RESUMO
Inhibition of the chemokine receptor CXCR4 in combination with blockade of the PD-1/PD-L1 T cell checkpoint induces T cell infiltration and anticancer responses in murine and human pancreatic cancer. Here we elucidate the mechanism by which CXCR4 inhibition affects the tumor immune microenvironment. In human immune cell-based chemotaxis assays, we find that CXCL12-stimulated CXCR4 inhibits the directed migration mediated by CXCR1, CXCR3, CXCR5, CXCR6, and CCR2, respectively, chemokine receptors expressed by all of the immune cell types that participate in an integrated immune response. Inhibiting CXCR4 in an experimental cancer medicine study by 1-wk continuous infusion of the small-molecule inhibitor AMD3100 (plerixafor) induces an integrated immune response that is detected by transcriptional analysis of paired biopsies of metastases from patients with microsatellite stable colorectal and pancreatic cancer. This integrated immune response occurs in three other examples of immune-mediated damage to noninfected tissues: Rejecting renal allografts, melanomas clinically responding to anti-PD1 antibody therapy, and microsatellite instable colorectal cancers. Thus, signaling by CXCR4 causes immune suppression in human pancreatic ductal adenocarcinoma and colorectal cancer by impairing the function of the chemokine receptors that mediate the intratumoral accumulation of immune cells.
Assuntos
Neoplasias Colorretais/metabolismo , Imunidade/imunologia , Pâncreas/metabolismo , Neoplasias Pancreáticas/metabolismo , Receptores CXCR4/efeitos dos fármacos , Receptores CXCR4/metabolismo , Idoso , Benzilaminas , Carcinoma Ductal Pancreático , Quimiocina CXCL12 , Neoplasias Colorretais/patologia , Ciclamos , Feminino , Compostos Heterocíclicos/antagonistas & inibidores , Humanos , Imunoterapia , Masculino , Pessoa de Meia-Idade , Neoplasias Pancreáticas/patologia , Receptores CCR2/metabolismo , Receptores CXCR3/metabolismo , Receptores CXCR5/metabolismo , Receptores CXCR6/metabolismo , Receptores de Interleucina-8A/metabolismo , Transdução de Sinais/efeitos dos fármacos , Microambiente Tumoral/imunologia , Neoplasias PancreáticasRESUMO
During the primary response, the commitment of the CD8(+) T cell to Blimp-1 expression and the terminal differentiation that Blimp-1 induces must be timed so as not to impair the process of clonal expansion. We determined whether the Hippo pathway, which links cell-cell contact to differentiation in other cell lineages, controls Blimp-1 expression. Activating the CD8(+) T cell with antigen and IL-2 causes expression of the core Hippo pathway components, including the pivotal transcriptional cofactor Yap. Contact between activated CD8(+) T cells induces Hippo pathway-mediated Yap degradation and Blimp-1 expression; a Hippo-resistant, stable form of Yap suppresses Blimp-1 expression. Cytotoxic T lymphocyte antigen 4 (CTLA-4) and CD80 comprise the receptor-ligand pair that mediates contact-dependent Hippo pathway activation. In vivo, CD8(+) T cells expressing Hippo resistant-Yap or lacking CTLA-4 have diminished expression of the senescence marker, KLRG1, during a viral infection. The CTLA-4/Hippo pathway/Blimp-1 system may couple terminal differentiation of CD8(+) T cell with the magnitude of clonal expansion.
Assuntos
Linfócitos T CD8-Positivos/enzimologia , Linfócitos T CD8-Positivos/imunologia , Antígeno CTLA-4/metabolismo , Ativação Linfocitária/imunologia , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/imunologia , Fatores de Transcrição/metabolismo , Animais , Antígeno B7-1/metabolismo , Antígeno B7-2/metabolismo , Linfócitos T CD8-Positivos/citologia , Diferenciação Celular/imunologia , Membrana Celular/metabolismo , Ativação Enzimática , Ligantes , Camundongos , Fator 1 de Ligação ao Domínio I Regulador PositivoAssuntos
Fator D do Complemento/química , Fator D do Complemento/deficiência , Fator D do Complemento/genética , Via Alternativa do Complemento , Síndromes de Imunodeficiência/genética , Feminino , Doenças da Deficiência Hereditária de Complemento , Humanos , Masculino , Mutação , Linhagem , Conformação Proteica , Adulto JovemRESUMO
The Ataxia telangiectasia and Rad3-related (ATR) inhibitor ceralasertib in combination with the PD-L1 antibody durvalumab demonstrated encouraging clinical benefit in melanoma and lung cancer patients who progressed on immunotherapy. Here we show that modelling of intermittent ceralasertib treatment in mouse tumor models reveals CD8+ T-cell dependent antitumor activity, which is separate from the effects on tumor cells. Ceralasertib suppresses proliferating CD8+ T-cells on treatment which is rapidly reversed off-treatment. Ceralasertib causes up-regulation of type I interferon (IFNI) pathway in cancer patients and in tumor-bearing mice. IFNI is experimentally found to be a major mediator of antitumor activity of ceralasertib in combination with PD-L1 antibody. Improvement of T-cell function after ceralasertib treatment is linked to changes in myeloid cells in the tumor microenvironment. IFNI also promotes anti-proliferative effects of ceralasertib on tumor cells. Here, we report that broad immunomodulatory changes following intermittent ATR inhibition underpins the clinical therapeutic benefit and indicates its wider impact on antitumor immunity.
Assuntos
Linfócitos T CD8-Positivos , Indóis , Morfolinas , Neoplasias , Pirimidinas , Sulfonamidas , Humanos , Animais , Camundongos , Antígeno B7-H1 , Microambiente Tumoral , Linhagem Celular Tumoral , Imunoterapia , Modelos Animais de Doenças , Proteínas Mutadas de Ataxia TelangiectasiaRESUMO
BACKGROUND: The Regulatory T cell (Treg) lineage is defined by the transcription factor FOXP3, which controls immune-suppressive gene expression profiles. Tregs are often recruited in high frequencies to the tumor microenvironment where they can suppress antitumor immunity. We hypothesized that pharmacological inhibition of FOXP3 by systemically delivered, unformulated constrained ethyl-modified antisense oligonucleotides could modulate the activity of Tregs and augment antitumor immunity providing therapeutic benefit in cancer models and potentially in man. METHODS: We have identified murine Foxp3 antisense oligonucleotides (ASOs) and clinical candidate human FOXP3 ASO AZD8701. Pharmacology and biological effects of FOXP3 inhibitors on Treg function and antitumor immunity were tested in cultured Tregs and mouse syngeneic tumor models. Experiments were controlled by vehicle and non-targeting control ASO groups as well as by use of multiple independent FOXP3 ASOs. Statistical significance of biological effects was evaluated by one or two-way analysis of variance with multiple comparisons. RESULTS: AZD8701 demonstrated a dose-dependent knockdown of FOXP3 in primary Tregs, reduction of suppressive function and efficient target downregulation in humanized mice at clinically relevant doses. Surrogate murine FOXP3 ASO, which efficiently downregulated Foxp3 messenger RNA and protein levels in primary Tregs, reduced Treg suppressive function in immune suppression assays in vitro. FOXP3 ASO promoted more than 70% reduction in FOXP3 levels in Tregs in vitro and in vivo, strongly modulated Treg effector molecules (eg, ICOS, CTLA-4, CD25 and 4-1BB), and augmented CD8+ T cell activation and produced antitumor activity in syngeneic tumor models. The combination of FOXP3 ASOs with immune checkpoint blockade further enhanced antitumor efficacy. CONCLUSIONS: Antisense inhibitors of FOXP3 offer a promising novel cancer immunotherapy approach. AZD8701 is being developed clinically as a first-in-class FOXP3 inhibitor for the treatment of cancer currently in Ph1a/b clinical trial (NCT04504669).
Assuntos
Neoplasias , Oligonucleotídeos Antissenso , Animais , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Humanos , Terapia de Imunossupressão , Imunoterapia , Camundongos , Neoplasias/tratamento farmacológico , Neoplasias/genética , Linfócitos T Reguladores , Microambiente TumoralRESUMO
AZD4625 is a potent, selective, and orally bioavailable inhibitor of oncogenic KRASG12C as demonstrated in cellular assays and in vivo in preclinical cell line-derived and patient-derived xenograft models. In vitro and cellular assays have shown selective binding and inhibition of the KRASG12C mutant isoform, which carries a glycine to cysteine mutation at residue 12, with no binding and inhibition of wild-type RAS or isoforms carrying non-KRASG12C mutations. The pharmacology of AZD4625 shows that it has the potential to provide therapeutic benefit to patients with KRASG12C mutant cancer as either a monotherapy treatment or in combination with other targeted drug agents.
Assuntos
Antineoplásicos , Cisteína , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Glicina/farmacologia , Humanos , Mutação , Isoformas de Proteínas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
PURPOSE: Danvatirsen is a therapeutic antisense oligonucleotide (ASO) that selectively targets STAT3 and has shown clinical activity in two phase I clinical studies. We interrogated the clinical mechanism of action using danvatirsen-treated patient samples and conducted back-translational studies to further elucidate its immunomodulatory mechanism of action. EXPERIMENTAL DESIGN: Paired biopsies and blood samples from danvatirsen-treated patients were evaluated using immunohistochemistry and gene-expression analysis. To gain mechanistic insight, we used mass cytometry, flow cytometry, and immunofluorescence analysis of CT26 tumors treated with a mouse surrogate STAT3 ASO, and human immune cells were treated in vitro with danvatirsen. RESULTS: Within the tumors of treated patients, danvatirsen uptake was observed mainly in cells of the tumor microenvironment (TME). Gene expression analysis comparing baseline and on-treatment tumor samples showed increased expression of proinflammatory genes. In mouse models, STAT3 ASO demonstrated partial tumor growth inhibition and enhanced the antitumor activity when combined with anti-PD-L1. Immune profiling revealed reduced STAT3 protein in immune and stromal cells, and decreased suppressive cytokines correlating with increased proinflammatory macrophages and cytokine production. These changes led to enhanced T-cell abundance and function in combination with anti-PD-L1. CONCLUSIONS: STAT3 ASO treatment reverses a suppressive TME and promotes proinflammatory gene expression changes in patients' tumors and mouse models. Preclinical data provide evidence that ASO-mediated inhibition of STAT3 in the immune compartment is sufficient to remodel the TME and enhance the activity of checkpoint blockade without direct STAT3 inhibition in tumor cells. Collectively, these data provide a rationale for testing this combination in the clinic.
Assuntos
Antineoplásicos Imunológicos/farmacologia , Antígeno B7-H1/antagonistas & inibidores , Neoplasias do Colo/terapia , Neoplasias/terapia , Oligonucleotídeos/farmacologia , Fator de Transcrição STAT3/antagonistas & inibidores , Microambiente Tumoral/imunologia , Ensaios Clínicos Fase I como Assunto , Neoplasias do Colo/imunologia , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Quimioterapia Combinada , Humanos , Imunomodulação , Macrófagos/imunologia , Neoplasias/imunologia , Neoplasias/metabolismo , Neoplasias/patologia , Prognóstico , Fator de Transcrição STAT3/genética , Linfócitos T/imunologia , Células Tumorais CultivadasRESUMO
The induction of adaptive immunity is dependent on the structural organization of LNs, which is in turn governed by the stromal cells that underpin LN architecture. Using a novel fate-mapping mouse model, we trace the developmental origin of mesenchymal LN stromal cells (mLNSCs) to a previously undescribed embryonic fibroblast activation protein-α (FAP)+ progenitor. FAP+ cells of the LN anlagen express lymphotoxin ß receptor (LTßR) and vascular cell adhesion molecule (VCAM), but not intercellular adhesion molecule (ICAM), suggesting they are early mesenchymal lymphoid tissue organizer (mLTo) cells. Clonal labeling shows that FAP+ progenitors locally differentiate into mLNSCs. This process is also coopted in nonlymphoid tissues in response to infection to facilitate the development of tertiary lymphoid structures, thereby mimicking the process of LN ontogeny in response to infection.
Assuntos
Embrião de Mamíferos/imunologia , Gelatinases/imunologia , Linfonodos/imunologia , Proteínas de Membrana/imunologia , Células-Tronco Mesenquimais/imunologia , Modelos Imunológicos , Serina Endopeptidases/imunologia , Animais , Embrião de Mamíferos/citologia , Endopeptidases , Gelatinases/genética , Linfonodos/citologia , Receptor beta de Linfotoxina/genética , Receptor beta de Linfotoxina/imunologia , Proteínas de Membrana/genética , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Transgênicos , Serina Endopeptidases/genética , Molécula 1 de Adesão de Célula Vascular/genética , Molécula 1 de Adesão de Célula Vascular/imunologiaRESUMO
BACKGROUND: The ability to modulate immune-inhibitory pathways using checkpoint blockade antibodies such as αPD-1, αPD-L1, and αCTLA-4 represents a significant breakthrough in cancer therapy in recent years. This has driven interest in identifying small-molecule-immunotherapy combinations to increase the proportion of responses. Murine syngeneic models, which have a functional immune system, represent an essential tool for pre-clinical evaluation of new immunotherapies. However, immune response varies widely between models and the translational relevance of each model is not fully understood, making selection of an appropriate pre-clinical model for drug target validation challenging. METHODS: Using flow cytometry, O-link protein analysis, RT-PCR, and RNAseq we have characterized kinetic changes in immune-cell populations over the course of tumor development in commonly used syngeneic models. RESULTS: This longitudinal profiling of syngeneic models enables pharmacodynamic time point selection within each model, dependent on the immune population of interest. Additionally, we have characterized the changes in immune populations in each of these models after treatment with the combination of α-PD-L1 and α-CTLA-4 antibodies, enabling benchmarking to known immune modulating treatments within each model. CONCLUSIONS: Taken together, this dataset will provide a framework for characterization and enable the selection of the optimal models for immunotherapy combinations and generate potential biomarkers for clinical evaluation in identifying responders and non-responders to immunotherapy combinations.
Assuntos
Antineoplásicos Imunológicos/farmacologia , Descoberta de Drogas , Ensaios de Seleção de Medicamentos Antitumorais , Imunomodulação/efeitos dos fármacos , Animais , Biomarcadores Tumorais , Modelos Animais de Doenças , Sinergismo Farmacológico , Perfilação da Expressão Gênica , Humanos , Imunofenotipagem , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Camundongos , Neoplasias/tratamento farmacológico , Neoplasias/imunologia , Neoplasias/metabolismo , Neoplasias/patologia , Microambiente TumoralRESUMO
Highly polymorphic major histocompatibility complex (MHC) molecules are at the heart of adaptive immune responses, playing crucial roles in many kinds of disease and in vaccination. We report that breadth of peptide presentation and level of cell surface expression of class I molecules are inversely correlated in both chickens and humans. This relationship correlates with protective responses against infectious pathogens including Marek's disease virus leading to lethal tumours in chickens and human immunodeficiency virus infection progressing to AIDS in humans. We propose that differences in peptide binding repertoire define two groups of MHC class I molecules strategically evolved as generalists and specialists for different modes of pathogen resistance. We suggest that differences in cell surface expression level ensure the development of optimal peripheral T cell responses. The inverse relationship of peptide repertoire and expression is evidently a fundamental property of MHC molecules, with ramifications extending beyond immunology and medicine to evolutionary biology and conservation.
Assuntos
Imunidade Adaptativa , Herpesvirus Galináceo 2/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Doença de Marek/imunologia , Peptídeos/imunologia , Síndrome da Imunodeficiência Adquirida/imunologia , Síndrome da Imunodeficiência Adquirida/virologia , Alelos , Sequência de Aminoácidos , Animais , Apresentação de Antígeno , Sítios de Ligação , Linhagem Celular , Galinhas , Cristalografia por Raios X , Regulação da Expressão Gênica , HIV-1/imunologia , Haplótipos , Antígenos de Histocompatibilidade Classe I/química , Antígenos de Histocompatibilidade Classe I/genética , Doença de Marek/virologia , Modelos Moleculares , Dados de Sequência Molecular , Peptídeos/química , Peptídeos/genética , Ligação Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologiaRESUMO
The depletion of tumor stromal cells that are marked by their expression of the membrane protein fibroblast activation protein-α (FAP) overcomes immune suppression and allows an anticancer cell immune response to control tumor growth. In subcutaneous tumors established with immunogenic Lewis lung carcinoma cells expressing ovalbumin (LL2/OVA), the FAP(+) population is comprised of CD45(+) and CD45(-) cells. In the present study, we further characterize the tumoral FAP(+)/CD45(+) population as a minor subpopulation of F4/80(hi)/CCR2(+)/CD206(+) M2 macrophages. Using bone marrow chimeric mice in which the primate diphtheria toxin receptor is restricted either to the FAP(+)/CD45(+) or to the FAP(+)/CD45(-) subset, we demonstrate by conditionally depleting each subset that both independently contribute to the immune-suppressive tumor microenvironment. A basis for the function of the FAP(+)/CD45(+) subset is shown to be the immune inhibitory enzyme, heme oxygenase-1 (HO-1). The FAP(+)/CD45(+) cells are the major tumoral source of HO-1, and an inhibitor of HO-1, Sn mesoporphyrin, causes the same extent of immune-dependent arrest of LL2/OVA tumor growth as does the depletion of these cells. Because this observation of immune suppression by HO-1 expressed by the FAP(+)/CD45(+) stromal cell is replicated in a transplanted model of pancreatic ductal adenocarcinoma, we conclude that pharmacologically targeting this enzyme may improve cancer immunotherapy.
Assuntos
Carcinoma Pulmonar de Lewis/imunologia , Gelatinases/metabolismo , Heme Oxigenase-1/metabolismo , Macrófagos/metabolismo , Proteínas de Membrana/metabolismo , Serina Endopeptidases/metabolismo , Animais , Carcinoma Pulmonar de Lewis/terapia , Endopeptidases , Tolerância Imunológica , Imunoterapia/métodos , Macrófagos/imunologia , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Transplante de NeoplasiasRESUMO
The stromal microenvironment of tumors, which is a mixture of hematopoietic and mesenchymal cells, suppresses immune control of tumor growth. A stromal cell type that was first identified in human cancers expresses fibroblast activation protein-α (FAP). We created a transgenic mouse in which FAP-expressing cells can be ablated. Depletion of FAP-expressing cells, which made up only 2% of all tumor cells in established Lewis lung carcinomas, caused rapid hypoxic necrosis of both cancer and stromal cells in immunogenic tumors by a process involving interferon-γ and tumor necrosis factor-α. Depleting FAP-expressing cells in a subcutaneous model of pancreatic ductal adenocarcinoma also permitted immunological control of growth. Therefore, FAP-expressing cells are a nonredundant, immune-suppressive component of the tumor microenvironment.
Assuntos
Carcinoma Pulmonar de Lewis/imunologia , Carcinoma Ductal Pancreático/imunologia , Gelatinases/metabolismo , Tolerância Imunológica , Proteínas de Membrana/metabolismo , Serina Endopeptidases/metabolismo , Células Estromais/imunologia , Microambiente Tumoral/imunologia , Animais , Antígenos de Neoplasias/imunologia , Vacinas Anticâncer/administração & dosagem , Vacinas Anticâncer/imunologia , Carcinoma Pulmonar de Lewis/patologia , Carcinoma Pulmonar de Lewis/terapia , Carcinoma Ductal Pancreático/patologia , Hipóxia Celular , Linhagem Celular Tumoral , Sobrevivência Celular , Endopeptidases , Interferon gama/imunologia , Interferon gama/metabolismo , Camundongos , Camundongos Transgênicos , Necrose , Transplante de Neoplasias , Células Estromais/metabolismo , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Circulating neutrophils and monocytes form the first line of cellular defense against invading bacteria. Here, we describe a novel and specific mechanism of disabling and eliminating phagocytes by Staphylococcus aureus. Staphopain B (SspB) selectively cleaved CD11b on phagocytes, which rapidly acquired features of cell death. SspB-treated phagocytes expressed phosphatidylserine as well as annexin I and became permeable to propidium iodide, thus demonstrating distinctive features of both apoptosis and necrosis, respectively. The cell death observed was caspase and Syk tyrosine kinase independent, whilst cytochalasin D efficiently inhibited the staphopain-induced neutrophil killing. Neutrophil and monocyte cell death was not affected by integrin clustering ligands (ICAM-1 or fibrin) and was prevented, and even reversed, by IgG. This protective effect was dependent on the Fc fragment, collectively suggesting cooperation of the CD16 receptor and integrin Mac-1 (CD11b/CD18). We conclude that SspB, particularly in the presence of staphylococcal protein A, may reduce the number of functional phagocytes at infection sites, thus facilitating colonization and dissemination of S. aureus.