RESUMO
Externalized phosphatidylserine (PS) is a phospholipid and a selective marker of the tumor microenvironment (TME). It is exposed on the outer leaflet of the plasma membrane of tumor-associated endothelial cells, apoptotic tumor cells, and some viable tumor cells, where it functions in part to suppress immune responses by binding to PS receptors expressed on tumor-infiltrating myeloid cells. PS has been targeted with antibodies, such as bavituximab, that bind the phospholipid via a cofactor, ß2-glycoprotein 1 (ß2GP1); these antibodies showed excellent specificity for tumor vasculature and induce an immune stimulatory environment. We have advanced this concept by developing the next generation of PS targeting agent, a fusion protein (betabody) constructed by linking PS-binding domain V of ß2GP1 to the Fc of an IgG2a. Betabodies bind to externalized PS with high affinity (â¼1 nM), without the requirement of a co-factor and localize robustly to the TME. We demonstrate that betabodies are a direct PS-targeting agent that have the potential to be used as anti-tumor therapy, drug delivery vehicles, and tools for imaging the TME.
RESUMO
Metastatic cancer cells adapt to thrive in secondary organs. To investigate metastatic adaptation, we performed transcriptomic analysis of metastatic and non-metastatic murine breast cancer cells. We found that pleiotrophin (PTN), a neurotrophic cytokine, is a metastasis-associated factor that is expressed highly by aggressive breast cancers. Moreover, elevated PTN in plasma correlated significantly with metastasis and reduced survival of breast cancer patients. Mechanistically, we find that PTN activates NF-κB in cancer cells leading to altered cytokine production, subsequent neutrophil recruitment, and an immune suppressive microenvironment. Consequently, inhibition of PTN, pharmacologically or genetically, reduces the accumulation of tumor-associated neutrophils and reverts local immune suppression, resulting in increased T cell activation and attenuated metastasis. Furthermore, inhibition of PTN significantly enhanced the efficacy of immune checkpoint blockade and chemotherapy in reducing metastatic burden in mice. These findings establish PTN as a previously unrecognized driver of a prometastatic immune niche and thus represents a promising therapeutic target for the treatment of metastatic breast cancer.
Assuntos
Proteínas de Transporte , Neoplasias , Camundongos , Animais , Citocinas/metabolismo , NF-kappa B , Microambiente TumoralRESUMO
INTRODUCTION: Macrophage phenotype in the tumor microenvironment correlates with prognosis in NSCLC. Immunosuppressive macrophages promote tumor progression, whereas proinflammatory macrophages may drive an antitumor immune response. How individual NSCLCs affect macrophage phenotype is a major knowledge gap. METHODS: To systematically study the impact of lung cancer cells on macrophage phenotypes, we developed an in vitro co-culture model that consisted of molecularly and clinically annotated patient-derived NSCLC lines, human cancer-associated fibroblasts, and murine macrophages. Induced macrophage phenotype was studied through quantitative real-time polymerase chain reaction and validated in vivo using NSCLC xenografts through quantitative immunohistochemistry and clinically with The Cancer Genome Atlas (TCGA)-"matched" patient tumors. RESULTS: A total of 72 NSCLC cell lines were studied. The most frequent highly induced macrophage-related gene was Arginase-1, reflecting an immunosuppressive M2-like phenotype. This was independent of multiple clinicopathologic factors, which also did not affect M2:M1 ratios in matched TCGA samples. In vivo, xenograft tumors established from high Arginase-1-inducing lines (Arghi) had a significantly elevated density of Arg1+ macrophages. Matched TCGA clinical samples to Arghi NSCLC lines had a significantly higher ratio of M2:M1 macrophages (p = 0.0361). CONCLUSIONS: In our in vitro co-culture model, a large panel of patient-derived NSCLC lines most frequently induced high-expression Arginase-1 in co-cultured mouse macrophages, independent of major clinicopathologic and oncogenotype-related factors. Arghi cluster-matched TCGA tumors contained a higher ratio of M2:M1 macrophages. Thus, this in vitro model reproducibly characterizes how individual NSCLC modulates macrophage phenotype, correlates with macrophage polarization in clinical samples, and can serve as an accessible platform for further investigation of macrophage-specific therapeutic strategies.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Animais , Arginase/genética , Arginase/metabolismo , Arginase/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Técnicas de Cocultura , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Macrófagos/metabolismo , Camundongos , Fenótipo , Microambiente TumoralRESUMO
Pancreatic cancer is the third leading cause of cancer-related deaths in the United States with a 5-year survival less than 5%. Resistance to standard therapy and limited response to immune checkpoint blockade due to the immunosuppressive and stroma-rich microenvironment remain major challenges in the treatment of pancreatic cancer. A key cellular program involved in therapy resistance is epithelial plasticity, which is also associated with invasion, metastasis, and evasion of immune surveillance. The receptor tyrosine kinase AXL is a key driver of tumor cell epithelial plasticity. High expression and activity of AXL is associated with poor prognosis, metastasis, and therapy resistance in multiple types of cancer including pancreatic. Here, we show that an AXL inhibitor (TP-0903), has antitumor and therapy sensitizing effects in preclinical models of pancreatic ductal adenocarcinoma (PDA). We demonstrate that TP-0903 as a single agent or in combination with gemcitabine and/or anti-programmed cell death protein 1 (PD1) antibody has anti-metastatic and anti-tumor effects in PDA tumor bearing mice, leading to increased survival. In addition, gene expression analysis of tumors demonstrated upregulation of pro-inflammatory and immune activation genes in tumors from TP-0903-treated animals compared with the vehicle, indicating pharmacologic inhibition of AXL activation leads to an immunostimulatory microenvironment. This effect was augmented when TP-0903 was combined with gemcitabine and anti-PD1 antibody. These results provide clear rationale for evaluating TP-0903 in the treatment of pancreatic cancer.
Assuntos
Imunoterapia/métodos , Neoplasias Pancreáticas/tratamento farmacológico , Proteínas Proto-Oncogênicas/uso terapêutico , Pirimidinas/uso terapêutico , Receptores Proteína Tirosina Quinases/uso terapêutico , Sulfonamidas/uso terapêutico , Animais , Linhagem Celular Tumoral , Humanos , Camundongos , Metástase Neoplásica , Neoplasias Pancreáticas/mortalidade , Neoplasias Pancreáticas/patologia , Proteínas Proto-Oncogênicas/farmacologia , Pirimidinas/farmacologia , Receptores Proteína Tirosina Quinases/farmacologia , Sulfonamidas/farmacologia , Análise de Sobrevida , Microambiente Tumoral , Receptor Tirosina Quinase AxlRESUMO
Antimalarial antibody responses are essential for mediating the clearance of Plasmodium parasite-infected RBCs from infected hosts. However, the rapid appearance of large numbers of plasmablasts in Plasmodium-infected hosts can suppress the development and function of durable humoral immunity. Here, we identify that the formation of plasmablast populations in Plasmodium-infected mice is mechanistically linked to both hemolysis-induced exposure of phosphatidylserine on damaged RBCs and inflammatory cues. We also show that virus and Trypanosoma infections known to trigger hemolytic anemia and high-grade inflammation also induce exuberant plasmablast responses. The induction of hemolysis or administration of RBC membrane ghosts increases plasmablast differentiation. The phosphatidylserine receptor Axl is critical for optimal plasmablast formation, and blocking phosphatidylserine limits plasmablast expansions and reduces Plasmodium parasite burden in vivo. Our findings support that strategies aimed at modulating polyclonal B cell activation and phosphatidylserine exposure may improve immune responses against Plasmodium parasites and potentially other infectious diseases that are associated with anemia.
Assuntos
Diferenciação Celular/imunologia , Hemólise/imunologia , Fosfatidilserinas/imunologia , Plasmócitos/imunologia , Animais , Anticorpos Antiprotozoários/imunologia , Antimaláricos/imunologia , Linfócitos B/imunologia , Linfócitos B/parasitologia , Células Cultivadas , Eritrócitos/imunologia , Eritrócitos/parasitologia , Humanos , Imunidade Humoral/imunologia , Malária/imunologia , Malária/parasitologia , Camundongos , Camundongos Endogâmicos C57BL , Plasmócitos/parasitologia , Plasmodium yoelii/imunologiaRESUMO
Pancreatic ductal adenocarcinoma (PDA), a leading cause of cancer-related death in the United States, has a high metastatic rate, and is associated with persistent immune suppression. AXL, a member of the TAM (TYRO3, AXL, MERTK) receptor tyrosine kinase family, is a driver of metastasis and immune suppression in multiple cancer types. Here we use single-cell RNA-sequencing to reveal that AXL is expressed highly in tumor cells that have a mesenchymal-like phenotype and that AXL expression correlates with classic markers of epithelial-to-mesenchymal transition. We demonstrate that AXL deficiency extends survival, reduces primary and metastatic burden, and enhances sensitivity to gemcitabine in an autochthonous model of PDA. PDA in AXL-deficient mice displayed a more differentiated histology, higher nucleoside transporter expression, and a more active immune microenvironment compared with PDA in wild-type mice. Finally, we demonstrate that AXL-positive poorly differentiated tumor cells are critical for PDA progression and metastasis, emphasizing the potential of AXL as a therapeutic target in PDA. IMPLICATIONS: These studies implicate AXL as a marker of undifferentiated PDA cells and a target for therapy.
Assuntos
Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Plasticidade Celular/fisiologia , Metástase Neoplásica/patologia , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Proteínas Proto-Oncogênicas/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Animais , Biomarcadores Tumorais/metabolismo , Carcinoma Ductal Pancreático/tratamento farmacológico , Linhagem Celular Tumoral , Plasticidade Celular/efeitos dos fármacos , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacologia , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/fisiologia , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias Pancreáticas/tratamento farmacológico , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/fisiologia , Gencitabina , Receptor Tirosina Quinase AxlRESUMO
Cancer evolves through a multistep process that occurs by the temporal accumulation of genetic mutations. Tumor-derived exosomes are emerging contributors to tumorigenesis. To understand how exosomes might contribute to cell transformation, we utilized the classic two-step NIH/3T3 cell transformation assay and observed that exosomes isolated from pancreatic cancer cells, but not normal human cells, can initiate malignant cell transformation and these transformed cells formed tumors in vivo. However, cancer cell exosomes are unable to transform cells alone or to act as a promoter of cell transformation. Utilizing proteomics and exome sequencing, we discovered cancer cell exosomes act as an initiator by inducing random mutations in recipient cells. Cells from the pool of randomly mutated cells are driven to transformation by a classic promoter resulting in foci, each of which encode a unique genetic profile. Our studies describe a novel molecular understanding of how cancer cell exosomes contribute to cell transformation. Editorial note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that major issues remain unresolved (see decision letter).
Assuntos
Transformação Celular Neoplásica/patologia , Exossomos/metabolismo , Neoplasias Pancreáticas/patologia , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Exossomos/química , Genômica , Humanos , Camundongos , Células NIH 3T3 , Transplante de Neoplasias , ProteômicaRESUMO
Activation of the receptor tyrosine kinase Axl is associated with poor outcomes in pancreatic cancer (PDAC), where it coordinately mediates immune evasion and drug resistance. Here, we demonstrate that the selective Axl kinase inhibitor BGB324 targets the tumor-immune interface to blunt the aggressive traits of PDAC cells in vitro and enhance gemcitibine efficacy in vivo Axl signaling stimulates the TBK1-NFκB pathway and innate immune suppression in the tumor microenvironment. In tumor cells, BGB324 treatment drove epithelial differentiation, expression of nucleoside transporters affecting gemcitabine response, and an immune stimulatory microenvironment. Our results establish a preclinical mechanistic rationale for the clinical development of Axl inhibitors to improve the treatment of PDAC patients.Significance: These results establish a preclinical mechanistic rationale for the clinical development of AXL inhibitors to improve the treatment of PDAC patients. Cancer Res; 78(1); 246-55. ©2017 AACR.
Assuntos
Benzocicloeptenos/farmacologia , Carcinoma Ductal Pancreático/tratamento farmacológico , Neoplasias Pancreáticas/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Triazóis/farmacologia , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Benzocicloeptenos/administração & dosagem , Carcinoma Ductal Pancreático/imunologia , Linhagem Celular Tumoral , Desoxicitidina/administração & dosagem , Desoxicitidina/análogos & derivados , Feminino , Humanos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Terapia de Alvo Molecular , Neoplasias Pancreáticas/imunologia , Triazóis/administração & dosagem , Ensaios Antitumorais Modelo de Xenoenxerto , Gencitabina , Receptor Tirosina Quinase AxlRESUMO
Pancreatic carcinoma ranks among the most lethal of human cancers. Besides late detection, other factors contribute to its lethality, including a high degree of chemoresistance, invasion, and distant metastases. Currently, the mainstay of therapy involves resection of local disease in a minority of patients (Whipple procedure) and systemic gemcitabine. While systemic chemotherapy has some benefit, even with optimal treatment, the five year survival after diagnosis is dismal. Thus, treatment of pancreatic carcinoma remains a tremendous unmet need.The organometallic compound tris DBA palladium is a potent inhibitor of N-myristoyltransferase 1 (NMT1), an enzyme that catalyzes the transfer of myristate to protein substrates. This compound is highly effective in vivo against murine models of melanoma with both mutant and wild type b-RAF genotypes. Based upon the signaling similarities between melanoma and pancreatic carcinoma, we evaluated the efficacy of tris DBA palladium in vitro and in vivo against pancreatic carcinoma. We found that tris DBA palladium decreased proliferation and colony formation of pancreatic cancer cells in vitro. In an orthotopic mouse model, tris DBA palladium was highly active in inhibiting growth, ascites production, and distant metastases in vivo. Furthermore, tris DBA palladium impaired chemotaxis and inhibited cilia formation in Pan02 cells in a NMT1-dependent manner. We propose that NMT1 is a novel regulator of cilia formation and tris DBA palladium a novel inhibitor of cilia formation and metastasis in pancreatic cancer. Thus, further evaluation of tris DBA palladium for the treatment of pancreatic cancer is warranted.
Assuntos
Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Compostos Organometálicos/farmacologia , Neoplasias Pancreáticas/patologia , Animais , Linhagem Celular Tumoral , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Metástase Neoplásica , Compostos Organometálicos/uso terapêutico , Neoplasias Pancreáticas/tratamento farmacológico , Resultado do Tratamento , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Phosphatidylserine (PS) is an anionic phospholipid maintained on the inner-leaflet of the cell membrane and is externalized in malignant cells. We previously launched a careful unbiased selection targeting biomolecules (e.g. protein, lipid or carbohydrate) distinct to cancer cells by exploiting HCC4017 lung cancer and HBEC30KT normal epithelial cells derived from the same patient, identifying HCC4017 specific peptide-peptoid hybrid PPS1. In this current study, we identified PS as the target of PPS1. We validated direct PPS1 binding to PS using ELISA-like assays, lipid dot blot and liposome based binding assays. In addition, PPS1 recognized other negatively charged and cancer specific lipids such as phosphatidic acid, phosphatidylinositol and phosphatidylglycerol. PPS1 did not bind to neutral lipids such as phosphatidylethanolamine found in cancer and phosphatidylcholine and sphingomyelin found in normal cells. Further we found that the dimeric version of PPS1 (PPS1D1) displayed strong cytotoxicity towards lung cancer cell lines that externalize PS, but not normal cells. PPS1D1 showed potent single agent anti-tumor activity and enhanced the efficacy of docetaxel in mice bearing H460 lung cancer xenografts. Since PS and anionic phospholipid externalization is common across many cancer types, PPS1 may be an alternative to overcome limitations of protein targeted agents.
Assuntos
Neoplasias Pulmonares/tratamento farmacológico , Oligopeptídeos/farmacologia , Fosfatidilserinas/antagonistas & inibidores , Ensaios Antitumorais Modelo de Xenoenxerto , Animais , Linhagem Celular , Linhagem Celular Tumoral , Feminino , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Lipídeos de Membrana/antagonistas & inibidores , Lipídeos de Membrana/metabolismo , Camundongos Endogâmicos NOD , Camundongos SCID , Terapia de Alvo Molecular , Oligopeptídeos/química , Oligopeptídeos/metabolismo , Peptídeos/química , Peptoides/química , Fosfatidilserinas/metabolismo , Ligação Proteica , Carga Tumoral/efeitos dos fármacosRESUMO
Elevated oxidative stress is an aberration seen in many solid tumors, and exploiting this biochemical difference has the potential to enhance the efficacy of anticancer agents. Homeostasis of reactive oxygen species (ROS) is important for normal cell function, but excessive production of ROS can result in cellular toxicity, and therefore ROS levels must be balanced finely. Here, we highlight the relationship between the extracellular matrix and ROS production by reporting a novel function of the matricellular protein Fibulin-5 (Fbln5). We used genetically engineered mouse models of pancreatic ductal adenocarcinoma (PDAC) and found that mutation of the integrin-binding domain of Fbln5 led to decreased tumor growth, increased survival, and enhanced chemoresponse to standard PDAC therapies. Through mechanistic investigations, we found that improved survival was due to increased levels of oxidative stress in Fbln5-mutant tumors. Furthermore, loss of the Fbln5-integrin interaction augmented fibronectin signaling, driving integrin-induced ROS production in a 5-lipooxygenase-dependent manner. These data indicate that Fbln5 promotes PDAC progression by functioning as a molecular rheostat that modulates cell-ECM interactions to reduce ROS production, and thus tip the balance in favor of tumor cell survival and treatment-refractory disease.
Assuntos
Carcinoma Ductal Pancreático/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Neoplasias Pancreáticas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas Recombinantes/metabolismo , Animais , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Matriz Extracelular/metabolismo , Matriz Extracelular/patologia , Proteínas da Matriz Extracelular/biossíntese , Humanos , Camundongos , Camundongos Transgênicos , Estresse Oxidativo/fisiologia , Neoplasias Pancreáticas/patologia , Proteínas Recombinantes/biossíntese , Microambiente Tumoral/fisiologiaRESUMO
Repurposing "old" drugs can facilitate rapid clinical translation but necessitates novel mechanistic insight. Warfarin, a vitamin K "antagonist" used clinically for the prevention of thrombosis for more than 50 years, has been shown to have anticancer effects. We hypothesized that the molecular mechanism underlying its antitumor activity is unrelated to its effect on coagulation, but is due to inhibition of the Axl receptor tyrosine kinase on tumor cells. Activation of Axl by its ligand Gas6, a vitamin K-dependent protein, is inhibited at doses of warfarin that do not affect coagulation. Here, we show that inhibiting Gas6-dependent Axl activation with low-dose warfarin, or with other tumor-specific Axl-targeting agents, blocks the progression and spread of pancreatic cancer. Warfarin also inhibited Axl-dependent tumor cell migration, invasiveness, and proliferation while increasing apoptosis and sensitivity to chemotherapy. We conclude that Gas6-induced Axl signaling is a critical driver of pancreatic cancer progression and its inhibition with low-dose warfarin or other Axl-targeting agents may improve outcome in patients with Axl-expressing tumors.
Assuntos
Carcinoma Ductal Pancreático/tratamento farmacológico , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Proteínas de Neoplasias/fisiologia , Neoplasias Pancreáticas/tratamento farmacológico , Proteínas Proto-Oncogênicas/fisiologia , Receptores Proteína Tirosina Quinases/fisiologia , Varfarina/farmacologia , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Apoptose/efeitos dos fármacos , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Divisão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Desoxicitidina/administração & dosagem , Desoxicitidina/análogos & derivados , Desoxicitidina/uso terapêutico , Progressão da Doença , Sinergismo Farmacológico , Feminino , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos SCID , Invasividade Neoplásica , Metástase Neoplásica , Proteínas de Neoplasias/antagonistas & inibidores , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Proteínas Proto-Oncogênicas/genética , RNA Interferente Pequeno/farmacologia , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Receptores Proteína Tirosina Quinases/genética , Transdução de Sinais/efeitos dos fármacos , Organismos Livres de Patógenos Específicos , Varfarina/administração & dosagem , Varfarina/uso terapêutico , Ensaios Antitumorais Modelo de Xenoenxerto , Gencitabina , Receptor Tirosina Quinase AxlRESUMO
Modern cancer treatment employs many effective chemotherapeutic agents originally discovered from natural sources. The cyclic depsipeptide didemnin B has demonstrated impressive anticancer activity in preclinical models. Clinical use has been approved but is limited by sparse patient responses combined with toxicity risk and an unclear mechanism of action. From a broad-scale effort to match antineoplastic natural products to their cellular activities, we found that didemnin B selectively induces rapid and wholesale apoptosis through dual inhibition of PPT1 and EEF1A1. Furthermore, empirical discovery of a small panel of exceptional responders to didemnin B allowed the generation of a regularized regression model to extract a sparse-feature genetic biomarker capable of predicting sensitivity to didemnin B. This may facilitate patient selection in a fashion that could enhance and expand the therapeutic application of didemnin B against neoplastic disease.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Depsipeptídeos/farmacologia , Proteínas de Membrana/antagonistas & inibidores , Fator 1 de Elongação de Peptídeos/antagonistas & inibidores , Farmacogenética , Apoptose/genética , Biomarcadores/metabolismo , Linhagem Celular Tumoral , Estudo de Associação Genômica Ampla , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina , Proteínas de Membrana/genética , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Fator 1 de Elongação de Peptídeos/genética , Biossíntese de Proteínas/efeitos dos fármacos , Biossíntese de Proteínas/genética , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Tioléster Hidrolases , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genéticaRESUMO
There is growing evidence that antiangiogenic therapy stimulates cancer cell invasion and metastasis. However, the underlying molecular mechanisms responsible for these changes have not been fully defined. Here, we report that anti-VEGF therapy promotes local invasion and metastasis by inducing collagen signaling in cancer cells. We show that chronic VEGF inhibition in a genetically engineered mouse model of pancreatic ductal adenocarcinoma (PDA) induces hypoxia, a less differentiated mesenchymal-like tumor cell phenotype, TGF-ß expression, and collagen deposition and signaling. In addition, we show that collagen signaling is critical for protumorigenic activity of TGF-ß in vitro. To further model the impact of collagen signaling in tumors, we evaluated PDA in mice lacking Sparc, a protein that reduces collagen binding to cell surface receptors. Importantly, we show that loss of Sparc increases collagen signaling and tumor progression. Together, these findings suggest that collagen actively promotes PDA spread and that enhanced disease progression associated with anti-VEGF therapy can arise from elevated extracellular matrix-mediated signaling.
Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/patologia , Colágeno/fisiologia , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/patologia , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Animais , Bevacizumab , Linhagem Celular Tumoral , Modelos Animais de Doenças , Progressão da Doença , Camundongos , Camundongos Transgênicos , Invasividade Neoplásica , Transdução de Sinais/fisiologia , Falha de Tratamento , Fator A de Crescimento do Endotélio Vascular/imunologiaRESUMO
Context-specific molecular vulnerabilities that arise during tumor evolution represent an attractive intervention target class. However, the frequency and diversity of somatic lesions detected among lung tumors can confound efforts to identify these targets. To confront this challenge, we have applied parallel screening of chemical and genetic perturbations within a panel of molecularly annotated NSCLC lines to identify intervention opportunities tightly linked to molecular response indicators predictive of target sensitivity. Anchoring this analysis on a matched tumor/normal cell model from a lung adenocarcinoma patient identified three distinct target/response-indicator pairings that are represented with significant frequencies (6%-16%) in the patient population. These include NLRP3 mutation/inflammasome activation-dependent FLIP addiction, co-occurring KRAS and LKB1 mutation-driven COPI addiction, and selective sensitivity to a synthetic indolotriazine that is specified by a seven-gene expression signature. Target efficacies were validated in vivo, and mechanism-of-action studies informed generalizable principles underpinning cancer cell biology.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Indóis/farmacologia , Neoplasias Pulmonares/metabolismo , Triazinas/farmacologia , Animais , Proteína Reguladora de Apoptosis Semelhante a CASP8 e FADD/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Proteínas de Transporte , Linhagem Celular Tumoral , Proteína Coatomer/metabolismo , Feminino , Genes ras , Xenoenxertos , Humanos , Neoplasias Pulmonares/patologia , Lisossomos/metabolismo , Camundongos , Terapia de Alvo Molecular , Proteína 3 que Contém Domínio de Pirina da Família NLR , Transplante de Neoplasias , Fosforilação OxidativaRESUMO
In lung cancer, platelet-derived growth factor receptor α (PDGFRα) is expressed frequently by tumor-associated stromal cells and by cancer cells in a subset of tumors. We sought to determine the effect of targeting stromal PDGFRα in preclinical lung tumor xenograft models (human tumor, mouse stroma). Effects of anti-human (IMC-3G3) and anti-mouse (1E10) PDGFRα monoclonal antibodies (mAb) on proliferation and PDGFRα signaling were evaluated in lung cancer cell lines and mouse fibroblasts. Therapy studies were conducted using established PDGFRα-positive H1703 cells and PDGFRα-negative Calu-6, H1993, and A549 subcutaneous tumors in immunocompromised mice treated with vehicle, anti-PDGFRα mAbs, chemotherapy, or combination therapy. Tumors were analyzed for growth and levels of growth factors. IMC-3G3 inhibited PDGFRα activation and the growth of H1703 cells in vitro and tumor growth in vivo, but had no effect on PDGFRα-negative cell lines or mouse fibroblasts. 1E10 inhibited growth and PDGFRα activation of mouse fibroblasts, but had no effect on human cancer cell lines in vitro. In vivo, 1E10-targeted inhibition of murine PDGFRα reduced tumor growth as single-agent therapy in Calu-6 cells and enhanced the effect of chemotherapy in xenografts derived from A549 cells. We also identified that low expression cancer cell expression of VEGF-A and elevated expression of PDGF-AA were associated with response to stromal PDGFRα targeting. We conclude that stromal PDGFRα inhibition represents a means for enhancing control of lung cancer growth in some cases, independent of tumor cell PDGFRα expression.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Neoplasias Pulmonares/metabolismo , Terapia de Alvo Molecular , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Animais , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Feminino , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Camundongos , Fator de Crescimento Derivado de Plaquetas/metabolismo , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/antagonistas & inibidores , Especificidade da Espécie , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismo , Células Estromais/patologia , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
PURPOSE: COX-2 is expressed highly in pancreatic cancer and implicated in tumor progression. COX-2 inhibition can reduce tumor growth and augment therapy. The precise function of COX-2 in tumors remains poorly understood, but it is implicated in tumor angiogenesis, evasion of apoptosis, and induction of epithelial-to-mesenchymal transition (EMT). Current therapeutic regimens for pancreatic cancer are minimally effective, highlighting the need for novel treatment strategies. Here, we report that apricoxib, a novel COX-2 inhibitor in phase II clinical trials, significantly enhances the efficacy of gemcitabine/erlotinib in preclinical models of pancreatic cancer. EXPERIMENTAL DESIGN: Human pancreatic cell lines were evaluated in vitro and in vivo for response to apricoxib ± standard-of-care therapy (gemcitabine + erlotinib). Tumor tissue underwent posttreatment analysis for cell proliferation, viability, and EMT phenotype. Vascular parameters were also determined. RESULTS: COX-2 inhibition reduced the IC(50) of gemcitabine ± erlotinib in six pancreatic cancer cell lines tested in vitro. Furthermore, apricoxib increased the antitumor efficacy of standard combination therapy in several orthotopic xenograft models. In vivo apricoxib combination therapy was only effective at reducing tumor growth and metastasis in tumors with elevated COX-2 activity. In each model examined, treatment with apricoxib resulted in vascular normalization without a decrease in microvessel density and promotion of an epithelial phenotype by tumor cells regardless of basal COX-2 expression. CONCLUSIONS: Apricoxib robustly reverses EMT and augments standard therapy without reducing microvessel density and warrants further clinical evaluation in patients with pancreatic cancer.
Assuntos
Antineoplásicos/uso terapêutico , Inibidores de Ciclo-Oxigenase 2/uso terapêutico , Neoplasias Pancreáticas/tratamento farmacológico , Pirróis/uso terapêutico , Sulfonamidas/uso terapêutico , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Inibidores de Ciclo-Oxigenase 2/administração & dosagem , Inibidores de Ciclo-Oxigenase 2/farmacologia , Sinergismo Farmacológico , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Receptores ErbB/metabolismo , Cloridrato de Erlotinib , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Metástase Neoplásica , Neovascularização Patológica/tratamento farmacológico , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Pirróis/administração & dosagem , Pirróis/farmacologia , Quinazolinas/farmacologia , Sulfonamidas/administração & dosagem , Sulfonamidas/farmacologiaRESUMO
Although cyclooxygenase-2 (COX-2) inhibitors, such as the late stage development drug apricoxib, exhibit antitumor activity, their mechanisms of action have not been fully defined. In this study, we characterized the mechanisms of action of apricoxib in HT29 colorectal carcinoma. Apricoxib was weakly cytotoxic toward naive HT29 cells in vitro but inhibited tumor growth markedly in vivo. Pharmacokinetic analyses revealed that in vivo drug levels peaked at 2-4 µM and remained sufficient to completely inhibit prostaglandin E(2) production, but failed to reach concentrations cytotoxic for HT29 cells in monolayer culture. Despite this, apricoxib significantly inhibited tumor cell proliferation and induced apoptosis without affecting blood vessel density, although it did promote vascular normalization. Strikingly, apricoxib treatment induced a dose-dependent reversal of epithelial-mesenchymal transition (EMT), as shown by robust upregulation of E-cadherin and the virtual disappearance of vimentin and ZEB1 protein expression. In vitro, either anchorage-independent growth conditions or forced EMT sensitized HT29 and non-small cell lung cancer cells to apricoxib by 50-fold, suggesting that the occurrence of EMT may actually increase the dependence of colon and lung carcinoma cells on COX-2. Taken together, these data suggest that acquisition of mesenchymal characteristics sensitizes carcinoma cells to apricoxib resulting in significant single-agent antitumor activity.
Assuntos
Antineoplásicos/farmacologia , Inibidores de Ciclo-Oxigenase 2/farmacologia , Transição Epitelial-Mesenquimal , Pirróis/farmacologia , Sulfonamidas/farmacologia , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Dinoprostona/biossíntese , Feminino , Células HT29 , Humanos , Camundongos , Camundongos Nus , Fator A de Crescimento do Endotélio Vascular/fisiologiaRESUMO
Pancreatic adenocarcinoma, a desmoplastic disease, is the fourth leading cause of cancer-related death in the Western world due, in large part, to locally invasive primary tumor growth and ensuing metastasis. SPARC is a matricellular protein that governs extracellular matrix (ECM) deposition and maturation during tissue remodeling, particularly, during wound healing and tumorigenesis. In the present study, we sought to determine the mechanism by which lack of host SPARC alters the tumor microenvironment and enhances invasion and metastasis of an orthotopic model of pancreatic cancer. We identified that levels of active TGFß1 were increased significantly in tumors grown in SPARC-null mice. TGFß1 contributes to many aspects of tumor development including metastasis, endothelial cell permeability, inflammation and fibrosis, all of which are altered in the absence of stromal-derived SPARC. Given these results, we performed a survival study to assess the contribution of increased TGFß1 activity to tumor progression in SPARC-null mice using losartan, an angiotensin II type 1 receptor antagonist that diminishes TGFß1 expression and activation in vivo. Tumors grown in SPARC-null mice progressed more quickly than those grown in wild-type littermates leading to a significant reduction in median survival. However, median survival of SPARC-null animals treated with losartan was extended to that of losartan-treated wild-type controls. In addition, losartan abrogated TGFß induced gene expression, reduced local invasion and metastasis, decreased vascular permeability and altered the immune profile of tumors grown in SPARC-null mice. These data support the concept that aberrant TGFß1-activation in the absence of host SPARC contributes significantly to tumor progression and suggests that SPARC, by controlling ECM deposition and maturation, can regulate TGFß availability and activation.
Assuntos
Losartan/farmacologia , Osteonectina/deficiência , Neoplasias Pancreáticas/tratamento farmacológico , Fator de Crescimento Transformador beta/antagonistas & inibidores , Bloqueadores do Receptor Tipo 1 de Angiotensina II , Animais , Progressão da Doença , Matriz Extracelular/metabolismo , Losartan/uso terapêutico , Camundongos , Camundongos Knockout , Invasividade Neoplásica , Metástase Neoplásica , Taxa de Sobrevida , Fator de Crescimento Transformador beta/metabolismo , Resultado do TratamentoRESUMO
BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is highly resistant to conventional chemotherapy, in part due to the overexpression of inhibitors of apoptosis proteins (IAPs). Smac is an endogenous IAP-antagonist, which renders synthetic Smac mimetics attractive anticancer agents. We evaluated the benefits of combining a Smac mimetic, JP1201 (JP), with conventional chemotherapy agents used for PDAC management. METHODS: Cell viability assays and protein expression analysis were performed using WST-1 reagent and Western blotting, respectively. Apoptosis was detected by annexin V/propidium iodide staining. In vivo tumor growth and survival studies were performed in murine PDAC xenografts. RESULTS: JP and gemcitabine (Gem) inhibited PDAC cell proliferation with additive effects in combination. The percentage of early apoptotic cells in controls, JP, Gem and JP + Gem was 17%, 26%, 26% and 38%, respectively. JP-induced apoptosis was accompanied by PARP-1 cleavage. Similar additive anti-proliferative effects were seen for combinations of JP with doxorubicin (Dox) and docetaxel (DT). The JP + Gem combination caused a 30% decrease in tumor size in vivo compared to controls. Median animal survival was improved significantly in mice treated with JP + Gem (38 d) compared to controls (22 d), JP (28 d) or Gem (32 d) (p = 0.01). Animal survival was also improved with JP + DT treatment (32 d) compared to controls (16 d), JP (21 d) or DT alone (27 d). CONCLUSIONS: These results warrant further exploration of strategies that promote chemotherapy-induced apoptosis of tumors and highlight the potential of Smac mimetics in clinical PDAC therapy.