CMV-Specific T Cell Immunotherapy Promotes Restoration of Durable Functional Immunity in High Risk Patients Following Allogeneic Stem Cell Transplantation

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 352-352
Author(s):  
Karl S Peggs ◽  
Stephanie Verfuerth ◽  
Arnold Pizzey ◽  
Christine Chow ◽  
Kirsty Thomson ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
High Risk ◽  
Viral Infection ◽  
Cell Lines ◽  
Immune Reconstitution ◽  
Cellular Therapy ◽  
Ex Vivo ◽  
T Cell Lines ◽  
Cmv Disease

Abstract Adoptive cellular therapy with virus-specific T-cells offers the potential for accelerating pathogen-specific immune reconstitution, thus limiting the morbidity and mortality of viral infections following allogeneic haematopoietic stem cell transplantation (aHSCT). However, the logistics of production and the risk of inducing graft-versus-host disease (GvHD) secondary to the infusion of alloreactive clones have restricted their application. We examined the use of polyclonal cytomegalovirus (CMV)-specific mixed CD4+ and CD8+ T-cell lines, generated by short-term ex vivo culture of donor lymphocytes with donor monocyte-derived dendritic cells pulsed with virus-lysate. Following initial evaluation in a phase I study of pre-emptive cellular therapy in mainly matched related donor transplant recipients, we performed a phase I-II study including a higher risk population routinely receiving CMV-specific T cells 28 days following transplantation. Thirty patients received CMV-specific T cells in this study. The majority were at high risk of viral infection, being CMV seropositive recipients (24/30), having received a T-cell depleted graft (24/30), and/or having an unrelated or mismatched donor (14/30). T cell lines were administered prior to viral DNA detection (prophylaxis) in 10 patients, following a single viral DNA detection episode (pre-emptive) in 10 and concurrent with antiviral drug therapy in 10. There were no significant immediate toxicities. Acute GvHD > Grade I occurred in 2/24 T-cell depleted transplants (1 Grade II, 1 Grade III), and 2/6 T-cell replete transplants (2 Grade III), but in 0/14 unrelated or mismatched donor transplants, suggesting no excess of GvHD associated with cellular therapy. Only 3/10 treated prophylactically developed CMV infection requiring therapy (2 following introduction of systemic steroids). All 10 receiving cells pre-emptively required antiviral drugs. CMV-specific immune reconstitution was monitored using class I HLA-pentamers restricted by HLA-A*0201 (the NLV epitope of pp65, and the VLE epitope of IE-1) and HLA-B*0702 (the TPR and RPH epitopes of pp65). In vivo expansions of hCMV-specific T-lymphocytes were observed within days to weeks of adoptive transfer and temporally associated with periods of viral replication. Peak pp65-directed responses following infection ranged between 6.7–43% of CD8+ T cells, with absolute levels of 60.2–371.5 × 106/l, well in excess of those previously reported to offer protection against CMV (10–20 × 106/l), whilst those in patients not experiencing viral DNAemia reached maximal levels of 0.5–0.8% of CD8+ T cells equivalent to absolute levels of 0.2–0.8 × 106/l. Assuming transfused cells are largely responsible for the increase in CMV-specific T cell numbers, at least in the T-cell depleted unrelated donor setting which is associated with markedly impaired immune reconstitution within the first 100 days following transplantation, we estimate that NLVspecific T cells were expanded up to 5 log over a period as short as 10 days following transfer (assuming a 5 litre circulating volume and that 2% of the lymphocyte pool is located in the peripheral blood) with lymphocyte doubling times possibly as low as every 12 hours. Expanding populations maintained functional competence in terms of ability to produce IFNγ in ex vivo restimulation assays (up to 77% of pentamer-labelling cells secreted IFNγ). Following a primary post-transplantation treatment episode 20 patients were evaluable for subsequent viral infection. With a median follow-up of 953 days (range 250–1590) none developed a second episode requiring therapy (compared to 45/72 historical controls, P < 0.0001) and there were no cases of CMV disease. Combined with the absolute circulating levels attained and the functional competence of CMV-specific T cells following transfer, these data indicate that infusion of cell lines containing both CD4+ and CD8+ virus-specific T cells promotes reconstitution of durable functional CMV-specific immunity, effectively preventing recurrent viral infection and late CMV disease in a group of high risk aHSCT recipients.

scholarly journals Experimental Preemptive Immunotherapy of Murine Cytomegalovirus Disease with CD8 T-Cell Lines Specific for ppM83 and pM84, the Two Homologs of Human Cytomegalovirus Tegument Protein ppUL83 (pp65)

2001 ◽  
Vol 75 (14) ◽  
pp. 6584-6600 ◽  
Author(s):  
Rafaela Holtappels ◽  
Jürgen Podlech ◽  
Natascha K. A. Grzimek ◽  
Doris Thomas ◽  
Marcus-Folker Pahl-Seibert ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Cd8 T Cells ◽  
Productive Infection ◽  
Murine Cytomegalovirus ◽  
Tegument Protein ◽  
T Cell Lines ◽  
Cmv Disease

ABSTRACT CD8 T cells are the principal antiviral effectors controlling cytomegalovirus (CMV) infection. For human CMV, the virion tegument protein ppUL83 (pp65) has been identified as a source of immunodominant peptides and is regarded as a candidate for cytoimmunotherapy and vaccination. Two sequence homologs of ppUL83 are known for murine CMV, namely the virion protein ppM83 (pp105) expressed late in the viral replication cycle and the nonstructural protein pM84 (p65) expressed in the early phase. Here we show that ppM83, unlike ppUL83, is not delivered into the antigen presentation pathway after virus penetration before or in absence of viral gene expression, while other virion proteins of murine CMV are processed along this route. In cytokine secretion-based assays, ppM83 and pM84 appeared to barely contribute to the acute immune response and to immunological memory. Specifically, the frequencies of M83 and M84 peptide-specific CD8 T cells were low and undetectable, respectively. Nonetheless, in a murine model of cytoimmunotherapy of lethal CMV disease, M83 and M84 peptide-specific cytolytic T-cell lines proved to be highly efficient in resolving productive infection in multiple organs of cell transfer recipients. These findings demonstrate that proteins which fail to prime a quantitatively dominant immune response can nevertheless represent relevant antigens in the effector phase. We conclude that quantitative and qualitative immunodominance are not necessarily correlated. As a consequence of these findings, there is no longer a rationale for considering T-cell abundance as the key criterion for choosing specificities to be included in immunotherapy and immunoprophylaxis of CMV disease and of viral infections in general.

Ex vivo isolation of RCC-reactive CD8+ CTL clones from HLA-identical allogeneic donor T cell lines stimulated by CD80-transfected tumor cells

2007 ◽  
Vol 25 (18_suppl) ◽  
pp. 15625-15625 ◽  
Author(s):  
S. S. Tykodi ◽  
J. A. Thompson ◽  
B. M. Sandmaier ◽  
M. B. Maris ◽  
R. Storb ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Ex Vivo ◽  
Unrelated Donor ◽  
T Cell Antigens ◽  
Ifn Γ ◽  
T Cell Lines ◽  
Anti Tumor Activity

15625 Background: Regression of metastatic renal cell carcinoma (mRCC) is observed in a minority of patients treated by immunotherapies such as interleukin-2 (IL-2), interferon-a (IFN-a), or reduced-intensity allogeneic hematopoietic cell transplantation. However, the development of specific cellular immunotherapies for mRCC has been hindered by the lack of molecularly characterized T cell antigens with preferential expression on RCC cells. We have developed an ex vivo strategy for the isolation of RCC-reactive CD8+ CTL clones that may facilitate the identification of novel RCC-associated T cell antigens. Methods: RCC tumor lines were established from two patients with mRCC presenting to our institution for allogeneic HCT received from either an HLA-matched sibling or volunteer unrelated donor. Irradiated RCC tumor lines that were unmodified or transfected with a cDNA for human CD80 were used to stimulate responder CD8+ T cells isolated from pretransplant patient (autologous) or donor-derived (allogeneic) blood samples in mixed lymphocyte/tumor cell (MLTC) cultures supplemented with recombinant human IL-7 and IL-12 (stimulation #1) or IL-2 (2nd and subsequent stimulations). T cell lines with anti-tumor activity measured by IFN-γ ELISA were then cloned by limiting dilution. Results: After two or more in vitro stimulations, allogeneic CD8+ T cell lines stimulated by CD80- transfected RCC tumor cells, but not the other MLTC culture combinations tested demonstrated tumor-specific IFN-γ release. CD3+/CD8+/TCRaβ+ CTL clones with potent in vitro anti-tumor activity for unmodified RCC tumor were isolated from both sibling- and unrelated- donor derived T cell lines. Three such clones with unique specificities for allogeneic targets recognized the unmodified RCC tumor but not LCL or fibroblast target cells isolated from the same patient suggesting tumor-restricted expression of the target antigens. Conclusions: Ex vivo MLTC culture utilizing CD80-transfected RCC tumor and HLA- matched allogeneic responder CD8+ T cells warrants further study as a strategy to isolate CTL clones that may be used to identify novel RCC-associated T cell antigens. No significant financial relationships to disclose.

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

scholarly journals T Cell Immunity Toward Viral- and Tumor-Antigens Is Preserved in MDS Patients

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 4225-4225
Author(s):  
Hussein Hamad ◽  
Wingchi K Leung ◽  
Spyridoula Vasileiou ◽  
Shivani Mukhi ◽  
Quillan Huang ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Board Of Directors ◽  
Healthy Donor ◽  
Tumor Antigens ◽  
Ex Vivo ◽  
Advisory Committees ◽  
Cell Immunity ◽  
Healthy Donors ◽  
T Cell Lines

Myelodysplastic syndromes (MDS) are a heterogeneous group of disorders characterized by bone marrow failure and a propensity to progress to acute myeloid leukemia (AML). A core component of the underlying pathogenesis in MDS is deregulation of inflammatory cytokines, such as tumor growth factor-β (TGFβ), which impact the function of immune cells and hence their capacity to mount anti-infective or anti-tumor responses. However, little is known about antigen-specific T cell function in patients with MDS. We hypothesized that virus-specific T cell (VST) function might be preserved in patients with MDS, and that the functional capacity of T cells reactive against tumor-associated antigens aberrantly overexpressed by clonal MDS cells such as Cyclin A1 (CCNA1) and Proteinase (PR3) might also be preserved and exploited for immunotherapeutic purposes. Following informed consent, we collected peripheral blood samples from 10 patients (pts) with MDS and 17 healthy donors. Most pts (9 out of 10) were transfusion dependent and 3 subsequently underwent an allogeneic HSCT. Table 1 summarizes the other clinical characteristics, karyotypic and mutational profile at the time of blood collection. Compared with T cells isolated from healthy donors, MDS patient-derived T cells had a similar CD4 to CD8 ratio (1.5-2.5:1 for healthy donors and 3:1 for MDS pts), but displayed a more exhausted profile at baseline (CD3+TIM3+: 1% in healthy donors and 5% in MDS pts) and produced higher levels of inflammatory cytokines [IFNγ (18±3pg/ml vs 36±16pg/ml, healthy donor vs MDS; p=0.12), and IL-8 (56±32 vs 704±446 pg/ml, p=0.01)]. Next, to assess the capacity of MDS pts to mount ex vivo functional virus-directed responses, we stimulated patient-derived PBMCs (n=5) with overlapping peptide libraries (pepmixes) spanning immunogenic AdV, CMV, EBV, BK and HHV-6 antigens. Similar to healthy donor-derived T cell lines (n=5, 3 specific for 4 viruses and 2 for 5 viruses), all 5 MDS patient-derived lines demonstrated specificity for one or more of the target viruses (1 for 5 viruses, 1 for 4, 2 for 3 and 1 for 1 virus) as observed by IFNγ ELISpot assay with comparable magnitude (range Adv: 43-730 vs 384-941 in healthy donors, CMV: 0-1599 vs 0-3002, EBV: 0-1486 vs 0-541, BK: 0-839 vs 38-275 and HHV6: 0-794 vs 5-407 SFU/2x105 cells, respectively). We next examined the feasibility of expanding autologous MDS-antigen directed T cell products (n=10) to determine whether an adoptive immunotherapeutic approach might be applicable for MDS treatment. Thus, we exposed patient-derived PBMCs to autologous dendritic cells (DC) loaded with pepmixes spanning 6 MDS-associated antigens (CCNA1, survivin, WT1, PRAME, PR3 and NYESO1). After 3 rounds of stimulation, the products obtained were predominantly CD3+ T cells (mean 88±1.3%) that were polyclonal (CD4: 46±5% and CD8: 41±4%) containing predominantly memory T cells (TEM: 36±6% TCM: 37±5% and Tnaïve =13±3%). Six lines (60%) showed specific recognition to at least one of the target antigens: 4 lines specific for PRAME, 1 for CCNA1, 1 for WT1 and 1 for NYESO1 (range 0-40, 0-184, 0-1386 and 0-179 SFU/2x105 cells, respectively by IFNγ ELIspot). T cell lines were capable of specifically secreting multiple effector cytokines in response to targets (TNFα: 12% and IFNγ: 16% in response to PRAME in a representative patient-derived T cell line). Furthermore, this line was capable of killing PRAME+ targets in a 4hr 51Cr release assay [60% specific lysis, E:T 20:1]. In conclusion, functional virus-directed T cell immunity in patients with MDS is preserved, potentially explaining the lower rates of viral reactivation seen in these patients compared with other infections. Moreover, T cells specific for MDS-expressed tumor antigens can also be successfully expanded ex vivo from patients. Taken together this raises the possibility of applying an adoptive immunotherapeutic approach for the treatment of MDS. Disclosures Ramos: Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Tessa Therapeutics: Research Funding. Leen:Allovir: Consultancy, Other: Cofounder, Ownership Interest; Marker Therapeutics: Consultancy, Other: Cofounder, Ownership Interest.

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)

scholarly journals Patient-derived hepatitis C virus and JFH-1 clones differ in their ability to infect human hepatoma cells and lymphocytes

2012 ◽  
Vol 93 (11) ◽  
pp. 2399-2407 ◽  
Author(s):  
Mohammed A. Sarhan ◽  
Annie Y. Chen ◽  
Rodney S. Russell ◽  
Tomasz I. Michalak
Keyword(s):  
T Cells ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
T Cell ◽  
Cell Lines ◽  
Productive Infection ◽  
Wild Type Virus ◽  
Human Hepatoma ◽  
Naturally Occurring ◽  
T Cell Lines

Hepatitis C virus (HCV) is a hepatotropic virus that also infects cells of the immune system. HCV clones cultivated in human hepatoma Huh-7.5 cells have significantly advanced our understanding of HCV replication and candidate hepatocyte receptors. However, naturally occurring patient-derived HCV, in contrast to the HCV JFH-1 clone, is unable to infect Huh-7.5 cells, while it can replicate in human primary T-cells and selected T-cell lines. To better understand this incongruity, we examined the susceptibility of primary T-cells, PBMCs and T-cell lines to infection with patient-derived HCV, the classical HCV JFH-1 and a cell culture-adapted JFH1T known to be highly infectious to Huh-7.5 cells. We also tested whether Huh-7.5 cells are prone to virus readily infecting T-lymphocytes. The results revealed that while primary T-cells and Molt4 and Jurkat T-cell lines were susceptible to patient-derived HCV, they were resistant to infection with either JFH1T or JFH-1. However, the JFH1T clone interacted more firmly, although non-productively, with the cells than JFH-1. Further, Huh-7.5 cells robustly supported replication of JFH1T but not patient-derived, wild-type virus, despite using highly sensitive detection assays. In conclusion, JFH-1 and JFH1T clones were unable to establish productive infection in human primary T-cells, PBMCs and T-cell lines known to be prone to infection by patient-derived HCV, while Huh-7.5 cells were resistant to infection with naturally occurring virus infecting immune cells. The data showed that the ability to infect lymphocytes is a characteristic of native virus but not laboratory HCV clones.

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