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1.
Int J Oncol ; 50(6): 1947-1954, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28498437

RESUMO

In cervical cancer, the epidermal growth factor receptor (EGFR) is overexpressed in 70-90% of the cases and has been associated with poor prognosis. EGFR-based therapy is currently being explored in cervical cancer. We investigated which EGFR ligand is primarily expressed in cervical cancer and which cell type functions as the major source of this ligand. We hypothesized that macrophages are the main source of EGFR ligands and that a paracrine loop between tumor cells and macrophages is responsible for ligand expression. mRNA expression analysis was performed on 32 cervical cancer cases to determine the expression of the EGFR ligands amphiregulin, ß-cellulin, epidermal growth factor (EGF), epiregulin, heparin-binding EGF-like growth factor (HB­EGF) and transforming growth factor α (TGFα). Subsequently, protein expression was determined immunohistochemically on 36 additional cases. To assess whether macrophages are the major source of EGFR ligands, immunohistochemical double staining was performed on four representative tissue slides. Expression of the chemokines granulocyte-macrophage colony-stimulating factor (GM-CSF) and C-C motif ligand 2 (CCL2) was determined by mRNA in situ hybridization. Of the known EGFR ligands, HB­EGF had the highest mRNA expression and HB­EGF and EGFR protein expression were highly correlated. Tumor specimens with high EGFR expression showed higher numbers of macrophages, and higher expression of GM-CSF and CCL2, but only a small subset (9%) of macrophages was found to be HB­EGF-positive. Strikingly, 78% of cervical cancer specimens were found to express HB­EGF. Standardized assessment of staining intensity, using spectral imaging analysis, showed that HB­EGF expression was higher in the tumor compartment than in the stromal compartment. These results suggest that HB­EGF is an important EGFR ligand in cervical cancer and that cervical cancer cells are the predominant source of HB­EGF. Therefore, we propose an autocrine EGFR stimulation model in cervical carcinomas.


Assuntos
Receptores ErbB/genética , Fator de Crescimento Semelhante a EGF de Ligação à Heparina/genética , Neoplasias do Colo do Útero/genética , Anfirregulina/genética , Comunicação Autócrina/genética , Linhagem Celular Tumoral , Quimiocina CCL2/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Humanos , Ligantes , RNA Mensageiro , Transdução de Sinais/genética , Neoplasias do Colo do Útero/patologia
2.
Cancer Immunol Immunother ; 65(4): 393-403, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26899388

RESUMO

Patients with HPV-positive oropharyngeal squamous cell carcinomas (OPSCCs) have a better prognosis than patients with non-HPV-induced OPSCC. The role of the immune response in this phenomenon is yet unclear. We studied the number of T cells, regulatory T cells (Tregs), T helper 17 (Th17) cells and IL-17(+) non-T cells (mainly granulocytes) in matched HPV-positive and HPV-negative OPSCC cases (n = 162). Furthermore, the production of IFN-γ and IL-17 by tumor-infiltrating T cells was analyzed. The number of tumor-infiltrating T cells and Tregs was higher in HPV-positive than HPV-negative OPSCC (p < 0.0001). In contrast, HPV-negative OPSCC contained significantly higher numbers of IL-17(+) non-T cells (p < 0.0001). Although a high number of intra-tumoral T cells showed a trend toward improved survival of all OPSCC patients, their prognostic effect in patients with a low number of intra-tumoral IL-17(+) non-T cells was significant with regard to disease-specific (p = 0.033) and disease-free survival (p = 0.012). This suggests that a high frequency of IL-17(+) non-T cells was related to a poor immune response, which was further supported by the observation that a high number of T cells was correlated with improved disease-free survival in the HPV-positive OPSCC (p = 0.008). In addition, we detected a minor Th17 cell population. However, T cells obtained from HPV-positive OPSCC produced significantly more IL-17 than those from HPV-negative tumors (p = 0.006). The improved prognosis of HPV-positive OPSCC is thus correlated with higher numbers of tumor-infiltrating T cells, more active Th17 cells and lower numbers of IL-17(+) non-T cells.


Assuntos
Carcinoma de Células Escamosas/imunologia , Neoplasias Orofaríngeas/imunologia , Linfócitos T/imunologia , Células Th17/imunologia , Microambiente Tumoral/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Complexo CD3/imunologia , Complexo CD3/metabolismo , Carcinoma de Células Escamosas/complicações , Feminino , Fatores de Transcrição Forkhead/imunologia , Fatores de Transcrição Forkhead/metabolismo , Granulócitos/imunologia , Granulócitos/metabolismo , Humanos , Interleucina-17/imunologia , Interleucina-17/metabolismo , Estimativa de Kaplan-Meier , Contagem de Linfócitos , Masculino , Microscopia Confocal , Pessoa de Meia-Idade , Neoplasias Orofaríngeas/complicações , Papillomaviridae/imunologia , Infecções por Papillomavirus/complicações , Infecções por Papillomavirus/imunologia , Infecções por Papillomavirus/virologia , Linfócitos T/metabolismo , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Células Th17/metabolismo
3.
J Transl Med ; 13: 295, 2015 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-26357849

RESUMO

BACKGROUND: Deep invasion of the normal surrounding tissue by primary cervical cancers is a prognostic parameter for postoperative radiotherapy and relatively worse survival. However, patients with tumor-specific immunity in the blood at the time of surgery displayed a much better disease free survival. Here we analyzed if this was due to a more tumor-rejecting immune population in the tumor. METHODS: Tumor sections from a group of 58 patients with deep normal tissue-invading cervical tumors were stained for the presence of immune cells (CD45), IFNγ-producing cells (Tbet) and regulatory T cells (Foxp3) by immunohistochemistry. The slides were scanned and both the tumor area and the infiltration of the differently stained immune cells were objectively quantified using computer software. RESULTS: We found that an increased percentage of tumor occupied by CD45+ cells was strongly associated with an enhanced tumor-infiltration by Tbet+ cells and Foxp3+ cells. Furthermore, the area occupied by CD45+ immune cells, Tbet+ cells but not Foxp3+ cells within the tumor were, in addition to the lymph node status of patients, associated with a longer disease free survival and disease specific survival. Moreover, interaction analyses between these immune parameters and lymph node status indicated an independent prognostic effect of tumor infiltrating Tbet+ cells. This was confirmed in a multivariate Cox analysis. CONCLUSIONS: The area occupied by a preferentially type I oriented CD45+ cell infiltrate forms an independent prognostic factor for recurrence-free and disease-specific survival on top of the patient's lymph node status.


Assuntos
Linfócitos T Reguladores/citologia , Neoplasias do Colo do Útero/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Proliferação de Células , Intervalo Livre de Doença , Feminino , Fatores de Transcrição Forkhead/metabolismo , Humanos , Imuno-Histoquímica , Interferon gama/metabolismo , Antígenos Comuns de Leucócito/metabolismo , Linfócitos/citologia , Pessoa de Meia-Idade , Invasividade Neoplásica , Metástase Neoplásica , Infecções por Papillomavirus/metabolismo , Prognóstico , Modelos de Riscos Proporcionais , Resultado do Tratamento , Neoplasias do Colo do Útero/diagnóstico
4.
Oncotarget ; 6(36): 38681-94, 2015 Nov 17.
Artigo em Inglês | MEDLINE | ID: mdl-26299617

RESUMO

Cervical cancer is typically well infiltrated by immune cells. Because of the intricate relationship between cancer cells and immune cells, we aimed to identify both cancer cell and immune cell expressed biomarkers. Using a novel approach, we isolated RNA from flow-sorted viable EpCAM+ tumor epithelial cells and CD45+ tumor-infiltrating immune cells obtained from squamous cell cervical cancer samples (n = 24). Total RNA was sequenced and differential gene expression analysis of the CD45+ immune cell fractions identified TCL1A as a novel marker for predicting improved survival (p = 0.007). This finding was validated using qRT-PCR (p = 0.005) and partially validated using immunohistochemistry (p = 0.083). Importantly, TCL1A was found to be expressed in a subpopulation of B cells (CD3-/CD19+/CD10+/CD34-) using multicolor immunofluorescence. A high TCL1A/CD20 (B cell) ratio, determined in total tumor samples from a separate patient cohort using qRT-PCR (n = 52), was also correlated with improved survival (p = 0.027). This is the first study demonstrating the prognostic value of separating tumor epithelial cells from tumor-infiltrating immune cells and determining their RNA expression profile for identifying putative cancer biomarkers. Our results suggest that intratumoral TCL1A+ B cells are important for controlling cervical cancer development.


Assuntos
Linfócitos B/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Neoplasias do Colo do Útero/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Perfilação da Expressão Gênica , Humanos , Pessoa de Meia-Idade , Prognóstico , Análise de Sobrevida , Microambiente Tumoral , Neoplasias do Colo do Útero/mortalidade , Neoplasias do Colo do Útero/patologia
5.
PLoS One ; 10(6): e0129119, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26066796

RESUMO

Galectins are proteins that bind ß-galactoside sugars and provide a new type of potential biomarkers and therapeutic targets in cancer. Galectin-1, -3 and -9 have become the focus of different research groups, but their expression and function in cervical cancer is still unclear. The aim of this study was to determine the phenotype of galectin-1, -3 and -9 expressing cells and the association with clinico-pathological parameters in cervical cancer. Galectin expression was scored in tumor cells, tumor epithelium infiltrating immune cells and stromal cells in squamous cervical cancer (n = 160). Correlations with clinico-pathological parameters and survival were studied according to the REMARK recommendations. We additionally investigated whether the galectins were expressed by tumor cells, fibroblasts, macrophages and T cells. Galectin-1 and -9 were both expressed by tumor cells in 11% of samples, while 84% expressed galectin-3. Strong galectin-1 expression by tumor cells was an independent predictor for poor survival (hazard ratio: 8.02, p = 0.001) and correlated with increased tumor invasion (p = 0.032) and receiving post-operative radiotherapy (p = 0.020). Weak and positive tumor cell galectin-3 expression were correlated with increased and decreased tumor invasion, respectively (p = 0.012). Tumor cell expression of galectin-9 showed a trend toward improved survival (p = 0.087). The predominant immune cell type expressing galectin-1, -3 and -9 were CD163+ macrophages. Galectin-1 and -3 were expressed by a minor population of T cells. Galectin-1 was mainly expressed by fibroblasts in the tumor stroma. To conclude, while tumor cell expression of galectin-9 seemed to represent a beneficial response, galectin-1 expression might be used as a marker for a more aggressive anti-cancer treatment.


Assuntos
Biomarcadores Tumorais/metabolismo , Galectinas/metabolismo , Neoplasias do Colo do Útero/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Feminino , Fibroblastos/metabolismo , Galectinas/genética , Humanos , Macrófagos/metabolismo , Pessoa de Meia-Idade , Linfócitos T/metabolismo , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologia
6.
Oncoimmunology ; 4(1): e984539, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25949866

RESUMO

The role of interleukin (IL)-17 in cancer remains controversial. In view of the growing interest in the targeting of IL-17, knowing its cellular sources and clinical implications is crucial. In the present study, we unraveled the phenotype of IL-17 expressing cells in cervical cancer using immunohistochemical double and immunofluorescent triple stainings. In the tumor stroma, IL-17 was found to be predominantly expressed by neutrophils (66%), mast cells (23%), and innate lymphoid cells (8%). Remarkably, T-helper 17 (Th17) cells were a minor IL-17 expressing population (4%). A similar distribution was observed in the tumor epithelium. The Th17 and granulocyte fractions were confirmed in head and neck, ovarian, endometrial, prostate, breast, lung, and colon carcinoma. An above median number of total IL-17 expressing cells was an independent prognostic factor for poor disease-specific survival in early stage disease (p = 0.016). While a high number of neutrophils showed at trend toward poor survival, the lowest quartile of mast cells correlated with poor survival (p = 0.011). IL-17 expressing cells and neutrophils were also correlated with the absence of vaso-invasion (p < 0.01). IL-17 was found to increase cell growth or tightness of cervical cancer cell lines, which may be a mechanism for tumorigenesis in early stage disease. These data suggest that IL-17, primarily expressed by neutrophils, predominantly promotes tumor growth, correlated with poor prognosis in early stage disease. Strikingly, a high number of Th17 cells was an independent prognostic factor for improved survival (p = 0.026), suggesting Th17 cells are part of a tumor suppressing immune response.

7.
Oncoimmunology ; 4(2): e984547, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25949881

RESUMO

Both IL-17 and Th17 cells have been ascribed tumor promoting as well as tumor suppressing functions. We reviewed the literature on correlations between IL-17 versus Th17 cells and survival in human cancer, following the PRISMA guidelines. Serum, formalin-fixed, paraffin-embedded (FFPE) tissue and peripheral blood samples were most frequently studied. High IL-17 quantities were correlated with poor prognosis, whereas high Th17 cell frequencies were correlated with improved prognosis. Since Th17 cells are a subpopulation of IL-17+ cells and had a different correlation with prognosis than total IL-17, we substantiate that a distinction should be made between Th17 and other IL-17+ cells.

8.
Mol Cancer ; 14: 71, 2015 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-25889974

RESUMO

BACKGROUND: The tumour microenvironment comprises a network of immune response and vascularization factors. From this network, we identified immunological and vascularization gene expression clusters and the correlations between the clusters. We subsequently determined which factors were correlated with patient survival in cervical carcinoma. METHODS: The expression of 42 genes was investigated in 52 fresh frozen squamous cervical cancer samples by qRT-PCR. Weighted gene co-expression network analysis and mixed-model analyses were performed to identify gene expression clusters. Correlations and survival analyses were further studied at expression cluster and single gene level. RESULTS: We identified four immune response clusters: 'T cells' (CD3E/CD8A/TBX21/IFNG/FOXP3/IDO1), 'Macrophages' (CD4/CD14/CD163), 'Th2' (IL4/IL5/IL13/IL12) and 'Inflammation' (IL6/IL1B/IL8/IL23/IL10/ARG1) and two vascularization clusters: 'Angiogenesis' (VEGFA/FLT1/ANGPT2/ PGF/ICAM1) and 'Vessel maturation' (PECAM1/VCAM1/ANGPT1/SELE/KDR/LGALS9). The 'T cells' module was correlated with all modules except for 'Inflammation', while 'Inflammation' was most significantly correlated with 'Angiogenesis' (p < 0.001). High expression of the 'T cells' cluster was correlated with earlier TNM stage (p = 0.007). High CD3E expression was correlated with improved disease-specific survival (p = 0.022), while high VEGFA expression was correlated with poor disease-specific survival (p = 0.032). Independent predictors of poor disease-specific survival were IL6 (hazard ratio = 2.3, p = 0.011) and a high IL6/IL17 ratio combined with low IL5 expression (hazard ratio = 4.2, p = 0.010). CONCLUSIONS: 'Inflammation' marker IL6, especially in combination with low levels of IL5 and IL17, was correlated with poor survival. This suggests that IL6 promotes tumour growth, which may be suppressed by a Th17 and Th2 response. Measuring IL6, IL5 and IL17 expression may improve the accuracy of predicting prognosis in cervical cancer.


Assuntos
Carcinoma de Células Escamosas/imunologia , Expressão Gênica/imunologia , Neovascularização Patológica/imunologia , Neoplasias do Colo do Útero/imunologia , Adulto , Idoso , Feminino , Humanos , Inflamação/imunologia , Linfócitos do Interstício Tumoral/imunologia , Macrófagos/imunologia , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Análise de Sobrevida , Linfócitos T Reguladores/imunologia , Microambiente Tumoral/imunologia , Adulto Jovem
9.
Cancer Immunol Immunother ; 64(6): 745-53, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25795131

RESUMO

Cervical adenocarcinoma comprises approximately 15 % of cervical cancer cases. This histological subtype has different characteristics than cervical squamous cell carcinoma, which may influence disease progression. To study whether the infiltration of T cell subpopulations was correlated with cervical adenocarcinoma patient survival, similar to squamous cell carcinoma, the tumor-infiltrating T cells, Tregs, Th17 cells and IL-17(+) cell frequencies were analyzed in a cohort of cervical adenocarcinoma patients (n = 67). Intraepithelial, stromal and total cell frequencies were scored using triple immunofluorescence. The majority of Tregs were present in the tumor stroma, while other T cells and IL-17(+) cells infiltrated the tumor epithelium three times more frequently. A high total number of Tregs were significantly correlated with improved disease-specific and disease-free survival (p = 0.010, p = 0.007). Within the tumor epithelium, a high T cell frequency was significantly correlated with improved disease-free survival (p = 0.034). In particular, a low number of both Tregs and IL-17(+) cells were correlated with poor disease-specific survival (p = 0.007). A low number of Tregs combined with Th17 cells present were also correlated with poor survival (p = 0.018). An increased number of IL-17(+) cells were significantly correlated with the absence of vaso-invasion (p = 0.001), smaller tumor size (p = 0.030) and less infiltration depth (p = 0.021). These results suggest that Tregs and IL-17(+) cells represent a beneficial immune response, whereas Th17 cells might represent a poor response in cervical adenocarcinoma. This contrasts with the correlations described in squamous cell carcinoma, suggesting that the local immune response in cervical adenocarcinoma contributes differently to tumor growth than in squamous cell carcinoma.


Assuntos
Adenocarcinoma/imunologia , Interleucina-17/metabolismo , Neoplasias do Colo do Útero/imunologia , Adenocarcinoma/patologia , Progressão da Doença , Feminino , Fatores de Transcrição Forkhead , Humanos , Estimativa de Kaplan-Meier , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/patologia , Inclusão em Parafina , Prognóstico , Linfócitos T/imunologia , Linfócitos T/patologia , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia , Fixação de Tecidos , Microambiente Tumoral , Neoplasias do Colo do Útero/patologia
10.
Int J Cancer ; 134(9): 2061-73, 2014 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-24136650

RESUMO

Cervical carcinogenesis is a consequence of persistent infection with high-risk human papillomaviruses (HPVs). Recent studies indicate that HPV-transformed cells actively instruct their microenvironment to promote carcinogenesis. Here, we demonstrate that cervical cancer cells activate monocytes to produce their own CCL2 for further monocyte recruitment and reprogram their function during differentiation and maturation to dendritic cells (DCs). Our data show that cervical cancer cells suppress the induction of the chemokine receptor CCR7 in phenotypically mature DCs and impair their migration toward a lymph node homing chemokine, required to initiate adaptive immune responses. We confirmed the presence of CD83(+)CCR7(low) DCs in cancer biopsies. The second factor essential for DC migration, matrix-metalloproteinase MMP-9, which also has vasculogenic and protumorigenic properties, is not suppressed but upregulated in immature as well as mature DCs. We identified interleukin-6 (IL-6) as a crucial cervical cancer cell-derived mediator and nuclear factor kappaB (NF-jB) as the central signaling pathway targeted in DCs. Anti-IL-6 antibodies reverted not only NF-jB inhibition and restored CCR7-dependent migration but also blocked MMP-9 induction. This is the first report demonstrating the dissociation of CCR7 and MMP-9 expression in phenotypically mature CD83(+) DCs by cancer cells. Our results show that cervical cancer cells actively shape the local microenvironment. They induce the accumulation of myeloid cells and skew their function from immune activation to local production of protumorigenic MMP-9. Neutralizing anti-IL-6 antibodies can counteract this functional dysbalance and should therefore be considered for adjuvant cervical cancer therapy.


Assuntos
Células Dendríticas/metabolismo , Interleucina-6/metabolismo , Metaloproteinase 9 da Matriz/biossíntese , Receptores CCR7/metabolismo , Neoplasias do Colo do Útero/metabolismo , Diferenciação Celular , Movimento Celular/fisiologia , Células Dendríticas/citologia , Ensaio de Desvio de Mobilidade Eletroforética , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Imunofluorescência , Humanos , Imuno-Histoquímica , Monócitos/citologia , Monócitos/metabolismo , Infecções por Papillomavirus/complicações , Infecções por Papillomavirus/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Neoplasias do Colo do Útero/patologia , Neoplasias do Colo do Útero/virologia
11.
J Immunol ; 187(3): 1157-65, 2011 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-21709158

RESUMO

Monocytes attracted by tumor-induced chronic inflammation differentiate to APCs, the type of which depends on cues in the local tumor milieu. In this work, we studied the influence of human cervical cancer cells on monocyte differentiation and showed that the majority of cancer cells either hampered monocyte to dendritic cell differentiation or skewed their differentiation toward M2-like macrophages. Blocking studies revealed that M2 differentiation was caused by tumor-produced PGE(2) and IL-6. TGF-ß, IL-10, VEGF, and macrophage colony-stimulating factor did not play a role. Notably, these CD14(+)CD163(+) M2 macrophages were also detected in situ. Activation of cancer cell-induced M2-like macrophages by several TLR-agonists revealed that compared with dendritic cells, these M2 macrophages displayed a tolerogenic phenotype reflected by a lower expression of costimulatory molecules, an altered balance in IL-12p70 and IL-10 production, and a poor capacity to stimulate T cell proliferation and IFN-γ production. Notably, upon cognate interaction with Th1 cells, these tumor-induced M2 macrophages could be switched to activated M1-like macrophages that expressed high levels of costimulatory molecules, produced high amounts of IL-12 and low amounts of IL-10, and acquired the lymphoid homing marker CCR7. The effects of the interaction between M2 macrophages and Th1 cells could partially be mimicked by activation of these APCs via CD40 in the presence of IFN-γ. Our data on the presence, induction, and plasticity of tumor-induced tolerogenic APCs in cervical cancer suggest that tumor-infiltrated Th1 cells can stimulate a tumor-rejecting environment by switching M2 macrophages to classical proinflammatory M1 macrophages.


Assuntos
Dinoprostona/fisiologia , Interleucina-6/fisiologia , Ativação de Macrófagos/imunologia , Macrófagos/imunologia , Proteínas de Neoplasias/fisiologia , Infecções por Papillomavirus/imunologia , Células Th1/imunologia , Neoplasias do Colo do Útero/imunologia , Anticorpos Bloqueadores/farmacologia , Células Apresentadoras de Antígenos/imunologia , Células Apresentadoras de Antígenos/metabolismo , Linhagem Celular Tumoral , Movimento Celular/imunologia , Células Cultivadas , Técnicas de Cocultura , Feminino , Células HeLa , Papillomavirus Humano 16/imunologia , Papillomavirus Humano 18/imunologia , Humanos , Mediadores da Inflamação/antagonistas & inibidores , Mediadores da Inflamação/imunologia , Mediadores da Inflamação/fisiologia , Interleucina-12/biossíntese , Interleucina-6/antagonistas & inibidores , Interleucina-6/imunologia , Macrófagos/classificação , Macrófagos/patologia , Monócitos/imunologia , Monócitos/patologia , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/imunologia , Infecções por Papillomavirus/metabolismo , Infecções por Papillomavirus/patologia , Células Th1/metabolismo , Células Th1/virologia , Neoplasias do Colo do Útero/patologia , Neoplasias do Colo do Útero/prevenção & controle
12.
Mod Pathol ; 24(5): 720-8, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21252859

RESUMO

The epidermal growth factor receptor is overexpressed in 70-90% of cervical cancers. Previously, we have shown that epidermal growth factor receptor overexpression independently predicts poor prognosis in cervical cancer patients, which makes it a potential therapeutic target. The aim of this study was to systematically analyze the molecular mechanism leading to epidermal growth factor receptor overexpression in cervical cancer. All experiments were performed on archival paraffin-embedded material. In 166 cervical cancer patients, cytoplasmic, membrane and phosphorylated epidermal growth factor receptor protein expression were studied in association with patient survival. Membrane epidermal growth factor receptor overexpression was associated with poor disease-specific survival (P=0.027). This association was particularly present in human papillomavirus 16-positive patients (P=0.029). We analyzed whether epidermal growth factor receptor overexpression was caused by gene amplification using fluorescence in situ hybridization. Epidermal growth factor receptor gene copy number was linked to chromosome 7 ploidy, as no gene amplification could be detected when corrected for chromosome 7 centromeric signals. Chromosome 7 aneuploidy was associated with membrane epidermal growth factor receptor overexpression (P=0.013). Additional mutation analysis was performed by sequencing pure, flow-sorted tumor cells, but no mutations were detected. Furthermore, human papillomavirus 16 E5 and E6 oncogene mRNA expression was measured, using quantitative real-time polymerase chain reaction, to determine the association between the human papillomavirus and epidermal growth factor receptor overexpression. High human papillomavirus 16 E5 and E6 mRNA expression were associated with decreased survival (P=0.045 and 0.047, respectively). High human papillomavirus 16 E6 mRNA expression was associated with membrane epidermal growth factor receptor overexpression (P=0.013). This is the first study performed on cancer patient material showing that chromosome 7 aneuploidy and high human papillomavirus 16 E6 mRNA expression lead to membrane epidermal growth factor receptor overexpression in cervical cancer.


Assuntos
Adenocarcinoma/diagnóstico , Adenocarcinoma/genética , Receptores ErbB/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias do Colo do Útero/diagnóstico , Neoplasias do Colo do Útero/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/mortalidade , Aneuploidia , Cromossomos Humanos Par 7 , Estudos de Coortes , Receptores ErbB/metabolismo , Feminino , Dosagem de Genes , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/isolamento & purificação , Humanos , Hibridização In Situ , Mutação , Países Baixos/epidemiologia , Proteínas Oncogênicas Virais/genética , Infecções por Papillomavirus , Prognóstico , RNA Viral/análise , Proteínas Repressoras/genética , Taxa de Sobrevida , Neoplasias do Colo do Útero/metabolismo , Neoplasias do Colo do Útero/mortalidade
13.
Mod Pathol ; 23(12): 1605-15, 2010 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-20729814

RESUMO

Cervical carcinoma is the second most frequent cancer type in women worldwide. Both inflammatory cells and stromal cells are important for tumor progression. Stromal cells produce growth factors and extracellular matrix and provide an adequate environment for angiogenesis. Versican, a member of the extracellular matrix, has been shown to have a role in tumor progression. The aim of this study was to investigate versican expression, and its association with tumor-infiltrating inflammatory cell subsets and with clinicopathological parameters in human cervical cancers. We have studied the expression of versican in 149 cervical cancers using immunohistochemistry and mRNA in situ hybridization. Versican was predominantly expressed in the stroma (myofibroblasts). Using quantitative real-time-PCR, V0 was found to be the most prominent isoform. High stromal versican expression was significantly associated with a low number of tumor-infiltrating T cells (P=0.018) and particularly a low number of CD8-positive T cells (cytotoxic T cells; P=0.002). Stromal versican expression was significantly higher in patients with an infiltration depth >14 mm (P=0.004) and in patients with parametrial invasion (P=0.044). Stromal versican expression was not associated with survival. Our results suggest that versican expression in the stromal compartment of cervical cancers results in reduced numbers of intraepithelial CD8-positive T cells and enhanced local invasion.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Linfócitos do Interstício Tumoral/imunologia , Neoplasias do Colo do Útero/imunologia , Neoplasias do Colo do Útero/patologia , Versicanas/biossíntese , Biomarcadores Tumorais/análise , Feminino , Humanos , Imuno-Histoquímica , Hibridização In Situ , Prognóstico , Isoformas de Proteínas/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Estromais/metabolismo , Células Estromais/patologia , Neoplasias do Colo do Útero/metabolismo
14.
Clin Cancer Res ; 14(7): 2028-35, 2008 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-18381941

RESUMO

PURPOSE: To investigate the effect of intraepithelial tumor-infiltrating lymphocytes (ieTIL) and their ligands expressed by cervical tumor cells on the outcome of cervical cancer patients. EXPERIMENTAL DESIGN: The prognostic value of ieTILs was investigated in 115 cases of cervical cancer. T-cell subsets, CD57(+) cells, and regulatory T cells (Treg) were enumerated. The associations of these different ieTIL subtypes with human leukocyte antigen (HLA) class I and MHC class I chain-related molecule A (MICA) expression were determined in relation to clinical variables and patient survival. RESULTS: Survival analysis showed that a high number of intraepithelial Treg (FoxP3(+)), a low CD8(+)/regulatory T-cell ratio, and a weak HLA-A expression were all associated with worse survival (P=0.034, 0.025, and 0.033, respectively, log-rank test). Further stratification of patient groups based on HLA-A-MICA expression and HLA-A-MICA-CD8(+)/Treg ratio revealed an even poorer survival (P=0.005). In a multivariate Cox analysis, low CD8(+)/Treg ratio (P=0.047), weak HLA-A-MICA expression (P=0.003), and weak HLA-A-MICA expression combined with low CD8(+)/Treg ratio (P=0.002) were all found to be independent unfavorable prognostic predictors in cervical carcinoma (hazard ratios, 2.7, 4.0, and 4.9, respectively). CONCLUSION: Weak HLA-A-MICA expression combined with low CD8(+)/Treg ratio reveals a patient group with the poorest survival in cervical cancer. As a single variable, low CD8(+)/Treg ratio was a significant independent unfavorable prognostic factor.


Assuntos
Biomarcadores Tumorais/análise , Antígenos HLA-A/biossíntese , Antígenos de Histocompatibilidade Classe I/biossíntese , Linfócitos do Interstício Tumoral/imunologia , Neoplasias do Colo do Útero/mortalidade , Adulto , Idoso , Idoso de 80 Anos ou mais , Linfócitos T CD8-Positivos/imunologia , Feminino , Imunofluorescência , Humanos , Estimativa de Kaplan-Meier , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Prognóstico , Linfócitos T Reguladores/imunologia , Neoplasias do Colo do Útero/imunologia , Neoplasias do Colo do Útero/metabolismo
15.
Mod Pathol ; 21(7): 866-75, 2008 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-18425078

RESUMO

Alterations in transforming growth factor-beta signaling, due to a decrease in Smad2 and especially Smad4 expression, has primarily been reported in pancreatic and colorectal cancers, although loss of the chromosomal region 18q21.1, containing the loci of Smad2 and Smad4, is among the most frequent molecular alterations in cervical cancer. The aim of our study was to investigate whether decreased Smad2 and Smad4 protein expression in primary cervical cancers is associated with molecular alterations at 18q21.1, mutations in the functional domains of Smad2 and Smad4 or hypermethylation, and to assess the biological relevance of decreased Smad2 and Smad4 expression. Subsequently, Smad2, Smad4 and p21 protein expression was determined by immunohistochemistry in 117 primary cervical carcinomas, assembled in a tissue array. Smad signaling was shown to be associated with p21 mRNA expression. All the tumors expressed Smad2 or Smad4. Weak cytoplasmic Smad2 or weak cytoplasmic Smad4 expression could not be attributed to loss of heterozygosity at 18q21.1. Despite weak/moderate Smad2 expression and absent nuclear Smad4 expression, the coding regions of the functional MH1 and MH2 domains of Smad2 and Smad4 were unchanged, as assessed by sequence analysis. The Smad4 promoter region was unmethylated in tumor samples with weak/moderate cytoplasmic Smad4 expression. Remarkably, both weak cytoplasmic Smad4 expression and absent nuclear Smad4 expression significantly correlated with poor disease-free (P=0.003 and P=0.003, respectively) and overall 5-year survival (P=0.003 and P=0.010, respectively). Our findings support the hypothesis that Smad4 is a target molecule for functional inactivation in cervical cancer.


Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/metabolismo , Núcleo Celular/metabolismo , Proteína Smad2/metabolismo , Proteína Smad4/metabolismo , Neoplasias do Colo do Útero/metabolismo , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/mortalidade , Carcinoma de Células Escamosas/patologia , Núcleo Celular/patologia , Metilação de DNA , Análise Mutacional de DNA , DNA de Neoplasias/análise , Feminino , Inativação Gênica , Humanos , Histerectomia , Perda de Heterozigosidade , Países Baixos/epidemiologia , Prognóstico , Proteína Smad2/genética , Proteína Smad4/genética , Taxa de Sobrevida , Análise Serial de Tecidos , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/mortalidade , Neoplasias do Colo do Útero/patologia
16.
Mol Immunol ; 45(1): 152-9, 2008 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-17560652

RESUMO

Infection with oncogenic human papillomavirus (HPV) is considered to be the major etiologic event for cervical cancer. Tumor necrosis factor alpha (TNFalpha), a proinflammatory cytokine, may be involved in orchestrating an antitumor immune response against human papillomavirus expressing cervical cancer cells. Hence, loss of TNFalpha could be advantageous for tumor cells to escape immune clearance. The aim of our study was to investigate TNFalpha gene expression and epigenetic characteristics associated with the loss of TNFalpha expression in cervical cancer. To this end, we examined TNFalpha expression, loss of heterozygosity (LOH) at 6p21.3, the locus of TNFalpha, mutational status of the TNFalpha locus, loss of the TNFalpha promoter variant 2 allele and CpG hypermethylation of the TNFalpha promoter. RNA in situ hybridization showed absence of TNFalpha expression in 45% of 63 tumors. LOH occurred in 57% of the tumors and was not concordant with absence of TNFalpha mRNA. No mutations in the TNFalpha gene were identified in 15 cases deficient in TNFalpha expression exhibiting LOH. Furthermore, lack of TNFalpha expression did not correlate with promoter methylation. In conclusion, TNFalpha mRNA expression is absent in nearly half of the cervical tumors analyzed. Neither promoter methylation nor genetic causes for lack of expression were evident.


Assuntos
Cromossomos Humanos Par 6/genética , Metilação de DNA , Perda de Heterozigosidade/genética , Mutação/genética , Regiões Promotoras Genéticas/genética , Fator de Necrose Tumoral alfa/genética , Neoplasias do Colo do Útero/genética , Alelos , Linhagem Celular Tumoral , Ilhas de CpG/genética , Análise Mutacional de DNA , Éxons/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Repetições de Microssatélites/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Neoplasias do Colo do Útero/patologia
17.
Matrix Biol ; 27(3): 267-71, 2008 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18023162

RESUMO

Over-expression of matrilysin (MMP-7) is predominantly associated with epithelial (pre)malignant cells. In the present study MMP-7 expression is also found in endothelial cells in various human cancer types. Endothelial MMP-7 was associated with CD34 and/or CD105 expression. These immunohistochemical data were confirmed by RT-PCR on VEGF-stimulated endothelial cells. In addition, MMP-7 was also identified in sprouting endothelial cells in vitro. The potential clinical relevance of endothelial MMP-7 was assessed for cervical cancer patients by evaluating the association with overall survival. In contrast to MMP-7 in malignant epithelial cells, MMP-7 expression in endothelial cells showed a significant association with poor survival (LR 5.12, P=0.02, n=30). Our data suggest that MMP-7 is involved in tumor angiogenesis, thereby contributing to malignant growth and hence associated with decreased survival.


Assuntos
Carcinoma/enzimologia , Endotélio/enzimologia , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Metaloproteinase 7 da Matriz/biossíntese , Neoplasias do Colo do Útero/enzimologia , Linhagem Celular Tumoral , Feminino , Humanos , Invasividade Neoplásica , Neovascularização Patológica , Esferoides Celulares/patologia , Fatores de Tempo , Distribuição Tecidual , Veias Umbilicais/citologia
18.
BMC Genomics ; 8: 53, 2007 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-17311676

RESUMO

BACKGROUND: Cervical carcinoma develops as a result of multiple genetic alterations. Different studies investigated genomic alterations in cervical cancer mainly by means of metaphase comparative genomic hybridization (mCGH) and microsatellite marker analysis for the detection of loss of heterozygosity (LOH). Currently, high throughput methods such as array comparative genomic hybridization (array CGH), single nucleotide polymorphism array (SNP array) and gene expression arrays are available to study genome-wide alterations. Integration of these 3 platforms allows detection of genomic alterations at high resolution and investigation of an association between copy number changes and expression. RESULTS: Genome-wide copy number and genotype analysis of 10 cervical cancer cell lines by array CGH and SNP array showed highly complex large-scale alterations. A comparison between array CGH and SNP array revealed that the overall concordance in detection of the same areas with copy number alterations (CNA) was above 90%. The use of SNP arrays demonstrated that about 75% of LOH events would not have been found by methods which screen for copy number changes, such as array CGH, since these were LOH events without CNA. Regions frequently targeted by CNA, as determined by array CGH, such as amplification of 5p and 20q, and loss of 8p were confirmed by fluorescent in situ hybridization (FISH). Genome-wide, we did not find a correlation between copy-number and gene expression. At chromosome arm 5p however, 22% of the genes were significantly upregulated in cell lines with amplifications as compared to cell lines without amplifications, as measured by gene expression arrays. For 3 genes, SKP2, ANKH and TRIO, expression differences were confirmed by quantitative real-time PCR (qRT-PCR). CONCLUSION: This study showed that copy number data retrieved from either array CGH or SNP array are comparable and that the integration of genome-wide LOH, copy number and gene expression is useful for the identification of gene specific targets that could be relevant for the development and progression in cervical cancer.


Assuntos
Perda de Heterozigosidade , Hibridização de Ácido Nucleico/métodos , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Polimorfismo de Nucleotídeo Único , Neoplasias do Colo do Útero/genética , Linhagem Celular Tumoral , Cromossomos Humanos Par 3/genética , Cromossomos Humanos Par 8/genética , Feminino , Dosagem de Genes , Genoma Humano , Células HeLa , Humanos , Hibridização in Situ Fluorescente , Reação em Cadeia da Polimerase Via Transcriptase Reversa
19.
Cancer ; 109(3): 556-65, 2007 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-17177206

RESUMO

BACKGROUND: Proinflammatory cytokines are important in modifying the activity, differentiation, and migration of antigen-presenting cells and may influence the survival of cancer patients. The study assessed whether GM-CSF, TNF-alpha, and IL-12, produced by cervical cancer cells, are important for the activity, differentiation, and migration of antigen-presenting cells. METHODS: In 90 patients with cervical carcinoma the number of monocytes/tumor-associated macrophages (TAM), mature dendritic cells (DC), and Langerhans cells (LHC) was determined using immunohistochemistry. An RNA in situ hybridization technique was used to measure the expression level of GM-CSF, TNF-alpha, IL-12p35, and IL-12p40. RESULTS: TAM were detected intraepithelial as well as in the stroma of the tumor. LHC were only detected intraepithelial and mature DC only in the tumor stroma. The number of TAM correlated positively with the number of mature DC. The expression levels of GM-CSF and TNF-alpha correlated positively with the number of TAM and DC. TNF-alpha showed a negative correlation with the number of LHC. A significant correlation between the expression of functional IL-12 (IL-12p40) and stromal TAM was found. The expression of GM-CSF, TNF-alpha, and IL-12p40 did not correlate significantly with disease-free survival. However, high IL-12p40 expression was associated with a favorable cumulative overall survival. CONCLUSIONS: The results suggest that GM-CSF as well as TNF-alpha, produced by cervical carcinoma cells, may play a role in the differentiation of monocytes into mature DC. Furthermore, TNF-alpha may influence the migration of LHC from the tumor.


Assuntos
Células Apresentadoras de Antígenos/imunologia , Diferenciação Celular , Movimento Celular , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Subunidade p40 da Interleucina-12/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Neoplasias do Colo do Útero/imunologia , Adenocarcinoma/imunologia , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Adulto , Idoso , Carcinoma Adenoescamoso/imunologia , Carcinoma Adenoescamoso/metabolismo , Carcinoma Adenoescamoso/patologia , Carcinoma de Células Escamosas/imunologia , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Células Dendríticas/imunologia , Feminino , Humanos , Técnicas Imunoenzimáticas , Hibridização In Situ , Células de Langerhans/imunologia , Macrófagos/imunologia , Pessoa de Meia-Idade , Monócitos/imunologia , Sondas RNA , Taxa de Sobrevida , Neoplasias do Colo do Útero/metabolismo , Neoplasias do Colo do Útero/patologia
20.
Eur J Immunol ; 36(4): 977-84, 2006 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-16525993

RESUMO

Complement (C) regulatory proteins decrease the effectiveness of immunotherapeutic anti-cancer antibodies. Bispecific mAb (bi-mAb) that target a tumor antigen and simultaneously inhibit a C regulator increase the effectiveness of such a therapy. Here we investigated the mechanism by which bi-mAb increase tumor cell lysis. Apart from C-dependent cytotoxicity, C activation can lead to complement receptor 3 (CR3)-dependent cellular cytotoxicity (CR3-DCC) by CR3-positive effector cells in the presence of beta-glucan. Here we show that an anti-Ep-CAM*anti-CD55 bi-mAb induced more than threefold higher CR3-DCC (71%) of human colorectal cancer cells compared with anti-Ep-CAM alone (20%). This CR3-DCC was dependent on the binding of the anti-CD55 arm of tumor-bound anti-Ep-CAM*anti-CD55 bi-mAb to effector cell CD55, CR3 priming by beta-glucan and the presence of iC3b on the target cell. Comparable lysis could be obtained in the absence of iC3b, when CR3 and CD55 were cross-linked on the effector cells, suggesting cooperation between CD55 and CR3 in signal transduction. Tumor cells with low antigen expression were effectively lysed via this mechanism in contrast to direct C-dependent cytotoxicity. These data imply that the effectiveness of mAb immunotherapy can be improved using anti-tumor antigen*anti-CD55 bi-mAb and beta-glucan, thereby initiating CR3-DCC as an additional effector mechanism that is efficient for eradication of tumor cells with lower antigen expression.


Assuntos
Anticorpos Biespecíficos/imunologia , Antígenos de Neoplasias/imunologia , Antígenos CD55/imunologia , Moléculas de Adesão Celular/imunologia , Citotoxicidade Imunológica , Antígeno de Macrófago 1/imunologia , beta-Glucanas/imunologia , Antígenos CD55/metabolismo , Comunicação Celular/imunologia , Linhagem Celular Tumoral , Neoplasias Colorretais/imunologia , Reagentes de Ligações Cruzadas , Molécula de Adesão da Célula Epitelial , Citometria de Fluxo , Humanos , Antígeno de Macrófago 1/metabolismo , beta-Glucanas/metabolismo
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