Immunosuppression Modulates Immune Responses to AAV Capsid in Human Subjects Undergoing Intramuscular Gene Transfer for Lipoprotein Lipase Deficiency

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 822-822 ◽  
Author(s):  
Daniel J Hui ◽  
Federico Mingozzi ◽  
Annemarie Kleefstra ◽  
Janneke M Meulenberg ◽  
Shyrie Edmonson ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
T Cell ◽  
Gene Transfer ◽  
Immune Responses ◽  
T Cell Responses ◽  
T Cell Response ◽  
Safety And Efficacy ◽  
Cell Responses ◽  
Ifn Γ

Abstract Administration of adeno-associated viral vectors (AAV) has resulted in long-term therapeutic gene transfer in multiple large animal models of disease, but attempts to translate systemic administration of AAV to humans have been limited in some cases by an immune response to the vector capsid (Nature Med12:342–7, 2006; Nature Med13:419–422, 2007). To overcome this obstacle, we have proposed that a short course of immunosuppression (IS) be administered with vector injection. Here we report the safety and efficacy results of this maneuver in a trial of AAV-1 administered to skeletal muscle. Lipoprotein lipase (LPL) deficiency is a familial disorder in which insufficient levels of LPL enzyme result in the accumulation of triglycerides in plasma. In a clinical study to correct this disorder, an AAV-1 vector encoding the therapeutic transgene LPL was administered to the skeletal muscle of affected individuals. Eight subjects were assigned to two dose cohorts, receiving 1×1011 genome copies (gc)/kg or 3×1011gc/kg. In this study, one subject receiving the high vector dose experienced a transient, asymptomatic increase in the muscle enzyme creatinine phosphokinase beginning 4 weeks after gene transfer, persisting for several weeks. This was associated with capsid-specific CD4+ and CD8+ T cell activation detectable by IFN-γ ELISPOT and intracellular cytokine staining on PBMC. In total, a T cell response to the AAV capsid, but not to the LPL transgene, was detectable in 4/8 subjects. In some of these subjects, T cell responses were detectable in peripheral blood up to 2 years after gene transfer. To prevent potentially harmful immune responses directed to the AAV capsid, a follow up study in LPL deficient subjects was initiated in which a 12-week regimen of mycophenolate mofetil and cyclosporine A was administered orally starting at the time of AAV-1 intramuscular gene transfer. Two additional subjects were administered AAV-1-LPL in the absence of immunosuppression, to compare the safety and efficacy of two different vector production methods. Overall, IS was well tolerated and no adverse events were reported. At a dose of 3×1011 gc/kg, IS effectively blocked T cell responses to capsid, which were undetectable by IFN-γ ELISPOT in 4/4 subjects, even after IS was discontinued. However, at a dose of 1×1012gc/kg, a delayed IFN-γ response to capsid antigen was observed in 3/5 subjects. In two subjects the T cell response was still detectable after IS was discontinued. T cell responses did not correlate with pre-existing antibody titers in any of the subjects, as positivity for antibodies against the AAV capsid was not predictive of ELISPOT results. Antibody analysis revealed that IS did not have any effect on the development of antibodies against AAV-1 capsid, as all subjects developed humoral immunity against capsid, with predominance of IgG1 antibody subclass. None of the subjects receiving IS developed humoral or cellular immunity to the LPL transgene product. In conclusion, the use of IS in the context of AAV-1 gene transfer for LPL deficiency is safe and at least partially effective in blocking T cell responses directed to the capsid antigen. Ongoing long-term evaluation of transgene expression in these subjects will allow further assessment of the effects of IS on efficacy of gene transfer.

scholarly journals T Cell-Mediated Immune Responses to AAV and AAV Vectors

2021 ◽  
Vol 12 ◽  
Author(s):  
Hildegund C. J. Ertl
Keyword(s):  
T Cell ◽  
Gene Transfer ◽  
Immune Responses ◽  
De Novo ◽  
T Cell Responses ◽  
Adeno Associated Virus ◽  
Effector Functions ◽  
Cell Responses ◽  
High Doses

Adeno-associated virus (AAV)-mediated gene transfer has benefited patients with inherited diseases, such as hemophilia B, by achieving long-term expression of the therapeutic transgene. Nevertheless, challenges remain due to rejection of AAV-transduced cells, which in some, but not all, patients can be prevented by immunosuppression. It is assumed that CD8+ T cells induced by natural infections with AAVs are recalled by the AAV vector’s capsid and upon activation eliminate cells expressing the degraded capsid antigens. Alternatively, it is feasible that AAV vectors, especially if given at high doses, induce de novo capsid- or transgene product-specific T cell responses. This chapter discusses CD8+ T cell responses to AAV infections and AAV gene transfer and avenues to prevent their activation or block their effector functions.

scholarly journals AAV Vector Dose Determines TLR9 Dependence of CD8+ T Cell Response to Transgene Product

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 3-3
Author(s):  
Ning Li ◽  
Thais Bertolini ◽  
Roland W Herzog
Keyword(s):  
T Cells ◽  
T Cell ◽  
Gene Transfer ◽  
Immune Responses ◽  
Cell Response ◽  
T Cell Responses ◽  
Cd8 T Cell ◽  
T Cell Response ◽  
Innate Immune ◽  
Cell Responses

Adeno-associated viral (AAV) vectors are currently evaluated in multiple Phase III clinical trial for the treatment of hemophilia and neuromuscular disorders. A major concern is the potential for immune responses. Viral vectors are initially sensed by the innate immune system, which shapes subsequent adaptive immune responses. Particularly, toll-like receptors (TLRs) have been reported as major sensors of pathogens during innate immune response. TLRs recognize pathogen-associated molecular patterns (PAMPs). Our previous studies found that cross-priming of AAV capsid-specific CD8+ T cells depended on TLR9-MyD88 pathway. TLR9 is an endosomal DNA receptor that responds most potently to unmethylated CpG motifs as found in bacterial and viral DNA. Similarly, others documented TLR9-dependent CD8+ T cell responses against non-secreted transgene products such as LacZ and hemagglutinin upon muscle-directed AAV gene transfer. Similarly, we published that CD8+ T cell responses to a secreted ovalbumin (ova) transgene product were substantially reduced (although not entirely eliminated) upon muscle gene transfer in TLR9-deficient mice [J Innate Immun. 7:302-14]. For those studies, we had used a self-complementary scAAV genomes, which we found to more strongly activate TLR9 than conventional single-stranded ssAAV vectors. Here, we performed intramuscular injections of 3 doses of ssAAV1-CMV-ova vector (2X1010, 2X1011 and1X1012 vg) in wild-type (WT), TLR9-/-, or MYD88-/- C57BL/6 mice. Using MHC tetramer (H2-Kb -SIINFEKL), ova-specific CD8+ T cell frequencies were monitored in peripheral blood for up to 6 weeks. As expected from prior studies, TLR9-/- mice showed a substantially reduced response (1.2% tetramer+ of CD8) at the low dose when compared to WT (12% tetramer+ of CD8) animals (p<0.0001, n=5/group). To our surprise, CD8+ T cell responses were similar in TLR9-/- and WT mice at the 2 higher doses. TLR9-/- mice displayed 16% and 3.3% tetramer+ of CD8 frequencies at the median and the high doses, respectively; which was comparable to WT mice, where 15% and 4.8% tetramer+ of CD8 frequencies were observed (n=5/group). Therefore, sensing of the AAV genome by TLR9 is more critical for the CD8+ T cell response to the secreted transgene product at lower vector doses (possibly related to the lower levels of transgene expression). Interestingly, transgene product-specific CD8+ T cell responses were much reduced in MyD88-/- mice, in which 0.2% and 1.7% tetramer+ of CD8 frequencies were found for low and median doses. Therefore, an alternative signaling pathway that includes the MyD88 adaptor molecule likely exists that is more critical than TLR9 above a certain level of expression. The reduced strength of the CD8+ T cell response seen at the highest vector dose compared to the medium dose may be explained by a transient increase in FoxP3+ Treg and in PD-1+ T cells that we observed 1 week after gene transfer and that was significantly greater at the highest vector dose. In related experiments, we performed intramuscular gene transfer using a ssAAV1-EF1a-FIX vector in hemophilia B mice (C3H/HeJ F9-/-, 1x1011 vg/mouse). Here, we used either a vector with native sequences or with an expression cassette that was entirely devoid of CpG motifs (and there stimulates TLR9 less effectively). CpG depletion did not have substantial effects on antibody formation against human FIX or the viral capsid. However, CD8+ T cell infiltrates in skeletal muscle were markedly reduced but not entirely eliminated when tissue sections were examined 1 month after gene transfer. In conclusion, TLR9 signaling is one important factor in the activation of transgene product-specific CD8+ T cells in AAV gene transfer, but other pathways exist that may be more critical depending on vector dose or levels of expression. Disclosures Herzog: Takeda Pharmaceuticals: Patents & Royalties.

scholarly journals Potent HIV-1-Specific CD8 T Cell Responses Induced in Mice after Priming with a Multiepitopic DNA-TMEP and Boosting with the HIV Vaccine MVA-B

Viruses ◽  
2018 ◽  
Vol 10 (8) ◽  
pp. 424 ◽  
Author(s):  
Beatriz Perdiguero ◽  
Suresh C. Raman ◽  
Cristina Sánchez-Corzo ◽  
Carlos Oscar S. Sorzano ◽  
José Ramón Valverde ◽  
...  
Keyword(s):  
T Cell ◽  
Immune Responses ◽  
T Cell Responses ◽  
Cd8 T Cell ◽  
T Cell Response ◽  
Effective Vaccine ◽  
T Cell Epitopes ◽  
Cell Responses ◽  
Modified Vaccinia Virus Ankara ◽  
Hiv 1

An effective vaccine against Human Immunodeficiency Virus (HIV) still remains the best solution to provide a sustainable control and/or eradication of the virus. We have previously generated the HIV-1 vaccine modified vaccinia virus Ankara (MVA)-B, which exhibited good immunogenicity profile in phase I prophylactic and therapeutic clinical trials, but was unable to prevent viral rebound after antiretroviral (ART) removal. To potentiate the immunogenicity of MVA-B, here we described the design and immune responses elicited in mice by a new T cell multi-epitopic B (TMEP-B) immunogen, vectored by DNA, when administered in homologous or heterologous prime/boost regimens in combination with MVA-B. The TMEP-B protein contained conserved regions from Gag, Pol, and Nef proteins including multiple CD4 and CD8 T cell epitopes functionally associated with HIV control. Heterologous DNA-TMEP/MVA-B regimen induced higher HIV-1-specific CD8 T cell responses with broader epitope recognition and higher polyfunctional profile than the homologous DNA-TMEP/DNA-TMEP or the heterologous DNA-GPN/MVA-B combinations. Moreover, higher HIV-1-specific CD4 and Tfh immune responses were also detected using this regimen. After MVA-B boost, the magnitude of the anti-VACV CD8 T cell response was significantly compromised in DNA-TMEP-primed animals. Our results revealed the immunological potential of DNA-TMEP prime/MVA-B boost regimen and supported the application of these combined vectors in HIV-1 prevention and/or therapy.

scholarly journals Sustained Specific and Cross-Reactive T Cell Responses to Zika and Dengue Virus NS3 in West Africa

2018 ◽  
Vol 92 (7) ◽  
Author(s):  
Bobby Brooke Herrera ◽  
Wen-Yang Tsai ◽  
Charlotte A. Chang ◽  
Donald J. Hamel ◽  
Wei-Kung Wang ◽  
...  
Keyword(s):  
T Cell ◽  
Immune Responses ◽  
Dengue Virus ◽  
Zika Virus ◽  
Cell Response ◽  
Vaccine Development ◽  
T Cell Responses ◽  
T Cell Response ◽  
Zikv Infection ◽  
Cell Responses

ABSTRACT Recent studies on the role of T cells in Zika virus (ZIKV) infection have shown that T cell responses to Asian ZIKV infection are important for protection, and that previous dengue virus (DENV) exposure amplifies the protective T cell response to Asian ZIKV. Human T cell responses to African ZIKV infection, however, remain unexplored. Here, we utilized the modified anthrax toxin delivery system to develop a flavivirus enzyme-linked immunosorbent spot (ELISPOT) assay. Using human ZIKV and DENV samples from Senegal, West Africa, our results demonstrate specific and cross-reactive T cell responses to nonstructural protein 3 (NS3). Specifically, we found that T cell responses to NS3 protease are ZIKV and DENV specific, but responses to NS3 helicase are cross-reactive. Sequential sample analyses revealed immune responses sustained many years after infection. These results have important implications for African ZIKV/DENV vaccine development, as well as for potential flavivirus diagnostics based on T cell responses. IMPORTANCE The recent Zika virus (ZIKV) epidemic in Latin America and the associated congenital microcephaly and Guillain-Barré syndrome have raised questions as to why we have not recognized these distinct clinical diseases in Africa. The human immunologic response to ZIKV and related flaviviruses in Africa represents a research gap that may shed light on the mechanisms contributing to protection. The goal of our study was to develop an inexpensive assay to detect and characterize the T cell response to African ZIKV and DENV. Our data show long-term specific and cross-reactive human immune responses against African ZIKV and DENV, suggesting the usefulness of a diagnostic based on the T cell response. Additionally, we show that prior flavivirus exposure influences the magnitude of the T cell response. The identification of immune responses to African ZIKV and DENV is of relevance to vaccine development.

scholarly journals Priming with Recombinant BCG Expressing HTI Enhances the Magnitude and Breadth of the T-Cell Immune Responses Elicited by MVA.HTI in BALB/c Mice

Vaccines ◽  
2020 ◽  
Vol 8 (4) ◽  
pp. 678
Author(s):  
Narcís Saubi ◽  
Athina Kilpeläinen ◽  
Yoshiki Eto ◽  
Chun-Wei Chen ◽  
Àlex Olvera ◽  
...  
Keyword(s):  
T Cell ◽  
Immune Responses ◽  
T Cell Responses ◽  
Vaccine Platform ◽  
Cell Responses ◽  
Total Magnitude ◽  
T Cell Immune Responses ◽  
Ifn Γ ◽  
Hiv 1 ◽  
Recombinant Bcg

The use of Mycobacterium bovis bacillus Calmette–Guérin (BCG) as a live vaccine vehicle is a promising approach for HIV-1-specific T-cell induction. In this study, we used recombinant BCG expressing HIVACAT T-cell immunogen (HTI), BCG.HTI2auxo.int. BALB/c mice immunization with BCG.HTI2auxo.int prime and MVA.HTI boost was safe and induced HIV-1-specific T-cell responses. Two weeks after boost, T-cell responses were assessed by IFN-γ ELISpot. The highest total magnitude of IFN-γ spot-forming cells (SFC)/106 splenocytes was observed in BCG.HTI2auxo.int primed mice compared to mice receiving MVA.HTI alone or mice primed with BCGwt, although the differences between the vaccination regimens only reached trends. In order to evaluate the differences in the breadth of the T-cell immune responses, we examined the number of reactive peptide pools per mouse. Interestingly, both BCG.HTI2auxo.int and BCGwt primed mice recognized an average of four peptide pools per mouse. However, the variation was higher in BCG.HTI2auxo.int primed mice with one mouse recognizing 11 peptide pools and three mice recognizing few or no peptide pools. The recognition profile appeared to be more spread out for BCG.HTI2auxo.int primed mice and mice only receiving MVA.HTI. Here, we describe a useful vaccine platform for priming protective responses against HIV-1/TB and other prevalent infectious diseases.

Characterization of the Immune Response to Canine Factor IX Following AAV-Mediated Intravascular Gene Delivery to Skeletal Muscle in Hemophilia B Dogs.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1297-1297
Author(s):  
Daniel J. Hui ◽  
Federico Mingozzi ◽  
Denise E. Sabatino ◽  
Stephanie McCorquodale ◽  
Aaron Dillow ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Immune Response ◽  
T Cell ◽  
T Cell Responses ◽  
Route Of Administration ◽  
Factor Ix ◽  
Hemophilia B ◽  
Cell Responses ◽  
Chapel Hill ◽  
Ifn Γ

Abstract Progress towards an effective gene therapy for hemophilia B (HB) has been facilitated by large animal studies. Previous work has shown intravascular delivery of an adeno-associated viral (AAV) serotype 2 vector expressing canine factor IX (cF.IX) to skeletal muscle in HB dogs resulted in long-term expression of cF.IX at levels of 4–20% of normal, which nearly corrected the disease phenotype. However, occurrence of inhibitors to F.IX in some animals raises concerns of a potential immune response to the transgene product, prompting a more thorough examination of T cell responses in this setting. Early work revealed that transient immunosuppression (IS) with cyclophosphamide was required to avoid inadvertent antibody formation to F.IX. Here we report in detail the nature and the duration of T cell responses against the transgene product. Six HB dogs from the Chapel Hill colony received AAV2 vector at three different doses (1x1012 vg/kg, n=3; 3 x 1012 vg/kg, n=2; 8 x 1012 vg/kg, n=1) in addition to weekly infusion with cyclophosphamide (6 doses total). PBMCs were isolated from whole blood prior to vector infusion, during IS and after removal from IS, and used to measure the T cell response to F.IX by ELISpot assay for IL-10 and IFN-γ secretion using a cF.IX peptide library composed of 15-mers overlapping by 10 amino acids, spanning the entire protein sequence. Peptides were arranged into a matrix of pools, such that each peptide was contained in two orthogonal pools. Interestingly, in the IL-10 assay, one common T cell epitope corresponding to peptide 68 in the cF.IX library was found in all intravascularly-administered dogs from each of the three dose groups. The same epitope was also detectable in naïve HB dogs. Another epitope, corresponding to peptide 84, was found in a dog injected with the highest dose of vector after it developed a non-neutralizing antibody response against the cF.IX transgene product. Peptide 84 spans the region of the protein that contains the missense mutation responsible for HB (Chapel Hill mutation, Glu379 → Gly), which is a key difference in the newly introduced transgene product. Furthermore, the lack of any IFN-γ secretion coupled with the marked IL-10 response gives a cytokine profile that is characteristic of a Th2 response. This is in contrast with the Th1 response seen in previous studies with direct intramuscular injection of an AAV serotype 1 expressing cF.IX in dogs from the same colony. IS successfully reduced T cell responses to undetectable levels, while IL-10 secretion was detectable in PBMCs before vector delivery and one month after IS was discontinued. Overall circulating cF.IX levels did not seem to be affected by late restoration of T cells responses. No T cell responses against AAV capsid were detectable by ELISpot on PBMCs in any of the dogs studied. Interestingly, the relatively mild IS regimen also appeared to reduce the formation of neutralizing antibodies against AAV capsid, regardless of the route of administration. In summary, the nature of T cell responses (Th2 vs. Th1) suggests that route of administration, and/or AAV serotype, may play a role in the determination of the immune response elicited. We conclude that IS may provide a means to decrease T cell responses to the transgene following intravascular delivery of AAV-F.IX to skeletal muscle.

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.

EBV-positive lymphoma patients have a selective deficiency in EBV immunity

2007 ◽  
Vol 25 (18_suppl) ◽  
pp. 21032-21032
Author(s):  
K. N. Heller ◽  
P. G. Steinherz ◽  
C. S. Portlock ◽  
C. Münz
Keyword(s):  
Immune Response ◽  
T Cells ◽  
T Cell ◽  
Immune Responses ◽  
Healthy Volunteers ◽  
Cell Response ◽  
T Cell Responses ◽  
T Cell Response ◽  
Specific Immune Response ◽  
Cell Responses

21032 Background: Epstein-Barr virus (EBV) asymptomatically establishes persistent infections in more than 90% of the adult population. However, due to effective immune control, only a minority of infected carriers develops spontaneous EBV-associated lymphomas. Since EBV nuclear antigen-1 (EBNA1) is the only protein expressed in all proliferating EBV infected cells we hypothesize that EBNA1 specific immune response is critical in preventing EBV-positive lymphomas. Methods: After informed consent, peripheral blood from healthy volunteers and lymphoma patients (prior to therapy- no evidence of cytopenia) were stimulated (ex vivo) with overlapping peptides covering the immunogenic EBNA1 (aa400–641) sequence. Frequency of EBNA1-specific T-cells were assessed by intracellular cytokine staining and flow cytometric proliferation assays. Cytokine pattern, surface marker phenotype and functional reactivity against EBV specific and control antigens were analyzed. Results: Patient and volunteer immune responses to control antigens and other viruses were assessed and statistically indistinguishable. EBNA1 specific CD4+ T cell responses were detected among 18 of 20 healthy carriers, and among 10 of 16 patients with EBV-negative lymphoma (relative to healthy volunteers p=0.145 via paired student T test). None of the patients with EBV-positive lymphomas (n=8) had a detectable EBNA1-specific CD4+ T-cell response (p<0.003 relative to healthy volunteers and patients with EBV-negative lymphomas). Conclusions: Healthy volunteers and patients with EBV-negative lymphoma have statistically similar EBNA1-specific CD4+ T cell responses. Although patients with EBV-positive lymphoma have intact immune responses to common viruses and antigens, they selectively lack an EBNA1-specific CD4+ T cell response. An intact EBNA1 specific immune response among patients with EBV-negaitve lymphoma implies that lymphoma is not a cause of a selective immune deficiency. On the contrary, these findings suggest that EBNA1-specific CD4+ T cells are critical in the prevention of EBV mediated lymphomas, and a defect in EBNA1 specific immunity may leave EBV carriers suseptible to EBV-positive lymphomas. EBNA1- specific CD4+ T cell function may be a new target for therapies of EBV-associated malignancies. No significant financial relationships to disclose.

Allogeneic partially HLA-matched dendritic cells (DC) as a vaccine in metastatic renal cell cancer (mRCC): Final analysis of a clinical phase I/II study.

2012 ◽  
Vol 30 (15_suppl) ◽  
pp. e15053-e15053 ◽  
Author(s):  
Anne Flörcken ◽  
Joachim Kopp ◽  
Kamran Movassaghi ◽  
Antje van Lessen ◽  
Anna Takvorian ◽  
...  
Keyword(s):  
T Cell ◽  
Immune Responses ◽  
Clinical Response ◽  
Cell Cancer ◽  
T Cell Responses ◽  
T Cell Response ◽  
Delayed Type Hypersensitivity ◽  
Autologous Tumor ◽  
Tumor Lysate ◽  
Cell Responses

e15053 Background: Despite novel kinase inhibitors, prognosis of metastatic RCC remains poor and new experimental approaches are warranted. Our aim was to evaluate a DC-based vaccine, which exploits alloreactivity as a means to amplify specific anti-tumor immune responses. Methods: Allogeneic, partially HLA-matched DC were generated in our GMP facility. DC were loaded with autologous tumor lysate. 8 patients with progressive mRCC were included, 7 patients were immunized repetitively with 107 DC s.c. over 20 weeks. Low-dose IL-2 (3 Mio U s.c. qd) was used concomitantly. Endpoints of the study were feasibility, safety, immunological and clinical responses. T cell responses against HLA-A2-restricted RCC-associated antigens were evaluated by proliferation assays, ELISpot and cytokine bead array (CBA). T cell repertoire was analysed by T cell receptor γ and –β PCR. Results: Vaccination was feasible and safe, no treatment-related grade 3/4 toxicity or clinically relevant autoimmunity was observed. No objective responses were observed, however, 2/7 patients showed stable disease, one a minimal clinical response. The mean TTP was 24.6 weeks (range 5 to 96). Delayed-type hypersensitivity was detected in 3/7 and HLA antibodies were induced in 3/7 patients. In 3/7 patients T cell responses against RCC-associated antigens such as TYMS, G250, vimentin, surviving and cyclin-D1 were induced by vaccination. These antigen-specific T cells showed a predominant TH1-cytokine profile. Interestingly, a clonally expanded T cell population could be detected by γ- and –β PCR in one patient with both a minimal clinical response and a T cell response. This clone is currently persisting for more than 80 months, its specificity is under investigation. Conclusions: Vaccination with allogeneic tumor-lysate-loaded DC was feasible, safe and was able to induce TH1-polarized immune responses against RCC-associated antigens. Tumor vaccination might be a promising approach in minimal residual disease, possibly in combination with antibodies against CTLA-4 or PD-1.

scholarly journals T-Cell Immune Response Assessment as a Complement to Serology and Intranasal Protection Assays in Determining the Protective Immunity Induced by Acellular Pertussis Vaccines in Mice

2003 ◽  
Vol 10 (4) ◽  
pp. 637-642 ◽  
Author(s):  
C. M. Ausiello ◽  
R. Lande ◽  
P. Stefanelli ◽  
C. Fazio ◽  
G. Fedele ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cell ◽  
Immune Responses ◽  
Pertussis Toxin ◽  
Protective Immunity ◽  
T Cell Responses ◽  
Additional Tool ◽  
Cell Responses ◽  
T Cell Immune Responses ◽  
Ifn Γ

ABSTRACT The relative value of antibodies and/or T-cell immune responses to Bordetella pertussis antigens in the immunity induced by acellular pertussis (aP) vaccines is still an open issue, probably due to the incomplete knowledge on the mechanisms of protective immunity to pertussis. The relevance of T-cell immune responses in protection from pertussis has been demonstrated in murine and human models of infection; thus, in this study, the ability of different vaccine preparations of three component (pertussis toxin, filamentous hemagglutinin, and pertactin) aP vaccines to induce T-cell responses was investigated in mice. All vaccine preparations examined passed the immunogenicity control test, based on antibody titer assessment, according to European Pharmacopoeia standards, and protected mice from B. pertussis intranasal challenge, but not all preparations were able to prime T cells to pertussis toxin, the specific B. pertussis antigen. In particular, one vaccine preparation was unable to induce proliferation and gamma interferon (IFN-γ) production while the other two gave borderline results. The evaluation of T-cell responses to pertussis toxin antigen may provide information on the protective immunity induced by aP vaccines in animal models. Considering the critical role of the axis interleukin-12-IFN-γ for protection from pertussis, our results suggest that testing the induction of a key protective cytokine such as IFN-γ could be an additional tool for the evaluation of the immune response induced by aP vaccines.

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