RESUMO
Tumor-infiltrating dendritic cells (DCs) assume varied functional states that impact anti-tumor immunity. To delineate the DC states associated with productive anti-tumor T cell immunity, we compared spontaneously regressing and progressing tumors. Tumor-reactive CD8+ T cell responses in Batf3-/- mice lacking type 1 DCs (DC1s) were lost in progressor tumors but preserved in regressor tumors. Transcriptional profiling of intra-tumoral DCs within regressor tumors revealed an activation state of CD11b+ conventional DCs (DC2s) characterized by expression of interferon (IFN)-stimulated genes (ISGs) (ISG+ DCs). ISG+ DC-activated CD8+ T cells ex vivo comparably to DC1. Unlike cross-presenting DC1, ISG+ DCs acquired and presented intact tumor-derived peptide-major histocompatibility complex class I (MHC class I) complexes. Constitutive type I IFN production by regressor tumors drove the ISG+ DC state, and activation of MHC class I-dressed ISG+ DCs by exogenous IFN-ß rescued anti-tumor immunity against progressor tumors in Batf3-/- mice. The ISG+ DC gene signature is detectable in human tumors. Engaging this functional DC state may present an approach for the treatment of human disease.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Células Dendríticas/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Interferon Tipo I/imunologia , Linfócitos do Interstício Tumoral/imunologia , Animais , Antígenos de Neoplasias/imunologia , Antígeno CD11b/imunologia , Apresentação Cruzada , Células Dendríticas/efeitos dos fármacos , Interferon beta/administração & dosagem , Interferon beta/farmacologia , Camundongos , Neoplasias/imunologia , Receptores de Interferon/imunologia , Transdução de Sinais/imunologia , Microambiente Tumoral/imunologiaRESUMO
INTRODUCTION: Tumors are characterized by alterations in the epithelial and stromal compartments, which both contribute to tumor promotion. However, where, when, and how the tumor stroma develops is still poorly understood. We previously demonstrated that DNA damage or telomere malfunction induces an activin A-dependent epithelial stress response that activates cell-intrinsic and cell-extrinsic consequences in mortal, nontumorigenic human mammary epithelial cells (HMECs and vHMECs). Here we show that this epithelial stress response also induces protumorigenic phenotypes in neighboring primary fibroblasts, recapitulating many of the characteristics associated with formation of the tumor stroma (for example, desmoplasia). METHODS: The contribution of extrinsic and intrinsic DNA damage to acquisition of desmoplastic phenotypes was investigated in primary human mammary fibroblasts (HMFs) co-cultured with vHMECs with telomere malfunction (TRF2-vHMEC) or in HMFs directly treated with DNA-damaging agents, respectively. Fibroblast reprogramming was assessed by monitoring increases in levels of selected protumorigenic molecules with quantitative polymerase chain reaction, enzyme-linked immunosorbent assay, and immunocytochemistry. Dependence of the induced phenotypes on activin A was evaluated by addition of exogenous activin A or activin A silencing. In vitro findings were validated in vivo, in preinvasive ductal carcinoma in situ (DCIS) lesions by using immunohistochemistry and telomere-specific fluorescent in situ hybridization. RESULTS: HMFs either cocultured with TRF2-vHMEC or directly exposed to exogenous activin A or PGE2 show increased expression of cytokines and growth factors, deposition of extracellular matrix (ECM) proteins, and a shift toward aerobic glycolysis. In turn, these "activated" fibroblasts secrete factors that promote epithelial cell motility. Interestingly, cell-intrinsic DNA damage in HMFs induces some, but not all, of the molecules induced as a consequence of cell-extrinsic DNA damage. The response to cell-extrinsic DNA damage characterized in vitro is recapitulated in vivo in DCIS lesions, which exhibit telomere loss, heightened DNA damage response, and increased activin A and cyclooxygenase-2 expression. These lesions are surrounded by a stroma characterized by increased expression of α smooth muscle actin and endothelial and immune cell infiltration. CONCLUSIONS: Thus, synergy between stromal and epithelial interactions, even at the initiating stages of carcinogenesis, appears necessary for the acquisition of malignancy and provides novel insights into where, when, and how the tumor stroma develops, allowing new therapeutic strategies.
Assuntos
Ativinas/metabolismo , Ciclo-Oxigenase 2/metabolismo , Dano ao DNA , Glândulas Mamárias Humanas/citologia , Telômero/patologia , Actinas/biossíntese , Ativinas/genética , Ativinas/farmacologia , Carcinoma Intraductal não Infiltrante , Movimento Celular/fisiologia , Transformação Celular Neoplásica , Células Cultivadas , Técnicas de Cocultura , Ciclo-Oxigenase 2/biossíntese , Reparo do DNA , Dinoprostona/farmacologia , Células Epiteliais/citologia , Fibroblastos/citologia , Glicólise/fisiologia , Humanos , Estresse Fisiológico , Homeostase do TelômeroRESUMO
BACKGROUND: For effective tumor elimination, cytotoxic CD8+ T cells must recognize tumor-derived antigens presented on class I major histocompatibility complex (MHC-I). Despite a general association between the expression of immunogenic antigens, typically neoantigens, and response to immunotherapy, the majority of patients lack strong endogenous responses to most putative neoantigens due to mechanisms that are not well understood. Cytotoxic CD8+ T-cell responses are induced by dendritic cells (DCs) cross-presenting tumor-derived peptides on MHC-I. We hypothesized that cross presentation may form an unappreciated source of bias in the induction of cytotoxic T-cell responses. METHODS: We used stable isotope labeling of amino acids combined with immunopeptidomics to distinguish cross-presented from endogenous MHC-I peptides on DCs. To test impacts on T-cell activation, we targeted the model antigen SIINFEKL to specific subcellular compartments in tumor cells, which were used as sources for cross presentation to T cells. In vitro observations were validated using DNA and RNA sequencing data from two cohorts of patients with melanoma undergoing checkpoint blockade therapy. We used a novel quantitative mass spectrometry approach to measure the levels of model antigen on cross-presenting DCs following various means of tumor cell death. RESULTS: DCs exhibited a strong bias for cross-presenting peptides derived from cytoplasmic proteins and against those from plasma membrane proteins, which was confirmed using the model antigen SIINFEKL. In patients with melanoma, the proportion of membrane-derived neoantigens was correlated with reduced survival and failure to respond to therapy. Quantification of cross-presented SIINFEKL revealed that the mode of cell death could overcome DCs' bias against plasma membrane proteins. CONCLUSIONS: Cross presentation of cellular antigens by DCs may impose constraints on the range of peptides available to activate CD8+ T cells that have previously gone unappreciated. The share of neoantigens arising from membrane-derived sources may render some tumors less immunogenic due to inefficient cross presentation. These observations carry important implications for the encounter and intracellular processing of cellular antigens by DCs and merit further clinical studies for their therapeutic potential in stratifying patient populations and design of vaccine-based therapies.Sorry this seems to be the only funciton that works yes I confirm TBF, LES and FC are joint first authors. Please that away the line TFB and FC contributed equally. thanks!!
Assuntos
Apresentação Cruzada , Células Dendríticas , Melanoma , Proteínas de Membrana , Apresentação de Antígeno , Linfócitos T CD8-Positivos/imunologia , Apresentação Cruzada/imunologia , Células Dendríticas/imunologia , Humanos , Melanoma/imunologia , Proteínas de Membrana/imunologia , Peptídeos/metabolismoRESUMO
Checkpoint blockade immunotherapy (CBT) has revolutionized cancer treatment; however, the cellular and molecular factors that govern responsiveness to immunotherapy remain poorly understood. One emerging area of clinical importance is differential responsiveness to CBT across different tissue sites of tumor growth. Each tissue site in the body can contain unique tissue-resident immune cells from both the lymphoid and the myeloid compartment and differences in tissue-specific immune cell composition might predispose tumors in certain tissue sites to be more or less responsive to immunotherapy. Understanding the interplay between tissue-resident and systemic immune responses against tumors will help to determine how to better therapeutically target the immune system to fight cancer. This review summarizes clinical and preclinical investigations of tissue-specific antitumor immune responses and how they influence the tumor immune microenvironment and the efficacy of immunotherapy.
Assuntos
Imunidade , Neoplasias/imunologia , Neoplasias/terapia , Animais , Biomarcadores Tumorais , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Metabolismo Energético , Humanos , Imunidade Inata , Imunoterapia , Terapia de Alvo Molecular , Células Mieloides/imunologia , Células Mieloides/metabolismo , Estadiamento de Neoplasias , Neoplasias/metabolismo , Neoplasias/patologia , Especificidade de Órgãos/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Linfócitos T/patologia , Microambiente Tumoral/imunologiaRESUMO
Developing tissues change shape and tumors initiate spreading through collective cell motility. Conserved mechanisms by which tissues initiate motility into their surroundings are not known. We investigated cytoskeletal regulators during collective invasion by mouse tumor organoids and epithelial Madin-Darby canine kidney (MDCK) acini undergoing branching morphogenesis in collagen. Use of the broad-spectrum formin inhibitor SMIFH2 prevented the formation of migrating cell fronts in both cell types. Focusing on the role of the formin Dia1 in branching morphogenesis, we found that its depletion in MDCK cells does not alter planar cell motility either within the acinus or in two-dimensional scattering assays. However, Dia1 was required to stabilize protrusions extending into the collagen matrix. Live imaging of actin, myosin, and collagen in control acini revealed adhesions that deformed individual collagen fibrils and generated large traction forces, whereas Dia1-depleted acini exhibited unstable adhesions with minimal collagen deformation and lower force generation. This work identifies Dia1 as an essential regulator of tissue shape changes through its role in stabilizing focal adhesions.
Assuntos
Neoplasias da Mama/metabolismo , Proteínas de Transporte/metabolismo , Adesão Celular , Movimento Celular , Células Epiteliais/metabolismo , Glândulas Mamárias Animais/metabolismo , Actinas/metabolismo , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Proteínas de Transporte/genética , Forma Celular , Cães , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Feminino , Proteínas Fetais/metabolismo , Colágenos Fibrilares/metabolismo , Forminas , Fator de Crescimento de Hepatócito/farmacologia , Células Madin Darby de Rim Canino , Glândulas Mamárias Animais/patologia , Camundongos , Morfogênese , Miosinas/metabolismo , Proteínas Nucleares/metabolismo , Transdução de Sinais , Fatores de Tempo , Células Tumorais CultivadasRESUMO
Enzalutamide (MDV3100) is a second generation Androgen Receptor (AR) antagonist with proven efficacy in the treatment of castration resistant prostate cancer (CRPC). The majority of treated patients, however, develop resistance and disease progression and there is a critical need to identify novel targetable pathways mediating resistance. The purpose of this study was to develop and extensively characterize a series of enzalutamide-resistant prostate cancer cell lines. Four genetically distinct AR-positive and AR-pathway dependent prostate cancer cell lines (CWR-R1, LAPC-4, LNCaP, VCaP) were made resistant to enzalutamide by long-term culture (> 6 months) in enzalutamide. Extensive characterization of these lines documented divergent in vitro growth characteristics and AR pathway modulation. Enzalutamide-resistant LNCaP and CWR-R1 cells, but not LAPC-4 and VCAP cells, demonstrated increased castration-resistant and metastatic growth in vivo. Global gene expression analyses between short-term enzalutamide treated vs. enzalutamide-resistant cells identified both AR pathway and non-AR pathway associated changes that were restored upon acquisition of enzalutamide resistance. Further analyses revealed very few common gene expression changes between the four resistant cell lines. Thus, while AR-mediated pathways contribute in part to enzalutamide resistance, an unbiased approach across several cell lines demonstrates a greater contribution toward resistance via pleiotropic, non-AR mediated mechanisms.