Use of Gene-Modified T-Cells as Antigen Presenting Cells (T-APC) for Vaccination Against Myeloma Antigens.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2758-2758
Author(s):  
Larry D. Anderson ◽  
Danielle R. Cook ◽  
Carolina Berger ◽  
David G. Maloney ◽  
Stanley R. Riddell
Keyword(s):  
T Cells ◽  
T Cell ◽  
Transgenic Mouse ◽  
Cd8 T Cells ◽  
Genetically Modified ◽  
Patient Specific ◽  
High Avidity ◽  
Vaccine Strategy ◽  
Myeloma Cells

Abstract Multiple myeloma is incurable with currently available standard treatments, and novel therapies with lower toxicity, including immunological therapies, are needed. Myeloma cells express class I MHC molecules and several potentially immunogenic proteins. Candidate target antigens, other than the patient-specific immunoglobulin idiotype, include cancer-testis antigens, such as MAGE-C1 (CT-7), and tissue differentiation antigens, such as B-Cell Maturation Antigen (BCMA). We have isolated, from normal donors, CD8+ T-cells that are CT-7 and BCMA peptide-specific, suggesting that immunologic tolerance to these proteins is incomplete. A subset of these CD8+ T-cells with the highest avidity are capable of recognizing myeloma cells. However, the frequency of T-cells recognizing these peptides is very low. Vaccination offers a potential approach for augmenting the frequency of tumor-reactive T-cells, provided that a vaccine strategy can be developed that elicits high-avidity T-cells. We have previously shown that adoptive transfer of autologous gene-modified T-cells leads to the rapid development of potent CD8+ T-cell immunity against transgene products expressed in the transferred T-cells. The aims of this study were to determine if vaccination with T-cells modified to express the CT-7 or BCMA genes (T-APC) could induce anti-myeloma immunity in an HLA-A2 transgenic mouse model, and to map additional immunogenic epitopes of these proteins presented by HLA-A2. To address this we genetically modified HLA-A2 transgenic mouse T-cells with retroviral vectors encoding the candidate myeloma antigens CT-7 and BCMA, or with a control vector encoding green fluorescent protein (GFP). The gene-modified T-APC were injected intravenously into HLA-A2 transgenic mice, and ELISPOT and cytotoxicity assays were performed on splenocytes from immunized mice to evaluate T-cell recognition of target cells expressing CT-7 or BCMA (including human myeloma cell lines). T-cell responses specific for CT-7, BCMA, and control antigens were elicited following gene-modified T-APC immunization, and in some cases these T-cells were capable of recognizing human myeloma cells. Furthermore, responses to CT-7 have been mapped to two different dominant CT-7 peptides. CD8+ T-cells from some normal A2+ donors also recognized these two epitopes after in vitro peptide stimulation, suggesting that these epitopes may be applicable to human myeloma therapy. Characterization of the BCMA target peptide(s) is ongoing. The ease with which T-cells can be obtained, genetically-modified, and expanded in vitro, together with data showing potent immunogenicity, suggests that T-APC vaccination should be evaluated as an alternative cell based vaccine strategy for human cancer therapy.

scholarly journals 711 IGM-7354 is an anti-PD-L1 IgM antibody and IL-15 cytokine fusion that enhances NK and CD8+ T cell proliferation and tumor cytotoxicity plus potently reverses T cell exhaustion

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A740-A740
Author(s):  
Thierry Giffon ◽  
Melanie Desbois ◽  
Dean Ng ◽  
Poonam Yakkundi ◽  
Marigold Manlusoc ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cynomolgus Monkey ◽  
Cd8 T Cell ◽  
High Avidity ◽  
T Cell Exhaustion ◽  
Cell Exhaustion ◽  
High Affinity

BackgroundWhile approved PD-1/PD-L1 inhibitory antibodies have demonstrated clinical efficacy in certain cancer patients, relapse following a primary response is often observed. Enhancing anti-tumor immune responses with an immunostimulatory cytokine, IL-15 is an attractive combination strategy to enhance anti-tumor NK and memory CD8+ T cell expansion and survival. We have developed IGM-7354, a high affinity, high avidity anti-PD-L1 pentameric IgM antibody with an IL-15Rα chain and IL-15 fused to the joining (J) chain, designed to deliver IL-15 to PD-L1 expressing tumors for enhancing anti-tumor immune responses.MethodsIGM-7354 was generated by grafting heavy chain variable regions of a high affinity humanized anti-PD-L1 IgG onto the IgM heavy chain framework, co-expressed with the light chain and the J chain which included a single IL-15Rα and IL-15 fusion. Binding ELISAs were performed using recombinant antigens. Human and cynomolgus monkey PBMCs were used for potency testing. Reversal of T cell exhaustion was tested using in vitro MLR. In vitro cytotoxicity assays were performed with luciferase-tagged MDA-MB-231 cells and PBMCs. In vivo pharmacodynamic studies were conducted in mice and cynomolgus monkeys.ResultsIGM-7354 bound human and cynomolgus monkey PD-L1 with the same affinity but did not bind to rat or mouse PD-L1. In addition, the IL-15 component of IGM-7354 bound to human and cynomolgus β chain of the trimeric IL-15 receptor with similar affinities, but with weaker binding affinity to rodent IL-15Rβ. Using in vitro assays with PBMCs, IGM-7354 dose dependently enhanced the proliferation of human and cynomolgus monkey NK and CD8+ T cells. Furthermore, IGM-7354 was able to reverse T cell exhaustion in an in vitro MLR beyond that of an IL-15/IL15Rα complex or anti-PD-L1 IgM or IgG alone, as demonstrated by an increase in activation and effector cytokine secretion. IGM-7354 also enhanced in vitro killing of PD-L1-expressing MDA-MB-231 breast cancer cells by human PBMCs. Pharmacodynamic studies in an MDA-MB-231 xenograft mouse model showed dose-dependent increases in circulating NK and CD8+ T cells and tumor infiltrating lymphocytes, which correlated with tumor regression. In cynomolgus monkeys, intravenous administration of IGM-7354 was well tolerated and dose dependently induced the proliferation of NK and CD8+ T cells.ConclusionsIGM-7354 stimulates NK and CD8+ T cell expansion in vitro and in vivo plus induces tumor regressions in mouse tumor models. This approach may enhance tumor localization of the immunostimulatory cytokine IL-15 through high affinity and high avidity binding to PD-L1 thereby improving anti-tumor responses and minimizing toxicity.Ethics ApprovalAll animal studies were conducted according to approved Institutional Animal Care and Use Committee (IACUC) protocols of the testing facilities.

scholarly journals Immunodominant Cytomegalovirus Epitopes Suppress Subdominant Epitopes in the Generation of High-Avidity CD8 T Cells

Pathogens ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 956
Author(s):  
Kirsten Freitag ◽  
Sara Hamdan ◽  
Matthias J. Reddehase ◽  
Rafaela Holtappels
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Immune Escape ◽  
Cd8 T Cell ◽  
Antigenic Drift ◽  
Murine Cytomegalovirus ◽  
High Avidity ◽  
Infected Cells ◽  
Cd8 T Cell Memory

CD8+ T-cell responses to pathogens are directed against infected cells that present pathogen-encoded peptides on MHC class-I molecules. Although natural responses are polyclonal, the spectrum of peptides that qualify for epitopes is remarkably small even for pathogens with high coding capacity. Among those few that are successful at all, a hierarchy exists in the magnitude of the response that they elicit in terms of numbers of CD8+ T cells generated. This led to a classification into immunodominant and non-immunodominant or subordinate epitopes, IDEs and non-IDEs, respectively. IDEs are favored in the design of vaccines and are chosen for CD8+ T-cell immunotherapy. Using murine cytomegalovirus as a model, we provide evidence to conclude that epitope hierarchy reflects competition on the level of antigen recognition. Notably, high-avidity cells specific for non-IDEs were found to expand only when IDEs were deleted. This may be a host’s back-up strategy to avoid viral immune escape through antigenic drift caused by IDE mutations. Importantly, our results are relevant for the design of vaccines based on cytomegaloviruses as vectors to generate high-avidity CD8+ T-cell memory specific for unrelated pathogens or tumors. We propose the deletion of vector-encoded IDEs to avoid the suppression of epitopes of the vaccine target.

scholarly journals CD28 Costimulation Is Required for In Vivo Induction of Peripheral Tolerance in CD8 T Cells

2002 ◽  
Vol 197 (1) ◽  
pp. 19-26 ◽  
Author(s):  
Melanie S. Vacchio ◽  
Richard J. Hodes
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Tolerance Induction ◽  
Peripheral Tolerance ◽  
Cd8 Cells ◽  
Costimulatory Signal ◽  
Cd28 Costimulation

Whereas ligation of CD28 is known to provide a critical costimulatory signal for activation of CD4 T cells, the requirement for CD28 as a costimulatory signal during activation of CD8 cells is less well defined. Even less is known about the involvement of CD28 signals during peripheral tolerance induction in CD8 T cells. In this study, comparison of T cell responses from CD28-deficient and CD28 wild-type H-Y–specific T cell receptor transgenic mice reveals that CD8 cells can proliferate, secrete cytokines, and generate cytotoxic T lymphocytes efficiently in the absence of CD28 costimulation in vitro. Surprisingly, using pregnancy as a model to study the H-Y–specific response of maternal T cells in the presence or absence of CD28 costimulation in vivo, it was found that peripheral tolerance does not occur in CD28KO pregnants in contrast to the partial clonal deletion and hyporesponsiveness of remaining T cells observed in CD28WT pregnants. These data demonstrate for the first time that CD28 is critical for tolerance induction of CD8 T cells, contrasting markedly with CD28 independence of in vitro activation, and suggest that the role of CD28/B7 interactions in peripheral tolerance of CD8 T cells may differ significantly from that of CD4 T cells.

scholarly journals DNAM-1 promotes activation of cytotoxic lymphocytes by nonprofessional antigen-presenting cells and tumors

2008 ◽  
Vol 205 (13) ◽  
pp. 2965-2973 ◽  
Author(s):  
Susan Gilfillan ◽  
Christopher J. Chan ◽  
Marina Cella ◽  
Nicole M. Haynes ◽  
Aaron S. Rapaport ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Nk Cells ◽  
Adhesion Molecules ◽  
Cd8 T Cells ◽  
Nk Cell ◽  
Cd8 T Cell ◽  
Antigen Presenting

Natural killer (NK) cells and CD8 T cells require adhesion molecules for migration, activation, expansion, differentiation, and effector functions. DNAX accessory molecule 1 (DNAM-1), an adhesion molecule belonging to the immunoglobulin superfamily, promotes many of these functions in vitro. However, because NK cells and CD8 T cells express multiple adhesion molecules, it is unclear whether DNAM-1 has a unique function or is effectively redundant in vivo. To address this question, we generated mice lacking DNAM-1 and evaluated DNAM-1–deficient CD8 T cell and NK cell function in vitro and in vivo. Our results demonstrate that CD8 T cells require DNAM-1 for co-stimulation when recognizing antigen presented by nonprofessional antigen-presenting cells; in contrast, DNAM-1 is dispensable when dendritic cells present the antigen. Similarly, NK cells require DNAM-1 for the elimination of tumor cells that are comparatively resistant to NK cell–mediated cytotoxicity caused by the paucity of other NK cell–activating ligands. We conclude that DNAM-1 serves to extend the range of target cells that can activate CD8 T cell and NK cells and, hence, may be essential for immunosurveillance against tumors and/or viruses that evade recognition by other activating or accessory molecules.

A Randomized Phase II Trial of Idiotype Vaccination and Adoptive Autologous T-Cell Transfer in Multiple Myeloma patients

Blood ◽  
2021 ◽  
Author(s):  
Muzaffar H Qazilbash ◽  
Neeraj Y Saini ◽  
Cha Soung-chul ◽  
Zhe Wang ◽  
Edward Stadtmauer ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Phase Ii ◽  
T Cell Activation ◽  
Cd8 T Cells ◽  
Cell Activation ◽  
Phase Ii Trial ◽  
Ex Vivo ◽  
Vaccine Strategy ◽  
Randomized Phase Ii

We hypothesized that combining adoptively transferred autologous T cells with a cancer vaccine strategy would enhance therapeutic efficacy by adding anti-myeloma idiotype-keyhole limpet hemocyanin (Id-KLH) vaccine to vaccine-specific co-stimulated T cells. In this randomized, phase II trial, eligible patients received either the control (KLH only) or Id-KLH vaccine, an auto-transplant, vaccine-specific co-stimulated T-cells expanded ex-vivo, and two booster doses of the assigned vaccine. In 36 patients (20 in KLH, 16 in Id-KLH) enrolled, no dose-limiting toxicity was seen in either arm. At last evaluation, 6 (30%) and 8 (50%) had achieved complete remission in KLH-only and Id-KLH, respectively (p=0.22) and no difference in 3-year progression-free survival was observed (59% and 56%, respectively; p=0.32). In a 594 Nanostring nCounter gene panel analyzed for immune reconstitution (IR), compared with KLH-only patients, there was a greater change in IR genes in T-cells in Id-KLH patients relative to baseline. Specifically, upregulation of genes associated with activation, induction of effector function, and generation of memory CD8+ T cells after Id-KLH, but not after KLH control vaccination, was observed. Similarly, responding patients across both arms were associated with upregulation of genes associated with T-cell activation. At baseline, all patients had greater expression of CD8+ T-cell exhaustion markers. These changes were associated with functional Id-specific immune responses in a subset of Id-KLH patients analyzed. In conclusion, in this combination immunotherapy approach, we observed a significantly more robust IR in CD4+ and CD8+ T cells in the Id-KLH arm, supporting further investigation of vaccine and adoptive immunotherapy strategies.

scholarly journals 708 Novel ways to exploit IL-21 to augment adoptive T cell transfer therapy against tumors

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A737-A737
Author(s):  
Anna Cole ◽  
Guillermo Rangel RIvera ◽  
Aubrey Smith ◽  
Megan Wyatt ◽  
Brandon Ware ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Immune Cells ◽  
Striking Difference ◽  
T Cell Therapy ◽  
Cell Immunity ◽  
Emory University

BackgroundIL-21 enhances the anti-tumor capacity of adoptively transferred CD8+ T cells, while IL-2 and IL-15 impair T cell immunity by driving their expansion to a more differentiated status. Yet, these cytokines can act on many different immune cells. Given the potency of IL-21, we tested if this cytokine directly augments T cells or rather if it enhances other immune cells in the culture that indirectly improves T cell therapy.MethodsTo test this question, splenocytes from pmel-1 transgenic mice were used, as all CD8+ T cells express a transgenic TCR specific for tumor-antigen gp10025–33 overexpressed on melanoma. We then peptide activated naïve CD8+ T cells enriched or not from the spleen of pmel-1 mice and expanded them in the presence of IL-21 or IL-2 (10 ng/mL) for four days. Expanded pmel-1 from these various cultures were then restimulated with irradiated splenocytes pulsed with gp10025–33 and grown an additional seven days with IL-2 (10 ng/mL), irrespective of their initial cytokine condition. The in vitro memory phenotype, exhaustion profile, and cytokine secretion of these cultures were then assayed. Furthermore, mice bearing B16KVP melanoma tumors were infused with pmel-1 T cells expanded via these various approaches and compared for their relative capacity to engraft, persist, and regress tumor in vivo.ResultsInterestingly, we discovered that IL-21-treated T cells generated from bulk splenocytes are phenotypically and functionally distinct from IL-21-treated isolated T cells. Upon restimulation, IL-21-treated T cells from bulk splenocytes exhibited an exhausted phenotype that was like anergic IL-2-treated T cells. Moreover, few cells expressed CD62L but expressed heightened markers of suppression, including TIM3, PD-1, and EOMES. Moreover, they produced more effector molecules, including granzyme B and IFN-gamma. In vivo IL-21-treated T cells expanded from bulk splenocytes engrafted and persisted poorly, in turn mediating suboptimal regression of melanoma. Conversely, IL-21 dramatically bolstered the engraftment and antitumor activity of T cells only if they were first isolated from the spleen prior to their expansion and infusion into the animal.ConclusionsCollectively, our data shows that IL-21 may improve ACT therapy best when used directly on antitumor CD8+ T cells. Further studies will illuminate the mechanism behind this striking difference and determine whether other cell subsets reactive to IL-21 cause T cell dysfunction and/or reduced bioavailability. These findings are important for defining the best culture conditions in which to use IL-21 for ACT.AcknowledgementsWe would like to acknowledge Emory University, The Winship Cancer Institute, and the Pediatrics/Winship Flow Cytometry Core.Ethics ApprovalAll animal procedures were approved by the Institutional Animal Care and Use Committee of Emory University, protocol number 201900225.

scholarly journals 663 Media based on the metabolic composition of tumor interstitial fluid reveals persistent T cell dysfunction induced through arginine deprivation and exposure to the oncometabolite phosphoethanolamine

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A691-A691
Author(s):  
Yupeng Wang ◽  
Chufan Cai ◽  
Dayana Rivadeneira ◽  
Alexander Muir ◽  
Greg Delgoffe
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Interstitial Fluid ◽  
Cytokine Production ◽  
Cell Dysfunction ◽  
Kennedy Pathway ◽  
T Cell Dysfunction ◽  
Tumor Interstitial Fluid

BackgroundWhile CD8 T cells are crucial for anti-tumor immunity, tumor infiltrating CD8 T cells encounter stressors which deviate their differentiation to a dysfunctional, exhausted phenotype. T cell functions are closely regulated by T cell metabolism, and the dysfunctional vasculature in tumor tissues and the deregulated metabolism of tumor cells lead to depletion of nutrients and accumulation of metabolic wastes in the tumor microenvironment (TME). Thus, the unbalanced levels of the nutrients and the metabolic wastes might skew the metabolism of T cells thus contributing to T cell dysfunction.MethodsOvalbumin-specific OT-I cells were activated with SIINFEKL/IL2 and cultured with IL2. The tumor interstitial fluid media (TIFM) was formulated based on the concentrations of the metabolites measured in the tumor interstitial fluid of pancreatic ductal adenocarcinoma.1 Purified arginine and phosphoethanolamine (PEtn) were used to change their levels in TIFM/RPMI1640 culture. Expression level of cytokines and PD-1 was measured by flow cytometry.ResultsWe sought to determine how T cells would differentiate, in vitro, if they were exposed only to the metabolites present in the TME. Using media formulated to model the metabolic composition of tumor interstitial fluid (TIFM),1 we show that CD8 T cells develop features of exhausted T cells in the TIFM culture: reduced proliferation, increased expression of PD-1 and decreased cytokine production. Using 'dropout' and 'add-back' approaches, we found arginine levels as a major contributor to the proliferation defect observed in TIFM-cultured T cells. Arginine was sufficient to restore proliferative capacity to T cells cultured in TIFM, but had no effect on the inhibited cytokine production. We then asked which metabolites were enriched in the TIFM, finding that PEtn, an intermediate in the ethanolamine branch of the Kennedy pathway and an oncometabolite enriched in the interstitial of many solid tumors, up-regulates PD-1 expression and compromises the cytokine production of the cells in culture. Depletion of Pcyt2, the metabolizing enzyme of PEtn and the rate limiting enzyme in the Kennedy pathway, makes CD8 T cells resistant to the effects of PEtn.ConclusionsOur data shows that the metabolic environment in the TME can be recapitulated in vitro and is sufficient to drive T cell dysfunction. Arginine depletion acts as a major inhibitor of T cell proliferation in the TME, but the oncometabolite PEtn drives a hypofunctional effector fate of T cells. Targeting PEtn metabolism via Pcyt2 depletion or inhibition is a potential target to reinvigorate T cells and enhance anti-tumor immunity.ReferenceSullivan MR, Danai LV, Lewis CA, Chan SH, Gui DY, Kunchok T, Dennstedt EA, Vander Heiden MG, Muir A. Quantification of microenvironmental metabolites in murine cancers reveals determinants of tumor nutrient availability. Elife 2019;;8:e44235. doi: 10.7554/eLife.44235. PMID: 30990168; PMCID: PMC6510537.

scholarly journals Infection of CD8+CD45RO+ Memory T-Cells by HIV-1 and Their Proliferative Response

2008 ◽  
Vol 2 (1) ◽  
pp. 43-57 ◽  
Author(s):  
Naveed Gulzar ◽  
Sowyma Balasubramanian ◽  
Greg Harris ◽  
Jaime Sanchez-Dardon ◽  
Karen F.T. Copeland
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Proliferative Response ◽  
Cell Infection ◽  
Cellular Target ◽  
Potential Reservoir ◽  
Infected Cells ◽  
Hiv 1

CD8+ T-cells are involved in controlling HIV-1 infection by eliminating infected cells and secreting soluble factors that inhibit viral replication. To investigate the mechanism and significance of infection of CD8+ T-cells by HIV-1in vitro, we examined the susceptibility of these cells and their subsets to infection. CD8+ T-cells supported greater levels of replication with T-cell tropic strains of HIV-1, though viral production was lower than that observed in CD4+ T-cells. CD8+ T-cell infection was found to be productive through ELISA, RT-PCR and flow cytometric analyses. In addition, the CD8+CD45RO+ memory T-cell population supported higher levels of HIV-1 replication than CD8+CD45RA+ naïve T-cells. However, infection of CD8+CD45RO+ T-cells did not affect their proliferative response to the majority of mitogens tested. We conclude, with numerous lines of evidence detecting and measuring infection of CD8+ T-cells and their subsets, that this cellular target and potential reservoir may be central to HIV-1 pathogenesis.

scholarly journals Transcription factor Etv6 regulates functional differentiation of cross-presenting classical dendritic cells

2018 ◽  
Vol 215 (9) ◽  
pp. 2265-2278 ◽  
Author(s):  
Colleen M. Lau ◽  
Ioanna Tiniakou ◽  
Oriana A. Perez ◽  
Margaret E. Kirkling ◽  
George S. Yap ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
Transcription Factor ◽  
T Cell ◽  
Cd8 T Cells ◽  
Functional Differentiation ◽  
Specific Gene ◽  
Hematopoietic Stem

An IRF8-dependent subset of conventional dendritic cells (cDCs), termed cDC1, effectively cross-primes CD8+ T cells and facilitates tumor-specific T cell responses. Etv6 is an ETS family transcription factor that controls hematopoietic stem and progenitor cell (HSPC) function and thrombopoiesis. We report that like HSPCs, cDCs express Etv6, but not its antagonist, ETS1, whereas interferon-producing plasmacytoid dendritic cells (pDCs) express both factors. Deletion of Etv6 in the bone marrow impaired the generation of cDC1-like cells in vitro and abolished the expression of signature marker CD8α on cDC1 in vivo. Moreover, Etv6-deficient primary cDC1 showed a partial reduction of cDC-specific and cDC1-specific gene expression and chromatin signatures and an aberrant up-regulation of pDC-specific signatures. Accordingly, DC-specific Etv6 deletion impaired CD8+ T cell cross-priming and the generation of tumor antigen–specific CD8+ T cells. Thus, Etv6 optimizes the resolution of cDC1 and pDC expression programs and the functional fitness of cDC1, thereby facilitating T cell cross-priming and tumor-specific responses.

scholarly journals A new in vitro model of polymyositis reveals CD8+ T cell invasion into muscle cells and its cytotoxic role

Rheumatology ◽  
2019 ◽  
Vol 59 (1) ◽  
pp. 224-232
Author(s):  
Mari Kamiya ◽  
Fumitaka Mizoguchi ◽  
Akito Takamura ◽  
Naoki Kimura ◽  
Kimito Kawahata ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Muscle Injury ◽  
In Vitro Model ◽  
Muscle Cells ◽  
Cd8 T Cell ◽  
Class I ◽  
Vitro Model

Abstract Objectives The hallmark histopathology of PM is the presence of CD8+ T cells in the non-necrotic muscle cells. The aim of this study was to clarify the pathological significance of CD8+ T cells in muscle cells. Methods C2C12 cells were transduced retrovirally with the genes encoding MHC class I (H2Kb) and SIINFEKL peptide derived from ovalbumin (OVA), and then differentiated to myotubes (H2KbOVA-myotubes). H2KbOVA-myotubes were co-cultured with OT-I CD8+ T cells derived from OVA-specific class I restricted T cell receptor transgenic mice as an in vitro model of PM to examine whether the CD8+ T cells invade into the myotubes and if the myotubes with the invasion are more prone to die than those without. Muscle biopsy samples from patients with PM were examined for the presence of CD8+ T cells in muscle cells. The clinical profiles were compared between the patients with and without CD8+ T cells in muscle cells. Results Analysis of the in vitro model of PM with confocal microscopy demonstrated the invasion of OT-I CD8+ T cells into H2KbOVA-myotubes. Transmission electron microscopic analysis revealed an electron-lucent area between the invaded CD8+ T cell and the cytoplasm of H2KbOVA-myotubes. The myotubes invaded with OT-I CD8+ T cells died earlier than the uninvaded myotubes. The level of serum creatinine kinase was higher in patients with CD8+ T cells in muscle cells than those without these cells. Conclusion CD8+ T cells invade into muscle cells and contribute to muscle injury in PM. Our in vitro model of PM is useful to examine the mechanisms underlying muscle injury induced by CD8+ T cells.

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