CD4+/CD8+ selection of anti-AML engager T cells alters their manufacturing environment, transcriptome, and functional phenotype
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PROVIDER: PRJNA1160783 | ENA |
REPOSITORIES: ENA
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Project description:Immunotherapies, including cell therapies, are effective anti-cancer agents. However, cellular product persistence can be limiting with short functional duration of activity contributing to disease relapse. A variety of manufacturing protocols are used to generate therapeutic engineered T-cells; these differ in techniques used for T-cell isolation, activation, genetic modification, and other methodology. We sought to determine how preselection affected the phenotype of T cells engineered to secrete a CD123xCD3 bispecific engager (ENG-T). These cells were designed to treat acute myeloid leukemia (AML). We evaluated the effect of T-cell selection on transduction efficiency, T-cell activation, short- and long-term anti-AML cytotoxicity, and gene transcription. Unselected, CD4, CD8, and CD4/CD8 pre-selected ENG-T cells have minor differences in T-cell subset components, equivalent activation, and equal cytotoxicity in short-term assays. While unselected and CD4/CD8-selected ENG-T cells have identical CD4:CD8 composition prior to target cell exposure, serial stimulation in vitro showed CD4/CD8 pre-selection supports ENG-T cell survival and long-term activity. Likewise, CD4 and CD4/CD8 pre-selected ENG-T cells display superior anti-tumor efficacy and prolong murine survival in AML xenografts. Unselected ENG-T cells are exposed to cytokines during early manufacture that imprint upregulation of intracellular inflammatory pathways. This early activation likely underpins long-term observed functional differences. Pre-selection of T cells from banked patient biospecimens decreased blast contamination, exposure to inflammatory cytokines, and may improve T-cell expansion during manufacture. Pre-selection of T-cell products should continue to be performed to enhance the quality of clinical cellular therapeutics.
2025-09-12 | GSE277096 | GEO
Project description:Bispecific dendritic-T Cell Engager potentiates anti-tumor immunity
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Project description:Immune checkpoint inhibition treatment using aPD-1 monoclonal antibodies is a promising cancer immunotherapy approach. However, its effect on tumor immunity is narrow, as most patients do not respond to the treatment or suffer from recurrence. We show that the crosstalk between conventional type I dendritic cells (cDC1) and T cells is essential for an effective aPD-1-mediated anti-tumor response. Accordingly, we developed a Bispecific DC-T Cell Engager (BiCE), a reagent that facilitates physical interactions between PD-1+ T-cells and cDC1. BiCE treatment promoted the formation of active dendritic/T cell crosstalk in the tumor and tumor-draining lymph nodes. In vivo, single cell and physical interacting cell analysis demonstrated the distinct and superior immune reprogramming of the tumors and tumor-draining lymph nodes treated with BiCE, as compared to conventional aPD-1 treatment. BiCE introduces a new concept in immunotherapy, bridging of immune cells to potentiate cell circuits and communication pathways needed for effective anti-tumor immunity.
2024-02-06 | GSE249283 | GEO
Project description:To date, bispecific antibodies designed to engage T cells have failed to show sustained responses in relapsed/refractory acute myeloid leukemia (R/R AML). Bispecific antibodies, including bispecific T-cell-engager (BiTE) molecules, predominantly recruit T cells from the local immune microenvironment. Because they are mainly applied in the R/R setting, understanding the fitness of T cells from bone marrow in AML progression, is fundamental for advancing this platform. Here, by flow cytometry analysis of bone marrow-derived T cells, we identified distinct differentiation stages and expression of inhibitory receptors at initial diagnosis (ID) and relapse (RL). Longitudinal transcriptional analyses of paired ID and RL T cells showed a senescent phenotype at ID, whereas an exhausted and memory phenotype was dominant at RL. In line with these observations, T cells at RL exhibited higher BiTE-mediated cytotoxicity than ID T cells. Finally, we provide evidence that T cells, both from ID and RL, exhibit limited functional capacity following continuous BiTE exposure compared to T cells from complete remission. Our study provides insights into the different stages of T-cell phenotype and function during AML evolution. Correlative biomarker studies in patients treated with T-cell-based immunotherapies are needed, to better understand resistance mechanisms and to advance the platform.
2024-08-22 | GSE261000 | GEO
Project description:To date, bispecific antibodies designed to engage T cells have failed to show sustained responses in relapsed/refractory acute myeloid leukemia (R/R AML). Bispecific antibodies, including bispecific T-cell-engager (BiTE) molecules, predominantly recruit T cells from the local immune microenvironment. Because they are mainly applied in the R/R setting, understanding the fitness of T cells from bone marrow in AML progression, is fundamental for advancing this platform. Here, by flow cytometry analysis of bone marrow-derived T cells, we identified distinct differentiation stages and expression of inhibitory receptors at initial diagnosis (ID) and relapse (RL). Longitudinal transcriptional analyses of paired ID and RL T cells showed a senescent phenotype at ID, whereas an exhausted and memory phenotype was dominant at RL. In line with these observations, T cells at RL exhibited higher BiTE-mediated cytotoxicity than ID T cells. Finally, we provide evidence that T cells, both from ID and RL, exhibit limited functional capacity following continuous BiTE exposure compared to T cells from complete remission. Our study provides insights into the different stages of T-cell phenotype and function during AML evolution. Correlative biomarker studies in patients treated with T-cell-based immunotherapies are needed, to better understand resistance mechanisms and to advance the platform.
2024-08-22 | GSE260999 | GEO
Project description:Glioblastoma (GBM) is the most lethal primary brain tumor in adults. No treatment provides durable relief for the vast majority of GBM patients. In this study, we've tested a bispecific antibody comprised of single-chain variable fragments (scFvs) against T cell CD3ε and GBM cell interleukin 13 receptor alpha 2 (IL13Rα2). We demonstrate that this bispecific T cell engager (BiTE) (BiTELLON) engages peripheral and tumor-infiltrating lymphocytes harvested from patients' tumors and, in so doing, exerts anti-GBM activity ex vivo. The interaction of BiTELLON with T cells and IL13Rα2-expressing GBM cells stimulates T cell proliferation and the production of proinflammatory cytokines interferon γ (IFNγ) and tumor necrosis factor α (TNFα). We have modified neural stem cells (NSCs) to produce and secrete the BiTELLON (NSCLLON). When injected intracranially in mice with a brain tumor, NSCLLON show tropism for tumor, secrete BiTELLON, and remain viable for over 7 d. When injected directly into the tumor, NSCLLON provide a significant survival benefit to mice bearing various IL13Rα2+ GBMs. Our results support further investigation and development of this therapeutic for clinical translation.
| S-EPMC7936285 | biostudies-literature
Project description:Cholangiocarcinoma (CCA) is a lethal cancer of bile duct epithelial cells with a high mortality rate and limited therapeutic options. An effective treatment is, therefore, urgently needed to improve treatment outcomes for these patients. To develop a new therapeutic option, we engineered T cells secreting αCD133-αCD3 bispecific T-cell engager and evaluated their antitumor effects against CD133-expressing CCA cells. The cDNA encoding αCD133-αCD3 bispecific T-cell engager (αCD133-αCD3-ENG) was cloned into pCDH lentiviral construct and its expression was tested in Lenti-X 293T cells. T cells from healthy donors were then transduced with engineered lentiviruses to create T cells secreting αCD133-αCD3 engager to evaluate their antitumor activities. The average transduction efficiency into T cells was approximately 60.03±21.65%. In the co-culture system containing T cells secreting αCD133-αCD3 engager (as effector cells) and mWasabi-luciferase-expressing CCA cells (KKU-100 and KKU-213A; as target cells), the effector T cells exhibited significantly higher cytolytic activities against the target CCA cells (49.0±9.76% and 64.10±13.18%, respectively) than those observed against the untransduced T cells (10.97±10.65%; p = 0.0103 and 9.80±11.05%; p = 0.0054) at an effector-to-target ratio of 5:1. In addition, the secreted αCD133-αCD3 engager significantly redirected both transduced T cells and bystander T cells to kill the target CCA cells (up to 73.20±1.68%; p<0.05). Moreover, the transduced and bystander T cells could kill the target CCA spheroids at a rate approximately 5-fold higher than that of the no treatment control condition (p = 0.0011). Our findings demonstrate proof-of-principle that T cells secreting αCD133-αCD3 engager can be an alternative approach to treating CD133-positive CCA, and they pave the way for future in vivo study and clinical trials.
| S-EPMC8936442 | biostudies-literature
Project description:Bispecific T-cell engager (BiTE)-based cancer therapies that activate the cytotoxic T cells of a patient’s own immune system have gained momentum with the recent FDA approval of Blinatumomab for treating B cell malignancies. However, this approach has had limited success in targeting solid tumors. We have reported the development of BiTE-sialidase fusion proteins that enhance tumor cell susceptibility to BiTE-mediated cytolysis by T cells via targeted desialylation at the BiTE-induced T cell-tumor cell interface. Targeted desialylation results in better immunological synapse formation, T-cell activation and effector function. As a result, BiTE-sialidase fusion proteins show remarkably increased efficacy in inducing T-cell-dependent tumor cell cytolysis in response to target antigens compared to the parent BiTE molecules alone. This enhanced function is seen both in vitro and in in vivo xenograft and syngeneic solid tumor mouse models. Our findings highlight BiTE-sialidase fusion proteins as promising candidates for the development of next-generation bispecific T-cell engaging molecules for cancer immunotherapy. This transcriptomic dataset documents the effect of BiTE-sialidase vs uncojugated BiTE on T cells including the upregulation of effector associated genes.
2024-06-05 | GSE245991 | GEO
Project description:BackgroundOncolytic virus (OV)-based therapies have an emerging role in the treatment of solid tumors, involving both direct cell lysis and immunogenic cell death. Nonetheless, tumor-associated stroma limits the efficacy of oncolytic viruses by forming a barrier that blocks efficient viral penetration and spread. The stroma also plays a critical role in progression, immunosuppression and invasiveness of cancer. Fibroblast activation protein-α (FAP) is highly overexpressed in cancer-associated fibroblasts (CAFs), the main cellular component of tumor stroma, and in this study we assessed whether arming oncolytic adenovirus (OAd) with a FAP-targeting Bispecific T-cell Engager (FBiTE) could retarget infiltrated lymphocytes towards CAFs, enhancing viral spread and T cell-mediated cytotoxicity against the tumor stroma to improve therapeutic activity.MethodsThe bispecific T-cell Engager against FAP was constructed using an anti-human CD3 single-chain variable fragment (scFv) linked to an anti-murine and human FAP scFv. This FBiTE was inserted in the oncolytic adenovirus ICOVIR15K under the control of the major late promoter, generating the ICO15K-FBiTE. ICO15K-FBiTE replication and potency were assessed in HT1080 and A549 tumor cell lines. The expression of the FBiTE and the activation and proliferation of T cells that induced along with the T cell-mediated cytotoxicity of CAFs were evaluated by flow cytometry in vitro. In vivo, T-cell biodistribution and antitumor efficacy studies were conducted in NOD/scid/IL2rg-/- (NSG) mice.ResultsFBiTE expression did not decrease the infectivity and replication potency of the armed virus. FBiTE-mediated binding of CD3+ effector T cells and FAP+ target cells led to T-cell activation, proliferation, and cytotoxicity of FAP-positive cells in vitro. In vivo, FBiTE expression increased intratumoral accumulation of T cells and decreased the level of FAP, a marker of CAFs, in tumors. The antitumor activity of the FBiTE-armed adenovirus was superior to the parental virus.ConclusionsCombination of viral oncolysis of cancer cells and FBiTE-mediated cytotoxicity of FAP-expressing CAFs might be an effective strategy to overcome a key limitation of oncolytic virotherapy, encouraging its further clinical development.
| S-EPMC6347837 | biostudies-literature
Project description:Transcriptional profiling of human peripheral B cell subsets sorted by flow cytometry based on the secretion of IL-10 in response to CpG2006 stimulation. Goal was to identify molecular markers to distinguish IL-10-producing B cells form non-IL-10-producing B cells. Two-condition experiment, non-IL-10-secreting B cells vs. IL-10-secreting B cells. Healthy donors. Biological replicates: 6
2013-03-04 | E-GEOD-35002 | biostudies-arrayexpress