Cell Responses In Patients Research Articles

Optimal timing of recombinant herpes zoster virus vaccination for a JAK inhibitor treatment in rheumatoid arthritis: a multicentre, open-label, randomised comparative study (STOP-HZ study): study protocol

IntroductionJanus kinase (JAK) inhibitors are an important therapeutic option in the treatment of rheumatoid arthritis, but increase the risk of developing herpes zoster. Although a dry recombinant zoster vaccine (RZV) that can be used under immunosuppressive conditions has recently been developed, its optimal use and appropriate timing in patients scheduled to start JAK inhibitors is still unclear. The present study is designed to clarify the appropriate timing of JAK inhibitor initiation to measure varicella zoster virus (VZV)-specific IgG titers and VZV-specific T cell response in patients with rheumatoid arthritis who start tofacitinib at the first RZV vaccination or at the second one.Methods and analysisSTOP HZ (Effectiveness andSafeTyOfProphylactic RecombinantHerpesZoster Virus Vaccination for Rheumatoid Arthritis Patients with Tofacitinib Treatment) study is a multicentre, open-label, randomised, comparative study in patients with rheumatoid arthritis who are scheduled to start tofacitinib. This study enrols 60 study subjects in 12 sites. Enrolled subjects receive RZV two times on day 1 and week 8 and initiate tofacitinib 5 mg two times a day at the time of their first RZV (day 1, group A) or second RZV (week 8, group B) based on randomisation. The random assignment is performed centrally in a 1:1 ratio. Patients in Group B continue the same treatment until the start of tofacitinib treatment. Primary endpoint is VZV-specific IgG antibody titers at week 12 compared with those at baseline in each group. Secondary endpoints include comparison of VZV-specific IgG antibody between the groups, changes in disease activity of rheumatoid arthritis, VZV-specific T cell response and adverse events.Ethics and disseminationThe study has been approved by the Certified Review Board of Keio (No. 2022008), and conforms to the Declaration of Helsinki and good clinical practice guidelines. Written informed consent is obtained from participants prior to enrolment. The results of this study are planned to be submitted for publishment in relevant peer-review journals.Trial registration numberjRCTs031230329.

Characterize GAD65-specific T cell Responses in Patients with Stiff-Person Syndrome and Type-I Diabetes.

Characterize GAD65-specific T cell Responses in Patients with Stiff-Person Syndrome and Type-I Diabetes.

Methotrexate treatment hampers induction of vaccine-specific CD4 T cell responses in patients with IMID

ObjectivesMethotrexate (MTX) is one of the most commonly used medications to treat rheumatoid arthritis (RA). However, the effect of MTX treatment on cellular immune responses remains incompletely understood. This raises...

Abstract C020: Identification of KRAS-specific TCRs from human peripheral blood

Abstract Mutant KRAS is the predominant driver oncogene in pancreatic cancer accounting for approximately 90% of KRAS mutations. However, these mutant KRAS peptides exhibit weak binding affinity to most human leukocyte antigens (HLAs), impeding their recognition by the immune system and the subsequent generation of a corresponding T cell response. Identification of KRAS-mutant specific T cells would be useful to track tumor-specific T cell responses in patients and could also be used for adoptive transfer of TCR-transduced cells. Current efforts to identify KRAS-specific T cells have been limited by their restriction to specific HLA haplotypes or the provision of only a few T cell receptor (TCR) sequences. In this study, we developed a facile means of detecting KRAS-specific T cells from peripheral blood at frequencies as low as 1 in 1,000,000. We employ an α41BB agonist-selective lentivirus system combined with a KRAS peptide pool to facilitate the high-throughput discovery of KRAS-specific TCRs in peripheral blood mononuclear cells (PBMCs) derived from both patients with pancreatic cancer and healthy donors. Through in vitro stimulation and single-cell TCR analysis, we identified a series of T cell expansions sharing the same clonotype, indicating a response to KRAS-mutant peptide stimulation. We have quantified CD4 and CD8 KRAS-specific TCRs from patients with a diverse range of HLA haplotypes. Citation Format: Jiao Shen, Blake E. Smith, Winiffer Conce Alberto, Ellen J. Kim, Michael L. Dougan, Harshabad Singh, Michael E. Birnbaum, Stephanie K. Dougan. Identification of KRAS-specific TCRs from human peripheral blood [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr C020.

Distinct mucosal endotypes as initiators and drivers of rheumatoid arthritis.

Rheumatoid arthritis (RA) is a potentially devastating autoimmune disease. The great majority of patients with RA are seropositive for anti-citrullinated protein antibodies (ACPAs), rheumatoid factors, or other autoantibodies. The onset of clinically apparent inflammatory arthritis meeting classification criteria (clinical RA) is preceded by ACPA seropositivity for an average of 3-5 years, a period that is designated as 'at-risk' of RA for ACPA-positive individuals who do not display signs of arthritis, or 'pre-RA' for individuals who are known to have progressed to developing clinical RA. Prior studies of individuals at-risk of RA have associated pulmonary mucosal inflammation with local production of ACPAs and rheumatoid factors, leading to development of the 'mucosal origins hypothesis'. Recent work now suggests the presence of multiple distinct mucosal site-specific mechanisms that drive RA evolution. Indicatively, subsets of individuals at-risk of RA and patients with RA harbour a faecal bacterial strain that has exhibited arthritogenic activity in animal models and that favours T helper 17 (TH17) cell responses in patients. Periodontal inflammation and oral microbiota have also been suggested to promote the development of arthritis through breaches in the mucosal barrier. Herein, we argue that mucosal sites and their associated microbial strains can contribute to RA evolution via distinct pathogenic mechanisms, which can be considered causal mucosal endotypes. Future therapies instituted for prevention in the at-risk period, or, perhaps, during clinical RA as therapeutics for active arthritis, will possibly have to address these individual mechanisms as part of precision medicine approaches.

MRNA-1273 vaccination induces polyfunctional memory CD4 and CD8 T cell responses in patients with solid cancers undergoing immunotherapy or/and chemotherapy.

Research has confirmed the safety and comparable seroconversion rates following SARS-CoV-2 vaccination in patients with solid cancers. However, the impact of cancer treatment on vaccine-induced T cell responses remains poorly understood. In this study, we expand on previous findings within the VOICE trial by evaluating the functional and phenotypic composition of mRNA-1273-induced T cell responses in patients with solid tumors undergoing immunotherapy, chemotherapy, or both, compared to individuals without cancer. We conducted an ELISpot analysis on 386 participants to assess spike-specific T cell responses 28 days after full vaccination. Further in-depth characterization of using flow cytometry was performed on a subset of 63 participants to analyze the functional phenotype and differentiation state of spike-specific T cell responses. ELISpot analysis showed robust induction of spike-specific T cell responses across all treatment groups, with response rates ranging from 75% to 80%. Flow cytometry analysis revealed a distinctive cytokine production pattern across cohorts, with CD4 T cells producing IFNγ, TNF, and IL-2, and CD8 T cells producing IFNγ, TNF, and CCL4. Variations were observed in the proportion of monofunctional CD4 T cells producing TNF, particularly higher in individuals without cancer and patients treated with chemotherapy alone, while those treated with immunotherapy or chemoimmunotherapy predominantly produced IFNγ. Despite these differences, polyfunctional spike-specific memory CD4 and CD8 T cell responses were comparable across cohorts. Notably, immunotherapy-treated patients exhibited an expansion of spike-specific CD4 T cells with a terminally differentiated effector memory phenotype. These findings demonstrate that systemic treatment in patients with solid tumors does not compromise the quality of polyfunctional mRNA-1273-induced T cell responses. This underscores the importance of COVID-19 vaccination in patients with solid cancers undergoing systemic treatment.

Development of an arenavirus-based immunotherapy for treatment of KRAS mutant cancer.

e14672 Background: Oncogenic mutations in KRAS are prevalent in more than 80% of pancreatic ductal adenocarcinomas (PDAC), approximately 30% of colorectal cancer (CRC) and 15-20% of lung adenocarcinoma (LUAD). It has been demonstrated previously that CD8+ T cell responses specific for KRAS mutations can lead to tumor control and regression of metastasis. Replicating arenavirus vectors are currently evaluated in multiple clinical trials in infectious diseases and oncology and have demonstrated potent induction of target antigen specific CD8+ T cell responses of up to almost 50% of the total CD8+ T cell compartment in peripheral blood. To augment KRAS mutation specific CD8+ T cell responses in patients, we are using replicating arenavirus vectors targeting five prevalent KRAS mutations which are found in approximately 95% (PDAC), 74% (CRC), and 83% (LUAD) of the KRAS mutated tumors, representing a promising off-the-shelf immunotherapy for these indications. Here, we systematically investigate immune responses induced by mutant KRAS targeted arenavirus-vector-based cancer vaccines in HLA transgenic mice and characterize the efficiency of target cell killing of activated KRAS mutation specific CD8+ T cells. Methods: A transgene cassette consisting of peptide stretches including KRAS mutations G12D, G12V, G12C, G12R and G13D was generated by in silico aided antigen design. Vectors encoding peptide stretches of single KRAS mutations as well as the corresponding wildtype KRAS sequence served as controls in preclinical experiments. KRAS mutation specific CD8+ T cell expansion was evaluated in HLA transgenic mice and functionality of induced CD8+ T cells was evaluated by assessing CD8+ T cell mediated killing of mutant KRAS peptide loaded target cells in vivo. Results: All treatment regimens were well tolerated, and no mortalities or major adverse events were observed. Following vector administration, HLA I restricted antigen-specific CD8+ T cell responses were detected against 4 encoded mutant KRAS epitopes in HLA-A*11:01 (KRAS G12D and G12V), or HLA-B*07:02 (KRAS G12C and G12R) transgenic mice. These cells did not cross-react with wildtype KRAS epitopes presented in the same HLA I molecules. Specific cytotoxicity against KRAS G12D/V or KRAS G12C/R pulsed target cells was observed in vivo in HLA transgenic animals, indicating functionality of the induced KRAS mutation specific T cells. No specific cytotoxicity towards target cells pulsed with KRAS WT peptides was observed in any of the groups. Conclusions: This study indicates efficient induction of KRAS mutation specific T cell responses in HLA transgenic mice using an arenavirus vector based vaccine. Expanded CD8+ T cells were capable of killing cells loaded with KRAS mutation specific epitopes in vivo indicating functionality of the induced T cell responses. Based on these results, initiation of a Phase I study for the treatment of KRAS mutated cancers is planned.

Erratum for the Research Article "A functional identification platform reveals frequent, spontaneous neoantigen-specific T cell responses in patients with cancer" by A. M. Miller et al.

Erratum for the Research Article "A functional identification platform reveals frequent, spontaneous neoantigen-specific T cell responses in patients with cancer" by A. M. Miller et al.

Stealth transgenes enable CAR-T cells to evade host immune responses

BackgroundAdoptive cell therapy, such as chimeric antigen receptor (CAR)-T cell therapy, has improved patient outcomes for hematological malignancies. Currently, four of the six FDA-approved CAR-T cell products use the FMC63-based...

Dose escalation study of a personalized peptide-based neoantigen vaccine (EVX-01) in patients with metastatic melanoma

BackgroundNeoantigens can serve as targets for T cell-mediated antitumor immunity via personalized neopeptide vaccines. Interim data from our clinical study NCT03715985 showed that the personalized peptide-based neoantigen vaccine EVX-01, formulated...

Histochemical Evidence for Reduced Immune Response in Nasal Mucosa of Patients with COVID-19.

The primary entry point of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the nasal mucosa, where viral-induced inflammation occurs. When the immune response fails against SARS-CoV-2, understanding the altered response becomes crucial. This study aimed to compare SARS-CoV-2 immunological responses in the olfactory and respiratory mucosa by focusing on epithelia and nerves. Between 2020 and 2022, we obtained post mortem tissues from the olfactory cleft from 10 patients with histologically intact olfactory epithelia (OE) who died with or from COVID-19, along with four age-matched controls. These tissues were subjected to immunohistochemical reactions using antibodies against T cell antigens CD3, CD8, CD68, and SARS spike protein for viral evidence. Deceased patients with COVID-19 exhibited peripheral lymphopenia accompanied by a local decrease in CD3+ cells in the OE. However, SARS-CoV-2 spike protein was sparsely detectable in the OE. With regard to the involvement of nerve fibers, the present analysis suggested that SARS-CoV-2 did not significantly alter the immune response in olfactory or trigeminal fibers. On the other hand, SARS spike protein was detectable in both nerves. In summary, the post mortem investigation demonstrated a decreased T cell response in patients with COVID-19 and signs of SARS-CoV-2 presence in olfactory and trigeminal fibers.

A functional identification platform reveals frequent, spontaneous neoantigen-specific T cell responses in patients with cancer.

The clinical impact of tumor-specific neoantigens as both immunotherapeutic targets and biomarkers has been impeded by the lack of efficient methods for their identification and validation from routine samples. We have developed a platform that combines bioinformatic analysis of tumor exomes and transcriptional data with functional testing of autologous peripheral blood mononuclear cells (PBMCs) to simultaneously identify and validate neoantigens recognized by naturally primed CD4+ and CD8+ T cell responses across a range of tumor types and mutational burdens. The method features a human leukocyte antigen (HLA)-agnostic bioinformatic algorithm that prioritizes mutations recognized by patient PBMCs at a greater than 40% positive predictive value followed by a short-term in vitro functional assay, which allows interrogation of 50 to 75 expressed mutations from a single 50-ml blood sample. Neoantigens validated by this method include both driver and passenger mutations, and this method identified neoantigens that would not have been otherwise detected using an in silico prediction approach. These findings reveal an efficient approach to systematically validate clinically actionable neoantigens and the T cell receptors that recognize them and demonstrate that patients across a variety of human cancers have a diverse repertoire of neoantigen-specific T cells.

Immune Cell Response of the Spleen in COVID-19

Objective. To study the morphometric characteristics and splenic immune cell response in patients with COVID-19.Material and methods. A prospective observational study included 70 patients. Of these, 45 patients admitted to the infectious diseases hospital with Coronavirus infection caused by the SARS-CoV-2 virus diagnosis were included in the COVID-19 group, and 25 patients were included in the acute respiratory viral infection (ARVI) comparison group. Spleen linear dimensions, including length, width, and thickness were assessed using ultrasound imaging, and calculations of the spleen weight and spleen weight coefficient (SWC) were obtained. Additionally leukocyte count and formula, erythrocyte sedimentation rate (ESR) were estimated, and the leukocyte index (LI) and neutrophil-to-lymphocyte ratio (NLR) were calculated.Results. Microsplenia was common in the acute period of COVID-19 with mean SWC value 1.6±0.2. In 17 (37.8%) patients the SWC varied from 1.0 to 1.5, and in 9 (20%) microsplenia was critical with SWC <1.0. Leukocyte count was lower, and ESR — higher in patients with COVID-19, compared to ARVI group (5.4±2.1×10⁹/l and 10.8±4.8×109/l, respectively P<0.00001, and ESR — 36.1±13.8 mm/h and 23.0±5.1 mm/h, respectively P=0.03). The course of COVID-19 was characterized by a slight decrease in LI — from 0.29±0.02 to 0.22±0.01 (P=0.19), and significant increase in NLR from 3.7±0.1 to 4.3±0.12 (P=0.002). Opposite trends were documented in patients with ARVI. On Day 5 since initiation of treatment LI was significantly lower in the COVID-19 vs ARVI group (0.22 [0.16; 0.39] vs. 0.48 [0.29; 0.93], P=0.003), and NLR was significantly higher (4.3 [2.5; 6.1] vs. 2.1 [0.9; 2.9], P=0.002).Conclusion. The course of coronavirus infection caused by the SARS-CoV-2 virus is characterized by significant immunological shifts. Microsplenia verified by ultrasonography stays as one of the pathognomonic signs. This phenomenon is explained by rapid «depletion» of the spleen as a secondary immune organ, and is associated with a high risk of developing acute immune deficiency.

ECCO Grant Dissecting the intestinal mucosa-associated HUMoral immune response in ileal Crohn’s Disease: the HUM-CD study

Abstract Background and Aims The pathophysiology of Crohn's disease (CD) is complex and not fully understood. While CD can affect the entire gastrointestinal tract, it primarily affects the terminal ileum. The underlying mechanistic reasons for this preferential location are unknown. Patients with isolated ileal CD have higher levels of circulating IgG antibodies targeting flagellins. These antibodies are associated with the ileal location of CD as well as with a complicated phenotype (intestinal stenosis and fistulae). Cellular determinants of this humoral response in ileal CD are yet to be unraveled. Techniques to study B cells at the single-cell levels have tremendously evolved these recent years and our laboratory has developed expertise in most of these techniques. The objectives of our study are to comprehensively characterize the humoral B cell response in patients with ileal CD. Methods To fulfill our aim, we will use different approaches: 1. Extensive characterization of the B cell cellular and molecular landscape of the intestinal ileum in CD as compared to homeostasis states at the single-cell level 2. Dissection of the intestinal B cell antigen-specific response in CD and assess differences between inflamed and non-inflamed, and ileal and colonic mucosa and the cross-reactivity of the flagellin-specific response 3. Identification of the antigenic reactivity and clonal dynamics of memory B cells extracted from the terminal ileum of patients with CD. We will work from human samples, collected from ileal, colonic biopsies, and blood of patients with CD and healthy controls using Anticipated Impact Using advanced B-cell-focused single-cell techniques, we will unravel the humoral response at the ileal level and its association with clinical outcomes. By doing so, we aim to identify the main antigenic drivers and putative therapeutic targets of ileal CD.

Imbalance of Follicular Helper and Follicular Regulatory T Cells in Chronic Rhinosinusitis with Nasal Polyps.

Data regarding the imbalance in follicular helper T (Tfh) and follicular regulatory T (Tfr) cell responses in patients having chronic rhinosinusitis with nasal polyps (CRSwNP) is so far limited. Thus, we aimed to assess the changes in circulating Tfh and Tfr in CRSwNP patients. This case-control study included 21 patients having CRSwNP and 20 age and sex-matched healthy blood donors as a control group. Lund-Mackay staging system was used for radiologic scoring of chronic rhinosinusitis. Two milliliters of peripheral blood samples were collected from all participants into EDTA-containing vacutainer tubes to assess the levels of Tfh and Tfr cells using flow cytometry. Patients having CRSwNP did not show significant differences in the percentages of CD4+ T cells and total CD4+CXCR5+ T cells from healthy controls. Meanwhile, levels of both activated circulating Tfh and Tfr showed a marked rise in patients than controls. In addition, a positive correlation was observed between the levels of both activated Tfh and Tfr cells. An imbalance in circulating Tfh/Tfr levels was detected in patients having CRSwNP. A significant rise in the levels of Tfh and Tfr was detected in patients proposing a possible role of this imbalance in disease pathogenesis.

Autoreactive T cells targeting type II pneumocyte antigens in COVID-19 convalescent patients

BackgroundThe role of autoreactive T cells on the course of Coronavirus disease-19 (COVID-19) remains elusive. Type II pneumocytes represent the main target cells of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Autoimmune responses against antigens highly expressed in type II pneumocytes may influence the severity of COVID-19 disease. ObjectiveThe aim of this study was to investigate autoreactive T cell responses against self-antigens highly expressed in type II pneumocytes in the blood of COVID-19 patients with severe and non-severe disease. MethodsWe collected blood samples of COVID-19 patients with varying degrees of disease severity and of pre-pandemic controls. T cell stimulation assays with peptide pools of type II pneumocyte antigens were performed in two independent cohorts to analyze the autoimmune T cell responses in patients with non-severe and severe COVID-19 disease. Target cell lysis assays were performed with lung cancer cell lines to determine the extent of cell killing by type II PAA-specific T cells.Results: We identified autoreactive T cell responses against four recently described self-antigens highly expressed in type II pneumocytes, known as surfactant protein A, surfactant protein B, surfactant protein C and napsin A, in the blood of COVID-19 patients. These antigens were termed type II pneumocyte-associated antigens (type II PAAs). We found that patients with non-severe COVID-19 disease showed a significantly higher frequency of type II PAA-specific autoreactive T cells in the blood when compared to severely ill patients. The presence of high frequencies of type II PAA-specific T cells in the blood of non-severe COVID-19 patients was independent of their age. We also found that napsin A-specific T cells from convalescent COVID-19 patients could kill lung cancer cells, demonstrating the functional and cytotoxic role of these T cells. ConclusionsOur data suggest that autoreactive type II PAA-specific T cells have a protective role in SARS-CoV-2 infections and the presence of high frequencies of these autoreactive T cells indicates effective viral control in COVID-19 patients. Type II-PAA-specific T cells may therefore promote the killing of infected type II pneumocytes and viral clearance.

Selection of cross-reactive T cells by commensal and food-derived yeasts drives cytotoxic TH1 cell responses in Crohn’s disease

Aberrant CD4+ T cell reactivity against intestinal microorganisms is considered to drive mucosal inflammation in inflammatory bowel diseases. The disease-relevant microbial species and the corresponding microorganism-specific, pathogenic T cell phenotypes remain largely unknown. In the present study, we identified common gut commensal and food-derived yeasts, as direct activators of altered CD4+ T cell reactions in patients with Crohn’s disease (CD). Yeast-responsive CD4+ T cells in CD display a cytotoxic T helper cell (TH1 cell) phenotype and show selective expansion of T cell clones that are highly cross-reactive to several commensal, as well as food-derived, fungal species. This indicates cross-reactive T cell selection by repeated encounter with conserved fungal antigens in the context of chronic intestinal disease. Our results highlighted a role of yeasts as drivers of aberrant CD4+ T cell reactivity in patients with CD and suggest that both gut-resident fungal commensals and daily dietary intake of yeasts might contribute to chronic activation of inflammatory CD4+ T cell responses in patients with CD.

Immune monitoring of SARS-CoV-2-specific T cell and B cell responses in patients with multiple sclerosis treated with ocrelizumab.

Since the development of the coronavirus disease (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), there has been significant interest in determining the effectiveness of SARS-CoV-2 vaccines in patients under immunomodulatory or immunosuppressive therapies. The aim of this study was to evaluate the impact of ocrelizumab, a monoclonal anti-CD20 antibody, on SARS-CoV-2-specific T cell and B cell responses in patients with relapsing-remitting multiple sclerosis (RRMS). To this end, peripheral blood mononuclear cells (PBMCs) were isolated from n = 23 patients with RRMS. Of these patients, n = 17 were tested before (time point t0) and one month after (time point t1) their first dose of ocrelizumab. In addition, we studied n = 9 RRMS patients that got infected with SARS-CoV-2 over the course of ocrelizumab therapy (time point t2). PBMCs were also isolated from n = 19 age- and gender-matched healthy controls (HCs) after vaccination or infection with SARS-CoV-2, respectively. Interferon-γ (IFN-γ)/interleukin-2 (IL-2) and granzyme B (GzB)/perforin (PFN) double-color enzyme-linked immunospot (ELISPOT) assays or single-color ELISPOT assays were performed to measure SARS-CoV-2 antigen-specific T cell and B cell responses. Anti-viral antibody titers were quantified in the serum by chemiluminescence immunoassay. Our data indicate a significant difference in the SARS-CoV-2 specific IFN-γ (P = 0.0119) and PFN (P = 0.0005) secreting T cell compartment in the MS cohort at t0 compared to HCs. Following the first dose of ocrelizumab treatment, a significant decrease in the number of SARS-CoV-2 spike protein-specific B cells was observed (P = 0.0012). Infection with SARS-CoV-2 in MS patients under ocrelizumab therapy did not significantly alter their existing immune response against the virus. Kaplan-Meier survival analysis suggested that the spike S1 protein-specific immunoglobulin (Ig)G response might be a key parameter for predicting the probability of (re)infection with SARS-CoV-2. Our results call for a critical discussion regarding appropriate vaccination intervals and potential biomarkers for the prediction of (re)infection with SARS-CoV-2 in patients with MS receiving ocrelizumab. DRKS00029110; URL: http://apps.who.int/trialsearch/.

PL01.4. A LATE BREAKING ABSTRACT: A LONG PEPTIDE VACCINE TARGETING THE CLONAL DRIVER MUTATION H3K27M IN ADULT PATIENTS WITH DIFFUSE MIDLINE GLIOMA

Abstract BACKGROUND Clonal and mutually exclusive substitution of lysine 27 to methionine in histone H3-3A (H3K27M) defines an aggressive subtype of diffuse glioma. These diffuse midline gliomas (DMG) are aggressive, incurable primary brain tumors that occur predominantly in a midline location in children and young adults. Surgical treatment options remain limited, response to chemoradiation is poor and palliative radiotherapy remains the only intervention with proven benefit, resulting in a median overall survival between 10 and 15 months after initial diagnosis. Previous studies have shown that a H3K27M-specific long peptide vaccine (H3K27M-vac) induces mutation-specific immune responses capable of controlling H3K27M+ tumors in a major histocompatibility complex (MHC)-humanized mouse model. MATERIAL AND METHODS Here we describe the first-in-human treatment with H3K27M-vac of eight adult patients with recurrent H3K27M+ DMG. Induction of mutation-specific immune responses was determined in peripheral blood using Interferon-γ ELISpot assays. Class II HLA-restricted peripheral H3K27M-specific T cells were expanded in vitro to characterize and track unique H3K27M-reactive T cell responses in blood and cerebrospinal fluid (CSF). RESULTS Repeat vaccinations with H3K27M-vac were safe and induced mutation-specific immune responses in five of eight patients after a median of two vaccinations. Median progression free survival after start of vaccination was 6.2 months and median overall survival was 12.8 months. In vitro restimulation of peripheral CD4+ and CD8+ T cells with H3 mutant peptide or wild type peptide revealed that H3K27M specific immune responses are CD4-mediated and can be suppressed by blocking antibodies against MHC II, but not MHC I. Furthermore, proximity ligation assay of formalin-fixed paraffin-embedded primary tumor tissue revealed that H3K27M neoepitope co-localizes with HLA class II-DR. One patient showed radiographic pseudoprogression (PsPD) according to iRANO criteria within six weeks after first detection of mutation-specific peripheral immune response. Following PsPD, three out of the top ten vaccine-induced, K27M-expanded CD4+ TCR clonotypes from peripheral blood were detectable in the CSF of this patient who went into sustained complete remission for > 31 months. CONCLUSION H3K27M-vac is safe and induces H3K27M-specific CD4 T cell responses in patients with H3K27M+ DMG. An ongoing multicenter, phase 1 clinical trial for adult patients with newly diagnosed H3K27M+ DMG will assess the safety and immunogenicity of H3K27M-vac in combination with atezolizumab in standard-of-care radiotherapy (NCT04808245).

Elucidating the direct effects of the novel HDAC inhibitor bocodepsin (OKI-179) on T cells to rationally design regimens for combining with immunotherapy.

Histone deacetylase inhibitors (HDACi) are currently being explored for the treatment of both solid and hematological malignancies. Although originally thought to exert cytotoxic responses through tumor-intrinsic mechanisms by increasing expression of tumor suppressor genes, several studies have demonstrated that therapeutic responses depend on an intact adaptive immune system: particularly CD8 T cells. It is therefore critical to understand how HDACi directly affects T cells in order to rationally design regimens for combining with immunotherapy. In this study, we evaluated T cell responses to a novel class-selective HDACi (OKI-179, bocodepsin) by assessing histone acetylation levels, which revealed rapid responsiveness accompanied by an increase in CD4 and CD8 T cell frequencies in the blood. However, these rapid responses were transient, as histone acetylation and frequencies waned within 24 hours. This contrasts with in vitro models where high acetylation was sustained and continuous exposure to HDACi suppressed cytokine production. In vivo comparisons demonstrated that stopping OKI-179 treatment during PD-1 blockade was superior to continuous treatment. These findings provide novel insight into the direct effects of HDAC inhibitors on T cells and that treatment schedules that take into account acute T cell effects should be considered when combined with immunotherapies in order to fully harness the tumor-specific T cell responses in patients.