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1.
Cell ; 178(1): 135-151.e19, 2019 06 27.
Artigo em Inglês | MEDLINE | ID: mdl-31251913

RESUMO

Loss of BRCA1 p220 function often results in basal-like breast cancer (BLBC), but the underlying disease mechanism is largely opaque. In mammary epithelial cells (MECs), BRCA1 interacts with multiple proteins, including NUMB and HES1, to form complexes that participate in interstrand crosslink (ICL) DNA repair and MEC differentiation control. Unrepaired ICL damage results in aberrant transdifferentiation to a mesenchymal state of cultured, human basal-like MECs and to a basal/mesenchymal state in primary mouse luminal MECs. Loss of BRCA1, NUMB, or HES1 or chemically induced ICL damage in primary murine luminal MECs results in persistent DNA damage that triggers luminal to basal/mesenchymal transdifferentiation. In vivo single-cell analysis revealed a time-dependent evolution from normal luminal MECs to luminal progenitor-like tumor cells with basal/mesenchymal transdifferentiation during murine BRCA1 BLBC development. Growing DNA damage accompanied this malignant transformation.


Assuntos
Proteína BRCA1/genética , Neoplasias da Mama/genética , Transdiferenciação Celular/genética , Dano ao DNA/genética , Reparo do DNA/genética , Glândulas Mamárias Animais/patologia , Animais , Proteína BRCA1/metabolismo , Neoplasias da Mama/induzido quimicamente , Neoplasias da Mama/patologia , Diferenciação Celular/genética , Transformação Celular Neoplásica , Modelos Animais de Doenças , Células Epiteliais/metabolismo , Feminino , Células HEK293 , Humanos , Células MCF-7 , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Transgênicos , Proteínas do Tecido Nervoso/metabolismo , Fatores de Transcrição HES-1/metabolismo , Transfecção
2.
Nat Rev Mol Cell Biol ; 20(2): 69-84, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30459476

RESUMO

Epithelial-mesenchymal transition (EMT) is a cellular programme that is known to be crucial for embryogenesis, wound healing and malignant progression. During EMT, cell-cell and cell-extracellular matrix interactions are remodelled, which leads to the detachment of epithelial cells from each other and the underlying basement membrane, and a new transcriptional programme is activated to promote the mesenchymal fate. In the context of neoplasias, EMT confers on cancer cells increased tumour-initiating and metastatic potential and a greater resistance to elimination by several therapeutic regimens. In this Review, we discuss recent findings on the mechanisms and roles of EMT in normal and neoplastic tissues, and the cell-intrinsic signals that sustain expression of this programme. We also highlight how EMT gives rise to a variety of intermediate cell states between the epithelial and the mesenchymal state, which could function as cancer stem cells. In addition, we describe the contributions of the tumour microenvironment in inducing EMT and the effects of EMT on the immunobiology of carcinomas.


Assuntos
Transição Epitelial-Mesenquimal/fisiologia , Neoplasias/patologia , Animais , Matriz Extracelular/patologia , Humanos , Células-Tronco Neoplásicas/patologia , Microambiente Tumoral/fisiologia
3.
Proc Natl Acad Sci U S A ; 116(34): 16971-16980, 2019 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-31375632

RESUMO

Immunotherapy using checkpoint-blocking antibodies against PD-1 has produced impressive results in a wide range of cancers. However, the response remains heterogeneous among patients. We used noninvasive immuno-positron emission tomography (PET), using 89Zr-labeled PEGylated single-domain antibody fragments (nanobodies or VHHs), to explore the dynamics and distribution of intratumoral CD8+ T cells and CD11b+ myeloid cells in response to anti-PD-1 treatment in the MC38 colorectal mouse adenocarcinoma model. Responding and nonresponding tumors showed consistent differences in the distribution of CD8+ and CD11b+ cells. Anti-PD-1 treatment mobilized CD8+ T cells from the tumor periphery to a more central location. Only those tumors fully infiltrated by CD8+ T cells went on to complete resolution. All tumors contained CD11b+ myeloid cells from the outset of treatment, with later recruitment of additional CD11b+ cells. As tumors grew, the distribution of intratumoral CD11b+ cells became more heterogeneous. Shrinkage of tumors in responders correlated with an increase in the CD11b+ population in the center of the tumors. The changes in distribution of CD8+ and CD11b+ cells, as assessed by PET, served as biomarkers to gauge the efficacy of anti-PD-1 treatment. Single-cell RNA sequencing of RNA from intratumoral CD45+ cells showed that CD11b+ cells in responders and nonresponders were markedly different. The responders exhibited a dominant population of macrophages with an M1-like signature, while the CD45+ population in the nonresponders displayed an M2-like transcriptional signature. Thus, by using immuno-PET and single-cell RNA sequencing, we show that anti-PD-1 treatment not only affects interactions of CD8+ T cells with the tumor but also impacts the intratumoral myeloid compartment.


Assuntos
Adenocarcinoma , Antígenos de Neoplasias/imunologia , Linfócitos T CD8-Positivos , Neoplasias Colorretais , Proteínas de Neoplasias/imunologia , Neoplasias Experimentais , Tomografia por Emissão de Pósitrons , Receptor de Morte Celular Programada 1/imunologia , Adenocarcinoma/diagnóstico por imagem , Adenocarcinoma/imunologia , Adenocarcinoma/patologia , Adenocarcinoma/terapia , Animais , Antígeno CD11b/imunologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Linhagem Celular Tumoral , Neoplasias Colorretais/diagnóstico por imagem , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/patologia , Neoplasias Colorretais/terapia , Feminino , Camundongos , Neoplasias Experimentais/diagnóstico por imagem , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/patologia , Neoplasias Experimentais/terapia , Microambiente Tumoral/imunologia
4.
Mol Diagn Ther ; 27(4): 433-444, 2023 07.
Artigo em Inglês | MEDLINE | ID: mdl-37193859

RESUMO

Immune checkpoint blockade therapies have generated efficacious responses in certain tumor types; however, the responses of breast carcinomas have been largely limited. Moreover, the identity of various parameters that can predict responses to immunotherapies, and at the same time, serve as putative biomarkers that can be therapeutically targeted to enhance the effectiveness of immunotherapies for breast cancers, remains to be comprehensively delineated. Activation of epithelial-mesenchymal plasticity in cancer cells, including those of the breast, increases their tumor-initiating potential and promotes their aggressiveness and resistance to multiple treatment regimens. Moreover, the residence of cancer cells in alternating epithelial or mesenchymal plastic phenotypic states can also influence their immuno-modulatory properties and susceptibilities to immune checkpoint blockade therapies. In this current opinion, we discuss the lessons that can be learnt from epithelial-mesenchymal transition to potentiate the efficacy of immunotherapy for breast cancers. We also discuss strategies to sensitize more-mesenchymal cancer cells to anti-tumor immunity and immune checkpoint blockade therapies, with the hope that these can serve as new translational avenues for the treatment of human breast tumors.


Assuntos
Neoplasias da Mama , Humanos , Feminino , Neoplasias da Mama/tratamento farmacológico , Inibidores de Checkpoint Imunológico , Transição Epitelial-Mesenquimal , Biomarcadores , Imunoterapia
5.
J Chem Phys ; 137(4): 045101, 2012 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-22852657

RESUMO

Low energy electrons (LEEs) are produced in copious amounts by the primary radiation used in radiation therapy. The damage caused to the DNA by these secondary electrons in the energy range 5-22 eV has been studied to understand their possible role in radiation induced damage. Electrons are irradiated on dried films of plasmid DNA (pQE30) and analysed using agarose gel electrophoresis. Single strand breaks (SSBs) induced by LEE to supercoiled plasmid DNA show resonance structures at 7, 12, and 15 eV for low doses and 6, 10, and ∼18 eV at saturation doses. The present measurements have an overall agreement with the literature that LEEs resonantly induce SSBs in DNA. Resonant peaks in the SSBs induced by LEEs at 7, 12, and 15 eV with the lowest employed dose in the current study are somewhat different from those reported earlier by two groups. The observed differences are perhaps related to the irradiation dose, conditions and the nature of DNA employed, which is further elaborated.


Assuntos
DNA Bacteriano/química , DNA Super-Helicoidal/química , Elétrons , Plasmídeos/química , Escherichia coli/química
6.
Eur J Cancer ; 169: 106-122, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35550950

RESUMO

INTRODUCTION: Epithelial-to-mesenchymal transition (EMT) is associated with tumor aggressiveness, drug resistance, and poor survival in non-small cell lung cancer (NSCLC) and other cancers. The identification of immune-checkpoint ligands (ICPLs) associated with NSCLCs that display a mesenchymal phenotype (mNSCLC) could help to define subgroups of patients who may benefit from treatment strategies using immunotherapy. METHODS: We evaluated ICPL expression in silico in 130 NSCLC cell lines. In vitro, CRISPR/Cas9-mediated knockdown and lentiviral expression were used to assess the impact of ZEB1 expression on CD70. Gene expression profiles of lung cancer samples from the TCGA (n = 1018) and a dataset from MD Anderson Cancer Center (n = 275) were analyzed. Independent validation was performed by immunohistochemistry and targeted-RNA sequencing in 154 NSCLC whole sections, including a large cohort of pulmonary sarcomatoid carcinomas (SC, n = 55). RESULTS: We uncover that the expression of CD70, a regulatory ligand from the tumor necrosis factor ligand family, is enriched in mNSCLC in vitro models. Mechanistically, the EMT-inducer ZEB1 impacted CD70 expression and fostered increased activity of the CD70 promoter. CD70 overexpression was also evidenced in mNSCLC patient tumor samples and was particularly enriched in SC, a lung cancer subtype associated with poor prognosis. In these tumors, CD70 expression was associated with decreased CD3+ and CD8+ T-cell infiltration and increased T-cell exhaustion markers. CONCLUSION: Our results provide evidence on the pivotal roles of CD70 and ZEB1 in immune escape in mNSCLC, suggesting that EMT might promote cancer progression and metastasis by not only increasing cancer cell plasticity but also reprogramming the immune response in the local tumor microenvironment.


Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Ligante CD27/genética , Ligante CD27/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Imuno-Histoquímica , Ligantes , Neoplasias Pulmonares/patologia , Microambiente Tumoral
7.
Cancer Discov ; 11(5): 1286-1305, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33328216

RESUMO

The epithelial-to-mesenchymal transition, which conveys epithelial (E) carcinoma cells to quasi-mesenchymal (qM) states, enables them to metastasize and acquire resistance to certain treatments. Murine tumors composed of qM mammary carcinoma cells assemble an immunosuppressive tumor microenvironment (TME) and develop resistance to anti-CTLA4 immune-checkpoint blockade (ICB) therapy, unlike their E counterparts. Importantly, minority populations of qM cells within a tumor can cross-protect their more E neighbors from immune attack. The underlying mechanisms of immunosuppression and cross-protection have been unclear. We demonstrate that abrogation of qM carcinoma cell-derived factors (CD73, CSF1, or SPP1) prevents the assembly of an immunosuppressive TME and sensitizes otherwise refractory qM tumors partially or completely to anti-CTLA4 ICB. Most strikingly, mixed tumors in which minority populations of carcinoma cells no longer express CD73 are now sensitized to anti-CTLA4 ICB. Finally, loss of CD73 also enhances the efficacy of anti-CTLA4 ICB during the process of metastatic colonization. SIGNIFICANCE: Minority populations of qM carcinoma cells, which likely reside in human breast carcinomas, can cross-protect their E neighbors from immune attack. Understanding the mechanisms by which qM carcinoma cells resist antitumor immune attack can help identify signaling channels that can be interrupted to potentiate the efficacy of checkpoint blockade immunotherapies.This article is highlighted in the In This Issue feature, p. 995.


Assuntos
Antineoplásicos Imunológicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Antineoplásicos Imunológicos/uso terapêutico , Neoplasias da Mama/imunologia , Modelos Animais de Doenças , Feminino , Humanos , Terapia de Imunossupressão , Camundongos , Camundongos Endogâmicos C57BL , Microambiente Tumoral
8.
Nat Cell Biol ; 20(9): 1084-1097, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-30154549

RESUMO

Lack of insight into mechanisms governing breast cancer metastasis has precluded the development of curative therapies. Metastasis-initiating cancer cells (MICs) are uniquely equipped to establish metastases, causing recurrence and therapeutic resistance. Using various metastasis models, we discovered that certain primary tumours elicit a systemic inflammatory response involving interleukin-1ß (IL-1ß)-expressing innate immune cells that infiltrate distant MIC microenvironments. At the metastatic site, IL-1ß maintains MICs in a ZEB1-positive differentiation state, preventing MICs from generating highly proliferative E-cadherin-positive progeny. Thus, when the inherent plasticity of MICs is impeded, overt metastases cannot be established. Ablation of the pro-inflammatory response or inhibition of the IL-1 receptor relieves the differentiation block and results in metastatic colonization. Among patients with lymph node-positive breast cancer, high primary tumour IL-1ß expression is associated with better overall survival and distant metastasis-free survival. Our data reveal complex interactions that occur between primary tumours and disseminated MICs that could be exploited to improve patient survival.


Assuntos
Neoplasias da Mama/metabolismo , Inflamação/metabolismo , Interleucina-1beta/metabolismo , Neoplasias Pulmonares/metabolismo , Células Mieloides/metabolismo , Microambiente Tumoral , Animais , Anti-Inflamatórios/farmacologia , Antígenos CD/genética , Antígenos CD/metabolismo , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Caderinas/genética , Caderinas/metabolismo , Comunicação Celular , Diferenciação Celular , Linhagem Celular Tumoral , Plasticidade Celular , Proliferação de Células , Feminino , Humanos , Inflamação/imunologia , Inflamação/patologia , Inflamação/prevenção & controle , Interleucina-1beta/genética , Interleucina-1beta/farmacologia , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Metástase Linfática , Camundongos Nus , Células Mieloides/efeitos dos fármacos , Células Mieloides/imunologia , Células Mieloides/patologia , Transdução de Sinais , Fatores de Tempo , Ensaios Antitumorais Modelo de Xenoenxerto , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo
10.
Cancer Res ; 77(15): 3982-3989, 2017 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-28428275

RESUMO

The epithelial-to-mesenchymal transition (EMT) is a cell biological program that confers mesenchymal traits on carcinoma cells and drives their metastatic dissemination. It is unclear, however, whether the activation of EMT in carcinoma cells can change their susceptibility to immune attack. We demonstrate here that mammary tumor cells arising from more epithelial carcinoma cell lines expressed high levels of MHC-I, low levels of PD-L1, and contained within their stroma CD8+ T cells and M1 (antitumor) macrophages. In contrast, tumors arising from more mesenchymal carcinoma cell lines exhibiting EMT markers expressed low levels of MHC-I, high levels of PD-L1, and contained within their stroma regulatory T cells, M2 (protumor) macrophages, and exhausted CD8+ T cells. Moreover, the more mesenchymal carcinoma cells within a tumor retained the ability to protect their more epithelial counterparts from immune attack. Finally, epithelial tumors were more susceptible to elimination by immunotherapy than corresponding mesenchymal tumors. Our results identify immune cells and immunomodulatory markers that can be potentially targeted to enhance the susceptibility of immunosuppressive tumors to various therapeutic regimens. Cancer Res; 77(15); 3982-9. ©2017 AACR.


Assuntos
Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Transição Epitelial-Mesenquimal/imunologia , Evasão Tumoral/imunologia , Animais , Western Blotting , Modelos Animais de Doenças , Feminino , Citometria de Fluxo , Imunofluorescência , Camundongos , Camundongos Endogâmicos C57BL
11.
J Exp Med ; 214(8): 2243-2255, 2017 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-28666979

RESUMO

Immunotherapy using checkpoint-blocking antibodies against targets such as CTLA-4 and PD-1 can cure melanoma and non-small cell lung cancer in a subset of patients. The presence of CD8 T cells in the tumor correlates with improved survival. We show that immuno-positron emission tomography (immuno-PET) can visualize tumors by detecting infiltrating lymphocytes and, through longitudinal observation of individual animals, distinguish responding tumors from those that do not respond to therapy. We used 89Zr-labeled PEGylated single-domain antibody fragments (VHHs) specific for CD8 to track the presence of intratumoral CD8+ T cells in the immunotherapy-susceptible B16 melanoma model in response to checkpoint blockade. A 89Zr-labeled PEGylated anti-CD8 VHH detected thymus and secondary lymphoid structures as well as intratumoral CD8 T cells. Animals that responded to CTLA-4 therapy showed a homogeneous distribution of the anti-CD8 PET signal throughout the tumor, whereas more heterogeneous infiltration of CD8 T cells correlated with faster tumor growth and worse responses. To support the validity of these observations, we used two different transplantable breast cancer models, yielding results that conformed with predictions based on the antimelanoma response. It may thus be possible to use immuno-PET and monitor antitumor immune responses as a prognostic tool to predict patient responses to checkpoint therapies.


Assuntos
Linfócitos T CD8-Positivos/fisiologia , Antígeno CTLA-4/antagonistas & inibidores , Neoplasias Mamárias Experimentais/terapia , Animais , Antígeno CTLA-4/fisiologia , Feminino , Fragmentos de Imunoglobulinas/imunologia , Fragmentos de Imunoglobulinas/uso terapêutico , Imunoterapia/métodos , Neoplasias Mamárias Experimentais/diagnóstico por imagem , Neoplasias Mamárias Experimentais/imunologia , Camundongos , Transplante de Neoplasias , Tomografia por Emissão de Pósitrons/métodos , Resultado do Tratamento
13.
Cancer Res ; 76(23): 6778-6784, 2016 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-27530323

RESUMO

The emergence of metastatic disease in cancer patients many years or decades after initial successful treatment of primary tumors is well documented but poorly understood at the molecular level. Recent studies have begun exploring the cell-intrinsic programs, causing disseminated tumor cells to enter latency and the cellular signals in the surrounding nonpermissive tissue microenvironment that maintain the latent state. However, relatively little is known about the mechanisms that enable disseminated tumor cells to escape cancer dormancy or tumor latency. We describe here an in vivo model of solitary metastatic latency in the lung parenchyma. The induction of a localized inflammation in the lungs, initiated by lipopolysaccharide treatment, triggers the awakening of these cells, which develop into macroscopic metastases. The escape from latency is dependent on the expression of Zeb1, a key regulator of the epithelial-to-mesenchymal transition (EMT). Furthermore, activation of the EMT program on its own, as orchestrated by Zeb1, is sufficient to incite metastatic outgrowth by causing carcinoma cells to enter stably into a metastasis-initiating cell state. Cancer Res; 76(23); 6778-84. ©2016 AACR.


Assuntos
Inflamação/genética , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética , Animais , Linhagem Celular Tumoral , Proliferação de Células , Humanos , Camundongos , Metástase Neoplásica , Transdução de Sinais , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo
14.
Front Immunol ; 5: 54, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24611064

RESUMO

Cleavage of the Notch receptor via a γ-secretase, results in the release of the active intra-cellular domain of Notch that migrates to the nucleus and interacts with RBP-Jκ, resulting in the activation of downstream target genes. This canonical Notch signaling pathway has been documented to influence T cell development and function. However, the mechanistic details underlying this process remain obscure. In addition to RBP-Jκ, the intra-cellular domain of Notch also interacts with other proteins in the cytoplasm and nucleus, giving rise to the possibility of an alternate, RBP-Jκ independent Notch pathway. However, the contribution of such RBP-Jκ independent, "non-canonical" Notch signaling in regulating peripheral T cell responses is unknown. In this report, we specifically demonstrate the requirement of Notch1 for regulating signal strength and signaling events distal to the T cell receptor in peripheral CD4(+) T cells. By using mice with a conditional deletion in Notch1 or RBP-Jκ, we show that Notch1 regulates activation and proliferation of CD4(+) T cells independently of RBP-Jκ. Furthermore, differentiation to TH1 and iTreg lineages although Notch dependent, is RBP-Jκ independent. Our striking observations demonstrate that many of the cell-intrinsic functions of Notch occur independently of RBP-Jκ. Such non-canonical regulation of these processes likely occurs through NF-κ B. This reveals a previously unknown, novel role of non-canonical Notch signaling in regulating peripheral T cell responses.

15.
J Exp Med ; 210(7): 1311-29, 2013 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-23733784

RESUMO

Severe aplastic anemia (AA) is a bone marrow (BM) failure (BMF) disease frequently caused by aberrant immune destruction of blood progenitors. Although a Th1-mediated pathology is well described for AA, molecular mechanisms driving disease progression remain ill defined. The NOTCH signaling pathway mediates Th1 cell differentiation in the presence of polarizing cytokines, an action requiring enzymatic processing of NOTCH receptors by γ-secretase. Using a mouse model of AA, we demonstrate that expression of both intracellular NOTCH1(IC) and T-BET, a key transcription factor regulating Th1 cell differentiation, was increased in spleen and BM-infiltrating T cells during active disease. Conditionally deleting Notch1 or administering γ-secretase inhibitors (GSIs) in vivo attenuated disease and rescued mice from lethal BMF. In peripheral T cells from patients with untreated AA, NOTCH1(IC) was significantly elevated and bound to the TBX21 promoter, showing NOTCH1 directly regulates the gene encoding T-BET. Treating patient cells with GSIs in vitro lowered NOTCH1(IC) levels, decreased NOTCH1 detectable at the TBX21 promoter, and decreased T-BET expression, indicating that NOTCH1 signaling is responsive to GSIs during active disease. Collectively, these results identify NOTCH signaling as a primary driver of Th1-mediated pathogenesis in AA and may represent a novel target for therapeutic intervention.


Assuntos
Anemia Aplástica/imunologia , Anemia Aplástica/terapia , Medula Óssea/imunologia , Medula Óssea/patologia , Receptor Notch1/fisiologia , Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Anemia Aplástica/patologia , Animais , Modelos Animais de Doenças , Inibidores Enzimáticos/uso terapêutico , Feminino , Transplante de Células-Tronco Hematopoéticas , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Receptor Notch1/deficiência , Receptor Notch1/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia
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