ◽  
Jianfei Qian ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Antitumor Activity ◽  
Effector Memory ◽  
T Cell Therapy ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T ◽  
Ifn Γ

AbstractCAR-T cell therapy is effective for hematologic malignancies. However, considerable numbers of patients relapse after the treatment, partially due to poor expansion and limited persistence of CAR-T cells in vivo. Here, we demonstrate that human CAR-T cells polarized and expanded under a Th9-culture condition (T9 CAR-T) have an enhanced antitumor activity against established tumors. Compared to IL2-polarized (T1) cells, T9 CAR-T cells secrete IL9 but little IFN-γ, express central memory phenotype and lower levels of exhaustion markers, and display robust proliferative capacity. Consequently, T9 CAR-T cells mediate a greater antitumor activity than T1 CAR-T cells against established hematologic and solid tumors in vivo. After transfer, T9 CAR-T cells migrate effectively to tumors, differentiate to IFN-γ and granzyme-B secreting effector memory T cells but remain as long-lived and hyperproliferative T cells. Our findings are important for the improvement of CAR-T cell-based immunotherapy for human cancers.

Generation of Cytomegalovirus (CMV)-Specific CD4 and CD8 T Cell Lines Using Protein-Spanning Pools of pp65 and IE1 Derived Peptides.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 477-477
Author(s):  
Erica Dander ◽  
Giuseppina Li Pira ◽  
Ettore Biagi ◽  
Fabrizio Manca ◽  
Andrea Biondi ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Cd8 T Cell ◽  
Effector Memory ◽  
Target Cells ◽  
Pulsed Dc ◽  
Ifn Γ ◽  
T Cell Lines

Abstract BACKGROUND: Reactivation of latent CMV in immunocompromised recipients of allogeneic stem cell transplantation remains a major cause of morbidity and mortality. Reconstitution of immunity by CMV specific immunotherapy is an attractive alternative to drugs currently used, which show high toxicity and are sometimes ineffective. It has been demonstrated that CD4 helper T-cell function is crucial for the persistence of in vivo transferred CD8 CMV-specific CTL. Based on this finding, we have explored the feasibility of generating both anti-CMV CD4 and anti-CMV CD8 T-cell lines. METHODS: Dendritic Cells (DC) were generated from donor peripheral blood (PB) monocytes after a 7-day culture in the presence of GM-CSF plus IL-4 and matured with TNF-α, IFN-α, IFN-γ, IL1-β, POLI I:C. Matured-DC were then pulsed with a pool of 50 peptides spanning pp65 and IE1 proteins which are recognised by both CD4 and CD8 T lymphocytes. Donor T cells were stimulated three times at a T cell/DC ratio of 1:6 on day 0, +7 and +14 with mature peptide pulsed-DC. At the end of the culture the specificity of generated T cells was determined as percentage of pentamer-positive cells and intracellular IFN-γ production after incubation with peptide pulsed-DC. Cultured T cells were also analysed for their ability to proliferate in response to peptide pulsed-target cells, to kill them in a standard citotoxicity assay and to migrate in response to inflammatory (CXCL9, CCL3 and CCL5) and constitutive (CXCL12) chemokines. RESULTS: CMV-specific T cell lines were generated from five CMV seropositive donors. In four cases CD4 and CD8 CMV-specific T cell lines were expanded successfully. Cultured T cells expressed CD8 (mean= 70%, range 60–81%) and CD4 (mean= 20%, range 15–28%) and showed a CD45RA- CCR7- Effector Memory phenothype (mean=26%, range 19–30%) or a CD45RA+ CCR7- T Effector Memory RA-Positive phenothype (mean=67%, range 59–77%). An enriched CMV-specific T cell population was observed after staining with pentamers (7–45% pentamer-positive T cells). Furthermore, 90% of CD8+ and 40% of CD4+ T cells expressed high levels of intracytoplasmatic perforin and granzyme. In 4/5 cases tested, cutured T cells showed a cytolitic activity against CD8-peptide pulsed target cells (average lysis=50%, range 40–55%) and to a lesser extent against CD4-peptide pulsed target cells (average lysis=35%, range 30–40%). In addition, cultured T lymphocytes were able to proliferate and to produce intracytoplasmic IFN-γ (average production=50%, range 35–60%) after exposure to peptide-pulsed DC. Finally, Cultured T cells strongly migrated in response to chemokines (CXCL9, CCL3 and CCL5) involved in the recruitment of effector cells during viral infection. DISCUSSION: In conclusion, a great advantage of this method is represented by the possibility to generate anti-CMV CD4+ T cells, which could support in vivo the persistence of re-infused CMV-specific CTL. Moreover, the possibility of generating peptides under GMP conditions would facilitate the translation of this approach into clinical intervention.

scholarly journals Mycobacteria-specific CD4+IFN-γ+ cell expresses naïve-surface markers and confers superior protection against tuberculosis infection compared to central and effector memory CD4+ T cell subsets

2018 ◽  
Author(s):  
Jinyun Yuan ◽  
Janice Tenant ◽  
Thomas Pacatte ◽  
Christopher Eickhoff ◽  
Azra Blazevic ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Population ◽  
Vaccine Trial ◽  
Memory Cell ◽  
T Cell Subsets ◽  
Effector Memory ◽  
Surface Markers ◽  
Activation Markers ◽  
Ifn Γ

AbstractFailure of the most recent tuberculosis (TB) vaccine trial to boost BCG mediated anti-TB immunity despite highly durable Th1-specific central (TCM) and effector (TEM) memory cell responses, highlights the importance of identifying optimal T cell targets for protective vaccines. Here we describe a novel, Mycobacterium tuberculosis (Mtb)-specific IFN-γ+CD4+ T cell population expressing surface markers characteristic of naïve T cells (TNLM), that were induced in both human (CD45RA+CCR7+CD27+CD95-) and murine (CD62L+CD44-Sca-1+CD122-) systems in response to mycobacteria. In BCG vaccinated subjects and those with latent TB infection, TNLM cells, compared to bonafide naïve CD4+ T cells were identified by absence of CD95 expression and had increased expression CCR7 and CD27, the activation markers T-bet, CD69 and PD-1 and the survival marker CD74. Increased TNLM frequencies were noted in the lung and spleen of wild type C57BL6 mice at 2 weeks after infection with Mtb, and progressively decreased at later time points, a pattern not seen in TNF-α+CD4+ T cells expressing naïve cell surface markers. Importantly, adoptive transfer of highly purified TNLM from vaccinated ESAT-61-20-specific TCR transgenic mice conferred superior protection against Mtb infection in Rag-/- mice when compared with total meory populations (central and effector memory cells). Thus, TNLM cells may represent a memory T cell population that if optimally targeted may significantly improve future TB vaccine responses.

scholarly journals Interplay between IL-10, IFN-γ, IL-17A and PD-1 Expressing EBNA1-Specific CD4+ and CD8+ T Cell Responses in the Etiologic Pathway to Endemic Burkitt Lymphoma

Cancers ◽  
2021 ◽  
Vol 13 (21) ◽  
pp. 5375
Author(s):  
Catherine S. Forconi ◽  
David H. Mulama ◽  
Priya Saikumar Lakshmi ◽  
Joslyn Foley ◽  
Juliana A. Otieno ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Burkitt Lymphoma ◽  
Cd4 T Cells ◽  
Cd8 T Cell ◽  
T Cell Response ◽  
T Cell Subsets ◽  
Effector Memory ◽  
Healthy Children ◽  
Ifn Γ

Children diagnosed with endemic Burkitt lymphoma (eBL) are deficient in interferon-γ (IFN-γ) responses to Epstein–Barr Nuclear Antigen1 (EBNA1), the viral protein that defines the latency I pattern in this B cell tumor. However, the contributions of immune-regulatory cytokines and phenotypes of the EBNA1-specific T cells have not been characterized for eBL. Using a bespoke flow cytometry assay we measured intracellular IFN-γ, IL-10, IL-17A expression and phenotyped CD4+ and CD8+ T cell effector memory subsets specific to EBNA1 for eBL patients compared to two groups of healthy children with divergent malaria exposures. In response to EBNA1 and a malaria antigen (PfSEA-1A), the three study groups exhibited strikingly different cytokine expression and T cell memory profiles. EBNA1-specific IFN-γ-producing CD4+ T cell response rates were lowest in eBL (40%) compared to children with high malaria (84%) and low malaria (66%) exposures (p < 0.0001 and p = 0.0004, respectively). However, eBL patients did not differ in CD8+ T cell response rates or the magnitude of IFN-γ expression. In contrast, eBL children were more likely to have EBNA1-specific CD4+ T cells expressing IL-10, and less likely to have polyfunctional IFN-γ+IL-10+ CD4+ T cells (p = 0.02). They were also more likely to have IFN-γ+IL-17A+, IFN-γ+ and IL-17A+ CD8+ T cell subsets compared to healthy children. Cytokine-producing T cell subsets were predominantly CD45RA+CCR7+ TNAIVE-LIKE cells, yet PD-1, a marker of persistent activation/exhaustion, was more highly expressed by the central memory (TCM) and effector memory (TEM) T cell subsets. In summary, our study suggests that IL-10 mediated immune regulation and depletion of IFN-γ+ EBNA1-specific CD4+ T cells are complementary mechanisms that contribute to impaired T cell cytotoxicity in eBL pathogenesis.

scholarly journals Gamma Interferon Regulates Contraction of the Influenza Virus-Specific CD8 T Cell Response and Limits the Size of the Memory Population

2013 ◽  
Vol 87 (23) ◽  
pp. 12510-12522 ◽  
Author(s):  
Nayana Prabhu ◽  
Adrian W. Ho ◽  
Kenneth H. S. Wong ◽  
Paul Edward Hutchinson ◽  
Yen Leong Chua ◽  
...  
Keyword(s):  
T Cells ◽  
Influenza Virus ◽  
T Cell ◽  
Cell Population ◽  
Cd8 T Cells ◽  
Cell Response ◽  
Memory Cell ◽  
Cd8 T Cell ◽  
T Cell Response ◽  
Ifn Γ

The factors that regulate the contraction of the CD8 T cell response and the magnitude of the memory cell population against localized mucosal infections such as influenza are important for generation of efficient vaccines but are currently undefined. In this study, we used a mouse model of influenza to demonstrate that the absence of gamma interferon (IFN-γ) or IFN-γ receptor 1 (IFN-γR1) leads to aberrant contraction of antigen-specific CD8 T cell responses. The increased accumulation of the effector CD8 T cell population was independent of viral load. Reduced contraction was associated with an increased fraction of CD8 T cells expressing the interleukin-7 receptor (IL-7R) at the peak of the response, resulting in enhanced numbers of memory/memory precursor cells in IFN-γ−/−and IFN-γR−/−compared to wild-type (WT) mice. Blockade of IL-7 within the lungs of IFN-γ−/−mice restored the contraction of influenza virus-specific CD8 T cells, indicating that IL-7R is important for survival and is not simply a consequence of the lack of IFN-γ signaling. Finally, enhanced CD8 T cell recall responses and accelerated viral clearance were observed in the IFN-γ−/−and IFN-γR−/−mice after rechallenge with a heterologous strain of influenza virus, confirming that higher frequencies of memory precursors are formed in the absence of IFN-γ signaling. In summary, we have identified IFN-γ as an important regulator of localized viral immunity that promotes the contraction of antigen-specific CD8 T cells and inhibits memory precursor formation, thereby limiting the size of the memory cell population after an influenza virus infection.

scholarly journals Effects of pregnancy-specific β-1-glycoprotein on the helper T-cell response

2019 ◽  
Vol 71 (2) ◽  
pp. 369-378 ◽  
Author(s):  
Valeria Timganova ◽  
Maria Bochkova ◽  
Pavel Khramtsov ◽  
Sofia Kochurova ◽  
Mikhail Rayev ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
Suppressive Effect ◽  
T Cell Response ◽  
Effector Memory ◽  
T Cell Proliferation ◽  
Helper T Cell ◽  
Innate And Adaptive Immunity

Pregnancy-specific ?-1-glycoproteins (PSGs) are capable of regulating innate and adaptive immunity. As fetal antigens circulate in the blood of pregnant women, it is of particular interest to reveal the effects of PSGs on the differentiation of memory T cells in the context of maternal-fetal tolerance formation. We studied if, native PSG preparation affects helper T-cell proliferation, the frequencies of CD4+CD45R0+, CD4+CD45RA+, CD4+CD45RA+CD45R0+cells, naive CD45RA+CD45R0-CD62L+ cells (NAIVE), central memory CD45RA-CD45R0+CD62L+ cells (TCM), effector memory helper T cells (CD45RA-CD45R0+CD62L- (TEM) and CD45RA+CD45R0-CD62L- cells (TEMRA), together with IL-4 and IFN-? production by all ?D4+ T cells. The suppressive effect of PSG on helper T-cell proliferation was established. It was found that PSG does not influence the frequencies of CD45RA+ and CD45R0+ cells, while it decreased the percentage of CD45RA+CD45R0+ cells. PSG increased the percentage of NAIVE cells in culture, and prevented the conversion of these cells into TEMRA, without affecting the levels of TCM and TEM. In addition, PSG lowered the amount of IL-4 and IFN-? in the supernatants of helper T-cell cultures. As TEMRA exhibit cytotoxic activity that is unfavorable during pregnancy, the revealed PSG effects may play a fetoprotective role in vivo.

scholarly journals Vaccination and Infection of Swine With Salmonella Typhimurium Induces a Systemic and Local Multifunctional CD4+ T-Cell Response

2021 ◽  
Vol 11 ◽  
Author(s):  
Selma Schmidt ◽  
Elena L. Sassu ◽  
Eleni Vatzia ◽  
Alix Pierron ◽  
Julia Lagler ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cells ◽  
T Cell ◽  
Lymph Nodes ◽  
T Cell Response ◽  
Effector Memory ◽  
Cell Immune Response ◽  
Control Group ◽  
Tnf Α ◽  
Ifn Γ

The gram-negative facultative intracellular bacteria Salmonella Typhimurium (STM) often leads to subclinical infections in pigs, but can also cause severe enterocolitis in this species. Due to its high zoonotic potential, the pathogen is likewise dangerous for humans. Vaccination with a live attenuated STM strain (Salmoporc) is regarded as an effective method to control STM infections in affected pig herds. However, information on the cellular immune response of swine against STM is still scarce. In this study, we investigated the T-cell immune response in pigs that were vaccinated twice with Salmoporc followed by a challenge infection with a virulent STM strain. Blood- and organ-derived lymphocytes (spleen, tonsils, jejunal and ileocolic lymph nodes, jejunum, ileum) were stimulated in vitro with heat-inactivated STM. Subsequently, CD4+ T cells present in these cell preparations were analyzed for the production of IFN-γ, TNF-α, and IL-17A by flow cytometry and Boolean gating. Highest frequencies of STM-specific cytokine-producing CD4+ T cells were found in lamina propria lymphocytes of jejunum and ileum. Significant differences of the relative abundance of cytokine-producing phenotypes between control group and vaccinated + infected animals were detected in most organs, but dominated in gut and lymph node-residing CD4+ T cells. IL-17A producing CD4+ T cells dominated in gut and gut-draining lymph nodes, whereas IFN-γ/TNF-α co-producing CD4+ T cells were present in all locations. Additionally, the majority of cytokine-producing CD4+ T cells had a CD8α+CD27- phenotype, indicative of a late effector or effector memory stage of differentiation. In summary, we show that Salmonella-specific multifunctional CD4+ T cells exist in vaccinated and infected pigs, dominate in the gut and most likely contribute to protective immunity against STM in the pig.

The Importance of Be(ginn)Ing Naïve: Implications for Cancer Immune-Gene Therapy

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2052-2052
Author(s):  
Nicoletta Cieri ◽  
Barbara Camisa ◽  
Fabienne Cocchiarella ◽  
Elena Provasi ◽  
Zulma Magnani ◽  
...  
Keyword(s):  
Gene Therapy ◽  
T Cells ◽  
T Cell ◽  
Cell Population ◽  
Cell Subset ◽  
T Cell Response ◽  
Immune Gene ◽  
Self Renewal ◽  
T Cell Subset

Abstract Abstract 2052 T cell engineering against tumor antigens aims at ameliorating current immunotherapeutic strategies. To date, however, the suboptimal persistence of the transferred cells represents a serious limitation of this approach. The most appropriate T cell subset to be infused should ensure optimal in vivo persistence and yet appropriate anti-tumor activity. Here we report that culturing highly purified naïve T (TN) cells with beads conjugated to anti-CD3 and anti-CD28 antibodies, allows the retrieval of a novel post-mitotic CD45RA+ CD62L+ CCR7+ T cell population, which requires IL-7 and IL-15 for expansion and maintenance. This population is highly proliferative and sensitive to RV and LV transduction, expresses low levels of γIFN and cytotoxic molecules and is best defined as IL-7Rα+ CXCR4+ c-kit+ CCR5− HLA-DR− PD-1−. When infused in immunodeficient mice, genetically manipulated and in vitro expanded TN proved superior engraftment and longer persistence than transduced central memory (TCM) cells, and xenoreactivity comparable to that of unmanipulated lymphocytes. Engineered TN, but not TCM, maintained engraftment and xenoreactivity in serial transplantation experiments, indicating unique self-renewal abilities. Given the great potentials of this novel TN-derived cell population for immune-gene therapy, we further characterized it by molecular profiling. The gene expression signature is typical of antigen-experienced lymphocytes and classifies these cells between naturally occurring TN and TCM lymphocytes. Because of this and of the self-renewal abilities displayed in vivo, we termed them as precursor to TCM (TpreCM). We next sought to identify the natural counterpart of this TpreCM population in healthy donors, exploiting some of the markers present in the TpreCM signature, such as CD95 which is expressed by all memory T subsets but not by TN, and we selected the pp65 protein of cytomegalovirus (CMV) as a model antigen. CMV persistent infection induces a T-cell response that is maintained throughout life, indicating that a self-renewing memory T cells are generated. We studied the phenotype of CMV pp65-specific CD8+ T cells in seropositive donors and identified antigen-specific CD45RA+CD62L+. Among CD45RA+CD62L+ cells CMV-specific cells were enriched for CD45ROdim and CD95+ lymphocytes, which represent bona fide TpreCM. To functionally characterize natural TpreCM, we sorted CD45RA+CD62L+ cells according to their CD95 expression and challenged them with increasing doses anti-CD3 antibody, with or without a costimulatory signal. We found that among CD45RA+CD62L+ cells, only CD95+ lymphocytes were responsive to TCR-triggering alone, while CD95− cells required costimulation to proliferate. In conclusion, we identified a novel memory T cell subset, and identified conditions able to gene-modify and expand these memory lymphocytes while preserving their functional characteristics. Exploitation of these concepts might improve cancer adoptive immunotherapy. Disclosures: Bonini: MolMed: Consultancy.

Safety and immunogenicity of tyrosinase DNA vaccines in patients with melanoma

2007 ◽  
Vol 25 (18_suppl) ◽  
pp. 3005-3005 ◽  
Author(s):  
J. D. Wolchok ◽  
H. Gallardo ◽  
M. Perales ◽  
T. Rasalan ◽  
J. Wang ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Dna Vaccines ◽  
T Cell Responses ◽  
Cd8 T Cell ◽  
T Cell Response ◽  
Antibody Responses ◽  
Effector Memory ◽  
Cell Responses ◽  
Ifn Γ

3005 Background: T-cell and antibody responses to self antigens on cancer are usually constrained by immunologic tolerance and ignorance. We found that DNA vaccines encoding xenogeneic differentiation antigens, such as tyrosinase (TYR), can mediate tumor protection and regression in implantable mouse models and dogs with spontaneously arising melanoma. Based on this, we conducted a trial of DNA vaccines encoding mouse and human TYR in patients with AJCC stage III/IV melanoma. Methods: HLA-A*0201+ melanoma patients were randomized to 2 different schedules: one group received 3 injections of mouse TYR DNA followed by 3 injections of human TYR DNA while the other group received 3 injections of human TYR DNA followed by 3 injections with the mouse gene. The study was conducted a three different dose levels: 100, 500 and 1,500 mcg DNA/injection, administered IM every 3 weeks. A total of 18 patients were treated, 6 at each dose level being randomized to one of the two schedules. Anti-TYR antibodies and CD8+ T cells recognizing the native human tyrosinase369-377 (YMDGTMSQV) peptide were measured at fixed time points. T-cell responses were monitored with MHC tetramer and intracytoplasmic IFN-γ staining assays using 10-day in vitro stimulation. Multiparametric flow cytometry was performed to further define the phenotype of responding cells. Results: Most toxicities were transient grade I injection site reactions. Seven patients had CD8+ T cell responses, defined as a >3 standard deviation increase in baseline reactivity to the TYR peptide in either the tetramer or intracellular IFN-γ assay. There was no relationship between dose level or assigned schedule and occurrence of T-cell response. Phenotypic characterization of responding T cells showed that most were consistent with an effector memory phenotype including the expression of granzyme B and surface expression of CD107a. No antibody responses were observed. At a median of 42 months of follow-up, median survival has not been reached and 6/18 patients have died from melanoma (1 in the group of patients who had a T cell response and 5 in the non-responders). Conclusions: Mouse and human TYR DNA vaccines were safe and induced CD8+ T cell responses in 7/18 patients. T cells recognizing a native TYR peptide had a phenotype consistent with that of effector memory cells. No significant financial relationships to disclose.

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