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1.
BMC Cancer ; 19(1): 959, 2019 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-31619200

RESUMO

BACKGROUND: HER3 mediates drug resistance against epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs), resulting in tumor relapse in lung cancers. Previously, we demonstrated that EGFR induces HER3 overexpression, which facilitates the formation of cancer stem-like tumorspheres. However, the cellular mechanism through which EGFR regulates HER3 expression remains unclear. We hypothesized that EGFR downstream of STAT3 participates in HER3 expression because STAT3 contributes to cancer stemness and survival of EGFR-TKI resistant cancers. METHODS: First, RNAseq was used to uncover potential genes involved in the formation of lung cancer HCC827-derived stem-like tumorspheres. EGFR-positive lung cancer cell lines, including HCC827, A549, and H1975, were individually treated with a panel containing 172 therapeutic agents targeting stem cell-associated genes to search for potential agents that could be applied against EGFR-positive lung cancers. In addition, gene knockdown and RNAseq were used to investigate molecular mechanisms through which STAT3 regulates tumor progression and the survival in lung cancer. RESULTS: BBI608, a STAT3 inhibitor, was a potential therapeutic agent that reduced the cell viability of EGFR-positive lung cancer cell lines. Notably, the inhibitory effects of BBI608 were similar with those associated with YM155, an ILF3 inhibitor. Both compounds reduced G9a-mediated HER3 expression. We also demonstrated that STAT3 upregulated G9a to silence miR-145-5p, which exacerbated HER3 expression in this study. CONCLUSIONS: The present study revealed that BBI608 could eradicate EGFR-positive lung cancers and demonstrated that STAT3 enhanced the expression of HER3 through miR-145-5p repression by G9a, indicating that STAT3 is a reliable therapeutic target against EGFR-TKI-resistant lung cancers.


Assuntos
Antígenos de Histocompatibilidade/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Neoplasias Pulmonares/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Receptor ErbB-3/metabolismo , Fator de Transcrição STAT3/metabolismo , Células A549 , Animais , Benzofuranos/farmacologia , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/metabolismo , Técnicas de Silenciamento de Genes , Antígenos de Histocompatibilidade/genética , Histona-Lisina N-Metiltransferase/genética , Humanos , Imidazóis/farmacologia , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , MicroRNAs/genética , MicroRNAs/metabolismo , Naftoquinonas/farmacologia , Proteínas do Fator Nuclear 90/antagonistas & inibidores , Proteínas do Fator Nuclear 90/genética , Inibidores de Proteínas Quinases/efeitos adversos , Receptor ErbB-3/antagonistas & inibidores , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/genética , Ensaios Antitumorais Modelo de Xenoenxerto
2.
J Clin Med ; 8(9)2019 Sep 12.
Artigo em Inglês | MEDLINE | ID: mdl-31547418

RESUMO

Radiotherapy is commonly used to treat patients with oral squamous cell carcinoma (OSCC), but a subpopulation of OSCC patients shows a poor response to irradiation treatment. Therefore, identifying a biomarker to predict the effectiveness of radiotherapy in OSCC patients is urgently needed. In silico analysis of public databases revealed that upregulation of CHRNA5, the gene encoding nicotinic acetylcholine receptor subunit alpha-5, is extensively detected in primary tumors compared to normal tissues and predicts poor prognosis in OSCC patients. Moreover, CHRNA5 transcript level was causally associated with the effective dose of irradiation in a panel of OSCC cell lines. Artificial silencing of CHRNA5 expression enhanced, but nicotine reduced, the radiosensitivity of OSCC cells. Gene set enrichment analysis demonstrated that the E2F signaling pathway is highly activated in OSCC tissues with high levels of CHRNA5 and in those derived from patients with cancer recurrence after radiotherapy. CHRNA5 knockdown predominantly suppressed E2F activity and decreased the phosphorylation of the Rb protein; however, nicotine treatment dramatically promoted E2F activity and increased Rb phosphorylation, which was mitigated after CHRNA5 knockdown in OSCC cells. Notably, the signature combining increased mRNA levels of CHRNA5 and the E2F signaling gene set was associated with worse recurrence-free survival probability in OSCC patients recorded to be receiving radiotherapy. Our findings suggest that CHRNA5 is not only a useful biomarker for predicting the effectiveness of radiotherapy but also a druggable target to enhance the cancericidal effect of irradiation on OSCC.

3.
J Cell Mol Med ; 23(8): 5692-5704, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31199053

RESUMO

Several biological effects of haem oxygenase (HO)-1, including anti-inflammatory, antiapoptotic and antioxidative properties were reported; however, the role of HO-1 in apoptosis is still unclear. In the presence of stimulation by cobalt protoporphyrin (CoPP), an HO-1 inducer, apoptotic characteristics were observed, including DNA laddering, hypodiploid cells, and cleavages of caspase (Casp)-3 and poly(ADP) ribose polymerase (PARP) proteins in human colon carcinoma COLO205, HCT-15, LOVO and HT-29 cells in serum-free (SF) conditions with increased HO-1, but not heat shock protein 70 (HSP70) or HSP90. The addition of 10% foetal bovine serum (FBS) or 1% bovine serum albumin accordingly inhibited CoPP-induced apoptosis and HO-1 protein expression in human colon cancer cells. CoPP-induced apoptosis of colon cancer cells was prevented by the addition of the pan-caspase inhibitor, Z-VAD-FMK (VAD), and the Casp-3 inhibitor, Z-DEVD-FMK (DEVD). N-Acetyl cysteine inhibited reactive oxygen species-generated H2 O2 -induced cell death with reduced intracellular peroxide production, but did not affect CoPP-induced apoptosis in human colorectal carcinoma (CRC) cells. Two CoPP analogs, ferric protoporphyrin and tin protoporphyrin, did not affect the viability of human CRC cells or HO-1 expression by those cells, and knockdown of HO-1 protein expression by HO-1 small interfering (si)RNA reversed the cytotoxic effect elicited by CoPP. Furthermore, the carbon monoxide (CO) donor, CORM, but not FeSO4 or biliverdin, induced DNA ladders, and cleavage of Casp-3 and PARP proteins in human CRC cells. Increased phosphorylated levels of the endoplasmic reticular (ER) stress proteins, protein kinase R-like ER kinase (PERK), and eukaryotic initiation factor 2α (eIF2α) by CORM and CoPP were identified, and the addition of the PERK inhibitor, GSK2606414, inhibited CORM- and CoPP-induced apoptosis. Increased GRP78 level and formation of the HO-1/GRP78 complex were detected in CORM- and CoPP-treated human CRC cells. A pro-apoptotic role of HO-1 against the viability of human CRC cells via induction of CO and ER stress was firstly demonstrated herein.

4.
Acta Histochem ; 121(5): 628-637, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31133374

RESUMO

The identification of prognostic markers for colorectal cancer (CRC) has important clinical implications. However, the association between meningioma 1 (MN1) expression and clinical outcomes of CRC has not been fully investigated. The aim of this study was to investigate the expression of MN1 in the clinical context of CRC. We first used immunohistochemistry (IHC) staining to examine and compare MN1 expression between multiple human cancer tissues and normal tissues. Initial screening revealed that the expression of MN1 proteins was significantly higher in tumor tissues of the breast, colon, and liver than in normal tissues. In further testing conducted on 59 paired CRC samples, we observed that the expression of MN1 in CRC tissue samples was significantly higher than in adjacent normal tissues. Moreover, high MN1 expression was not significantly associated with clinicopathological characteristics. Kaplan-Meier survival analysis revealed that high expression of MN1 mRNA or MN1 protein was significantly associated with poor CRC prognosis. Furthermore, univariate Cox analysis revealed that a high MN1 score was significantly associated with prognostic factors. Multivariate Cox analysis further indicated that gender, histologic grade, tumor-node-metastasis (TNM) stage, and a high MN1 score were independent factors of overall CRC survival rates. Finally, MN1 and PCNA protein levels were positively correlated, which suggests that MN1 may be involved in the cell proliferation process during CRC formation. Our results, which confirm those of other studies, indicate that (1) high levels of MN1 expression contribute to poor CRC prognosis and (2) MN1 can serve as a novel potential biomarker in predicting the prognosis of CRC patients.


Assuntos
Neoplasias Colorretais/mortalidade , Proteínas Supressoras de Tumor/metabolismo , Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Taxa de Sobrevida , Proteínas Supressoras de Tumor/genética
5.
J Biomed Sci ; 25(1): 60, 2018 Aug 02.
Artigo em Inglês | MEDLINE | ID: mdl-30068339

RESUMO

BACKGROUND: Cancer stem cells are capable of undergoing cell division after surviving cancer therapies, leading to tumor progression and recurrence. Inhibitory agents against cancer stem cells may be therapeutically used for efficiently eradicating tumors. Therefore, the aim of this study was to identify the relevant driver genes that maintain cancer stemness in epidermal growth factor receptor (EGFR)-positive colorectal cancer (CRC) cells and to discover effective therapeutic agents against these genes. METHODS: In this study, EGFR-positive cancer stem-like cells (CSLCs) derived from HCT116 and HT29 cells were used as study models for in vitro inductions. To identify the differential genes that maintain CSLCs, RNAseq analysis was conducted followed by bioinformatics analysis. Moreover, a panel containing 172 therapeutic agents targeting the various pathways of stem cells was used to identify effective therapeutics against CSLCs. RESULTS: RNAseq analysis revealed that 654 and 840 genes were significantly upregulated and downregulated, respectively, in the HCT116 CSLCs. Among these genes, notably, platelet-derived growth factor A (PDGFA) and signal transducer and activator of transcription 3 (STAT3) were relevant according to the cancer pathway analyzed using NetworkAnalyst. Furthermore, therapeutic screening revealed that the agents targeting STAT3 and Wnt signaling pathways were efficient in reducing the cell viabilities of both HCT116 and HT29 cells. Consequently, we discovered that STAT3 inhibition using homoharringtonine and STAT3 knockdown significantly reduced the formation and survival of HT29-derived tumorspheres. We also observed that STAT3 phosphorylation was regulated by epidermal growth factor (EGF) to induce PDGFA and Wnt signaling cascades. CONCLUSIONS: We identified the potential genes involved in tumorsphere formation and survival in selective EGFR-positive CRCs. The results reveal that the EGF-STAT3 signaling pathway promotes and maintains CRC stemness. In addition, a crosstalk between STAT3 and Wnt activates the Wnt/ß-catenin signaling pathway, which is also responsible for cancer stemness. Thus, STAT3 is a putative therapeutic target for CRC treatment.


Assuntos
Neoplasias Colorretais/genética , Receptores ErbB/genética , Células-Tronco Neoplásicas/patologia , Fator de Transcrição STAT3/genética , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/patologia , Detecção Precoce de Câncer , Fator de Crescimento Epidérmico/genética , Regulação Neoplásica da Expressão Gênica , Células HT29 , Humanos , Fosforilação , Fator de Crescimento Derivado de Plaquetas/genética , Análise de Sequência de RNA , Via de Sinalização Wnt
6.
Mol Carcinog ; 57(11): 1588-1598, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30035369

RESUMO

The epidermal growth factor (EGF) receptor (EGFR) overexpressed in many cancers, including lung and head and neck cancers, and is involved in cancer cell progression and survival. PD-L1, increases in tumor cells to evade and inhibit CD8+ T cells, is a clinical immunotherapeutic target. This study investigated the molecular mechanism of EGF on regulating PD-L1 in EGFR-positive cancers and determined potential agents to reduce PD-L1 expression. RNA sequencing (RNAseq) and bioinformatics analysis were performed to determine potential driver genes that regulate PD-L1 in tumor cells-derived tumorspheres which mimicking cancer stem cells. Then, the specific inhibitors targeting EGFR were applied to reduce the expression of PD-L1 in vitro and in vivo. We validated that EGF could induce PD-L1 expression in the selected EGFR-positive cancers. RNAseq results revealed that STAT1 increased as a driver gene in KOSC-3-derived tumorspheres; these data were analyzed using PANTHER followed by NetworkAnalyst. The blockade of EGFR by afatinib resulted in decreased STAT1 and IRF-1 levels, both are transcriptional factors of PD-L1, and disabled the IFNr-STAT1-mediated PD-L1 axis in vitro and in vivo. Moreover, STAT1 knockdown significantly reduced EGF-mediated PD-L1 expression, and ruxolitinib, a JAK1/JAK2 inhibitor, significantly inhibited STAT1 phosphorylation to reduce the IFNr-mediated PD-L1 axis. These results indicate that EGF exacerbates PD-L1 by increasing the protein levels of STAT1 to enforce the IFNr-JAK1/2-mediated signaling axis in selected EGFR-positive cancers. The inhibition of EGFR by afatinib significantly reduced PD-L1 and may be a potential strategy for enhancing immunotherapeutic efficacy.


Assuntos
Antígeno B7-H1/metabolismo , Fator de Crescimento Epidérmico/metabolismo , Receptores ErbB/metabolismo , Regulação Neoplásica da Expressão Gênica , Interferon gama/metabolismo , Neoplasias/genética , Neoplasias/metabolismo , Fator de Transcrição STAT1/genética , Afatinib/farmacologia , Animais , Antígeno B7-H1/antagonistas & inibidores , Biomarcadores , Linhagem Celular Tumoral , Receptores ErbB/antagonistas & inibidores , Humanos , Imunofenotipagem , Masculino , Camundongos , Inibidores de Proteínas Quinases/farmacologia , Fator de Transcrição STAT1/metabolismo
7.
Lung Cancer ; 116: 80-89, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29413056

RESUMO

OBJECTIVES: YM155, an inhibitor of interleukin enhancer-binding factor 3 (ILF3), significantly suppresses cancer stemness property, implying that ILF3 contributes to cell survival of cancer stem cells. However, the molecular function of ILF3 inhibiting cancer stemness remains unclear. This study aimed to uncover the potential function of ILF3 involving in cell survival of epidermal growth factor receptor (EGFR)-positive lung stem-like cancer, and to investigate the potential role to improve the efficacy of anti-EGFR therapeutics. MATERIALS AND METHODS: The association of EGFR and ILF3 in expression and regulations was first investigated in this study. Lung cancer A549 cells with deprivation of ILF3 were created by the gene-knockdown method and then RNAseq was applied to identify the putative genes regulated by ILF3. Meanwhile, HCC827- and A549-derived cancer stem-like cells were used to investigate the role of ILF3 in the formation of cancer stem-like tumorspheres. RESULTS: We found that EGFR induced ILF3 expression, and YM155 reduced EGFR expression. The knockdown of ILF3 reduced not only EGFR expression in mRNA and protein levels, but also cell proliferation in vitro and in vivo, demonstrating that ILF3 may play an important role in contributing to cancer cell survival. Moreover, the knockdown and inhibition of ILF3 by shRNA and YM155, respectively, reduced the formation and survival of HCC827- and A549-derived tumorspheres through inhibiting ErbB3 (HER3) expression, and synergized the therapeutic efficacy of afatinib, a tyrosine kinase inhibitor, against EGFR-positive A549 lung cells. CONCLUSION: This study demonstrated that ILF3 plays an oncogenic like role in maintaining the EGFR-mediated cellular pathway, and can be a therapeutic target to improve the therapeutic efficacy of afatinib. Our results suggested that YM155, an ILF3 inhibitor, has the potential for utilization in cancer therapy against EGFR-positive lung cancers.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Imidazóis/farmacologia , Neoplasias Pulmonares/metabolismo , Naftoquinonas/farmacologia , Células-Tronco Neoplásicas/metabolismo , Proteínas do Fator Nuclear 90/metabolismo , Células A549 , Afatinib/administração & dosagem , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Sinergismo Farmacológico , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/metabolismo , Humanos , Imidazóis/administração & dosagem , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Terapia de Alvo Molecular , Naftoquinonas/administração & dosagem , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Proteínas do Fator Nuclear 90/antagonistas & inibidores , Fosforilação , Inibidores de Proteínas Quinases/administração & dosagem , Distribuição Aleatória , Ensaios Antitumorais Modelo de Xenoenxerto
8.
APMIS ; 125(11): 985-995, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28913972

RESUMO

This study investigated the clinical implications of SETDB1 (also known as KMT1E) in human colon adenocarcinoma. Expression levels of SETDB1 proteins were analyzed by immunohistochemistry staining, and tissue microarrays were used to examine expression profiles in human patients. Our results revealed that SETDB1 protein expression was significantly higher in tumor tissue than in normal tissue for the breast, colon, liver, and lung (p < 0.05). Moreover, an analysis with SurvExpress software suggested that elevated expression of SETDB1 mRNA was significantly associated with the overall survival of colon adenocarcinoma patients (p < 0.05); and additional analysis involving 90 paired samples of colon adenocarcinoma tissue and normal tissue revealed that SETDB1 protein expression was 82% higher in cancerous cells (p < 0.001). High SETDB1 expression was also found to be significantly correlated with histological grade (p = 0.005), TNM stage (p = 0.003), T-class/primary tumor (p = 0.001), and N-class/regional lymph nodes (p = 0.017); and Kaplan-Meier survival curves indicated that SETDB1 protein expression was significantly associated with poor survival. Finally, univariate analysis demonstrated that SETDB1 protein expression was related to TNM stage (p = 0.004) and SETDB1 score (p = 0.001), whereas multivariate analysis showed that the influence of SETDB1 on overall colon adenocarcinoma survival was independent from other risk factors. Taken together, our results suggest that the SETDB1 protein could serve as a clinical prognostic indicator for colon adenocarcinoma.


Assuntos
Adenocarcinoma/diagnóstico , Biomarcadores Tumorais/genética , Neoplasias do Colo/diagnóstico , Metiltransferases de Proteína/genética , Adenocarcinoma/genética , Adenocarcinoma/mortalidade , Adenocarcinoma/patologia , Adulto , Idoso , Neoplasias do Colo/genética , Neoplasias do Colo/mortalidade , Neoplasias do Colo/patologia , Feminino , Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Gradação de Tumores , Estadiamento de Neoplasias , Prognóstico
9.
PLoS One ; 12(8): e0182149, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28787001

RESUMO

Cancer stem cell survival is the leading factor for tumor recurrence after tumor-suppressive treatments. Therefore, specific and efficient inhibitors of cancer stemness must be discovered for reducing tumor recurrence. YM155 has been indicated to significantly reduce stemness-derived tumorsphere formation. However, the pharmaceutical mechanism of YM155 against cancer stemness is unclear. This study investigated the potential mechanism of YM155 against cancer stemness in lung cancer. Tumorspheres derived from epidermal growth factor receptor (EGFR)-mutant HCC827 and EGFR wild-type A549 cells expressing higher cancer stemness markers (CD133, Oct4, and Nanog) were used as cancer stemness models. We observed that EGFR autophosphorylation (Y1068) was higher in HCC827- and A549-derived tumorspheres than in parental cells; this autophosphorylation induced tumorsphere formation by activating G9a-mediated stemness. Notably, YM155 inhibited tumorsphere formation by blocking the autophosphorylation of EGFR and the EGFR-G9a-mediated stemness pathway. The chemical and genetic inhibition of EGFR and G9a revealed the significant role of the EGFR-G9a pathway in maintaining the cancer stemness property. In conclusion, this study not only revealed that EGFR could trigger tumorsphere formation by elevating G9a-mediated stemness but also demonstrated that YM155 could inhibit this formation by simultaneously blocking EGFR autophosphorylation and G9a activity, thus acting as a potent agent against lung cancer stemness.


Assuntos
Antineoplásicos/farmacologia , Receptores ErbB/metabolismo , Antígenos de Histocompatibilidade/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Imidazóis/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Naftoquinonas/farmacologia , Afatinib , Linhagem Celular Tumoral , Avaliação Pré-Clínica de Medicamentos , Receptores ErbB/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/fisiologia , Histona-Lisina N-Metiltransferase/antagonistas & inibidores , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Metilação/efeitos dos fármacos , Fator 3 de Transcrição de Octâmero/metabolismo , Fosforilação/efeitos dos fármacos , Fosforilação/fisiologia , Quinazolinas/farmacologia , RNA Mensageiro/metabolismo
10.
Exp Hematol ; 44(10): 940-6, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27393574

RESUMO

Vaso-occlusive crisis (VOC) is one of the most common complications of sickle cell disease (SCD). Recurrent episodes of VOC may cause irreversible organ damage and early mortality in patients with SCD. Emerging evidence suggests that VOC arises from a complex cascade that involves interactions among multiple blood and endothelial cells in the vasculature. Previous studies have identified αMß2 integrin (Mac-1) as a critical molecule that mediates heterotypic interactions between red blood cells (RBCs) and adherent leukocytes and promotes VOC in SCD mice. Here, we show that RBC-leukocyte interactions are significantly diminished in Mac-1-deficient SCD mice, leading to an improvement of blood flow rates and prolonged survival time in a tumor necrosis factor-alpha and surgical-trauma-induced VOC model. Mac-1-deletion, however, was not sufficient to reduce SCD-related chronic organ damage. Our results thus suggest uncoupled mechanisms between acute VOC benefits and the long-term complications of SCD that should be considered in future clinical trials.


Assuntos
Anemia Falciforme/complicações , Anemia Falciforme/metabolismo , Eritrócitos/metabolismo , Leucócitos/metabolismo , Antígeno de Macrófago 1/metabolismo , Doenças Vasculares/etiologia , Doenças Vasculares/metabolismo , Anemia Falciforme/genética , Anemia Falciforme/mortalidade , Animais , Adesão Celular , Comunicação Celular , Modelos Animais de Doenças , Feminino , Deleção de Genes , Rim/metabolismo , Rim/patologia , Fígado/metabolismo , Fígado/patologia , Antígeno de Macrófago 1/genética , Camundongos , Camundongos Knockout , Baço/metabolismo , Baço/patologia , Doenças Vasculares/patologia
11.
J Biomed Sci ; 23: 18, 2016 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-26822586

RESUMO

BACKGROUND: Hypoxia in tumor niche is one of important factors to start regeneration of blood vessels, leading to increase survival, proliferation, and invasion in cancer cells. Under hypoxia microenvironment, furthermore, steadily increased hypoxia-inducible factor -1α (HIF-1α) is observed, and can increase vascular endothelial growth factor (VEGF) expression and promote angiogenesis. Zinc protoporphyrin (ZnPP), a heme oxygenase-1 (HO-1) inhibitor, is potential to inhibit tumor proliferation and progression. However, the mechanism of ZnPP in inhibition of tumor is not completely clear. We hypothesize that ZnPP may modulate HIF-1α through inhibiting HO-1, and then inhibit angiogenesis and tumor progression. This study aimed to dissect the mechanism of ZnPP in tumor suppression. RESULTS: We observed the amount of VEGF was increased in the sera of the colorectal cancer (CRC) patients (n = 34, p < 0.05). Furthermore, increased VEGF expression was also measured in colorectal cancer cells, HCT-15, culturing under mimicking hypoxic condition. It suggested that hypoxia induced VEGF production from cancer cells. VEGF production was significantly reduced from HCT-15 cells after exposure to HIF-1α inhibitor KC7F2, suggesting that HIF-1α regulated VEGF production. Moreover, we observed that the HO-1 inhibitor ZnPP inhibited the expressions of HIF-1α and VEGF coupled with cell proliferations of HCT-15 cells, suggesting that ZnPP blocked HIF-1α expression, and then inhibited the consequent VEGF production. In the xenograft model, we also observed that the animals exposed to ZnPP displayed much smaller tumor nodules and less degree of angiogenesis with decreased expression of the angiogenesis marker, αvß3 integrin, compared to that in normal control. CONCLUSIONS: This study demonstrated that VEGF level in serum was elevated in the patients with CRC. The HO-1 inhibitor, ZnPP, possessed the properties of anti-tumor agent by decreasing HIF-1α levels, blocking VEGF production, impairing tumor angiogenesis, and inhibiting tumor growth.


Assuntos
Neoplasias Colorretais , Heme Oxigenase-1/antagonistas & inibidores , Hipóxia , Metaloporfirinas/farmacologia , Proteínas de Neoplasias , Neovascularização Patológica/tratamento farmacológico , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Linhagem Celular Tumoral , Neoplasias Colorretais/irrigação sanguínea , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/enzimologia , Heme Oxigenase-1/metabolismo , Humanos , Camundongos , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/metabolismo , Neovascularização Patológica/enzimologia , Neovascularização Patológica/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
12.
Oncotarget ; 6(34): 36139-55, 2015 Nov 03.
Artigo em Inglês | MEDLINE | ID: mdl-26447758

RESUMO

Hypoxic microenvironment is a common situation in solid tumors. Carbonic anhydrase IX (CA9) is one of the reliable cellular biomarkers of hypoxia. The role of CA9 in colorectal cancer (CRC) remains to be clarified. CA9 inhibitor such as sulfonamides is known to block CA9 activation and reduce tumor growth consequently. Here, we aimed to investigate the CA9 expression in serum and tumor from different stages of CRC patients and utilize sulfonamide derivative with indium-111 labeling as a probe for CRC nuclear imaging detection in vivo. The serum CA9 was correlated with the tumor CA9 levels in different stages of CRC patients. Hypoxia increased cell viability and CA9 expression in colorectal cancer HCT-15 cells. Sulfonamide derivative 5-(2-aminoethyl)thiophene-2-sulfonamide (ATS) could bind with CA9 in vitro under hypoxia. Moreover, tumor tissues in HCT-15-induced xenograft mice possessed higher hypoxic fluorescence signal as compared with other organs. We also found that the radioisotope signal of indium-111 labeled ATS, which was utilized for CRC detection in HCT-15-induced xenograft mice, was markedly enhanced in tumors as compared with non-ATS control. Taken together, these findings suggest that CA9 is a potential hypoxic CRC biomarker and measurement of serum CA9 can be as a potential tool for diagnosing CA9 expressions in CRC clinical practice. The radioisotope-labeled sulfonamide derivative (ATS) may be useful to apply in CRC patients for nuclear medicine imaging.


Assuntos
Antígenos de Neoplasias/sangue , Anidrases Carbônicas/sangue , Neoplasias Colorretais/enzimologia , Diagnóstico por Imagem/métodos , Sulfonamidas/química , Animais , Anidrase Carbônica IX , Hipóxia Celular/fisiologia , Linhagem Celular Tumoral , Neoplasias Colorretais/diagnóstico por imagem , Neoplasias Colorretais/patologia , Xenoenxertos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Cintilografia , Sulfonamidas/farmacocinética
13.
Oncotarget ; 5(13): 4868-80, 2014 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-24947902

RESUMO

Colorectal cancer (CRC) is known as a common malignant neoplasm worldwide. The role of EGFR/HER2 in CRC is unclear. Afatinib is an irreversible EGFR/HER2 inhibitor. There were few studies of afatinib on CRC. Here, we investigated the protein levels/expressions of HER2 in sera and tumors from CRC patients and the therapeutic effect of afatinib on HER2-overexpressed CRC in vitro and in vivo. The increased HER2 levels were detected in the collected sera and tumors of patients with CRC. The serological HER2 levels were correlated with the tumor HER2 expressions in patients. Afatinib also inhibited the HER2-positive tumor cell growth and caused apoptosis in HER2-overexpressed human colorectal cancer HCT-15 cells but not in low HER2 expressed human gastric cancer MKN45 cells. In vivo study showed that afatinib reduced tumor growth in HER2-overexpressed xenografts. Moreover, afatinib-encapsulated micelles displayed higher cytotoxic activity in HCT-15 cells and were more effective for tumor growth suppression in HCT-15-induced tumor xenografts than afatinib performance alone. Taken together, these findings suggest that higher serum HER2 levels reflect the higher HER2 contents in tumors of CRC patients, and the improved afatinib-encapsulated micelles possess high therapeutic efficacy in HER2-overexpressed CRC in vitro and in vivo.


Assuntos
Neoplasias Colorretais/tratamento farmacológico , Micelas , Quinazolinas/farmacologia , Receptor ErbB-2/antagonistas & inibidores , Ensaios Antitumorais Modelo de Xenoenxerto , Adulto , Afatinib , Idoso , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Composição de Medicamentos , Feminino , Células Hep G2 , Humanos , Immunoblotting , Células MCF-7 , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Polímeros/química , Quinazolinas/química , Receptor ErbB-2/sangue , Receptor ErbB-2/metabolismo , Carga Tumoral/efeitos dos fármacos
14.
Oncotarget ; 5(2): 473-80, 2014 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-24457622

RESUMO

Neutrophil elastase (NE), a serine protease secreted by neutrophils, contributes to the progression of cancers to enhance tumor invasion and metastasis. It has been well reported that the regions surrounding the colorectal cancerous tissues usually are decorated with increased accumulation or aggregation of neutrophils coupled with a higher deposition/expression of NE. Therefore, we hypothesized that an increased expressional level of NE in patients with colorectal cancer (CRC) may represent as one of putative biomarkers for CRC. The aim of this study was to evaluate and assure our hypothesis by measurements of the expressional level of NE in the sera and tissues from CRC patients. Moreover, we also proposed a potential therapeutic strategy by blocking enzymatic activity of NE using sivelestat to inhibit the progression of tumor developments. The infiltrated numbers of neutrophils from specimen tissues of CRC patients, and the secreted forms of NE in the sera were quantitatively measured and compared. To evaluate the serum NE as one of putative biomarkers of CRC patients, the receiver operating characteristic (ROC) curve was made to determine the cut-off value of NE in sera for assurance of CRC diagnosis. To evaluate NE as therapeutic target for CRC, sivelestat, a NE inhibitor, was used and administrated into the CRC xenografts. NE expression level coupled with tumor volume were measured and compared between the control and sivelestat-treated xenografts. We found that more infiltrated neutrophils and an increased NE expression were detected in the cancerous tissues compared to the normal tissues. The serum NE concentration in CRC patients was statistically higher than that in the healthy controls (0.56 ± 0.08 µg/ml vs. 0.22 ± 0.03 µg/ml) (p<0.05), indicating that serum NE can potentially be a putative marker of CRC. To characterize the role of NE in tumorigenesis, the NE activity was detected in HCT-15-xenografts using in vivo imaging system (IVIS). Compare to normal mice, the amounts of active NE in xenografts are significantly higher than normal control animals. In the therapeutic characterizing studies, we found that sivelestat can inhibit tumor growth in the HCT-15-induced xenografts. This study suggests that NE is not only as a putative diagnostic biomarker of CRC, but also a potential therapeutic target for patients suffered with CRC.


Assuntos
Biomarcadores Tumorais/sangue , Neoplasias Colorretais/enzimologia , Elastase de Leucócito/sangue , Sequência de Aminoácidos , Animais , Neoplasias Colorretais/sangue , Neoplasias Colorretais/tratamento farmacológico , Glicina/análogos & derivados , Glicina/farmacologia , Humanos , Elastase de Leucócito/antagonistas & inibidores , Elastase de Leucócito/biossíntese , Camundongos , Camundongos Nus , Dados de Sequência Molecular , Terapia de Alvo Molecular , Inibidores de Serino Proteinase/farmacologia , Sulfonamidas/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Int J Nanomedicine ; 8: 1385-91, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23630420

RESUMO

Increased expression of cellular membrane bound glucose-regulated protein 78 (GRP78) is considered to be one of the biomarkers for gastric cancers. Therefore, peptides or molecules with specific recognition to GRP78 can act as a guiding probe to direct conjugated imaging agents to localized cancers. Based on this rationale, GRP78-guided polymeric micelles were designed and manufactured for nuclear imaging detection of tumors. Thiolated GRP78 binding peptide (GRP78BP) was first labeled with maleimide-terminated poly(ethylene glycol)- poly(ɛ-caprolactone) and then mixed with diethylenetriaminepentaacetic acid (DTPA)-linked poly(ethylene glycol)-poly(ɛ-caprolactone) to form DTPA/GRP78BP-conjugated micelles. The coupling efficiency of micelles with radioisotope indium-111 ((111)In) was measured and analyzed by instant thin layer chromatography. The coupling efficiency of DTPA-conjugated micelles and DTPA/GRP78BP-conjugated micelles with (111)In was 85% and 93%, respectively. For characterization and trace imaging, the radioisotope (111)In-targeting tumors were detected and imaged in a xenograft murine model using nano single photon emission computed tomography/computed tomography. The results revealed that the radioactive intensity measured in the animals administered with GRP78BP-guided (111)In-labeled micelles was statistically higher than that in animals administered with (111)In-labeled micelles, demonstrating that GRP78BP more than doubled the accumulation of micelles to the tumor tissue (P < 0.05). The results indicate that the gastric cancer biomarker GRP78 is a probing target in the application of nuclear imaging for tumor diagnosis. This novel GRP78BP-guided micelle agent may be applied in clinical practice to complement the histological diagnosis.


Assuntos
Proteínas de Choque Térmico/metabolismo , Radioisótopos de Índio , Micelas , Imagem Molecular/métodos , Compostos Radiofarmacêuticos , Neoplasias Gástricas/diagnóstico por imagem , Animais , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Portadores de Fármacos/química , Portadores de Fármacos/farmacocinética , Xenoenxertos , Humanos , Radioisótopos de Índio/química , Radioisótopos de Índio/farmacocinética , Camundongos , Camundongos Nus , Tamanho da Partícula , Peptídeos/química , Peptídeos/farmacocinética , Cintilografia , Compostos Radiofarmacêuticos/química , Compostos Radiofarmacêuticos/farmacocinética , Neoplasias Gástricas/metabolismo
16.
Proteomics ; 12(15-16): 2584-97, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22778057

RESUMO

The survivals of gastric cancer (GC) patients are associated with early diagnosis and effective treatments. Therefore, it is urgent for the discovery of early GC biomarkers and tumor-targeting therapeutics. The aim of this study was to uncover putative tissue biomarkers of GC using 2D DIGE and then apply one of these specific markers in GC treatment. We found three putative biomarkers of GC with significant differences in expression level compared to adjacent normal tissue, including glucose-regulated protein 78 (GRP78) and glutathione s-transferase pi (GSTpi) with increased expression level, and alpha-1 antitrypsin (A1AT) with reduced expression level. The overexpressed GRP78 was used as a targeted protein for guiding the drugs to tumor cells, leading to more effective treatment for GC xenografts. Our results demonstrated that the designated GRP78-binding peptide based on the sequence, WIFPWIQL, was selectively prone to recognize and bind to GC MKN45 cells in vitro, and also improve the delivery efficiency of polymeric micelles-encapsulated drugs into tumor cells and displayed better therapeutic outcome in experimental animals. This strategy of GRP78-mediated drug targeting system may bring chemotherapeutic drugs with more precise targeting to tumor cells, leading to minimize side effects on patients after chemotherapy.


Assuntos
Proteínas de Choque Térmico/metabolismo , Micelas , Neoplasias Gástricas/diagnóstico , Neoplasias Gástricas/tratamento farmacológico , Eletroforese em Gel Diferencial Bidimensional/métodos , Sequência de Aminoácidos , Animais , Biomarcadores Tumorais/metabolismo , Western Blotting , Linhagem Celular Tumoral , Doxorrubicina/farmacologia , Doxorrubicina/uso terapêutico , Proteínas de Choque Térmico/química , Humanos , Imuno-Histoquímica , Ligantes , Camundongos , Dados de Sequência Molecular , Poliésteres/química , Polietilenoglicóis/química , Reprodutibilidade dos Testes , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Resultado do Tratamento , Ensaios Antitumorais Modelo de Xenoenxerto
17.
Dis Markers ; 32(1): 21-31, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22297599

RESUMO

OBJECTIVE: Human neutrophil peptides (HNPs) -1, -2 and -3 are significantly upregulated and were reported as biomarkers in gastric cancer (GC). However, the tissue location and function of HNPs 1-3 are still unclear in GC, and the spatial distribution of the triad needs to be disclosed. The aims of this study were to investigate the distribution and relationships among HNPs-1, -2 and -3, and assess whether infiltrated neutrophils accumulate in gastric tumor. METHODS: In this study, paired samples (n=33) of the GC tissues and adjacent normal tissues from the same patients were obtained from surgery. Expression of HNPs 1-3 were detected by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). The distributions of the HNPs 1-3 in GC tissues were investigated. After verification of HNPs-1 by immunohistochemistry, infiltrated neutrophils were also detected. Then, an in vitro assay was used to observe the binding capacity and measure the cytotoxic effect of HNPs-1 against AGS cells. RESULTS: Comparing to neighboring normal tissue, expressional level of HNPs 1-3 were significantly higher and their distributions overlapped in cancerous tissues of GC patients with high abundance in the lamina propria, whereas HNPs-1 was identified as the highest major peak. Moreover, HNPs-1, -2 and -3 correlated with each other. Besides, we also observed that increased infiltrated neutrophils accumulating in GC tissues, indicating that a strong positive correlation between HNPs 1-3 and infiltrated neutrophils. In addition, the further investigated demonstrated that the major peptide, HNPs-1, was statistically increased with the advance of tumor development from the early to advanced stage of GC (p< 0.05). Moreover, we also noticed that HNPs-1 with a great binding capacity to GC AGS cells in vitro can inhibit tumor cell growth. CONCLUSIONS: Our results suggest that neutrophil secreted peptides, HNPs 1-3, increased in the GC tissues and could be used as potential biomarkers detected using MALDI-TOF MS, implying that elevated neutrophils may be used as a tumor target for tumor treatment. The binding capacity of HNPs-1 with GC cells implies that tracking molecules conjugated with HNPs-1 could be applied as a specific probe for GC diagnoses.


Assuntos
Biomarcadores Tumorais/análise , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Neoplasias Gástricas/metabolismo , alfa-Defensinas/análise , Idoso , Idoso de 80 Anos ou mais , Estudos de Casos e Controles , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Valores de Referência , Reprodutibilidade dos Testes , Neoplasias Gástricas/patologia , Neoplasias Gástricas/cirurgia , alfa-Defensinas/metabolismo
18.
J Biomed Sci ; 18: 52, 2011 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-21806828

RESUMO

BACKGROUND: Excessive consumption of alcohol contributes to alcoholic liver disease. Fatty liver is the early stage of alcohol-related liver disease. The aim of this study was to search for specific serological biomarkers of alcoholic fatty liver (AFL) compared to healthy controls, non-alcoholic fatty liver (NAFL) and liver fibrosis in a rodent model. METHODS: Serum samples derived from animals with AFL, NAFL, or liver fibrosis were characterized and compared using two-dimensional differential gel electrophoresis. A matrix-assisted laser desorption ionization-time of flight tandem mass spectrometer in conjunction with mascot software was used for protein identification. Subsequently, Western blotting and flexible multi-analyte profiling were used to measure the expressions of the putative biomarkers present in the serum of animals and clinical patients. RESULTS: Eight differential putative biomarkers were identified, and the two most differentiated proteins, including upregulated C-reactive protein (CRP) and downregulated haptoglobin (Hp), were further investigated. Western blotting validated that CRP was dramatically higher in the serum of AFL compared to healthy controls and other animals with liver disease of NAFL or liver fibrosis (p < 0.05). Moreover, we found that CRP and Hp were both lower in liver fibrosis of TAA-induced rats and clinical hepatitis C virus-infected patients. CONCLUSION: The results suggest that increased levels of CRP are an early sign of AFL in rats. The abnormally elevated CRP induced by ethanol can be used as a biomarker to distinguish AFL from normal or otherwise diseased livers.


Assuntos
Proteína C-Reativa/análise , Fígado Gorduroso Alcoólico/sangue , Animais , Biomarcadores/sangue , Western Blotting , Eletroforese em Gel Bidimensional , Fígado Gorduroso/sangue , Perfilação da Expressão Gênica , Cirrose Hepática/sangue , Hepatopatia Gordurosa não Alcoólica , Ratos , Ratos Wistar , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
19.
Blood ; 116(10): 1779-86, 2010 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-20508165

RESUMO

Leukocyte adhesion in the microvasculature influences blood rheology and plays a key role in vaso-occlusive manifestations of sickle cell disease. Notably, polymorphonuclear neutrophils (PMNs) can capture circulating sickle red blood cells (sRBCs) in inflamed venules, leading to critical reduction in blood flow and vaso-occlusion. Recent studies have suggested that E-selectin expression by endothelial cells plays a key role by sending activating signals that lead to the activation of Mac-1 at the leading edge of PMNs, thereby allowing RBC capture. Thus, the inhibition of E-selectin may represent a valuable target in this disease. Here, we have tested the biologic properties of a novel synthetic pan-selectin inhibitor, GMI-1070, with in vitro assays and in a humanized model of sickle cell vaso-occlusion analyzed by intravital microscopy. We have found that GMI-1070 predominantly inhibited E-selectin-mediated adhesion and dramatically inhibited sRBC-leukocyte interactions, leading to improved microcirculatory blood flow and improved survival. These results suggest that GMI-1070 may represent a valuable novel therapeutic intervention for acute sickle cell crises that should be further evaluated in a clinical trial.


Assuntos
Anemia Falciforme/prevenção & controle , Glicolipídeos/farmacologia , Selectinas/metabolismo , Doenças Vasculares/prevenção & controle , Doença Aguda , Anemia Falciforme/metabolismo , Anemia Falciforme/fisiopatologia , Animais , Velocidade do Fluxo Sanguíneo/efeitos dos fármacos , Adesão Celular/efeitos dos fármacos , Linhagem Celular , Selectina E/metabolismo , Eritrócitos/citologia , Eritrócitos/efeitos dos fármacos , Eritrócitos/metabolismo , Feminino , Glicolipídeos/química , Hemodinâmica/efeitos dos fármacos , Humanos , Estimativa de Kaplan-Meier , Selectina L/metabolismo , Migração e Rolagem de Leucócitos/efeitos dos fármacos , Leucócitos/citologia , Leucócitos/efeitos dos fármacos , Leucócitos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Selectina-P/metabolismo , Doenças Vasculares/metabolismo , Doenças Vasculares/fisiopatologia
20.
Nat Med ; 15(4): 384-91, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19305412

RESUMO

Selectins and their ligands mediate leukocyte rolling, allowing interactions with chemokines that lead to integrin activation and arrest. Here we show that E-selectin is crucial for generating a secondary wave of activating signals, transduced specifically by E-selectin ligand-1, that induces polarized, activated alpha(M)beta(2) integrin clusters at the leading edge of crawling neutrophils, allowing capture of circulating erythrocytes or platelets. In a humanized mouse model of sickle cell disease, the capture of erythrocytes by alpha(M)beta(2) microdomains leads to acute lethal vascular occlusions. In a model of transfusion-related acute lung injury, polarized neutrophils capture circulating platelets, resulting in the generation of oxidative species that produce vascular damage and lung injury. Inactivation of E-selectin or alpha(M)beta(2) prevents tissue injury in both inflammatory models, suggesting broad implications of this paradigm in thromboinflammatory diseases. These results indicate that endothelial selectins can influence neutrophil behavior beyond its canonical rolling step through delayed, organ-damaging, polarized activation.


Assuntos
Selectina E/fisiologia , Inflamação/prevenção & controle , Inflamação/fisiopatologia , Leucócitos/fisiologia , Microdomínios da Membrana/fisiologia , Neutrófilos/fisiologia , Animais , Plaquetas/fisiologia , Eritrócitos/fisiologia , Selectina L/genética , Selectina L/fisiologia , Leucócitos/efeitos dos fármacos , Antígeno de Macrófago 1/fisiologia , Camundongos , Microscopia de Fluorescência , Neutrófilos/efeitos dos fármacos , Fator de Necrose Tumoral alfa/farmacologia , Ferimentos e Lesões/fisiopatologia
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