RESUMO
Inflammatory neuropathies, which include chronic inflammatory demyelinating polyneuropathy (CIDP) and Guillain Barré syndrome (GBS), result from autoimmune destruction of the PNS and are characterized by progressive weakness and sensory loss. CD4+ T cells play a key role in the autoimmune destruction of the PNS. Yet, key properties of pathogenic CD4+ T cells remain incompletely understood. Here, we used paired single-cell RNA-Seq (scRNA-Seq) and single-cell T cell receptor-sequencing (scTCR-Seq) of peripheral nerves from an inflammatory neuropathy mouse model to identify IL-21-expressing CD4+ T cells that were clonally expanded and multifunctional. These IL-21-expressing CD4+ T cells consisted of 2 transcriptionally distinct expanded cell populations, which expressed genes associated with T follicular helper (Tfh) and T peripheral helper (Tph) cell subsets. Remarkably, TCR clonotypes were shared between these 2 IL-21-expressing cell populations, suggesting a common lineage differentiation pathway. Finally, we demonstrated that IL-21 receptor-KO (IL-21R-KO) mice were protected from neuropathy development and had decreased immune infiltration into peripheral nerves. IL-21 signaling upregulated CXCR6, a chemokine receptor that promotes CD4+ T cell localization in peripheral nerves. Together, these findings point to IL-21 signaling, Tfh/Tph differentiation, and CXCR6-mediated cellular localization as potential therapeutic targets in inflammatory neuropathies.
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Linfócitos T CD4-Positivos , Interleucinas , Animais , Camundongos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/patologia , Linhagem da Célula , Interleucinas/genética , Interleucinas/imunologia , Interleucinas/metabolismo , Camundongos Knockout , Masculino , FemininoRESUMO
In this randomized phase II clinical trial, we evaluated the effectiveness of adding the TLR agonists, poly-ICLC or resiquimod, to autologous tumor lysate-pulsed dendritic cell (ATL-DC) vaccination in patients with newly-diagnosed or recurrent WHO Grade III-IV malignant gliomas. The primary endpoints were to assess the most effective combination of vaccine and adjuvant in order to enhance the immune potency, along with safety. The combination of ATL-DC vaccination and TLR agonist was safe and found to enhance systemic immune responses, as indicated by increased interferon gene expression and changes in immune cell activation. Specifically, PD-1 expression increases on CD4+ T-cells, while CD38 and CD39 expression are reduced on CD8+ T cells, alongside an increase in monocytes. Poly-ICLC treatment amplifies the induction of interferon-induced genes in monocytes and T lymphocytes. Patients that exhibit higher interferon response gene expression demonstrate prolonged survival and delayed disease progression. These findings suggest that combining ATL-DC with poly-ICLC can induce a polarized interferon response in circulating monocytes and CD8+ T cells, which may represent an important blood biomarker for immunotherapy in this patient population.Trial Registration: ClinicalTrials.gov Identifier: NCT01204684.
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Linfócitos T CD8-Positivos , Vacinas Anticâncer , Carboximetilcelulose Sódica/análogos & derivados , Células Dendríticas , Glioma , Interferons , Poli I-C , Polilisina/análogos & derivados , Humanos , Células Dendríticas/imunologia , Células Dendríticas/efeitos dos fármacos , Glioma/imunologia , Glioma/terapia , Feminino , Masculino , Pessoa de Meia-Idade , Vacinas Anticâncer/imunologia , Vacinas Anticâncer/administração & dosagem , Vacinas Anticâncer/uso terapêutico , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Poli I-C/administração & dosagem , Poli I-C/farmacologia , Adulto , Receptores Toll-Like/agonistas , Imidazóis/farmacologia , Imidazóis/uso terapêutico , Idoso , Vacinação , Monócitos/imunologia , Monócitos/efeitos dos fármacos , Neoplasias Encefálicas/imunologia , Neoplasias Encefálicas/terapia , Neoplasias Encefálicas/tratamento farmacológico , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Imunoterapia/métodos , Agonistas do Receptor Semelhante a TollRESUMO
OBJECTIVE: Unravel the potential mechanism(s) of the on- and off-target actions of dopamine agonist therapy in both human prolactinoma tumors and neighboring stromal and immune cells. DESIGN AND METHODS: Five surgically resected prolactinomas (PRLomas) from 3 cabergoline (CBG)-treated patients and 2 treatment-naive patients were analyzed by using single-cell RNA sequencing (scRNA-seq) to compare the cellular composition and transcriptional landscape. RESULTS: Six major cell populations, namely tumor (88.2%), immune (5.6%), stromal (4.9%), progenitor cells (0.6%), proliferating cells (0.4%), and erythrocytes (0.2%), were observed. Tumor cells from CBG-treated patients expressed lower levels of genes that regulated hormone secretion, such as SCG2, VGF, TIMP1, NNAT, and CALD1, consistent with the inhibitory effects of CBG on hormone processing and secretion. Interestingly, we also observed an increased number of CD8+ T cells in the CBG-treated tissues. These cytotoxic CD8+ T cells expressed killing granule components such as perforin and the granzymes GZMB, GNLY, and KLRD1 as well as the inflammatory cytokine CCL5. Immune cell activation of these CD8+ T cells was further analyzed in a compartment-specific manner, and increased CD25 (IL2R) expression was noted in the CD8+ T cells from the CBG-treated samples. Additionally, and confirming prior reports, we noted a higher stromal cell population in the CBG-treated samples. CONCLUSIONS: Our scRNA-seq studies revealed key differences in the transcriptomic features of CBG-treated and CBG-untreated PRLomas in both tumor and microenvironment cellular constituents, and for the first time, describe the previously unknown activation of CD8+ T cells following CBG treatment, which may play a role in the tumoricidal actions of CBG.
Assuntos
Cabergolina , Neoplasias Hipofisárias , Prolactinoma , Humanos , Cabergolina/farmacologia , Cabergolina/uso terapêutico , Prolactinoma/tratamento farmacológico , Prolactinoma/metabolismo , Masculino , Feminino , Neoplasias Hipofisárias/metabolismo , Neoplasias Hipofisárias/tratamento farmacológico , Neoplasias Hipofisárias/patologia , Adulto , Pessoa de Meia-Idade , Fibrose , Prolactina/metabolismo , Agonistas de Dopamina/farmacologia , Agonistas de Dopamina/uso terapêutico , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/metabolismo , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismo , Adulto Jovem , Microambiente Tumoral/efeitos dos fármacosRESUMO
Inflammatory neuropathies, which include CIDP (chronic inflammatory demyelinating polyneuropathy) and GBS (Guillain Barre Syndrome), result from autoimmune destruction of the peripheral nervous system (PNS) and are characterized by progressive weakness and sensory loss. CD4+ T cells play a key role in the autoimmune destruction of the PNS. Yet, key properties of pathogenic CD4+ T cells remain incompletely understood. Here, we use paired scRNAseq and scTCRseq of peripheral nerves from an inflammatory neuropathy mouse model to identify IL-21 expressing CD4+ T cells that are clonally expanded and multifunctional. These IL-21-expressing CD4+ T cells are comprised of two transcriptionally distinct expanded populations, which express genes associated with Tfh and Tph subsets. Remarkably, TCR clonotypes are shared between these two IL-21-expressing populations, suggesting a common lineage differentiation pathway. Finally, we demonstrate that IL-21 signaling is required for neuropathy development and pathogenic T cell infiltration into peripheral nerves. IL-21 signaling upregulates CXCR6, a chemokine receptor that promotes CD4+ T cell localization in peripheral nerves. Together, these findings point to IL-21 signaling, Tfh/Tph differentiation, and CXCR6-mediated cellular localization as potential therapeutic targets in inflammatory neuropathies.
RESUMO
Autologous tumor lysate-pulsed dendritic cell (ATL-DC) vaccination is a promising immunotherapy for patients with high grade gliomas, but responses have not been demonstrated in all patients. To determine the most effective combination of autologous tumor lysate-pulsed DC vaccination, with or without the adjuvant toll-like receptor (TLR) agonists poly-ICLC or resiquimod, we conducted a Phase 2 clinical trial in 23 patients with newly diagnosed or recurrent WHO Grade III-IV malignant gliomas. We then performed deep, high-dimensional immune profiling of these patients to better understand how TLR agonists may influence the systemic immune responses induced by ATL-DC vaccination. Bulk RNAseq data demonstrated highly significant upregulation of type 1 and type 2 interferon gene expression selectively in patients who received adjuvant a TLR agonist together with ATL-DC. CyTOF analysis of patient peripheral blood mononuclear cells (PBMCs) showed increased expression of PD-1 on CD4+ T-cells, decreases in CD38 and CD39 on CD8+ T cells and elevated proportion of monocytes after ATL-DC + TLR agonist administration. In addition, scRNA-seq demonstrated a higher expression fold change of IFN-induced genes with poly-ICLC treatment in both peripheral blood monocytes and T lymphocytes. Patients who had higher expression of interferon response genes lived significantly longer and had longer time to progression compared to those with lower expression. The results suggest that ATL-DC in conjunction with adjuvant poly-ICLC induces a polarized interferon response in circulating monocytes and specific activation of a CD8+ T cell population, which may represent an important blood biomarker for immunotherapy in this patient population. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT01204684.
RESUMO
In comparison with responses in recurrent glioblastoma (rGBM), the intracranial response of brain metastases (BrM) to immune checkpoint blockade (ICB) is less well studied. Here, we present an integrated single-cell RNA-Seq (scRNA-Seq) study of 19 ICB-naive and 9 ICB-treated BrM samples from our own and published data sets. We compared them with our previously published scRNA-Seq data from rGBM and found that ICB led to more prominent T cell infiltration into BrM than rGBM. These BrM-infiltrating T cells exhibited a tumor-specific phenotype and displayed greater activated/exhausted features. We also used multiplex immunofluorescence and spatial transcriptomics to reveal that ICB reduced a distinct CD206+ macrophage population in the perivascular space, which may modulate T cell entry into BrM. Furthermore, we identified a subset of progenitor exhausted T cells that correlated with longer overall survival in BrM patients. Our study provides a comprehensive immune cellular landscape of ICB's effect on metastatic brain tumors and offers insights into potential strategies for improving ICB efficacy for brain tumor patients.
Assuntos
Neoplasias Encefálicas , Glioblastoma , Humanos , Inibidores de Checkpoint Imunológico , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Perfilação da Expressão Gênica , Glioblastoma/tratamento farmacológico , Glioblastoma/genética , Macrófagos , Microambiente TumoralRESUMO
Introduction: Increased T cell infiltration and interferon gamma (IFNγ) pathway activation are seen in tumors of melanoma patients who respond to ICI (immune checkpoint inhibitor) or MAPK pathway inhibitor (MAPKi) therapies. Yet, the rate of durable tumor control after ICI is almost twice that of MAPKi, suggesting that additional mechanisms may be present in patients responding to ICI therapy that are beneficial for anti-tumor immunity. Methods: We used transcriptional analysis and clinical outcomes from patients treated with ICI or MAPKi therapies to delineate immune mechanisms driving tumor response. Results: We discovered response to ICI is associated with CXCL13-driven recruitment of CXCR5+ B cells with significantly higher clonal diversity than MAPKi. Our in vitro data indicate that CXCL13 production was increased in human peripheral blood mononuclear cells by anti-PD1, but not MAPKi, treatment. Higher B cell infiltration and B cell receptor (BCR) diversity allows presentation of diverse tumor antigens by B cells, resulting in activation of follicular helper CD4 T cells (Tfh) and tumor reactive CD8 T cells after ICI therapy. Higher BCR diversity and IFNγ pathway score post-ICI are associated with significantly longer patient survival compared to those with either one or none. Conclusions: Response to ICI, but not to MAPKi, depends on the recruitment of CXCR5+ B cells into the tumor microenvironment and their productive tumor antigen presentation to follicular helper and cytotoxic, tumor reactive T cells. Our study highlights the potential of CXCL13 and B cell based strategies to enhance the rate of durable response in melanoma patients treated with ICI.
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Inibidores de Checkpoint Imunológico , Melanoma , Humanos , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Apresentação de Antígeno , Leucócitos Mononucleares , Linfócitos T CD8-Positivos , Receptores de Antígenos de Linfócitos B , Melanoma/tratamento farmacológico , Microambiente Tumoral , Receptores CXCR5RESUMO
Autoimmune toxicity occurs in up to 60% of patients treated with immune checkpoint inhibitor (ICI) therapy for cancer and represents an increasing clinical challenge for expanding the use of these treatments. To date, human immunopathogenic studies of immune-related adverse events (IRAEs) have relied on sampling of circulating peripheral blood cells rather than affected tissues. Here, we directly obtained thyroid specimens from individuals with ICI-thyroiditis, one of the most common IRAEs, and compared immune infiltrates with those from individuals with spontaneous autoimmune Hashimoto's thyroiditis (HT) or no thyroid disease. Single-cell RNA sequencing revealed a dominant, clonally expanded population of thyroid-infiltrating cytotoxic CXCR6+ CD8+ T cells (effector CD8+ T cells) present in ICI-thyroiditis but not HT or healthy controls. Furthermore, we identified a crucial role for interleukin-21 (IL-21), a cytokine secreted by intrathyroidal T follicular (TFH) and T peripheral helper (TPH) cells, as a driver of these thyrotoxic effector CD8+ T cells. In the presence of IL-21, human CD8+ T cells acquired the activated effector phenotype with up-regulation of the cytotoxic molecules interferon-γ (IFN-γ) and granzyme B, increased expression of the chemokine receptor CXCR6, and thyrotoxic capacity. We validated these findings in vivo using a mouse model of IRAEs and further demonstrated that genetic deletion of IL-21 signaling protected ICI-treated mice from thyroid immune infiltration. Together, these studies reveal mechanisms and candidate therapeutic targets for individuals who develop IRAEs.
Assuntos
Tireoidite , Humanos , Linfócitos T CD8-Positivos/metabolismo , Doença de Hashimoto , Interleucinas , Tireoidite Autoimune/genética , Tireoidite Autoimune/patologia , Tireoidite/induzido quimicamente , Tireoidite/imunologiaRESUMO
Stimulator of interferon genes (STING) is a mediator of immune recognition of cytosolic DNA, which plays important roles in cancer, cytotoxic therapies, and infections with certain pathogens. Although pharmacologic STING activation stimulates potent antitumor immune responses in animal models, clinically applicable pharmacodynamic biomarkers that inform of the magnitude, duration, and location of immune activation elicited by systemic STING agonists are yet to be described. We investigated whether systemic STING activation induces metabolic alterations in immune cells that can be visualized by PET imaging. Methods: C57BL/6 mice were treated with systemic STING agonists and imaged with 18F-FDG PET after 24 h. Splenocytes were harvested 6 h after STING agonist administration and analyzed by single-cell RNA sequencing and flow cytometry. 18F-FDG uptake in total splenocytes and immunomagnetically enriched splenic B and T lymphocytes from STING agonist-treated mice was measured by γ-counting. In mice bearing prostate or pancreas cancer tumors, the effects of STING agonist treatment on 18F-FDG uptake, T-lymphocyte activation marker levels, and tumor growth were evaluated. Results: Systemic delivery of structurally distinct STING agonists in mice significantly increased 18F-FDG uptake in the spleen. The average spleen SUVmax in control mice was 1.90 (range, 1.56-2.34), compared with 4.55 (range, 3.35-6.20) in STING agonist-treated mice (P < 0.0001). Single-cell transcriptional and flow cytometry analyses of immune cells from systemic STING agonist-treated mice revealed enrichment of a glycolytic transcriptional signature in both T and B lymphocytes that correlated with the induction of immune cell activation markers. In tumor-bearing mice, STING agonist administration significantly delayed tumor growth and increased 18F-FDG uptake in secondary lymphoid organs. Conclusion: These findings reveal hitherto unknown functional links between STING signaling and immunometabolism and suggest that 18F-FDG PET may provide a widely applicable approach toward measuring the pharmacodynamic effects of systemic STING agonists at a whole-body level and guiding their clinical development.
Assuntos
Fluordesoxiglucose F18 , Ativação Linfocitária , Masculino , Animais , Camundongos , Fluordesoxiglucose F18/metabolismo , Camundongos Endogâmicos C57BL , Tomografia por Emissão de Pósitrons , Transdução de SinaisRESUMO
Immune checkpoint inhibitor (ICI) immunotherapy leverages the body's own immune system to attack cancer cells but leads to unwanted autoimmune side effects in up to 60% of patients. Such immune-related adverse events (IrAEs) may lead to treatment interruption, permanent organ dysfunction, hospitalization, and premature death. Thyroiditis is one of the most common IrAEs, but the cause of thyroid IrAEs remains unknown. In this study, we use a new, physiologically relevant mouse model of ICI-associated autoimmunity to identify a key role for type 3 immune cells in the development of thyroid IrAEs. Multiple lineages of IL-17A-producing T cells expand in thyroid tissue with ICI treatment. Intrathyroidal IL-17A-producing innate-like γδT17 cells were increased in tumor-free mice, whereas adaptive Th17 cells were also prominent in tumor-bearing mice, following ICI treatment. Furthermore, Ab-based inhibition of IL-17A, a clinically available therapy, significantly reduced thyroid IrAE development in ICI-treated mice with and without tumor challenge. Finally, combination of IL-17A neutralization with ICI treatment in multiple tumor models did not reduce ICI antitumor efficacy. These studies suggest that targeting Th17 and γδT17 cell function via the IL-17A axis may reduce IrAEs without impairing ICI antitumor efficacy and may be a generalizable strategy to address type 3 immune-mediated IrAEs.
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Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Neoplasias , Animais , Imunoterapia , Interleucina-17 , Camundongos , Neoplasias/patologia , Glândula Tireoide/patologiaRESUMO
Purine nucleoside phosphorylase (PNP) enables the breakdown and recycling of guanine nucleosides. PNP insufficiency in humans is paradoxically associated with both immunodeficiency and autoimmunity, but the mechanistic basis for these outcomes is incompletely understood. Here, we identify two immune lineage-dependent consequences of PNP inactivation dictated by distinct gene interactions. During T cell development, PNP inactivation is synthetically lethal with downregulation of the dNTP triphosphohydrolase SAMHD1. This interaction requires deoxycytidine kinase activity and is antagonized by microenvironmental deoxycytidine. In B lymphocytes and macrophages, PNP regulates Toll-like receptor 7 signaling by controlling the levels of its (deoxy)guanosine nucleoside ligands. Overriding this regulatory mechanism promotes germinal center formation in the absence of exogenous antigen and accelerates disease in a mouse model of autoimmunity. This work reveals that one purine metabolism gene protects against immunodeficiency and autoimmunity via independent mechanisms operating in distinct immune lineages and identifies PNP as a potentially novel metabolic immune checkpoint.
Assuntos
Síndromes de Imunodeficiência , Purina-Núcleosídeo Fosforilase , Animais , Autoimunidade , Humanos , Camundongos , Nucleosídeos de Purina , Purina-Núcleosídeo Fosforilase/genética , Purina-Núcleosídeo Fosforilase/metabolismo , Linfócitos T , Receptor 7 Toll-LikeRESUMO
Objective: Provide insights into the defective POMC processing and invasive behavior in silent pituitary corticotroph tumors. Design and methods: Single-cell RNAseq was used to compare the cellular makeup and transcriptome of silent and active corticotroph tumors. Results: A series of transcripts related to hormone processing peptidases and genes involved in the structural organization of secretory vesicles were reduced in silent compared to active corticotroph tumors. Most relevant to their invasive behavior, silent corticotroph tumors exhibited several features of epithelial-to-mesenchymal transition, with increased expression of mesenchymal genes along with the loss of transcripts that regulate hormonal biogenesis and secretion. Silent corticotroph tumor vascular smooth muscle cell and pericyte stromal cell populations also exhibited plasticity in their mesenchymal features. Conclusions: Our findings provide novel insights into the mechanisms of impaired POMC processing and invasion in silent corticotroph tumors and suggest that a common transcriptional reprogramming mechanism simultaneously impairs POMC processing and activates tumor invasion.
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Corticotrofos , Neoplasias Hipofisárias , Corticotrofos/metabolismo , Humanos , Neoplasias Hipofisárias/patologia , Pró-Opiomelanocortina/genética , Pró-Opiomelanocortina/metabolismo , Análise de Sequência de RNARESUMO
Immune checkpoint blockade (ICB) has become well-known in cancer therapy, strengthening the body's antitumor immune response rather than directly targeting cancer cells. Therapies targeting immune inhibitory checkpoints, such as PD-1, PD-L1, and CTLA-4, have resulted in impressive clinical responses across different types of solid tumors. However, as with other types of cancer treatments, ICB-based immunotherapy is hampered by both innate and acquired drug resistance. We previously reported the enrichment of gene signatures associated with wound healing, epithelial-to-mesenchymal, and angiogenesis processes in the tumors of patients with innate resistance to PD-1 checkpoint antibody therapy; we termed these the Innate Anti-PD-1 Resistance Signatures (IPRES). The TGF-ß and VEGFA pathways emerge as the dominant drivers of IPRES-associated processes. Here, we review these pathways' functions, their roles in immunosuppression, and the currently available therapies that target them. We also discuss recent developments in the targeting of TGF-ß using a specific antibody class termed trap antibody. The application of trap antibodies opens the promise of localized targeting of the TGF-ß and VEGFA pathways within the tumor microenvironment. Such specificity may offer an enhanced therapeutic window that enables suppression of the IPRES processes in the tumor microenvironment while sparing the normal homeostatic functions of TGF-ß and VEGFA in healthy tissues.
Assuntos
Melanoma , Neoplasias , Humanos , Receptor de Morte Celular Programada 1 , Fator de Crescimento Transformador beta , Fator A de Crescimento do Endotélio Vascular , Neoplasias/terapia , Imunoterapia/métodos , Microambiente Tumoral , Melanoma/tratamento farmacológico , AnticorposRESUMO
Immune cells infiltrate the peripheral nervous system (PNS) after injury and with autoimmunity, but their net effect is divergent. After injury, immune cells are reparative, while in inflammatory neuropathies (e.g., Guillain Barré Syndrome and chronic inflammatory demyelinating polyneuropathy), immune cells are proinflammatory and promote autoimmune demyelination. An understanding of immune cell phenotypes that distinguish these conditions may, therefore, reveal new therapeutic targets for switching immune cells from an inflammatory role to a reparative state. In an autoimmune regulator (Aire)-deficient mouse model of inflammatory neuropathy, we used single-cell RNA sequencing of sciatic nerves to discover a transcriptionally heterogeneous cellular landscape, including multiple myeloid, innate lymphoid, and lymphoid cell types. Analysis of cell-cell ligand-receptor interactions uncovered a macrophage-mediated tumor necrosis factor-α (TNF-α) signaling axis that is induced by interferon-γ and required for initiation of autoimmune demyelination. Developmental trajectory visualization suggested that TNF-α signaling is associated with metabolic reprogramming of macrophages and polarization of macrophages from a reparative state in injury to a pathogenic, inflammatory state in autoimmunity. Autocrine TNF-α signaling induced macrophage expression of multiple genes (Clec4e, Marcksl1, Cxcl1, and Cxcl10) important in immune cell activation and recruitment. Genetic and antibody-based blockade of TNF-α/TNF-α signaling ameliorated clinical neuropathy, peripheral nerve infiltration, and demyelination, which provides preclinical evidence that the TNF-α axis may be effectively targeted to resolve inflammatory neuropathies.
Assuntos
Doenças Neuroinflamatórias/etiologia , Doenças Neuroinflamatórias/metabolismo , Doenças do Sistema Nervoso Periférico/etiologia , Doenças do Sistema Nervoso Periférico/metabolismo , Poliendocrinopatias Autoimunes/complicações , Fator de Necrose Tumoral alfa/metabolismo , Transferência Adotiva , Animais , Anticorpos Monoclonais/farmacologia , Comunicação Autócrina , Biomarcadores , Citocinas/metabolismo , Modelos Animais de Doenças , Suscetibilidade a Doenças , Perfilação da Expressão Gênica , Mediadores da Inflamação/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Camundongos Transgênicos , Doenças Neuroinflamatórias/tratamento farmacológico , Doenças Neuroinflamatórias/patologia , Comunicação Parácrina , Doenças do Sistema Nervoso Periférico/tratamento farmacológico , Doenças do Sistema Nervoso Periférico/patologia , Poliendocrinopatias Autoimunes/genética , Receptores do Fator de Necrose Tumoral/deficiência , Nervo Isquiático/imunologia , Nervo Isquiático/metabolismo , Nervo Isquiático/patologia , Transdução de Sinais , Fator de Necrose Tumoral alfa/antagonistas & inibidoresRESUMO
Primary brain tumors, such as glioblastoma (GBM), are remarkably resistant to immunotherapy, even though pre-clinical models suggest effectiveness. To understand this better in patients, here we take advantage of our recent neoadjuvant treatment paradigm to map the infiltrating immune cell landscape of GBM and how this is altered following PD-1 checkpoint blockade using high dimensional proteomics, single cell transcriptomics, and quantitative multiplex immunofluorescence. Neoadjuvant PD-1 blockade increases T cell infiltration and the proportion of a progenitor exhausted population of T cells found within the tumor. We identify an early activated and clonally expanded CD8+ T cell cluster whose TCR overlaps with a CD8+ PBMC population. Distinct changes are also observed in conventional type 1 dendritic cells that may facilitate T cell recruitment. Macrophages and monocytes still constitute the majority of infiltrating immune cells, even after anti-PD-1 therapy. Interferon-mediated changes in the myeloid population are consistently observed following PD-1 blockade; these also mediate an increase in chemotactic factors that recruit T cells. However, sustained high expression of T-cell-suppressive checkpoints in these myeloid cells continue to prevent the optimal activation of the tumor infiltrating T cells. Therefore, future immunotherapeutic strategies may need to incorporate the targeting of these cells for clinical benefit.
Assuntos
Neoplasias Encefálicas/tratamento farmacológico , Glioblastoma/terapia , Inibidores de Checkpoint Imunológico/farmacologia , Terapia Neoadjuvante/métodos , Recidiva Local de Neoplasia/terapia , Encéfalo/efeitos dos fármacos , Encéfalo/imunologia , Encéfalo/patologia , Encéfalo/cirurgia , Neoplasias Encefálicas/imunologia , Neoplasias Encefálicas/patologia , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Glioblastoma/imunologia , Glioblastoma/patologia , Humanos , Inibidores de Checkpoint Imunológico/uso terapêutico , Recidiva Local de Neoplasia/imunologia , Recidiva Local de Neoplasia/patologia , Procedimentos Neurocirúrgicos , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/metabolismo , RNA-Seq , Análise de Célula Única , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Evasão Tumoral/efeitos dos fármacos , Macrófagos Associados a Tumor/efeitos dos fármacos , Macrófagos Associados a Tumor/imunologiaRESUMO
Wound healing is a multi-step process to rapidly restore the barrier function. This process is often impaired in diabetic patients resulting in chronic wounds and amputation. We previously found that paradoxical activation of the mitogen-activated protein kinase (MAPK) pathway via topical administration of the BRAF inhibitor vemurafenib accelerates wound healing by activating keratinocyte proliferation and reepithelialization pathways in healthy mice. Herein, we investigated whether this wound healing acceleration also occurs in impaired diabetic wounds and found that topical vemurafenib not only improves wound healing in a murine diabetic wound model but unexpectedly promotes hair follicle regeneration. Hair follicles expressing Sox-9 and K15 surrounded by CD34+ stroma were found in wounds of diabetic and non-diabetic mice, and their formation can be prevented by blocking downstream MEK signaling. Thus, topically applied BRAF inhibitors may accelerate wound healing, and promote the restoration of improved skin architecture in both normal and impaired wounds.