Impact of AAV Capsid-Specific T-Cell Responses on Design and Outcome of Clinical Gene Transfer Trials with Recombinant Adeno-Associated Viral Vectors: An Evolving Controversy

2017 ◽  
Vol 28 (4) ◽  
pp. 328-337 ◽  
Author(s):  
Hildegund C.J. Ertl ◽  
Katherine A. High
Keyword(s):  
T Cell ◽  
Gene Transfer ◽  
Viral Vectors ◽  
T Cell Responses ◽  
Cell Responses

scholarly journals Optimising T cell (re)boosting strategies for adenoviral and modified vaccinia Ankara vaccine regimens in humans

npj Vaccines ◽  
2020 ◽  
Vol 5 (1) ◽  
Author(s):  
Stefania Capone ◽  
Anthony Brown ◽  
Felicity Hartnell ◽  
Mariarosaria Del Sorbo ◽  
Cinzia Traboni ◽  
...  
Keyword(s):  
T Cells ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
T Cell ◽  
Viral Vectors ◽  
Standard Dose ◽  
T Cell Responses ◽  
Effector Memory ◽  
Cell Responses ◽  
Wide Range

Abstract Simian adenoviral and modified vaccinia Ankara (MVA) viral vectors used in heterologous prime-boost strategies are potent inducers of T cells against encoded antigens and are in advanced testing as vaccine carriers for a wide range of infectious agents and cancers. It is unclear if these responses can be further enhanced or sustained with reboosting strategies. Furthermore, despite the challenges involved in MVA manufacture dose de-escalation has not been performed in humans. In this study, healthy volunteers received chimpanzee-derived adenovirus-3 and MVA vaccines encoding the non-structural region of hepatitis C virus (ChAd3-NSmut/MVA-NSmut) 8 weeks apart. Volunteers were then reboosted with a second round of ChAd3-NSmut/MVA-NSmut or MVA-NSmut vaccines 8 weeks or 1-year later. We also determined the capacity of reduced doses of MVA-NSmut to boost ChAd3-NSmut primed T cells. Reboosting was safe, with no enhanced reactogenicity. Reboosting after an 8-week interval led to minimal re-expansion of transgene-specific T cells. However, after a longer interval, T cell responses expanded efficiently and memory responses were enhanced. The 8-week interval regimen induced a higher percentage of terminally differentiated and effector memory T cells. Reboosting with MVA-NSmut alone was as effective as with ChAd3-NSmut/MVA-NSmut. A ten-fold lower dose of MVA (2 × 107pfu) induced high-magnitude, sustained, broad, and functional Hepatitis C virus (HCV)-specific T cell responses, equivalent to standard doses (2 × 108 pfu). Overall, we show that following Ad/MVA prime-boost vaccination reboosting is most effective after a prolonged interval and is productive with MVA alone. Importantly, we also show that a ten-fold lower dose of MVA is as potent in humans as the standard dose.

scholarly journals 730. T Cell Responses to AAV Vector Capsid in Normal Donors and Subjects Who Have Undergone Liver-Directed AAV-Mediated Gene Transfer

2006 ◽  
Vol 13 ◽  
pp. S282 ◽  
Author(s):  
Marcela V. Maus ◽  
Federico Mingozzi ◽  
Denise E. Sabatino ◽  
Daniel Hui ◽  
Margaret V. Ragni ◽  
...  
Keyword(s):  
T Cell ◽  
Gene Transfer ◽  
T Cell Responses ◽  
Aav Vector ◽  
Cell Responses

scholarly journals Activation-induced Markers Detect Vaccine-Specific CD4+ T Cell Responses Not Measured by Assays Conventionally Used in Clinical Trials

Vaccines ◽  
2018 ◽  
Vol 6 (3) ◽  
pp. 50 ◽  
Author(s):  
Georgina Bowyer ◽  
Tommy Rampling ◽  
Jonathan Powlson ◽  
Richard Morter ◽  
Daniel Wright ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Response ◽  
Viral Vectors ◽  
Comprehensive Evaluation ◽  
T Cell Responses ◽  
T Cell Response ◽  
Cell Responses ◽  
Vaccine Immunogenicity ◽  
Sensitivity Specificity

Immunogenicity of T cell-inducing vaccines, such as viral vectors or DNA vaccines and Bacillus Calmette-Guérin (BCG), are frequently assessed by cytokine-based approaches. While these are sensitive methods that have shown correlates of protection in various vaccine studies, they only identify a small proportion of the vaccine-specific T cell response. Responses to vaccination are likely to be heterogeneous, particularly when comparing prime and boost or assessing vaccine performance across diverse populations. Activation-induced markers (AIM) can provide a broader view of the total antigen-specific T cell response to enable a more comprehensive evaluation of vaccine immunogenicity. We tested an AIM assay for the detection of vaccine-specific CD4+ and CD8+ T cell responses in healthy UK adults vaccinated with viral vectored Ebola vaccine candidates, ChAd3-EBO-Z and MVA-EBO-Z. We used the markers, CD25, CD134 (OX40), CD274 (PDL1), and CD107a, to sensitively identify vaccine-responsive T cells. We compared the use of OX40+CD25+ and OX40+PDL1+ in CD4+ T cells and OX40+CD25+ and CD25+CD107a+ in CD8+ T cells for their sensitivity, specificity, and associations with other measures of vaccine immunogenicity. We show that activation-induced markers can be used as an additional method of demonstrating vaccine immunogenicity, providing a broader picture of the global T cell response to vaccination.

scholarly journals Virus-based vaccine vectors with distinct replication mechanisms differentially infect and activate dendritic cells

npj Vaccines ◽  
2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Carolina Chiale ◽  
Anthony M. Marchese ◽  
Yoichi Furuya ◽  
Michael D. Robek
Keyword(s):  
Dendritic Cells ◽  
T Cell ◽  
Virus Replication ◽  
T Cell Activation ◽  
Viral Vectors ◽  
Cell Activation ◽  
T Cell Responses ◽  
Cell Responses ◽  
Identical Cell

AbstractThe precise mechanism by which many virus-based vectors activate immune responses remains unknown. Dendritic cells (DCs) play key roles in priming T cell responses and controlling virus replication, but their functions in generating protective immunity following vaccination with viral vectors are not always well understood. We hypothesized that highly immunogenic viral vectors with identical cell entry pathways but unique replication mechanisms differentially infect and activate DCs to promote antigen presentation and activation of distinctive antigen-specific T cell responses. To evaluate differences in replication mechanisms, we utilized a rhabdovirus vector (vesicular stomatitis virus; VSV) and an alphavirus-rhabdovirus hybrid vector (virus-like vesicles; VLV), which replicates like an alphavirus but enters the cell via the VSV glycoprotein. We found that while virus replication promotes CD8+ T cell activation by VLV, replication is absolutely required for VSV-induced responses. DC subtypes were differentially infected in vitro with VSV and VLV, and displayed differences in activation following infection that were dependent on vector replication but were independent of interferon receptor signaling. Additionally, the ability of the alphavirus-based vector to generate functional CD8+ T cells in the absence of replication relied on cDC1 cells. These results highlight the differential activation of DCs following infection with unique viral vectors and indicate potentially discrete roles of DC subtypes in activating the immune response following immunization with vectors that have distinct replication mechanisms.

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.

Indoleamine 2,3-dioxygenase gene transfer prolongs cardiac allograft survival

2007 ◽  
Vol 293 (6) ◽  
pp. H3415-H3423 ◽  
Author(s):  
Jianping Li ◽  
Andrea Meinhardt ◽  
Marc-Estienne Roehrich ◽  
Dela Golshayan ◽  
Jean Dudler ◽  
...  
Keyword(s):  
T Cell ◽  
Gene Transfer ◽  
T Cell Responses ◽  
Cardiac Allograft ◽  
Cardiac Transplant ◽  
Donor Heart ◽  
Gene Encoding ◽  
Cell Responses ◽  
Transplant Model

Cells that express indoleamine 2,3-dioxygenase (IDO), the rate-limiting enzyme in the catabolism of tryptophan, suppress T cell responses and promote immunological tolerance. However, their role in solid organ transplantation is incompletely understood. We analyzed T cell responses to allogeneic dendritic cells (DCs) genetically modified to express the gene encoding IDO in vitro and IDO gene transfer into the donor heart in a cardiac transplant model in vivo. Bone marrow-derived DCs transduced with the gene encoding IDO produced active IDO protein. This was associated with decreased stimulation of allogeneic T cell proliferation in the mixed leukocyte reaction in vitro. In a cardiac transplant model, adenovirus-mediated IDO gene transfer into the donor heart resulted in transgene expression predominantly in cardiomyocytes. Fischer-344 rat donor hearts transduced with the gene encoding IDO survived for longer periods of time when placed in Lewis rat recipients compared with control vector or vehicle alone [median survival times of 17 (range: 12–22) days vs. 10 (range: 8–14) and 9 (range: 8–13) days, respectively, P < 0.0001]. IDO gene transfer combined with low-dose cyclosporin A (CsA) was more effective than CsA alone ( P < 0.05). Numbers of monocytes/macrophages, CD4+ cells, and CD8α+ cells infiltrating the graft as well as intragraft cytokine transcript levels for IFN-γ, IL-1, TNF-α, regulated upon secretion, normal T cell expressed, and secreted/chemokine (C-C motif) ligand 5 were decreased after IDO gene transfer ( P < 0.05). In conclusion, DCs genetically engineered to overexpress IDO modulate T cell alloresponses in vitro. IDO gene transfer into the donor heart attenuates acute cardiac allograft rejection. Regulation of tryptophan catabolism by means of IDO overexpression may be a useful approach in heart transplantation.

Cross Priming of Transgene Product-Specific CD8+ T Cells in Hepatic AAV Gene Transfer Depends on IL-1 Receptor and XCR1+ Dendritic Cells but Not TLR9

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 2-3
Author(s):  
Sandeep Kumar ◽  
Moanaro Biswas ◽  
Annie R Pineros ◽  
Ype P De Jong ◽  
Roland W Herzog
Keyword(s):  
T Cells ◽  
T Cell ◽  
Gene Transfer ◽  
Cd8 T Cells ◽  
Cell Response ◽  
Adaptor Protein ◽  
T Cell Responses ◽  
Cd8 T Cell ◽  
T Cell Response ◽  
Cell Responses

Introduction: Adeno-associated virus (AAV) mediated gene transfer is currently evaluated in multiple Phase I/II and Phase III studies for the treatment of hemophilia. However, immune responses to both the AAV capsid and encoded transgene remain major impediments to clinical translation. Several studies have implicated innate immune sensors such as Toll-like receptors (TLR) and their downstream adaptor molecule MyD88 in sensing viral structures. TLR9-MyD88 signaling has been linked to cross-priming of CD8+ T cell responses to capsid and also to transgene product-specific CD8+ T cell responses. However, little is known about other signaling pathways that may lead to immune activation. Previously, our lab has shown that while liver gene transfer is capable of inducing immunological tolerance to AAV encoded transgene products, vector dose and design play a critical role. For instance, low hepatic gene expression levels may elicit a CD8+ T cell response to the AAV encoded transgene, resulting in loss of the model antigen ovalbumin (OVA) in C57BL/6 mice or of FIX expression in hemophilia B mice. We investigated innate immune sensing pathways that may play a role in initiating transgene specific CD8+ T cell response in the hepatic microenvironment. Further, we determined the contribution of hepatic antigen presenting cells (APC) by selectively depleting/neutralizing APCs and evaluating their effect on presentation of transgene product-derived antigen following AAV8-OVA liver gene delivery. Methods: Wild-type (WT) C57BL6 and specific innate sensing knockout mice on the C57BL6 background were intravenously (IV) injected with a predetermined immunogenic dose (1x109vg) of hepatotropic AAV8-OVA vector (Mol Ther 25:880, 2017). PBMCs were quantified at 4 weeks for OVA-specific CD8+ T cells using a class I MHC tetramer. Hepatic APC types [Kupffer cells, neutrophils, CD103+ dendritic cell (DC), CD11c+ DC, XCR1+ DC] involved in transgene specific CD8+ T cell activation were selectively depleted/inactivated by pre-treatment with gadolinium chloride (GdCl3), Ly6G, CD103 antibody respectively, or by administering diphtheria toxin (DT) to CD11c-DTR and XCR1-DTR mice. This was followed by intravenous administration of AAV8-OVA and CellTrace violet labeled OT-1 cells. Results: Similar to WT mice, TLR9-/-, TLR2-/-, TRIF-/-, IFNaR-/- and MDA5-/- mice developed a CD8+ T cell response indicating that these sensors do not play a role in transgene specific CD8+ T cells response. Interestingly, adaptor protein MyD88-/- mice did not elicit CD8+ T cell response to OVA, implying a MyD88 dependent but TLR9 independent response. Since MyD88 is an essential adaptor protein not only for TLR but also for interleukin-1 (IL-1) signaling pathways, we next analyzed IL-1R-/- mice. Similar to MyD88-/- mice, IL-1R-/- mice did not show OVA specific CD8+ T cells (p=0.006, 0.007 respectively), indicating that transgene-specific adaptive responses are mediated by IL-1R/MyD88 signaling. Kupffer cells and DCs are principal APCs in liver and infiltrating neutrophils could also act as APCs under inflammatory conditions in liver microenvironment. Using proliferation of OT-I cells as readout we tested if any of these cell types are required for presentation to transgene specific CD8+ T cells. In naïve control, GdCl3 treated and a-Ly6G antibody treated mice, OT-I cell proliferation reached 60%, 65% and 48% on average, respectively. Depletion of CD11c DCs substantially reduced the proliferation of OT-I cells to ~6% (p&lt;0.0001) indicating a critical role for DCs in mediating transgene specific CD8+ T cell responses. Since XCR1+ DCs are the major cross-presenting DCs and hepatic resident CD103+ DCs are shown to have intrinsically enhanced capacity to process and present antigen to naïve CD8+ T cells, we further sought to assess if any of these DCs plays a role in activation of transgene specific CD8+ T cells. Neutralization of CD103+ DCs reduced OT-I proliferation to 39% (p=0.01) whereas depletion of XCR1+ DCs reduced the proliferation to ~20% (p&lt;0.0001) indicating a major role for XCR1+ DCs. Conclusions: In summary, we uncovered a novel-signaling pathway that can activate CD8+ T cell responses during AAV gene transfer independent of TLR9 sensing. The IL-1R/MyD88 pathway drives activation of transgene specific CD8+ T cell, and XCR1+ DCs are critically involved in cross-presenting transgene product-derived antigen to CD8+ T cells. Disclosures Herzog: Takeda Pharmaceuticals: Patents & Royalties.

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.

Peptide-Induced Antigen-Specific CD4+CD25+FoxP3+ T Cells Suppress Cytotoxicity T Cell Responses Directed Against the AAV Capsid.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3769-3769 ◽  
Author(s):  
Daniel J. Hui ◽  
Etiena Basner-Tschakarjan ◽  
Gary C. Pien ◽  
William D. Martin ◽  
Annie S. DeGroot ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Gene Transfer ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
T Cell Responses ◽  
Cd8 T Cell ◽  
Cell Responses ◽  
Equity Ownership

Abstract Abstract 3769 Recent advances in adeno-associated viral (AAV) vector-mediated gene transfer continue to offer hope for the correction of monogenic disorders such as hemophilia B. However, unanticipated T cell responses directed against viral capsid epitopes may limit the efficacy of AAV gene transfer. A phase I clinical study in which an AAV2 vector expressing human factor IX (FIX) was delivered systemically provided the first evidence that AAV vector administration at high doses may trigger the expansion of memory CD8+ T cells directed against AAV capsid epitopes. This response was associated with the loss of FIX transgene expression and a transient increase in liver enzymes. Additional studies in human subjects undergoing AAV gene transfer suggest that the capsid antigen load is an important determinant of capsid-specific T cell activation. Thus, strategies for the modulation of capsid T cell responses could contribute to achieving sustained transgene expression following high dose delivery of AAV. MHC class II peptide ligands identified within the human IgG Fc fragment (Tregitopes, Blood 2008;112:3303) have been shown to expand regulatory T cells (Tregs). Restimulation of human peripheral blood mononuclear cells (PBMC) in vitro with AAV capsid antigen in the presence of Tregitopes resulted in the suppression of capsid-specific CD8+ T cells and in the expansion of CD4+CD25+FoxP3+ Tregs. To better define the nature of Tregitope-induced Tregs, we depleted CD25+ cells from PBMC prior to in vitro restimulation. This completely prevented capsid-specific CTL suppression and the expansion of Tregs, suggesting that Tregitopes act by expanding natural Tregs. Cytokine ELISA on conditioned media from PBMC co-cultured with AAV antigen and Tregitopes showed a 50% decrease in IL-2 levels and a >500-fold increase in IL-10 levels. These results suggest that the effect of Tregitopes may be cytokine mediated. To test this hypothesis, we used a transwell system in which the CD4+ T cell fraction of Tregitope-restimulated PBMC was co-cultured with the capsid-specific CD8+ T cells. Without cell contact, a nearly 50% suppression of anti-capsid CD8+ T cell responses was observed. Further evidence supporting the role of cytokine-mediated suppression came from the observation that Tregitope-treated capsid-specific CD8+ T cells appeared to be anergic, and depletion of CD4+ T cells (Tregs) followed by a 24-hour incubation of CD8+CD4− T cells with IL-2 restored >80% of CTL activity. Finally, antigen specificity of Tregitope-induced Tregs was tested by expanding PBMC in vitro with HLA-B*0702-restricted epitopes from either the AAV capsid or the Epstein-Barr Virus (EBV). After in vitro restimulation, negatively-isolated CD4+ T cells expanded in the presence of EBV antigen and Tregitopes were co-incubated with either CD8+ T cells expanded against the AAV capsid or against EBV. Suppression of CTL activity was observed only when EBV Tregs were co-incubated with EBV CD8+ T cells. Similarly, Tregs isolated from AAV and Tregitope cultures suppressed AAV-specific CD8+ T cells but not EBV-specific CD8+T cells. These results suggest that inhibition of CD8+ T cell responses is antigen-specific. We conclude that Tregitopes induce the expansion of Tregs, which can mediate a potent antigen-specific inhibition of CD8+ T cell responses directed to the AAV capsid. Disclosures: Hui: Children's Hospital of Philadelphia: Patents & Royalties. Martin:EpiVax: Employment, Equity Ownership, Patents & Royalties. DeGroot:EpiVax: Employment, Equity Ownership. High:Children's Hospital of Philadelphia: Patents & Royalties. Mingozzi:Children's Hospital of Philadelphia: Patents & Royalties.

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