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1.
Cancer Res ; 79(7): 1465-1479, 2019 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-30482772

RESUMO

Given the integral role of stimulator of interferon genes (STING, TMEM173) in the innate immune response, its loss or impairment in cancer is thought to primarily affect antitumor immunity. Here we demonstrate a role for STING in the maintenance of cellular homeostasis through regulation of the cell cycle. Depletion of STING in human and murine cancer cells and tumors resulted in increased proliferation compared with wild-type controls. Microarray analysis revealed genes involved in cell-cycle regulation are differentially expressed in STINGko compared with WT MEFs. STING-mediated regulation of the cell cycle converged on NFκB- and p53-driven activation of p21. The absence of STING led to premature activation of cyclin-dependent kinase 1 (CDK1), early onset to S-phase and mitosis, and increased chromosome instability, which was enhanced by ionizing radiation. These results suggest a pivotal role for STING in maintaining cellular homeostasis and response to genotoxic stress. SIGNIFICANCE: These findings provide clear mechanistic understanding of the role of STING in cell-cycle regulation, which may be exploited in cancer therapy because most normal cells express STING, while many tumor cells do not.See related commentary by Gius and Zhu, p. 1295.


Assuntos
Imunidade Inata , Proteínas de Membrana/genética , Animais , Proliferação de Células , Instabilidade Cromossômica , Homeostase , Humanos , Camundongos
2.
Cancer Immunol Res ; 7(2): 306-320, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30538091

RESUMO

Transforming growth factor ß (TGFß) is an effector of immune suppression and contributes to a permissive tumor microenvironment that compromises effective immunotherapy. We identified a correlation between TGFB1 and genes expressed by myeloid cells, but not granulocytes, in The Cancer Genome Atlas lung adenocarcinoma data, in which high TGFB1 expression was associated with poor survival. To determine whether TGFß affected cell fate decisions and lineage commitment, we studied primary cultures of CD14+ monocytes isolated from peripheral blood of healthy donors. We discovered that TGFß was a survival factor for CD14+ monocytes, which rapidly executed an apoptotic program in its absence. Continued exposure to TGFß in combination with granulocyte-macrophage colony stimulating factor (GM-CSF) and interleukin 6 (IL6) amplified HLA-DRlowCD14+CD11b+CD33+ myeloid-derived suppressor cells (MDSCs) at the expense of macrophage and dendritic cell (DC) differentiation. MDSCs generated in the presence of TGFß were more effective in suppressing T-cell proliferation and promoted the T regulatory cell phenotype. In contrast, inhibition of TGFß signaling using a small-molecule inhibitor of receptor kinase activity in CD14+ monocytes treated with GM-CSF and IL6 decreased MDSC differentiation and increased differentiation to proinflammatory macrophages and antigen-presenting DCs. The effect of autocrine and paracrine TGFß on myeloid cell survival and lineage commitment suggests that pharmacologic inhibition of TGFß-dependent signaling in cancer would favor antitumor immunity.


Assuntos
Comunicação Autócrina , Diferenciação Celular/imunologia , Imunomodulação , Monócitos/imunologia , Monócitos/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Adenocarcinoma de Pulmão/etiologia , Adenocarcinoma de Pulmão/metabolismo , Adenocarcinoma de Pulmão/mortalidade , Adenocarcinoma de Pulmão/patologia , Apresentação de Antígeno/imunologia , Biomarcadores , Sobrevivência Celular/genética , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Perfilação da Expressão Gênica , Humanos , Monócitos/citologia , Células Supressoras Mieloides/citologia , Células Supressoras Mieloides/imunologia , Células Supressoras Mieloides/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta1/metabolismo
3.
Int J Mol Sci ; 19(12)2018 Dec 09.
Artigo em Inglês | MEDLINE | ID: mdl-30544863

RESUMO

An immunophenotyping analysis was performed in peripheral blood samples from seven patients with lung cancer unfit for surgery treated with stereotactic body radiotherapy (SBRT). The objective was to characterize the effect of SBRT on the host immune system. Four patients received 60 Gy (7.5 Gy × 8) and three 50 Gy (12.5 Gy × 4). Analyses were performed before SBRT, 72 h after SBRT, and at one, three, and six months after the end of SBRT. Of note, there was a specific increase of the immunoactive component of the immune system, with elevation of CD56+highCD16+ natural killer (NK) cells (0.95% at baseline to 1.38% at six months), and a decrease of the immunosuppressive component of the immune system, with decreases of CD4+CD25+Foxp3+CDA5RA- regulatory T cells (4.97% at baseline to 4.46% at six months), granulocytic myeloid-derived suppressor cells (G-MDSCs) (from 66.1% at baseline to 62.6% at six months) and monocytic (Mo-MDSCs) (8.2% at baseline to 6.2% at six months). These changes were already apparent at 72 h and persisted over six months. SBRT showed an effect on systemic immune cell populations, which is a relevant finding for supporting future combinations of SBRT with immunotherapy for treating lung cancer patients.


Assuntos
Sistema Imunitário/patologia , Imunofenotipagem , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/radioterapia , Radiocirurgia , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Neoplasias Pulmonares/cirurgia , Masculino , Células Supressoras Mieloides/imunologia , Linfócitos T Citotóxicos/imunologia , Linfócitos T Reguladores/imunologia
4.
Oncotarget ; 8(16): 26344-26355, 2017 Apr 18.
Artigo em Inglês | MEDLINE | ID: mdl-28412751

RESUMO

The development of stereotactic body radiation therapy (SBRT) has revolutionized radiation therapy for lung cancers and is an emerging treatment option for pancreatic cancers. However, there are many questions on how to optimize its use in chemoradiotherapy. The most relevant addition to radiotherapy regimens are inhibitors of DNA repair and DNA damage response pathways. One such class of agents are inhibitors of poly (ADP-ribose) polymerase (PARP). In this study we examined the effects of the PARP inhibitor LT626 in combination with ionizing radiation in lung and pancreatic cancers. Our study demonstrated that combination treatment with LT626 and radiation effectively inhibited growth in lung and pancreatic cancer cell lines, better than individual treatment alone. Combination treatment also increased expression of γH2AX and 53BP1 foci and upregulated expression of phosphorylated ATM, ATR and their respective kinases. Using in vivo lung cancer xenograft models we demonstrated that LT626 functioned as an effective radiosensitizer during fractionated radiation treatment, leading to significant decrease in tumor burden and doubling the median survival compared to control group. Overall our in vitro and in vivo studies showed that PARP inhibitor LT626 acted synergistically with radiation in lung and pancreatic cancers.


Assuntos
Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Tolerância a Radiação/efeitos dos fármacos , Radiossensibilizantes/farmacologia , Animais , Proteínas Mutadas de Ataxia Telangiectasia/genética , Biomarcadores Tumorais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/efeitos da radiação , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Relação Dose-Resposta à Radiação , Sinergismo Farmacológico , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos da radiação , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/terapia , Camundongos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/terapia , Doses de Radiação , Tolerância a Radiação/efeitos da radiação , Radiação Ionizante , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Int J Radiat Oncol Biol Phys ; 91(1): 91-9, 2015 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-25835621

RESUMO

PURPOSE: To determine whether transforming growth factor (TGF)-ß inhibition increases the response to radiation therapy in human and mouse non-small-cell lung carcinoma (NSCLC) cells in vitro and in vivo. METHODS AND MATERIALS: TGF-ß-mediated growth response and pathway activation were examined in human NSCLC NCI-H1299, NCI-H292, and A549 cell lines and murine Lewis lung cancer (LLC) cells. Cells were treated in vitro with LY364947, a small-molecule inhibitor of the TGF-ß type 1 receptor kinase, or with the pan-isoform TGF-ß neutralizing monoclonal antibody 1D11 before radiation exposure. The DNA damage response was assessed by ataxia telangiectasia mutated (ATM) or Trp53 protein phosphorylation, γH2AX foci formation, or comet assay in irradiated cells. Radiation sensitivity was determined by clonogenic assay. Mice bearing syngeneic subcutaneous LLC tumors were treated with 5 fractions of 6 Gy and/or neutralizing or control antibody. RESULTS: The NCI-H1299, A549, and LLC NSCLC cell lines pretreated with LY364947 before radiation exposure exhibited compromised DNA damage response, indicated by decreased ATM and p53 phosphorylation, reduced γH2AX foci, and increased radiosensitivity. The NCI-H292 cells were unresponsive. Transforming growth factor-ß signaling inhibition in irradiated LLC cells resulted in unresolved DNA damage. Subcutaneous LLC tumors in mice treated with TGF-ß neutralizing antibody exhibited fewer γH2AX foci after irradiation and significantly greater tumor growth delay in combination with fractionated radiation. CONCLUSIONS: Inhibition of TGF-ß before radiation attenuated DNA damage recognition and increased radiosensitivity in most NSCLC cells in vitro and promoted radiation-induced tumor control in vivo. These data support the rationale for concurrent TGF-ß inhibition and RT to provide therapeutic benefit in NSCLC.


Assuntos
Aminoácidos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/radioterapia , Dano ao DNA/efeitos dos fármacos , Neoplasias Pulmonares/radioterapia , Tolerância a Radiação/efeitos dos fármacos , Fator de Crescimento Transformador beta/antagonistas & inibidores , Xantenos/farmacologia , Animais , Anticorpos Monoclonais/farmacologia , Anticorpos Neutralizantes/farmacologia , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Western Blotting/métodos , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Quimioterapia Adjuvante , Ensaio Cometa , Histonas/metabolismo , Humanos , Técnicas In Vitro , Neoplasias Pulmonares/patologia , Camundongos , Fosforilação , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo I , Receptores de Fatores de Crescimento Transformadores beta/antagonistas & inibidores , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Proteína Supressora de Tumor p53/metabolismo
6.
J Nucl Med ; 50(3): 435-43, 2009 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-19223400

RESUMO

UNLABELLED: To optimize in vivo tissue uptake kinetics and clearance of engineered monoclonal antibody (mAb) fragments for radiotherapeutic and radiodiagnostic applications, we compared the biodistribution and tumor localization of four (111)In- and (86)Y-labeled antibody formats, derived from a single antimindin/RG-1 mAb, in a prostate tumor model. The IgG, diabody, single-chain variable domain (scFv), and novel miniantibody formats, composed of the human IgE-C(H)4 and a modified IgG1 hinge linked to scFv domains, were compared. METHODS: Antibodies were first derivatized with the bifunctional chelator CHX-A''-diethylenetriamine pentaacetic acid and then bound to the radiometal to create radiolabeled immunoconjugates. Human LNCaP xenografts were grown in nude mice, and (111)In- or (86)Y-labeled antibodies were administered intravenously. Tissues were harvested at different times, and the level of antibody deposition was determined by measuring radioactivity. Whole-body small-animal PET of mice receiving (86)Y-labeled antibodies was performed at 6 time points and colocalized with simultaneous micro-CT imaging. RESULTS: The biodistributions of (111)In and (86)Y antibodies were quite similar. The blood, tumor, kidney, and liver tissues contained varying levels of radioactivity. The antibody accumulation in the tumor correlated with molecular size. The IgG steadily increased with time to 24.1 percentage injected dose per gram (%ID/g) at 48 h. The miniantibody accumulated at a similar rate to reach a lower level (14.2 %ID/g) at 48 h but with a higher tumor-to-blood ratio than the IgG. Tumor accumulation of the diabody peaked at 3 h, reaching a much lower level (3.7 %ID/g). A combination of rapid clearance and lower relative affinity of the scFv precluded deposition in the tumor. Small-animal PET results correlated well with the biodistribution results, with similar tumor localization patterns. CONCLUSION: The larger antibody formats (IgG and miniantibody) gave higher tumor uptake levels than did the smaller formats (diabody and scFv). These larger formats may be more suitable for radioimmunotherapy applications, evidenced by the preclinical efficacy previously shown by a report on the IgG format. The smaller formats were rapidly cleared from circulation, and the diabody, which accumulated in the tumor, may be more suitable for radiodiagnostic applications.


Assuntos
Anticorpos Monoclonais/farmacocinética , Neoplasias da Próstata/metabolismo , Compostos Radiofarmacêuticos/farmacocinética , Animais , Anticorpos Monoclonais/química , Linhagem Celular Tumoral , Quelantes/química , Humanos , Radioisótopos de Índio , Isotiocianatos/química , Masculino , Camundongos , Camundongos Nus , Transplante de Neoplasias , Ácido Pentético/análogos & derivados , Ácido Pentético/química , Tomografia por Emissão de Pósitrons , Compostos Radiofarmacêuticos/química , Distribuição Tecidual , Tomografia Computadorizada por Raios X , Transplante Heterólogo , Radioisótopos de Ítrio
7.
Cancer Res ; 66(7): 3611-9, 2006 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-16585186

RESUMO

Hepsin is a type II transmembrane serine protease that is expressed in normal liver, and at lower levels in kidney, pancreas, and testis. Several studies have shown that hepsin mRNA is significantly elevated in most prostate tumors, as well as a significant fraction of ovarian and renal cell carcinomas and hepatomas. Although the overexpression of mRNA in these tumors has been extensively documented, there has been conflicting literature on whether hepsin plays a role in tumor cell growth and progression. Early literature implied a role for hepsin in human tumor cell proliferation, whereas recent studies with a transgenic mouse model for prostate cancer support a role for hepsin in tumor progression and metastases. To evaluate this issue further, we have expressed an activatable form of hepsin, and have generated a set of monoclonal antibodies that neutralize enzyme activity. The neutralizing antibodies inhibit hepsin enzymatic activity in biochemical and cell-based assays. Selected neutralizing and nonneutralizing antibodies were used in cell-based assays with tumor cells to evaluate the effect of antibodies on tumor cell growth and invasion. Neutralizing antibodies failed to inhibit the growth of prostate, ovarian, and hepatoma cell lines in culture. However, potent inhibitory effects of the antibodies were seen on invasion of ovarian and prostate cells in transwell-based invasion assays. These results support a role for hepsin in tumor cell progression but not in primary tumor growth. Consistent with this, immunohistochemical experiments with a mouse monoclonal antibody reveal progressively increased staining of prostate tumors with advanced disease, and in particular, extensive staining of bone metastatic lesions.


Assuntos
Anticorpos Monoclonais/farmacologia , Neoplasias Ovarianas/enzimologia , Neoplasias da Próstata/enzimologia , Serina Endopeptidases/metabolismo , Inibidores de Serina Proteinase/farmacologia , Sequência de Aminoácidos , Anticorpos Monoclonais/imunologia , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/fisiologia , Linhagem Celular Tumoral , Clonagem Molecular , Feminino , Humanos , Imuno-Histoquímica , Masculino , Dados de Sequência Molecular , Invasividade Neoplásica , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/patologia , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Serina Endopeptidases/biossíntese , Serina Endopeptidases/genética , Serina Endopeptidases/imunologia , Inibidores de Serina Proteinase/imunologia
8.
Cancer Res ; 65(18): 8397-405, 2005 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-16166318

RESUMO

Gene expression analysis showed that a human mindin homologue, mindin/RG-1, is expressed selectively in prostate tissues and that its expression level is elevated in some prostate tumors. Mindin/RG-1 protein expression is maintained in >80% of prostate cancers metastatic to bone or lymph nodes as well as in locally recurrent tumors in androgen-unresponsive patients. In contrast, mindin/RG-1 expression in other normal tissues is significantly lower than that seen in the prostate. A fully human antibody, 19G9, was generated against mindin/RG-1 protein and was shown to accumulate at high abundance in LNCaP tumor xenografts. Conjugates of this antibody with the chelator CHX-A''-DTPA were generated and radiolabeled with either 111In, 90Y, or 86Y. Small animal positron emission tomography imaging with the 86Y-radiolabeled conjugate showed very specific accumulation of the antibody in LNCaP tumor xenografts with clear tumor delineation apparent at 4 hours. The therapeutic efficacy of [90Y]-CHX-A''-DTPA-19G9 was evaluated in mice bearing LNCaP xenografts. A dose-finding study identified a nontoxic therapeutic dose to be approximately 75 microCi. Significant antitumor effects were seen with a single administration of radiolabeled antibody to animals bearing 200 to 400 mm3 tumors. Inhibition of tumor growth was observed in all treated animals over a 49-day period. At 49 days posttreatment, slow tumor growth recurred but this could be prevented for an additional 40-day period by a second administration of a 75 microCi dose at day 49. We conclude that [90Y]-CHX-A''-DTPA-19G9 is a novel antibody conjugate that has considerable promise for therapy of metastatic prostate cancer in androgen-unresponsive patients.


Assuntos
Anticorpos Monoclonais/imunologia , Proteínas da Matriz Extracelular/imunologia , Imunotoxinas/imunologia , Neoplasias da Próstata/radioterapia , Radioimunoterapia/métodos , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/farmacocinética , Anticorpos Monoclonais/farmacologia , Especificidade de Anticorpos , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/secundário , Células CHO , Cricetinae , Relação Dose-Resposta Imunológica , Proteínas da Matriz Extracelular/biossíntese , Proteínas da Matriz Extracelular/genética , Humanos , Imunotoxinas/farmacocinética , Imunotoxinas/farmacologia , Isotiocianatos/imunologia , Isotiocianatos/farmacocinética , Isotiocianatos/farmacologia , Masculino , Dados de Sequência Molecular , Ácido Pentético/análogos & derivados , Ácido Pentético/imunologia , Ácido Pentético/farmacocinética , Ácido Pentético/farmacologia , Tomografia por Emissão de Pósitrons , Neoplasias da Próstata/diagnóstico por imagem , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/metabolismo , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Distribuição Tecidual , Ensaios Antitumorais Modelo de Xenoenxerto , Radioisótopos de Ítrio/administração & dosagem , Radioisótopos de Ítrio/farmacocinética , Radioisótopos de Ítrio/farmacologia
9.
Cancer Res ; 65(7): 2846-53, 2005 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-15805286

RESUMO

Radiotherapy is an effective approach for the treatment of local prostate cancer. However, once prostate cancer metastasizes, radiotherapy cannot be used due to the distribution of multiple metastases to lymph nodes and bones. In contrast, radioimmunotherapy should still be efficacious in metastatic prostate cancer as radioisotopes are brought to tumor cells by targeting antibodies. Here we identify and validate a prostate-expressed molecule, tomoregulin, as a target for radioimmunotherapy of prostate cancer. Tomoregulin is a transmembrane protein selectively expressed in the brain, prostate, and prostate cancer, but not expressed in other normal tissues. Immunohistochemical studies of tomoregulin protein in clinical samples show its location in the luminal epithelium of normal prostate, benign prostatic hyperplasia, and prostatic intraepithelial neoplasia. More importantly, the tomoregulin protein is expressed in primary prostate tumors and in their lymph node and bone metastases. The nature of tomoregulin as a transmembrane protein and its tissue-specific expression make tomoregulin an attractive target for radioimmunotherapy, in which tomoregulin-specific antibodies will deliver a radioisotope to prostate tumor cells and metastases. Indeed, biodistribution studies using a prostate tumor xenograft model showed that the (111)In-labeled anti-tomoregulin antibody 2H8 specifically recognizes tomoregulin protein in vivo, leading to a strong tumor-specific accumulation of the antibody. In efficacy studies, a single i.p. dose of 150 microCi (163 microg) (90)Y-labeled 2H8 substantially inhibits the growth rate of established LNCaP human prostate tumor xenograft in nude mice but produces no overt toxicity despite cross-reactivity of 2H8 with mouse tomoregulin. Our data clearly validate tomoregulin as a target for radioimmunotherapy of prostate cancer.


Assuntos
Imunotoxinas/uso terapêutico , Radioisótopos de Índio/uso terapêutico , Proteínas de Membrana/biossíntese , Proteínas de Neoplasias/biossíntese , Neoplasias da Próstata/radioterapia , Radioisótopos/uso terapêutico , Itérbio/uso terapêutico , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacocinética , Anticorpos Monoclonais/uso terapêutico , Encéfalo/metabolismo , Linhagem Celular Tumoral , Humanos , Imunoglobulina G/imunologia , Imunoglobulina G/metabolismo , Imunotoxinas/imunologia , Imunotoxinas/farmacocinética , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Camundongos , Camundongos Nus , Metástase Neoplásica , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/imunologia , Próstata/metabolismo , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Radioimunoterapia , Compostos Radiofarmacêuticos/imunologia , Compostos Radiofarmacêuticos/farmacocinética , Compostos Radiofarmacêuticos/uso terapêutico , Distribuição Tecidual , Ensaios Antitumorais Modelo de Xenoenxerto
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