Reprogramming intratumoral Tregs by morpholino-mediated splicing of FOXP3 for cancer immunotherapy
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PROVIDER: PRJNA1207540 | ENA |
REPOSITORIES: ENA
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Project description:Regulatory T cells (Tregs) represent a primary barrier to the development of effective anti-tumor immunity. In the clinic, Tregs are a major contributor to the resistance of immune checkpoint blockade (ICB). Preclinical studies have reported complete tumor regression upon Treg depletion in mice. However, systemic depletion of Tregs is detrimental and sometimes lethal as it causes immune-related adverse effects, such as autoimmune diseases. Here, we report that reprogramming Treg functions, by shifting the expression of FOXP3 from its full-length isoform (FOXP3FL) to a short isoform with exon 2 skipped (FOXP3dE2), promotes the tumor-killing activity of T effectors while limiting the adverse effects of Treg depletion. FOXP3dE2 mRNA expression in triple negative breast cancer tissue positively correlates with overall survival of the patients. Mice expressing only the FOXP3dE2 isoform are resistant to the development of various types of tumors, while the littermate wildtype (WT) mice expressing FOXP3FL isoform exhibit significant tumor growth. Intratumoral injection of FOXP3dE2-directing morpholino (dE2 MO) in WT mice markedly increased IFN-.+ CD4 and CD8 T cells in tumors and consequently inhibited the tumor growth. We developed a novel mouse strain with a humanized Foxp3 exon 2 capable of expressing both FOXP3 isoforms through alternative splicing. This mouse strain showed better antitumor immunity than WT mice and dE2 MO treatment can further enhance their antitumor activity. The antitumor activity of dE2 MO was also confirmed using autologous tumor-infiltrating T cells and patient-derived tumor organoids. Taken together, our results suggest that enhancing FOXP3dE2 isoform expression will reprogram tumor infiltrating Tregs and convert these immune suppressive Tregs to helper-like T cells, thus promoting antitumor immunity.
2025-07-10 | GSE286047 | GEO
Project description:Though high frequency of Tregs in tumor obstructs anti-tumor immune responses, systemic depletion of Tregs caused severe autoimmune disease in mice model. Therefore, specifically targeting intratumoral Tregs is direly needed to fight against tumor. Our study provides a novel approach to target intratumoral Tregs to enhance anti-tumor immunotherapy.
2020-03-01 | GSE139325 | GEO
Project description:Treg cell therapy is a promising curative approach for a variety of immune-mediated conditions. CRISPR-based genome editing allows precise insertion of transgenes through homology-directed repair, but its use in human Tregs has been limited. We report an optimized protocol for CRISPR-mediated gene knock-in in human Tregs with high-yield expansion. To establish a benchmark of human Treg dysfunction, we target the master transcription factor FOXP3 in naive and memory Tregs. Although FOXP3-ablated Tregs upregulate cytokine expression, effects on suppressive capacity in vitro manifest slowly and primarily in memory Tregs. Moreover, FOXP3-ablated Tregs retain their characteristic protein, transcriptional, and DNA methylation profile. Instead, FOXP3 maintains DNA methylation at regions enriched for AP-1 binding sites. Thus, while FOXP3 is important for human Treg development, it has a limited role in maintaining mature Treg identity. Optimized gene knock-in with human Tregs will enable mechanistic studies and the development of tailored, next-generation Treg cell therapies.
2021-08-03 | GSE176191 | GEO
Project description:Immunotherapy provides an alternative approach for cancer treatment. However, in-depth analyses of the effects of immunotherapy on the tumor microenvironment (TME) have not been conducted in non-melanoma tumors. Here we describe changes in the pancreatic ductal adenocarcinoma (PDAC) TME following immunotherapy treatment, and show for the first time that vaccine-based immunotherapy directly alters the TME, inducing neogenesis of tertiary lymphoid structures that convert immunologically quiescent tumors into immunologically active tumors. Alterations in five pathways important for immune modulation and lymphoid structure development (TH17/Treg, NFkB, Ubiquitin-proteasome, Chemokines/chemokine receptors, and Integrins/adhesion molecules) in vaccine-induced intratumoral lymphoid aggregates were associated with improved post-vaccination responses. Additional studies in other cancers and patients treated with other forms of immunotherapy are warranted to further develop signatures defined in intratumoral lymphoid structures into biomarkers that predict effective anti-tumor immune responses. These signatures may also expose therapeutic targets for promoting more robust antitumor immune responses in the TME. Between July 2008 and September 2012, 59 patients were enrolled into an ongoing study of an irradiated, allogeneic GM-CSF-secreting pancreatic tumor vaccine (GVAX) administered intradermally either alone or in combination with immune modulatory doses of cyclophophamide (Cy) as neoadjuvant and adjuvant treatment for patients with resectable pancreatic ductal adenocarcinoma (PDAC). Patients were randomized 1:1:1 to 3 treatment arms. In Arm A, patients received GVAX alone; in Arm B, patients received GVAX plus a single intravenous dose of Cy at 200 mg/m2 1 day prior to each vaccination; in Arm C, patients received GVAX plus oral Cy at 100 mg once daily for 1 week on and 1 week off. Up to 6 GVAX treatments were administered and all of the patients remained in their initial treatment arms throughout the duration of the study. All 59 of the patients received the 1st GVAX treatment 2 weeks +/-4 days prior to surgery. Formalin-fixed paraffin-embedded (FFPE) tissue blocks of surgically resected PDAC were obtained from the pathology archive. FFPE tissue blocks from each subject were stained by H&E immediately before the vaccine therapy-induced lymphoid aggregates were microdissected . To better understand the functional status of these vaccine therapy induced lymphoid aggregate structures, gene microarray analysis on RNA isolated from microdissected lymphoid aggregates was performed. Gene expression was compared among samples grouped according to patient overall survival, post-vaccination induction of enhanced mesothelin-specific T cell responses in peripheral blood lymphocytes (PBL), and the intratumoral CD8+ T effector to FoxP3+ Treg ratio. Post-vaccination induction of enhanced mesothelin-specific T cell responses has been reported to correlate with longer survival in patients treated with Panc GVAX.
2014-08-05 | E-GEOD-52171 | biostudies-arrayexpress
Project description:Treg cell therapy is a promising curative approach for a variety of immune-mediated conditions. CRISPR-based genome editing allows precise insertion of transgenes through homology-directed repair, but its use in human Tregs has been limited. We report an optimized protocol for CRISPR-mediated gene knock-in in human Tregs with high-yield expansion. To establish a benchmark of human Treg dysfunction, we target the master transcription factor FOXP3 in naive and memory Tregs. Although FOXP3-ablated Tregs upregulate cytokine expression, effects on suppressive capacity in vitro manifest slowly and primarily in memory Tregs. Moreover, FOXP3-ablated Tregs retain their characteristic protein, transcriptional, and DNA methylation profile. Instead, FOXP3 maintains DNA methylation at regions enriched for AP-1 binding sites. Thus, while FOXP3 is important for human Treg development, it has a limited role in maintaining mature Treg identity. Optimized gene knock-in with human Tregs will enable mechanistic studies and the development of tailored, next-generation Treg cell therapies.
2021-08-03 | GSE164149 | GEO
Project description:Foxp3 expressing regulatory T cells (Tregs) are the central regulator of immune homeostasis and tolerance. As it is believed that proper Treg function is compromised under inflammatory conditions, exploring a pathway that enhances Treg function is of great importance. In this study, we report that IL-27, an IL-12 family cytokine known to play both pro- and anti-inflammatory role in T cells, plays a pivotal role in Treg function to control T cell-induced colitis. Unlike WT Tregs capable of inhibiting colitogenic T cell expansion and inflammatory cytokine expression, IL-27R-deficient Tregs were unable to downregulate inflammatory T cell responses. Tregs stimulated with IL-27 expressed substantially enhanced suppressive function both in vitro and in vivo. IL-27 stimulation of Tregs induced expression of LAG3, a surface molecule implicated in negatively regulating immune responses. LAG3 expression in IL-27-stimulated Tregs was critical to mediate suppressive Treg function. Finally, human Tregs also displayed enhanced suppressive function and LAG3 expression in response to IL-27 stimulation. Taken together, our results highlight a novel function of the IL-27/LAG3 axis in Treg regulation of inflammatory responses in the intestine. FACS purified Foxp3+ Tregs were stimulated in the presence of media or IL-27 to compare IL-27 induced gene profiles. Four samples (media stimulated or IL-27-stimulated) were collected from four independent experiments. Genes altered by IL-27 treatment were compared to those of media stimulated Tregs.
2015-11-01 | E-GEOD-63455 | biostudies-arrayexpress
Project description:Objectives. Immune homeostasis in the intestinal tract is tightly controlled by FOXP3+ regulatory T cells (Tregs), with loss of this Treg-mediated control linked to development of chronic conditions, such as inflammatory bowel disease (IBD). As a mechanism of immune evasion, several species of intestinal parasites strengthen intestinal Treg activity, leading to the notion that parasite-derived products could be harnessed and used as an immunoregulatory therapy for IBD. It has been previously demonstrated that the parasite Heligmosomoides polygyrus secretes a molecule (Hp-TGM) which mimics the ability of TGF-β to induce FOXP3 expression in CD4+ T cells. Our aim was to investigate whether Hp-TGM could induce human FOXP3+ Tregs as a potential therapeutic approach. Methods. Human CD4+ T cells from healthy volunteers were expanded in the presence of Hp-TGM or mammalian TGF-β. The induction of Tregs was measured by flow cytometric detection of FOXP3 and other Treg markers, such as CTLA-4 and CD25. Epigenetic changes were detected using ChIP-Seq and pyrosequencing of FOXP3. Treg phenotype stability was assessed following inflammatory cytokine challenge and Treg function was assessed by cellular co-culture suppression assays and secreted cytokines measured by cytometric bead array. Results. Hp-TGM efficiently induced FOXP3 expression (>60%), in addition to another Treg functional marker CTLA-4. Hp-TGM caused epigenetic modification of the FOXP3 loci to a greater extent than did TGF-β. Hp-TGM-induced Tregs also had superior suppressive function compared to TGF--induced Tregs, and retained their phenotype following exposure to inflammatory cytokines. Hp-TGM also induced a Treg phenotype in in vivo differentiated Th17 cells, indicating its potential to re-program memory cells to enhance immune tolerance. Conclusion. These data indicate Hp-TGM has the potential to be used to generate stable human FOXP3+ Tregs to treat IBD.
2021-04-29 | GSE164548 | GEO
Project description:Foxp3+ T-regulatory (Treg) cells maintain immune homeostasis and limit autoimmunity, but can also curtail host responses to cancers. Tregs are therefore promising targets to enhance anti-tumor immunity. Histone/protein acetyltransferases (HATs) promote chromatin accessibility, gene transcription and the function of multiple transcription factors and non-histone proteins. We found that conditional deletion or pharmacologic inhibition of one specific HAT, p300, in Foxp3+ Tregs, increased TCR-induced apoptosis in Tregs, impaired Treg suppressive function and iTreg peripheral conversion, and limited tumor growth in immunocompetent, but not in immunodeficient, hosts. Our data demonstrate that p300 is important for Foxp3+ Treg function and homeostasis in vivo and in vitro, and identify a novel mechanism to diminish Treg function without overtly impairing effector Tcell responses or inducing autoimmunity. Collectively, these data suggest a new approach for cancer immunotherapy. RNA from three independent samples from magnetically separated CD4+CD25+ Treg of fl-p300/Foxp3cre mice, compared to wild type (Foxp3cre) control (all C57Bl/6 background).
2013-08-18 | E-GEOD-47989 | biostudies-arrayexpress
Project description:TIGIT+ Tregs suppress Th1 and Th17 responses while sparing Th2 responses. Analysis of global gene expression of TIGIT+ vs. TIGIT- Tregs from naive mice reveled that TIGIT+ Tregs display an activated phenotype and are enriched for Treg signature genes including the Treg effector molecule Fgl2 which enables them to selectively spare Th2 responses. TIGIT+ and TIGIT- Tregs were sorted from naïve Foxp3-GFP KI mice (pooled spleen and lymph nodes) TIGIT: T cell immunoreceptor with Ig and ITIM domains
2014-12-12 | E-GEOD-56299 | biostudies-arrayexpress
Project description:BATF3-dependent IRF4 binding inhibits FOXP3 in differentiating Tregs
| PRJNA869441 | ENA