Contractile activity (CA) induces changes of cellular environment in cells, such as increased oxidative stress or inflammation. The changes are double‐ edged sword. When they are subtle and transient, they serve as signals of muscle growth induced by CA. However, cells could be damaged if CA‐induced oxidative stress and inflammation are excessive and prolonged. To date, it is unknown whether the responses to aerobic CA are fiber‐type specific. The aim of the study is to determine the effects of skeletal muscle types on inflammation and oxidative stress following aerobic CA. Type I (soleus) and Type II (white gastrocnemius) muscles from stimulated and contralateral non‐stimulated limbs were collected immediately and 6 hours after 21‐mintues of aerobic CA. Markers of oxidative stress (GSH, GSH/GSSG, oxidative‐modified proteins and lipids), inflammation (TNFα, IL6), growth factors (IGF1, MGF and myostatin), myogenesis (Myf5), and protein degradation (MuRF1) were evaluated. Independent t test was used to compare parameters between groups. Compared to type I muscles, type II muscles had lower basal level of oxidative stress (less accumulation of oxidative‐modified proteins and lipids, lower GSH level and GSH/GSSG), lower inflammation state (lower TNFα and IL6 levels), and lower levels of signals that stimulate the increase of muscle mass (lower levels of IGF1, MGF and Myf5, and higher level of myostatin and MuRF1). In response to aerobic CA, type II muscles showed earlier accumulation of oxidative damages than that in type I muscles. In addition, the Myf5 level in type I muscles elevated at 6 h after CA, but this change was not observed in type II muscles. Type II muscle is more susceptible to contractile activity‐induced injury. In addition, the regeneration capacity of type II muscle following aerobic CA is weaker than type I muscle.
Abstract BACKGROUND A human leukocyte antigen (HLA)-A*02-restricted tumor associated antigen in the neuroligin 4 X-linked (NLGN4X) protein was found to be specifically overexpressed in human gliomas. Individualized multipeptide vaccination targeted NLGN4X and induced specific cytotoxic T cells responses in patients with newly diagnosed glioblastoma. MATERIAL AND METHODS Post vaccination NLGN4X-tetramer-sorted T cells were subjected to single cell T cell receptor (TCR) sequencing for TCR discovery. The identified TCR was delivered to human T cells (NLGN4X-TCR-T) and functional profiling was performed by flow cytometry and in vitro cytotoxicity assays. NOD scid gamma (NSG) major histocompatibility complex (MHC) I/II knockout (KO) (NSG MHC I/II KO) mice were challenged with NLGN4X-expressing experimental gliomas and treated with intracerebroventricular injection of NLGN4X-TCR-T to assess its therapeutic potential. RESULTS We apply for the first time for therapeutic use of an HLA-A*02 restricted vaccine-induced TCR that binds to the NLGN4X antigen. We show the cytotoxic and polyfunctional phenotype of NLGN4X-TCR-T in various cellular models. Intracerebroventricular delivery of NLGN4X-TCR-T prolongs survival and leads to objective radiographic responses in experimental gliomas-bearing NSG MHC I/II KO mice. CONCLUSION NLGN4X-TCR-T demonstrates efficacy in a preclinical experimental glioblastoma model. On a global scale, we provide first evidence for the therapeutic retrieval of vaccine-induced human TCRs for the off-the-shelf treatment of glioblastoma patients.
Abstract BACKGROUND Neoepitopes are presented on major histocompatibility class II (MHCII) molecules. In glioma, for instance, the recurrent driver mutation IDH1R132H was shown to bear an MHCII-restricted epitope in preclinical and clinical vaccine studies. The general relevance of MHCII expression in glioma for antitumor immunity, however, remains unknown. Here we evaluate stromal and tumoral MHCII expression, functionality, and its association with survival in gliomas. MATERIAL AND METHODS Immunostaining of human glioma tissues was used to identify tumoral, endothelial, and microglial MHCII expression and to enumerate T cell infiltrates. To gain insights into tumoral MHCII expression, bulk transcriptomic data from TCGA and single-cell transcriptomic data from publicly available datasets were analyzed. MHC ligandome analyses of an MHCII+ glioma cell line and human glioma tissues were used to determine the functionality of MHCII in vitro and ex vivo. Functional in vitro co-culture assays with an HLA-DR-matched tetanus toxoid (TT) epitope-overexpressing glioma cell line and in vitro-expanded TT-reactive T cells from healthy donors were used to examine direct target recognition by T helper cells. CRISPR-Cas9-mediated knockout of MHCII in preclinical hypermutant glioblastoma cell line GL261 was employed to further validate the consequences of tumoral MHCII expression and to probe potential clinical intervention with existing therapies. RESULTS MHCII is expressed in the majority of gliomas and associated with increased infiltration of T cells. In 10% of the analyzed glioma tissues and a subset of single cells, tumoral MHCII expression is detected. Clinical and transcriptomic data reveal that tumoral MHCII is associated with poor prognosis, cytokine responses, immune inhibition and T cell differentiation. Ligandome analyses evidence presentation of peptides by MHCII molecules on glioma cells. In in vitro assays, TT-reactive T helper cells specifically produce IFNg when co-cultured with MHCII+ glioma cells upon the presence of co-stimulation. In agreement with the clinical data, preclinical murine models demonstrate that tumoral MHCII expression leads to reduced survival. Co-culture assay shows that tumoral MHCII results in upregulation of PD-1 on T helper cells antigen-specifically. Concordantly, immune checkpoint blockade (ICB) therapy slows the disease progression of mice carrying MHCII+ tumors. CONCLUSION MHCII is expressed in gliomas by a subset of tumor cells. Although tumoral MHCII is functional, it is associated with poor survival in both clinical data and preclinical models. T cell exhaustion induced by tumoral MHCII expression can, in part, be overcome by ICB in vivo. Further experiments are required to decipher tumor cell intrinsic and microenvironmental consequences of tumoral MHCII expression.
Abstract BACKGROUND Novel concepts in immunotherapy have evolved with the identification of potential (neo)epitopes and/or combinations with immune checkpoint inhibitors (ICI). In contrast to many other solid tumors, ICI have not improved outcome for patients with glioblastoma (GB) in phase 3 studies. However, antigen-specific T cells induced by vaccines or adoptively transferred as chimeric antigen receptor (CAR) T cells produced encouraging responses in early clinical trials and case series. From one of these trials, the Actively Personalized Vaccine Consortium (GAPVAC) European Clinical Trial 101, we have identified a human leukocyte antigen (HLA)-A2-restricted T cell receptor (TCR) targeting protein tyrosine phosphatase receptor type zeta 1 (PTPRZ1). PTPRZ1 is strongly overexpressed in malignant gliomas, especially GB, and analyses of intratumoral heterogeneity revealed that the level of PTPRZ1 overexpression is strongly associated with cancer stemness. The PTPRZ1 epitope is naturally presented on HLA-A2. MATERIAL AND METHODS INVENT4GB is a German, multicenter, phase 1, first-in-human, first-of-kind dose-escalation window-of-opportunity investigator-initiated trial in recurrent GB, eligible for re-resection, assessing safety and feasibility of therapy with intravenously (iv) and intraventricularly (icv) infused TCR-engineered T cells (TCR-T). FPI is scheduled for 2024. HLA-A*02:01 positive patients will receive one neoadjuvant iv followed by up to three adjuvant icv infusions of PTPRZ1-specific TCR-transgenic autologous T cell therapy (TCR-T-001) with cyclophosphamide- and fludarabine-based lymphodepletion prior to iv infusion. Patient autologous T cells will be engineered by non-integrating nano scaffold matrix attachment region (SMAR) TCR DNA vectors. Icv infusions will be performed via intraventricular catheter implanted during re-resection. Biomarker discovery includes magnetic resonance imaging (MRI) and longitudinal flow cytometry-based immune monitoring of blood and cerebrospinal fluid. Primary endpoints are feasibility and safety (determination of maximum tolerated dose/ MTD), key secondary endpoints are progression-free survival according to the Immunotherapy Response Assessment in Neurooncology (iRANO) criteria. RESULTS TCR-T-001 robustly recognizes target-expressing cells in vitro and is efficacious in preclinical tumor models. Preclinical data and the trial concept will be presented. CONCLUSION INVENT4GB is a first-of-kind TCR-engineered T cell therapy for GB patients.
Background: Previous studies have shown that geriatric insomnia can impact the quality of life and lead to various health risks and comorbidities, which may further result in out-of-hospital cardiac arrest (OHCA). Therefore, assessing the risk of OHCA in elderly patients with insomnia is crucial. Hypothesis: We hypothesized that a predictive model incorporating demographic, clinical, and treatment adherence factors can effectively estimate the risk of OHCA among geriatric patients with insomnia. Aim: This study aimed to develop and validate a predictive model for OHCA in geriatric patients with insomnia using a large-scale population-based analysis. Method: A retrospective cohort study was conducted using data from Taiwan's Health and Welfare Data Science Center spanning from 2011 to 2020. The study included elderly patients diagnosed with insomnia who received medication. Predictors of OHCA, such as age, sex, comorbidities, medication adherence, and psychotherapy, were analyzed using multivariable logistic regression. Model performance was assessed using the area under the receiver operating characteristic curve (AUROC) with both internal and temporal validation. Results: Among 438,147 eligible elderly patients with insomnia, 7,666 experienced OHCA. Using data from 2011 to 2018 for model development, the AUROC for predicting OHCA within 7 days, 30 days, and 90 days ranged from 0.749 to 0.762 in the training set and 0.768 to 0.784 in the validation set. Key predictors included older age, male sex, prior medical resource use, hemodialysis, pre-existing comorbidities, medication possession ratio, medication changes, and recent psychotherapy (Figure 1). The model was also tested in 2019 and 2020 dataset (began of the COVID-19 in Taiwan), respectively, demonstrating consistent predictive performance (Figure 2). Specifically, the 7-day model in 2019 showed a sensitivity of 0.776, specificity of 0.632, and accuracy of 0.632, with a positive likelihood ratio (PLR) of 2.11, and negative likelihood ratio (NLR) of 0.36. In 2020, it achieved a sensitivity of 0.739, specificity of 0.661, accuracy of 0.661, PLR of 2.18, NLR of 0.40. Conclusion: The study developed a reliable predictive model for OHCA in geriatric patients with insomnia, highlighting significant predictors such as age, comorbidities, medication adherence, and recent psychotherapy. This model can aid clinicians in identifying high-risk patients and improving preventative care strategies.
