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Semaphorin-Plexin signaling plays a major role in the tumor microenvironment (TME). In particular, Semaphorin 4D (SEMA4D) has been shown to promote tumor growth and metastasis; however, the role of its high-affinity receptor Plexin-B1 (PLXNB1), which is expressed in the TME, is poorly understood. In this study, we directly targeted PLXNB1 in the TME of triple-negative murine breast carcinoma to elucidate its relevance in cancer progression. We found that primary tumor growth, and metastatic dissemination were strongly reduced in PLXNB1-deficient mice, which showed longer survival. PLXNB1-loss in the TME induced a switch in the polarization of tumor-associated macrophages (TAMs) towards a pro-inflammatory M1 phenotype and enhanced the infiltration of CD8+ T lymphocytes both in primary tumors and in distant metastases. Moreover, PLXNB1-deficiency promoted a shift in the Th1/Th2 balance of the T-cell population and an antitumor gene signature, with the up-regulation of Icos, Perforin-1, Stat3 and Ccl5 in tumor infiltrating lymphocytes (TILs). We thus tested the translational relevance of TME re-programming driven by PLXNB1 inactivation for responsiveness to immunotherapy. Indeed, in the absence of PLXNB1, the efficacy of anti-PD-1 blockade was strongly enhanced, efficiently reducing tumor growth and distant metastasis. Consistent with this, pharmacological PLXNB1 blockade by systemic treatment with a specific inhibitor significantly hampered breast cancer growth and enhanced the antitumor activity of the anti-PD1 treatment in a preclinical model. Altogether, these data indicate that PLXNB1 signaling controls the antitumor immune response in the TME and highlight this receptor as a promising immune therapeutic target for metastatic breast cancers.
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Pancreatic cancer is a major cause of demise worldwide. Although key associated genetic changes have been discovered, disease progression is sustained by pathogenic mechanisms that are poorly understood at the molecular level. In particular, the tissue microenvironment of pancreatic adenocarcinoma (PDAC) is usually characterized by high stromal content, scarce recruitment of immune cells, and the presence of neuronal fibers. Semaphorins and their receptors, plexins and neuropilins, comprise a wide family of regulatory signals that control neurons, endothelial and immune cells, embryo development, and normal tissue homeostasis, as well as the microenvironment of human tumors. We focus on the role of these molecular signals in pancreatic cancer progression, as revealed by experimental research and clinical studies, including novel approaches for cancer treatment.
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Adenocarcinoma , Neoplasias Pancreáticas , Semaforinas , Humanos , Neuropilinas , Microambiente Tumoral , Neoplasias PancreáticasRESUMEN
Breast cancer (BC) is the most frequently diagnosed cancer and one of the major causes of cancer death. Despite enormous progress in its management, both from the therapeutic and early diagnosis viewpoints, still around 700,000 patients succumb to the disease each year, worldwide. Late recurrency is the major problem in BC, with many patients developing distant metastases several years after the successful eradication of the primary tumor. This is linked to the phenomenon of metastatic dormancy, a still mysterious trait of the natural history of BC, and of several other types of cancer, by which metastatic cells remain dormant for long periods of time before becoming reactivated to initiate the clinical metastatic disease. In recent years, it has become clear that cancers are best understood if studied as ecosystems in which the impact of non-cancer-cell-autonomous events-dependent on complex interaction between the cancer and its environment, both local and systemic-plays a paramount role, probably as significant as the cell-autonomous alterations occurring in the cancer cell. In adopting this perspective, a metabolic vision of the cancer ecosystem is bound to improve our understanding of the natural history of cancer, across space and time. In BC, many metabolic pathways are coopted into the cancer ecosystem, to serve the anabolic and energy demands of the cancer. Their study is shedding new light on the most critical aspect of BC management, of metastatic dissemination, and that of the related phenomenon of dormancy and fostering the application of the knowledge to the development of metabolic therapies.
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Neoplasias de la Mama , Humanos , Femenino , Neoplasias de la Mama/patología , EcosistemaRESUMEN
The genetic changes sustaining the development of cancers of unknown primary (CUP) remain elusive. The whole-exome genomic profiling of 14 rigorously selected CUP samples did not reveal specific recurring mutation in known driver genes. However, by comparing the mutational landscape of CUPs with that of most other human tumor types, it emerged a consistent enrichment of changes in genes belonging to the axon guidance KEGG pathway. In particular, G842C mutation of PlexinB2 (PlxnB2) was predicted to be activating. Indeed, knocking down the mutated, but not the wild-type, PlxnB2 in CUP stem cells resulted in the impairment of self-renewal and proliferation in culture, as well as tumorigenic capacity in mice. Conversely, the genetic transfer of G842C-PlxnB2 was sufficient to promote CUP stem cell proliferation and tumorigenesis in mice. Notably, G842C-PlxnB2 expression in CUP cells was associated with basal EGFR phosphorylation, and EGFR blockade impaired the viability of CUP cells reliant on the mutated receptor. Moreover, the mutated PlxnB2 elicited CUP cell invasiveness, blocked by EGFR inhibitor treatment. In sum, we found that a novel activating mutation of the axon guidance gene PLXNB2 sustains proliferative autonomy and confers invasive properties to stem cells isolated from cancers of unknown primary, in EGFR-dependent manner.
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Neoplasias Primarias Desconocidas , Células Madre Neoplásicas , Proteínas del Tejido Nervioso , Animales , Humanos , Ratones , Orientación del Axón , Receptores ErbB/genética , Mutación , Recurrencia Local de Neoplasia , Neoplasias Primarias Desconocidas/genética , Proteínas del Tejido Nervioso/genética , Células Madre Neoplásicas/patologíaRESUMEN
In the original publication [...].
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Background: The pancreatic ductal adenocarcinoma (PDAC) microenvironment is highly fibrotic and hypoxic, with poor immune cell infiltration. Recently, we showed that nucleolin (NCL) inhibition normalizes tumour vessels and impairs PDAC growth. Methods: Immunocompetent mouse models of PDAC were treated by the pseudopeptide N6L, which selectively inhibits NCL. Tumour-infiltrating immune cells and changes in the tumour microenvironment were analysed. Results: N6L reduced the proportion of regulatory T cells (Tregs) and myeloid-derived suppressor cells (MDSCs) and increased tumour-infiltrated T lymphocytes (TILs) with an activated phenotype. Low-dose anti-VEGFR2 treatment normalized PDAC vessels but did not modulate the immune suppressive microenvironment. RNAseq analysis of N6L-treated PDAC tumours revealed a reduction of cancer-associated fibroblast (CAF) expansion in vivo and in vitro. Notably, N6L treatment decreased IL-6 levels both in tumour tissues and in serum. Treating mPDAC by an antibody blocking IL-6 reduced the proportion of Tregs and MDSCs and increased the amount of TILs, thus mimicking the effects of N6L. Conclusions: These results demonstrate that NCL inhibition blocks the amplification of lymphoid and myeloid immunosuppressive cells and promotes T cell activation in PDAC through a new mechanism of action dependent on the direct inhibition of the tumoral stroma.
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The formation of a functional blood vessel network relies on the ability of endothelial cells (ECs) to dynamically rearrange their adhesive contacts in response to blood flow and guidance cues, such as vascular endothelial growth factor-A (VEGF-A) and class 3 semaphorins (SEMA3s). Neuropilin 1 (NRP1) is essential for blood vessel development, independently of its ligands VEGF-A and SEMA3, through poorly understood mechanisms. Grounding on unbiased proteomic analysis, we report here that NRP1 acts as an endocytic chaperone primarily for adhesion receptors on the surface of unstimulated ECs. NRP1 localizes at adherens junctions (AJs) where, interacting with VE-cadherin, promotes its basal internalization-dependent turnover and favors vascular permeability initiated by histamine in both cultured ECs and mice. We identify a splice variant of tryptophanyl-tRNA synthetase (mini-WARS) as an unconventionally secreted extracellular inhibitory ligand of NRP1 that, by stabilizing it at the AJs, slows down both VE-cadherin turnover and histamine-elicited endothelial leakage. Thus, our work shows a role for NRP1 as a major regulator of AJs plasticity and reveals how mini-WARS acts as a physiological NRP1 inhibitory ligand in the control of VE-cadherin endocytic turnover and vascular permeability.
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Neuropilina-1 , Triptófano-ARNt Ligasa , Uniones Adherentes/metabolismo , Animales , Antígenos CD , Cadherinas/genética , Permeabilidad Capilar , Células Endoteliales/metabolismo , Histamina , Ligandos , Ratones , Neuropilina-1/genética , Neuropilina-1/metabolismo , Proteómica , Triptófano-ARNt Ligasa/genética , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
The connection with acute myelogenous leukemia (AML) of dihydroorotate dehydrogenase (hDHODH), a key enzyme in pyrimidine biosynthesis, has attracted significant interest from pharma as a possible AML therapeutic target. We recently discovered compound 1, a potent hDHODH inhibitor (IC50 = 1.2 nM), able to induce myeloid differentiation in AML cell lines (THP1) in the low nM range (EC50 = 32.8 nM) superior to brequinar's phase I/II clinical trial (EC50 = 265 nM). Herein, we investigate the 1 drug-like properties observing good metabolic stability and no toxic profile when administered at doses of 10 and 25 mg/kg every 3 days for 5 weeks (Balb/c mice). Moreover, in order to identify a backup compound, we investigate the SAR of this class of compounds. Inside the series, 17 is characterized by higher potency in inducing myeloid differentiation (EC50 = 17.3 nM), strong proapoptotic properties (EC50 = 20.2 nM), and low cytotoxicity toward non-AML cells (EC30(Jurkat) > 100 µM).
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Compuestos de Bifenilo/química , Inhibidores Enzimáticos/química , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/antagonistas & inhibidores , Pirazoles/química , Piridinas/química , Animales , Apoptosis/efectos de los fármacos , Sitios de Unión , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Dihidroorotato Deshidrogenasa , Diseño de Fármacos , Inhibidores Enzimáticos/metabolismo , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/uso terapéutico , Femenino , Semivida , Humanos , Leucemia Mieloide Aguda/tratamiento farmacológico , Masculino , Ratones , Ratones Endogámicos BALB C , Microsomas Hepáticos/metabolismo , Simulación del Acoplamiento Molecular , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/metabolismo , Pirazoles/metabolismo , Pirazoles/farmacología , Pirazoles/uso terapéutico , Piridinas/metabolismo , Piridinas/farmacología , Piridinas/uso terapéutico , Ratas , Ratas Sprague-Dawley , Relación Estructura-ActividadRESUMEN
Aim: Resistance to chemotherapy is a major limiting factor that hamper the effectiveness of anticancer therapies. Doxorubicin is an antineoplastic agent used in the treatment of a wide range of cancers. However, it presents several limitations such as dose-dependent cardiotoxicity, lack of selectivity for tumor cells, and induced cell resistance. Nanotechnology represents a promising strategy to avoid these drawbacks. In this work, new albumin-based nanoparticles were formulated for the intracellular delivery of doxorubicin with the aim to overcome cancer drug resistance. Methods: Glycol chitosan-coated and uncoated albumin nanoparticles were prepared with a tuned coacervation method. The nanoformulations were in vitro characterized evaluating the physicochemical parameters, morphology, and in vitro release kinetics. Biological assays were performed on A2780res and EMT6 cells from human ovarian carcinoma and mouse mammary cell lines resistant for doxorubicin, respectively. Results: Cell viability assays showed that nanoparticles have higher cytotoxicity than the free drug. Moreover, at low concentrations, both doxorubicin-loaded nanoparticles inhibited the cell colony formation in a greater extent than drug solution. In addition, the cell uptake of the different formulations was investigated by confocal microscopy and by the HPLC determination of doxorubicin intracellular accumulation. The nanoparticles were rapidly internalized in greater extent compared to the free drug. Conclusion: Based on these results, doxorubicin-loaded albumin nanoparticles might represent a novel platform to overcome the mechanism of drug resistance in cancer cell lines and improve the drug efficacy.
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Antiretrovirals belonging to the human immunodeficiency virus (HIV) protease inhibitor (HIV-PI) class exert inhibitory effects across several cancer types by targeting tumor cells and its microenvironment. Cervical carcinoma represents a leading cause of morbidity and mortality, particularly in women doubly infected with high-risk human papillomaviruses (HR-HPV) and HIV; of note, combined antiretroviral therapy has reduced cervical carcinoma onset and progression in HIV-infected women. We evaluated the effectiveness and mechanism(s) of action of HIV-PI against cervical carcinoma using a transgenic model of HR-HPV-induced estrogen-promoted cervical carcinoma (HPV16/E2) and found that treatment of mice with ritonavir-boosted HIV-PI, including indinavir, saquinavir, and lopinavir, blocked the growth and promoted the regression of murine cervical carcinoma. This was associated with inhibition of tumor angiogenesis, coupled to downregulation of matrix metalloproteinase (MMP)-9, reduction of VEGF/VEGFR2 complex, and concomitant upregulation of tissue inhibitor of metalloproteinase-3 (TIMP-3). HIV-PI also promoted deposition of collagen IV at the epithelial and vascular basement membrane and normalization of both vessel architecture and functionality. In agreement with this, HIV-PI reduced tumor hypoxia and enhanced the delivery and antitumor activity of conventional chemotherapy. Remarkably, TIMP-3 expression gradually decreased during progression of human dysplastic lesions into cervical carcinoma. This study identified the MMP-9/VEGF proangiogenic axis and its modulation by TIMP-3 as novel HIV-PI targets for the blockade of cervical intraepithelial neoplasia/cervical carcinoma development and invasiveness and the normalization of tumor vessel functions. These findings may lead to new therapeutic indications of HIV-PI to treat cervical carcinoma and other tumors in either HIV-infected or uninfected patients.
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Inhibidores de la Angiogénesis/farmacología , Antineoplásicos/farmacología , Inhibidores de la Proteasa del VIH/farmacología , Inhibidores de la Metaloproteinasa de la Matriz/farmacología , Inhibidor Tisular de Metaloproteinasa-3/agonistas , Animales , Línea Celular Tumoral , Modelos Animales de Enfermedad , Femenino , Papillomavirus Humano 16 , Humanos , Metaloproteinasa 9 de la Matriz/metabolismo , Ratones , Ratones Transgénicos , Neoplasias del Cuello Uterino/tratamiento farmacológico , Neoplasias del Cuello Uterino/etiología , Neoplasias del Cuello Uterino/metabolismo , Neoplasias del Cuello Uterino/patología , Factor A de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Engineering T cells to express chimeric antigen receptors (CARs) specific for antigens on hematological cancers has yielded remarkable clinical responses, but with solid tumors, benefit has been more limited. This may reflect lack of suitable target antigens, immune evasion mechanisms in malignant cells, and/or lack of T cell infiltration into tumors. An alternative approach, to circumvent these problems, is targeting the tumor vasculature rather than the malignant cells directly. CLEC14A is a glycoprotein selectively overexpressed on the vasculature of many solid human cancers and is, therefore, of considerable interest as a target antigen. Here, we generated CARs from 2 CLEC14A-specific antibodies and expressed them in T cells. In vitro studies demonstrated that, when exposed to their target antigen, these engineered T cells proliferate, release IFN-γ, and mediate cytotoxicity. Infusing CAR engineered T cells into healthy mice showed no signs of toxicity, yet these T cells targeted tumor tissue and significantly inhibited tumor growth in 3 mouse models of cancer (Rip-Tag2, mPDAC, and Lewis lung carcinoma). Reduced tumor burden also correlated with significant loss of CLEC14A expression and reduced vascular density within malignant tissues. These data suggest the tumor vasculature can be safely and effectively targeted with CLEC14A-specific CAR T cells, offering a potent and widely applicable therapy for cancer.
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Carcinoma Pulmonar de Lewis/prevención & control , Carcinoma Ductal Pancreático/prevención & control , Moléculas de Adhesión Celular/metabolismo , Inmunoterapia Adoptiva/métodos , Lectinas Tipo C/metabolismo , Neovascularización Patológica/prevención & control , Receptores Quiméricos de Antígenos/inmunología , Linfocitos T/inmunología , Animales , Carcinoma Pulmonar de Lewis/inmunología , Carcinoma Pulmonar de Lewis/metabolismo , Carcinoma Pulmonar de Lewis/patología , Carcinoma Ductal Pancreático/inmunología , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patología , Moléculas de Adhesión Celular/genética , Femenino , Células Endoteliales de la Vena Umbilical Humana , Humanos , Lectinas Tipo C/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Neovascularización Patológica/inmunología , Neovascularización Patológica/metabolismo , Neovascularización Patológica/patología , Neoplasias Pancreáticas/inmunología , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Neoplasias Pancreáticas/prevención & controlRESUMEN
Metastasis formation requires active energy production and is regulated at multiple levels by mitochondrial metabolism. The hyperactive metabolism of cancer cells supports their extreme adaptability and plasticity and facilitates resistance to common anticancer therapies. In spite the potential relevance of a metastasis metabolic control therapy, so far, limited experience is available in this direction. Here, we evaluated the effect of the recently described α-ketoglutarate dehydrogenase (KGDH) inhibitor, (S)-2-[(2,6-dichlorobenzoyl) amino] succinic acid (AA6), in an orthotopic mouse model of breast cancer 4T1 and in other human breast cancer cell lines. In all conditions, AA6 altered Krebs cycle causing intracellular α-ketoglutarate (α-KG) accumulation. Consequently, the activity of the α-KG-dependent epigenetic enzymes, including the DNA demethylation ten-eleven translocation translocation hydroxylases (TETs), was increased. In mice, AA6 injection reduced metastasis formation and increased 5hmC levels in primary tumours. Moreover, in vitro and in vivo treatment with AA6 determined an α-KG accumulation paralleled by an enhanced production of nitric oxide (NO). This epigenetically remodelled metabolic environment efficiently counteracted the initiating steps of tumour invasion inhibiting the epithelial-to-mesenchymal transition (EMT). Mechanistically, AA6 treatment could be linked to upregulation of the NO-sensitive anti-metastatic miRNA 200 family and down-modulation of EMT-associated transcription factor Zeb1 and its CtBP1 cofactor. This scenario led to a decrease of the matrix metalloproteinase 3 (MMP3) and to an impairment of 4T1 aggressiveness. Overall, our data suggest that AA6 determines an α-KG-dependent epigenetic regulation of the TET-miR200-Zeb1/CtBP1-MMP3 axis providing an anti-metastatic effect in a mouse model of breast cancer-associated metastasis.
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Neoplasias de la Mama/complicaciones , Neoplasias de la Mama/metabolismo , Inhibidores Enzimáticos/uso terapéutico , Complejo Cetoglutarato Deshidrogenasa/metabolismo , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/etiología , Neoplasias Pulmonares/metabolismo , Ácido Succínico/uso terapéutico , Animales , Neoplasias de la Mama/tratamiento farmacológico , Adhesión Celular/efectos de los fármacos , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Inhibidores Enzimáticos/química , Femenino , Células Endoteliales de la Vena Umbilical Humana , Humanos , Ratones , Ratones Endogámicos BALB C , Ácido Succínico/químicaRESUMEN
We previously demonstrated that miR-214 is upregulated in malignant melanomas and triple-negative breast tumors and promotes metastatic dissemination by affecting a complex pathway including the anti-metastatic miR-148b. Importantly, tumor dissemination could be reduced by blocking miR-214 function or increasing miR-148b expression or by simultaneous interventions. Based on this evidence, with the intent to explore the role of miR-214 as a target for therapy, we evaluated the capability of new chemically modified anti-miR-214, R97/R98, to inhibit miR-214 coordinated metastatic traits. Relevantly, when melanoma or breast cancer cells were transfected with R97/R98, anti-miR-214 reduced miR-214 expression and impaired transendothelial migration were observed. Noteworthy, when the same cells were injected in the tail vein of mice, cell extravasation and metastatic nodule formation in lungs were strongly reduced. Thus, suggesting that R97/R98 anti-miR-214 oligonucleotides were able to inhibit tumor cell escaping through the endothelium. More importantly, when R97/R98 anti-miR-214 compounds were systemically delivered to mice carrying melanomas or breast or neuroendocrine pancreatic cancers, a reduced number of circulating tumor cells and lung or lymph node metastasis formation were detected. Similar results were also obtained when AAV8-miR-214 sponges were used in neuroendocrine pancreatic tumors. Based on this evidence, we propose miR-214 as a promising target for anti-metastatic therapies.
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Antagomirs/administración & dosificación , MicroARNs/genética , Neoplasias/tratamiento farmacológico , Regulación hacia Arriba/efectos de los fármacos , Animales , Antagomirs/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Carcinoma Neuroendocrino/tratamiento farmacológico , Carcinoma Neuroendocrino/genética , Línea Celular Tumoral , Progresión de la Enfermedad , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Silenciador del Gen , Humanos , Melanoma/tratamiento farmacológico , Melanoma/genética , Ratones , MicroARNs/antagonistas & inhibidores , Metástasis de la Neoplasia/tratamiento farmacológico , Neoplasias/genética , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/genética , Transfección , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Vascular normalizing strategies, aimed at ameliorating blood vessel perfusion and lessening tissue hypoxia, are treatments that may improve the outcome of cancer patients. Secreted class 3 semaphorins (SEMA3), which are thought to directly bind neuropilin (NRP) co-receptors that, in turn, associate with and elicit plexin (PLXN) receptor signaling, are effective normalizing agents of the cancer vasculature. Yet, SEMA3A was also reported to trigger adverse side effects via NRP1. We rationally designed and generated a safe, parenterally deliverable, and NRP1-independent SEMA3A point mutant isoform that, unlike its wild-type counterpart, binds PLXNA4 with nanomolar affinity and has much greater biochemical and biological activities in cultured endothelial cells. In vivo, when parenterally administered in mouse models of pancreatic cancer, the NRP1-independent SEMA3A point mutant successfully normalized the vasculature, inhibited tumor growth, curbed metastatic dissemination, and effectively improved the supply and anticancer activity of chemotherapy. Mutant SEMA3A also inhibited retinal neovascularization in a mouse model of age-related macular degeneration. In summary, mutant SEMA3A is a vascular normalizing agent that can be exploited to treat cancer and, potentially, other diseases characterized by pathological angiogenesis.
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Antineoplásicos/farmacología , Diseño de Fármacos , Proteínas Mutantes/metabolismo , Neuropilina-1/metabolismo , Semaforina-3A/agonistas , Animales , Antineoplásicos/uso terapéutico , Permeabilidad Capilar/efectos de los fármacos , Moléculas de Adhesión Celular/metabolismo , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Neovascularización Coroidal/tratamiento farmacológico , Neovascularización Coroidal/patología , Simulación por Computador , Células Endoteliales/citología , Células Endoteliales/efectos de los fármacos , Ratones Transgénicos , Proteínas Mutantes/química , Neoplasias/irrigación sanguínea , Neoplasias/patología , Proteínas del Tejido Nervioso/metabolismo , Unión Proteica/efectos de los fármacos , Semaforina-3A/químicaRESUMEN
Tumor invasion is a critical first step in the organismic dissemination of cancer cells and the formation of metastasis in distant organs, the most important prognostic factor and the actual cause of death in most of the cancer patients. We report herein that the cell surface protein podoplanin (PDPN), a potent inducer of cancer cell invasion, is conspicuously expressed by the invasive front of squamous cell carcinomas (SCCs) of the cervix in patients and in the transgenic human papillomavirus/estrogen mouse model of cervical cancer. Laser capture microscopy combined with gene expression profiling reveals that the expression of interferon-responsive genes is up-regulated in PDPN-expressing cells at the tumor invasive front, which are exposed to CD45-positive inflammatory cells. Indeed, PDPN expression can be induced in cultured SCC cell lines by single or combined treatments with interferon-γ, transforming growth factor-ß, and/or tumor necrosis factor-α. Notably, shRNA-mediated ablation of either PDPN or STAT1 in A431 SCC cells repressed cancer cell invasion on s.c. transplantation into immunodeficient mice. The results highlight the induction of tumor cell invasion by the inflammatory cytokine-stimulated expression of PDPN in the outermost cell layers of cervical SCC.
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Carcinoma de Células Escamosas/metabolismo , Citocinas/farmacología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Glicoproteínas de Membrana/metabolismo , Invasividad Neoplásica/genética , Neoplasias del Cuello Uterino/metabolismo , Animales , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patología , Línea Celular Tumoral , Femenino , Humanos , Glicoproteínas de Membrana/genética , Ratones , Invasividad Neoplásica/patología , Transcriptoma , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/patologíaRESUMEN
Molecular alterations in genes involved in DNA mismatch repair (MMR) promote cancer initiation and foster tumour progression. Cancers deficient in MMR frequently show favourable prognosis and indolent progression. The functional basis of the clinical outcome of patients with tumours that are deficient in MMR is not clear. Here we genetically inactivate MutL homologue 1 (MLH1) in colorectal, breast and pancreatic mouse cancer cells. The growth of MMR-deficient cells was comparable to their proficient counterparts in vitro and on transplantation in immunocompromised mice. By contrast, MMR-deficient cancer cells grew poorly when transplanted in syngeneic mice. The inactivation of MMR increased the mutational burden and led to dynamic mutational profiles, which resulted in the persistent renewal of neoantigens in vitro and in vivo, whereas MMR-proficient cells exhibited stable mutational load and neoantigen profiles over time. Immune surveillance improved when cancer cells, in which MLH1 had been inactivated, accumulated neoantigens for several generations. When restricted to a clonal population, the dynamic generation of neoantigens driven by MMR further increased immune surveillance. Inactivation of MMR, driven by acquired resistance to the clinical agent temozolomide, increased mutational load, promoted continuous renewal of neoantigens in human colorectal cancers and triggered immune surveillance in mouse models. These results suggest that targeting DNA repair processes can increase the burden of neoantigens in tumour cells; this has the potential to be exploited in therapeutic approaches.
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Antígenos de Neoplasias/genética , Antígenos de Neoplasias/inmunología , Reparación de la Incompatibilidad de ADN/genética , Inmunoterapia/métodos , Neoplasias/inmunología , Neoplasias/patología , Animales , Anticuerpos Antineoplásicos/inmunología , Anticuerpos Antineoplásicos/uso terapéutico , Línea Celular Tumoral , Proliferación Celular/genética , Femenino , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones SCID , Homólogo 1 de la Proteína MutL/deficiencia , Homólogo 1 de la Proteína MutL/genética , Neoplasias/genética , Neoplasias/terapia , Receptores de Antígenos de Linfocitos T/inmunología , Escape del Tumor/genética , Escape del Tumor/inmunologíaRESUMEN
In this study, a novel anticancer reagent based on polyplexes nanoparticles was developed. These nanoparticles are obtained by mixing negatively charged polyelectrolytes with the antitumour cationically-charged pseudopeptide N6L. Using two in vivo experimental tumor pancreatic models based upon PANC-1 and mPDAC cells, we found that the antitumour activity of N6L is significantly raised via its incorporation in polyplexed nanoparticles. Study of the mechanism of action using affinity isolation and si-RNA experiments indicated that N6L-polyplexes are internalized through their interaction with nucleolin. In addition, using a very aggressive model of pancreatic cancer in which gemcitabine, a standard of care for this type of cancer, has a weak effect on tumour growth, we observed that N6L-polyplexes administration has a stronger efficacy than gemcitabine. Biodistribution studies carried out in tumour-bearing mice indicated that N6L-polyplexes localises in tumour tissue, in agreement with its antitumour effect. These results support the idea that N6L nanoparticles could develop into a promising strategy for the treatment of cancer, especially hard-to-treat pancreatic cancers.
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OBJECTIVE: Molecular pathways governing blood vessel patterning are vital to vertebrate development. Because of their ability to counteract proangiogenic factors, antiangiogenic secreted Sema3 (class 3 semaphorins) control embryonic vascular morphogenesis. However, if and how Sema3 may play a role in the control of extraembryonic vascular development is presently unknown. APPROACH AND RESULTS: By characterizing genetically modified mice, here, we show that surprisingly Sema3F acts instead as a selective extraembryonic, but not intraembryonic proangiogenic cue. Both in vivo and in vitro, in visceral yolk sac epithelial cells, Sema3F signals to inhibit the phosphorylation-dependent degradation of Myc, a transcription factor that drives the expression of proangiogenic genes, such as the microRNA cluster 17/92. In Sema3f-null yolk sacs, the transcription of Myc-regulated microRNA 17/92 cluster members is impaired, and the synthesis of Myc and microRNA 17/92 foremost antiangiogenic target Thbs1 (thrombospondin 1) is increased, whereas Vegf (vascular endothelial growth factor) signaling is inhibited in yolk sac endothelial cells. Consistently, exogenous recombinant Sema3F inhibits the phosphorylation-dependent degradation of Myc and the synthesis of Thbs1 in mouse F9 teratocarcinoma stem cells that were in vitro differentiated in visceral yolk sac epithelial cells. Sema3f-/- mice placentas are also highly anemic and abnormally vascularized. CONCLUSIONS: Sema3F functions as an unconventional Sema3 that promotes extraembryonic angiogenesis by inhibiting the Myc-regulated synthesis of Thbs1 in visceral yolk sac epithelial cells.
Asunto(s)
Células Epiteliales/metabolismo , Proteínas de la Membrana/metabolismo , Neovascularización Fisiológica , Proteínas del Tejido Nervioso/metabolismo , Placenta/irrigación sanguínea , Saco Vitelino/irrigación sanguínea , Animales , Línea Celular Tumoral , Células Madre de Carcinoma Embrionario/metabolismo , Células Endoteliales/metabolismo , Femenino , Regulación del Desarrollo de la Expresión Génica , Genotipo , Edad Gestacional , Proteínas de la Membrana/deficiencia , Proteínas de la Membrana/genética , Ratones Endogámicos C57BL , Ratones Noqueados , MicroARNs/genética , MicroARNs/metabolismo , Proteínas del Tejido Nervioso/deficiencia , Proteínas del Tejido Nervioso/genética , Fenotipo , Fosforilación , Embarazo , Proteolisis , Proteínas Proto-Oncogénicas c-myc/metabolismo , Transducción de Señal , Trombospondina 1/genética , Trombospondina 1/metabolismo , Factor A de Crecimiento Endotelial Vascular/genética , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Pancreatic cancer is a highly aggressive tumor, mostly resistant to the standard treatments. Nucleolin is overexpressed in cancers and its inhibition impairs tumor growth. Herein, we showed that nucleolin was overexpressed in human specimens of pancreatic ductal adenocarcinoma (PDAC) and that the overall survival significantly increased in patients with low levels of nucleolin. The nucleolin antagonist N6L strongly impaired the growth of primary tumors and liver metastasis in an orthotopic mouse model of PDAC (mPDAC). Similar antitumor effect of N6L has been observed in a highly angiogenic mouse model of pancreatic neuroendocrine tumor RIP-Tag2. N6L significantly inhibited both human and mouse pancreatic cell proliferation and invasion. Notably, the analysis of tumor vasculature revealed a strong increase of pericyte coverage and vessel perfusion both in mPDAC and RIP-Tag2 tumors, in parallel to an inhibition of tumor hypoxia. Nucleolin inhibition directly affected endothelial cell (EC) activation and changed a proangiogenic signature. Among the vascular activators, nucleolin inhibition significantly decreased angiopoietin-2 (Ang-2) secretion and expression in ECs, in the tumor and in the plasma of mPDAC mice. As a consequence of the observed N6L-induced tumor vessel normalization, pre-treatment with N6L efficiently improved chemotherapeutic drug delivery and increased the antitumor properties of gemcitabine in PDAC mice. In conclusion, nucleolin inhibition is a new anti-pancreatic cancer therapeutic strategy that dually blocks tumor progression and normalizes tumor vasculature, improving the delivery and efficacy of chemotherapeutic drugs. Moreover, we unveiled Ang-2 as a potential target and suitable response biomarker for N6L treatment in pancreatic cancer. Cancer Res; 76(24); 7181-93. ©2016 AACR.
Asunto(s)
Carcinoma Ductal Pancreático/patología , Neovascularización Patológica/patología , Neoplasias Pancreáticas/patología , Péptidos/farmacología , Fosfoproteínas/antagonistas & inhibidores , Proteínas de Unión al ARN/antagonistas & inhibidores , Animales , Proliferación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Humanos , Ratones , Análisis de Matrices Tisulares , NucleolinaRESUMEN
Secreted class 3 semaphorins (Sema3), which signal through holoreceptor complexes that are formed by different subunits, such as neuropilins (Nrps), proteoglycans, and plexins, were initially characterized as fundamental regulators of axon guidance during embryogenesis. Subsequently, Sema3A, Sema3C, Sema3D, and Sema3E were discovered to play crucial roles in cardiovascular development, mainly acting through Nrp1 and Plexin D1, which funnels the signal of multiple Sema3 in vascular endothelial cells. Mechanistically, Sema3 proteins control cardiovascular patterning through the enzymatic GTPase-activating-protein activity of the cytodomain of Plexin D1, which negatively regulates the function of Rap1, a small GTPase that is well-known for its ability to drive vascular morphogenesis and to elicit the conformational activation of integrin adhesion receptors.