scholarly journals 913 A murine gastric cancer YTN16 model for the rational design of combination immunotherapy

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A959-A959
Author(s):  
Koji Nagaoka ◽  
Changbo Sun ◽  
Yukari Kobayashi ◽  
Kazuhiro Kakimi
Keyword(s):  
Gastric Cancer ◽  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Adoptive Transfer ◽  
Rational Design ◽  
Gastric Cancer Cell ◽  
Missense Mutations ◽  
Combination Immunotherapy ◽  
T Cell Lines

BackgroundAlthough Immune checkpoint blockade (ICB) has changed the standard of care of cancer, ICB monotherapy is largely ineffective in patients with solid cancer. Therefore, the rational combination of ICB with other treatment modalities is warranted. We have recently established chemically induced gastric cancer cell line YTN16, transplantable in immune-competent mice,1 and showed that inhibition of FGFR4 signaling suppressed the YTN16 tumor growth. We also demonstrated the depletion of IL-17-producing T cells in YTN16 tumors enhanced the anti-tumor effects of anti-PD-1 treatment.2 In this study, we identified the neoantigens of YTN16 to monitor the dynamics of tumor-specific CD8< sup >+</sup > T cells in response to combination immunotherapy.MethodsWhole-exome and transcriptome sequencing analyses were performed to identify missense mutations. As a result, candidate neoepitopes were predicted as follows; FPKM≧25, variant allele frequency in RNAseq≧0.04, IC50 of NetMHCpan≦250nM, EL rank of NetMHCpan≦0.5 and presentation percentile of MHCflurry≦0.5.ResultsExome sequencing identified 3,347 missense mutations in the YTN16 tumor. We synthesized 11 candidate neoepitope peptides and screened their reactivity to YTN16-reactive CD8< sup >+</sup > T cell lines established from YTN16-rejected mice by ICB. Out of 11 peptides, five peptides (3 neoantigens, m(mutated)Cdt1, mScarb2 and mZfp106) induced IFNγ production in YTN16-reactive CD8< sup >+</sup > T cells. MHC class I dimer assay identified these three neoantigen-specific T cells in YTN16 tumors. Anti-CTLA-4, but not anti-PD-1 increased neoantigen-specific T cells and completely eradicated tumors. Adoptive transfer of neoantigen-reactive CD8< sup >+</sup > T cell lines and therapeutic vaccines of DCs pulsed with neoantigen short or long peptides inhibited YTN16 growth. Neoantigen-specific TCRs were cloned from neoantigen-reactive CD8< sup >+</sup > T cell lines and retrovirally transduced to activated CD8< sup >+</sup > T cells. TCR-transduced T cells killed YTN16 in vitro and adoptive transfer of TCR-transduced T cells showed anti-tumor effects. These results indicated that these three neoantigens were de fact neoantigens.ConclusionsWe identified three neoantigens that induced CD8< sup >+</sup > T cell response with anti-tumor effects in YTN16. Thus, YTN16 is a well-characterized murine gastric cancer model for rational design and optimization of combination immunotherapy.ReferencesYamamoto M, Nomura S, Hosoi A, Nagaoka K, Iino T, Yasuda T, Saito T, Matsushita H, Uchida E, Seto Y, Goldenring JR, Kakimi K, Tatematsu M, Tsukamoto T. Established gastric cancer cell lines transplantable into C57BL/6 mice show fibroblast growth factor receptor 4 promotion of tumor growth. Cancer Sci 2018;109(5):1480–1492.Nagaoka K, Shirai M, Taniguchi K, Hosoi A, Sun C, Kobayashi Y, Maejima K, Fujita M, Nakagawa H, Nomura S, Kakimi K. Deep immunophenotyping at the single-cell level identifies a combination of anti-IL-17 and checkpoint blockade as an effective treatment in a preclinical model of data-guided personalized immunotherapy. J Immunother Cancer 2020;8(2):e001185.

Rapid generation of combined CMV-specific CD4+ and CD8+ T-cell lines for adoptive transfer into recipients of allogeneic stem cell transplants

Blood ◽  
2004 ◽  
Vol 103 (9) ◽  
pp. 3565-3572 ◽  
Author(s):  
Georg Rauser ◽  
Hermann Einsele ◽  
Christian Sinzger ◽  
Dorothee Wernet ◽  
Gabriele Kuntz ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Cell Lines ◽  
Adoptive Transfer ◽  
Cd8 T Cell ◽  
Cell Transplants ◽  
Ifn Γ ◽  
Rapid Generation ◽  
T Cell Lines

Abstract Adoptive transfer of cytomegalovirus (CMV)-specific T cells can restore long-lasting, virus-specific immunity and clear CMV viremia in recipients of allogeneic stem cell transplants if CD4+ and CD8+ CMV-specific T cells are detected in the recipient after transfer. Current protocols for generating virus-specific T cells use live virus, require leukapheresis of the donor, and are time consuming. To circumvent these limitations, a clinical-scale protocol was developed to generate CMV-specific T cells by using autologous cellular and serum components derived from a single 500-mL blood draw. CMV-specific T cells were stimulated simultaneously with CMV-specific major histocompatibility complex class I (MHC I)- restricted peptides and CMV antigen. Activated T cells were isolated with the interferon-γ (IFN-γ) secretion assay and expanded for 10 days. In 8 randomly selected, CMV-seropositive donors, 1.34 × 108 combined CD4+ and CD8+ CMV-specific T cells, on average, were generated, as determined by antigen-triggered IFN-γ production. CMV-infected fibroblasts were efficiently lysed by the generated T cells, and CMV-specific CD4+ and CD8+ T cells expanded if they were stimulated with natural processed antigen. On the other hand, CD4+ and CD8+ T cell-mediated alloreactivity of generated CMV-specific T-cell lines was reduced compared with that of the starting population. In conclusion, the culture system developed allowed the rapid generation of allodepleted, highly enriched, combined CD4+ and CD8+ CMV-specific T cells under conditions mimicking good manufacturing practice. (Blood. 2004; 103:3565-3572)

scholarly journals Identification of Hexon-Specific CD4 and CD8 T-Cell Epitopes for Vaccine and Immunotherapy

2007 ◽  
Vol 82 (1) ◽  
pp. 546-554 ◽  
Author(s):  
Ann M. Leen ◽  
Anne Christin ◽  
Mariam Khalil ◽  
Heidi Weiss ◽  
Adrian P. Gee ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Adoptive Transfer ◽  
Cytotoxic T Cell ◽  
Cell Immune Response ◽  
T Cell Epitopes ◽  
Human Pathogens ◽  
T Cell Lines ◽  
Cytotoxic T Cell Lines

ABSTRACT Adenoviral infections in the immunocompromised host are associated with significant morbidity and mortality. Although the adoptive transfer of adenovirus-specific T cells may prevent and treat such infections, the T-cell immune response to the multiplicity of adenovirus serotypes and subspecies that infect humans has not been well characterized, impeding the development of such approaches. We have, therefore, analyzed the specificities of T-cell responses to the viral capsid hexon antigen, since this structure is highly conserved in human pathogens. We screened 25 human cytotoxic T-cell lines with adenovirus specificity to extensively characterize their responses to adenoviral hexon and to identify a panel of novel CD4+ and CD8+ T-cell epitopes. Using a peptide library spanning the entire sequence of the hexon protein, we confirmed the responsiveness of these cytotoxic T-cell lines to seven peptides described previously and also identified 33 new CD4- or CD8-restricted hexon epitopes. Importantly, the majority of these epitopes were shared among different adenovirus subspecies, suggesting that T cells with such specificities could recognize and be protective against multiple serotypes, simplifying the task of effective adoptive transfer or vaccine-based immunotherapy for treating infection by this virus.

scholarly journals Immune suppressed tumor microenvironment by exosomes derived from gastric cancer cells via modulating immune functions

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Juan Liu ◽  
Shaoxian Wu ◽  
Xiao Zheng ◽  
Panpan Zheng ◽  
Yuanyuan Fu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gastric Cancer ◽  
T Cells ◽  
T Cell ◽  
Tumor Microenvironment ◽  
Cancer Cells ◽  
Cell Lines ◽  
Gastric Cancer Cell ◽  
Gastric Cancer Cells ◽  
Gastric Cancer Cell Lines

Abstract Gastric cancer is one of the leading causes of cancer-related death due to late diagnosis with high metastatic frequency. In this study, the impact of tumor secreted exosomes on immune function in the tumor environment was investigated using exosomes isolated from gastric cancer cell lines MKN-28, MKN-45, and SGC-7901. Results show that exosomes derived from all of these cell lines changed the gene expression and cytokine secretion levels of CD8+ T cells. They also block cell cycle progression, induced apoptosis in CD8+ T cells. Image analysis of fluorescent labeled exosomes derived from three cell lines injected systemically into C57BL/6 mice revealed these exosomes primarily localize to the lungs. We further showed exosomes were mainly taken up by natural killer cells and macrophages in the lung. After long-term exposure to inject exosomes from MKN-45 cells, mice developed an immunosuppressive tumor microenvironment in the lung with increased frequency of effector memory CD4+ T and MDSC, decreased CD8+ T cell and NK frequency. This immune suppressive environment promotes gastric cancer lung metastasis. Lung metastasis sites developed after mice were exposed to exosomes isolated from all three gastric cancer cell lines when the mice were injected with MFC cells. Results suggest that exosomes derived from gastric cancer cells (especially MKN-45 and MKN-28) changed CD8+ T cell gene expression and cytokine secretion patterns to create an immunosuppressive condition for metastatic niche formation in the lung. Overall, this study provides new insights into how gastric cancer derived exosomes modulate the immune response to promote lung tumor metastasis.

scholarly journals Banked, GMP Grade Third Party T-Cell Lines Specific for CMVpp65 Epitopes Presented By Certain Prevalent HLA Alleles More Consistently Clear CMV Infections in a Genetically Heterogeneous Population of HSCT Recipients

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 309-309 ◽  
Author(s):  
Aisha N. Hasan ◽  
Susan E. Prockop ◽  
Guenther Koehne ◽  
Ekaterina Doubrovina ◽  
Richard J. O'Reilly
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Adoptive Transfer ◽  
Third Party ◽  
Clinical Activity ◽  
Allelic Variants ◽  
Hla Alleles ◽  
Allogeneic Hsct ◽  
T Cell Lines

Abstract Adoptive transfer of virus-specific T-cells (CTLs) derived from allogeneic HSCT donors or HLA partially matched third party donors can eradicate EBV, CMV or adenovirus infections in a high proportion of allogeneic HSCT recipients. At our center, the response rate to third party CMVpp65-specific T-cells (CMVCTLs) has been about 60%. The specific characteristics of CTLs from third party donors that can predict efficacy after adoptive transfer have not been fully defined. However, current evidence indicates that for third party CTLs to be effective, CTL lines selected for treatment must at least be restricted by an HLA allele shared by the cells infected in the patient and the T-cell donor. We have developed a CMV CTL bank containing 132 CTL lines that was generated using a pool of overlapping peptides spanning the sequence of CMVpp65. Each CMV CTL line in the bank has been characterized as to T-cell memory phenotype, cytokine profile, epitope specificity and the HLA allele restriction of the cytotoxic T-cells. Although our pool of CTL donors has inherited a diverse group of widely prevalent HLA alleles, the cytotoxic activity of the T-cell lines in the bank is restricted by only 11 alleles. Nevertheless, these T-cells provided an appropriately restricted CTL line for 51 of 56 requested cases. In the present study, we examined 52 HSCT recipients who had received CMV CTLs of defined HLA restriction from either their HSCT donor (n=23) or a third party donor (n= 29) to determine whether and to what degree specific immunodominant T-cells restricted by specific HLA alleles exhibited differential clinical activity following adoptive transfer. We did this analysis because in analyzing responses to epitopes presented by these alleles, we noted that epitopes presented by certain alleles elicited quantitatively greater in-vitro responses as measured by the number of IFNγ+ or tetramer+ T-cells/106T-cells administered. For 9 CTL lines used in our series, the epitope eliciting the immunodominant T-cell response was an 11-15 aa peptide that was presented by both an HLA class-I and class-II allele. These class-I and class-II shared epitopes elicited more robust, multifunctional CD8+ and CD4+ T-cells that generated IFNγ, TNFα and IL-2. 8 of 9 patients treated with one of these CTL lines rapidly cleared viremia and end organ disease. Another group of 4 CTL lines responded to 2 epitopes, 1 dominant and 1 subdominant, that were restricted by one HLA A and one B allele; specifically A01 and B08, A 01 and B35 and A24 and B35. Treatment with such dual epitope responsive CTL lines also yielded complete responses in 4/4 cases. Among lines specific for single immunodominant epitopes, HLA restricted CMV CTLs responding to 4 CMVpp65 epitopes also consistently induced rapid clearance of viremia and organ disease including colitis, retinitis and encephalitis; NLVPMVATV (14/16 Complete responses CR), TPRVTGGGAM/RPHERNGFTV (7/7 CR) and HERNGFTV (6/6 CR) presented by HLA A0201, B0702 and allelic variants of B40 or B44 (B4001-06 and/B4401-03 ) respectively. In contrast, none of the 10 patients treated CTL lines responding to epitopes presented by HLA A2601, A2407, A2902, B0705, B5001, or allelic variants of B35 responded to CTL treatment and progressed. These findings suggest a hierarchy of epitopes presented by prevalent HLA alleles that may exhibit more consistent clinical activity in-vivo. Selection strategies utilizing such data may permit the development of a CTL selection algorithm for CMV CTLs derived from third party donors for treatment of CMV infections that would, more consistenly, induce clearance of CMV viremia and disease. These data also offer the potential to focus strategies for generation of CMV CTLs using a limited peptide pool or a panel of AAPCs expressing specific HLA alleles, thereby making this treatment approach more financially and ligisticaly feasible. These data could also be applied toward the development of an effective CMV vaccine for high risk individuals. Disclosures No relevant conflicts of interest to declare.

EBV-Specific Cytotoxic T Lymphocytes by LMP2a- and EBNA3a-Specific T Cell Receptor Gene Transfer for Adoptive Therapy in EBV-Associated Hodgkin’s Disease and Post-Transplant Lymphoma.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2334-2334
Author(s):  
Tuan D. Nguyen ◽  
Haike Gelfort ◽  
Kathrin Sebelin-Wulf ◽  
Oliver Schmetzer ◽  
Wolfgang Uckert ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Adoptive Transfer ◽  
Adoptive Therapy ◽  
T Cell Clones ◽  
Cell Clones ◽  
Human T Cells ◽  
Ifn Γ ◽  
T Cell Lines

Abstract Adoptive transfer of polyclonal EBV-specific T cell lines has been used as prophylaxis and therapy in patients with EBV-associated malignancies. However, this strategy is time-consuming and demands difficult generation of lymphoblastoid cell lines (LCLs) and corresponding T cells for each individual patient. We applied an alternative strategy to confer T cell immunity against EBV-antigens by isolating EBV antigen-specific T cell receptors (TCRs) for transduction of primary human T cells for adoptive therapy. Previously, we have demonstrated the feasibility of using peptide-pulsed dendritic cells (DC) for generating high-affinity EBV antigen-specific T cell lines and T cell clones. Based on this strategy, T cell clones directed against LMP2a and EBNA3a were generated and functionally analyzed. Monospecificity was demonstrated by homogeneous double staining with CD8 and appropriate tetramers. High avidity of T cell clones (&lt; 0.01 μM) was shown by peptide titration in an ELISPOT assay for IFN-γ secretion. In addition, the cytokine secretion profiles of some of the T cell clones were tested by cytokine bead array assay. High secretion levels of IFN-γ, IL-2 as well as TNF-α after stimulation with the EBNA3a- or LMP2a-peptide were shown for the corresponding T cell clones. Potent TCRs from one LMP2a-specific, HLA-A2-restricted and one EBNA3a-specific, HLA-B8-restricted T cell clone were isolated and cloned into the retroviral vector MP71. Transduction efficiency of TCR-deficient T cell lines was &gt; 40% (TCR-LMP2a) and &gt; 30% (TCR-EBNA3a) as measured by tetramer staining. Both TCR-LMP2a- and TCR-EBNA3a-redirected T cell lines were functional as indicated by NFAT-mediated luciferase expression upon TCR-MHC-peptide ligation. Primary human T cells were successfully transduced with TCR-LMP2a (∼ 12% tetramer-positive) and TCR-EBNA3a (∼ 3% tetramer-positive). Importantly, both TCRs conferred similar cytolytic activity against EBV-transformed B cell lines. Our data support the development of TCR-transduced T cells for adoptive transfer in EBV-associated malignancies, including Hodgkin′ s disease and nasopharyngeal carcinoma in which only subdominant EBV antigens are expressed. The feasibility and the therapeutic potential of TCR-transduced T cells for adoptive transfer have already been shown in a clinical phase I trial in patients with metastatic melanoma. We believe that redirecting human PBLs is a rapid and efficient tool toward adoptive transfer in EBV-associated malignancies.

scholarly journals Unique Gene Expression Patterns in Human T-cell Lines Generated from Multiple Sclerosis Patients by Stimulation with a Synthetic MOG Peptide

2005 ◽  
Vol 12 (3) ◽  
pp. 203-209 ◽  
Author(s):  
Mathilda Mandel ◽  
Michael Gurevich ◽  
Gad Lavie ◽  
Irun R. Cohen ◽  
Anat Achiron
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Healthy Subjects ◽  
Expression Patterns ◽  
Gene Expression Pattern ◽  
Gene Expression Patterns ◽  
T Cell Lines ◽  
Myelin Antigens

Multiple sclerosis (MS) is an autoimmune disease where T-cells activated against myelin antigens are involved in myelin destruction. Yet, healthy subjects also harbor T-cells responsive to myelin antigens, suggesting that MS patient-derived autoimmune T-cells might bear functional differences from T-cells derived from healthy individuals. We addressed this issue by analyzing gene expression patterns of myelin oligodendrocytic glycoprotein (MOG) responsive T-cell lines generated from MS patients and healthy subjects. We identified 150 transcripts that were differentially expressed between MS patients and healthy controls. The most informative 43 genes exhibited >1.5-fold change in expression level. Eighteen genes were up-regulated including BCL2, lifeguard, IGFBP3 and VEGF. Twenty five genes were down-regulated, including apoptotic activators like TNF and heat shock protein genes. This gene expression pattern was unique to MOG specific T-cell lines and was not expressed in T-cell lines reactive to tetanus toxin (TTX). Our results indicate that activation in MS that promotes T-cell survival and expansion, has its own state and that the unique gene expression pattern that characterize autoreactive T-cells in MS represent a constellation of factors in which the chronicity, timing and accumulation of damage make the difference between health and disease.

scholarly journals Nonmalignant T cells stimulate growth of T-cell lymphoma cells in the presence of bacterial toxins

Blood ◽  
2006 ◽  
Vol 109 (8) ◽  
pp. 3325-3332 ◽  
Author(s):  
Anders Woetmann ◽  
Paola Lovato ◽  
Karsten W. Eriksen ◽  
Thorbjørn Krejsgaard ◽  
Tord Labuda ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Mhc Class Ii ◽  
Interleukin 2 ◽  
Class Ii ◽  
Bacterial Toxins ◽  
Dependent Manner ◽  
Malignant Cells ◽  
T Cell Lines

AbstractBacterial toxins including staphylococcal enterotoxins (SEs) have been implicated in the pathogenesis of cutaneous T-cell lymphomas (CTCLs). Here, we investigate SE-mediated interactions between nonmalignant T cells and malignant T-cell lines established from skin and blood of CTCL patients. The malignant CTCL cells express MHC class II molecules that are high-affinity receptors for SE. Although treatment with SE has no direct effect on the growth of the malignant CTCL cells, the SE-treated CTCL cells induce vigorous proliferation of the SE-responsive nonmalignant T cells. In turn, the nonmalignant T cells enhance proliferation of the malignant cells in an SE- and MHC class II–dependent manner. Furthermore, SE and, in addition, alloantigen presentation by malignant CTCL cells to irradiated nonmalignant CD4+ T-cell lines also enhance proliferation of the malignant cells. The growth-promoting effect depends on direct cell-cell contact and soluble factors such as interleukin-2. In conclusion, we demonstrate that SE triggers a bidirectional cross talk between nonmalignant T cells and malignant CTCL cells that promotes growth of the malignant cells. This represents a novel mechanism by which infections with SE-producing bacteria may contribute to pathogenesis of CTCL.

Establishment of Neospora caninum antigen-specific T cell lines of primarily CD4+ T cells

2004 ◽  
Vol 26 (5) ◽  
pp. 243-246 ◽  
Author(s):  
W. Tuo ◽  
W. C. Davis ◽  
R. Fetterer ◽  
M. Jenkins ◽  
P. C. Boyd ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Cd4 T Cells ◽  
Neospora Caninum ◽  
T Cell Lines

Activation of interleukin-13 expression in T cells from HTLV-1-infected individuals and in chronically infected cell lines

Blood ◽  
2003 ◽  
Vol 102 (12) ◽  
pp. 4130-4136 ◽  
Author(s):  
Hye-Kyung Chung ◽  
Howard A. Young ◽  
Peter K. C. Goon ◽  
Gisela Heidecker ◽  
Gerald L. Princler ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Infected Cell ◽  
Cell Lines ◽  
Regulatory Protein ◽  
Immunofluorescent Staining ◽  
Cell Leukemia Virus Type ◽  
Interleukin 13 ◽  
Human T Cell ◽  
T Cell Lines

Abstract Human T-cell leukemia virus type 1 (HTLV-1) infection profoundly alters T-cell gene expression, and the dysregulated synthesis of cytokines could influence the course and pathologic consequences of infection. In the process of screening T-cell lines for T helper 1 (Th1) and Th2 cytokine mRNAs, we observed that interleukin-13 (IL-13) mRNA was highly expressed in HTLV-1-infected, IL-2-dependent T-cell lines. IL-9 and interferon gamma (IFN-γ) mRNAs were also expressed at high levels in chronically infected cell lines. IL-5 mRNA was detected in 60% of the HTLV-1-infected cell lines, but mRNAs for IL-4, IL-10, IL-2, and IL-15 were either below detection limits or did not correlate with HTLV-1 infection. Transcriptional activation of the IL-13 promoter by the HTLV-1 Tax trans-regulatory protein was demonstrated in Jurkat T cells transiently transfected with an IL-13 promoter-reporter plasmid. The clinical relevance of these observations was demonstrated by immunofluorescent staining and flow cytometry of lymphocytes obtained from HTLV-1-infected patients. These studies revealed that IL-13 production was directly related to the level of Tax expression in the infected CD4+ T cells soon after in vitro culture. As IL-13 plays key roles in tumor immunosurveillance, asthma, and central nervous system inflammation, it may contribute to the pathophysiology of HTLV-1-associated diseases. (Blood. 2003;102:4130-4136)

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