RESUMEN
Recurrent disease and treatment-associated chemoresistance are the two main factors accounting for poor clinical outcomes of ovarian cancer (OC) patients. Both can be associated with cancer stem cells (CSCs), which contribute to cancer formation, progression, chemoresistance, and recurrence. Hence, this study investigated whether the expression of known CSC-associated markers ALDH1A, CD44, and CD133 may predict OC patient prognosis. We analyzed their expression in primary epithelial ovarian cancer (EOC) patients using immunohistochemistry and related them to clinicopathological data, including overall survival (OS) and progression-free survival (PFS). Expression of ALDH1A1 was detected in 32%, CD133 in 28%, and CD44 in 33% of cases. While Kaplan-Meier analysis revealed no association of the expression of CD133 and CD44 with PFS and OS, ALDH1A1-positive patients were characterized with both significantly shorter OS (p = 0.00022) and PFS (p = 0.027). Multivariate analysis demonstrated that the expression of ALDH1A1, FIGO stage III-IV, and residual disease after suboptimal debulking or neoadjuvant chemotherapy correlated with shorter OS. The results of this study identify ALDH1A1 as a potential independent prognostic factor of shorter OS and PFS in EOC patients. Therefore, targeting ALDH1A1-positive cancer cells may be a promising therapeutic strategy to influence the disease course and treatment response.
Asunto(s)
Receptores de Hialuranos , Neoplasias Ováricas , Femenino , Humanos , Familia de Aldehído Deshidrogenasa 1/metabolismo , Biomarcadores de Tumor/metabolismo , Carcinoma Epitelial de Ovario/patología , Estudios de Seguimiento , Receptores de Hialuranos/metabolismo , Células Madre Neoplásicas/metabolismo , Neoplasias Ováricas/diagnóstico , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/metabolismo , Pronóstico , Retinal-Deshidrogenasa/metabolismoRESUMEN
Cancer stem cells (CSCs) may contribute to an increased risk of recurrence in ovarian cancer (OC). Further research is needed to identify associations between CSC markers and OC patients' clinical outcomes with greater certainty. If they prove to be correct, in the future, the CSC markers can be used to help predict survival and indicate new therapeutic targets. This study aimed to determine the CSC markers at mRNA and protein levels and their association with clinical presentation, outcome, and risk of recurrence in HGSOC (High-Grade Serous Ovarian Cancer). TCGA (The Cancer Genome Atlas) database with 558 ovarian cancer tumor samples was used for the evaluation of 13 CSC markers (ALDH1A1, CD44, EPCAM, KIT, LGR5, NES, NOTCH3, POU5F1, PROM1, PTTG1, ROR1, SOX9, and THY1). Data on mRNA and protein levels assessed by microarray and mass spectrometry were retrieved from TCGA. Models to predict chemotherapy response and survival were built using multiple variables, including epidemiological data, expression levels, and machine learning methodology. ALDH1A1 and LGR5 mRNA expressions indicated a higher platinum sensitivity (p = 3.50 × 10-3; p = 0.01, respectively). POU5F1 mRNA expression marked platinum-resistant tumors (p = 9.43 × 10-3). CD44 and EPCAM mRNA expression correlated with longer overall survival (OS) (p = 0.043; p = 0.039, respectively). THY1 mRNA and protein levels were associated with worse OS (p = 0.019; p = 0.015, respectively). Disease-free survival (DFS) was positively affected by EPCAM (p = 0.004), LGR5 (p = 0.018), and CD44 (p = 0.012). In the multivariate model based on CSC marker expression, the high-risk group had 9.1 months longer median overall survival than the low-risk group (p < 0.001). ALDH1A1, CD44, EPCAM, LGR5, POU5F1, and THY1 levels in OC may be used as prognostic factors for the primary outcome and help predict the treatment response.
Asunto(s)
Ascomicetos , Neoplasias Ováricas , Humanos , Femenino , Pronóstico , Molécula de Adhesión Celular Epitelial , Relevancia Clínica , Neoplasias Ováricas/genéticaRESUMEN
Ovarian cancer is the most lethal gynecological malignancy. The high mortality results from late diagnosis and the development of drug resistance. Drug resistance results from changes in the expression of different drug-resistance genes that may be regulated miRNA. The main aim of our study was to detect changes in miRNA expression levels in two cisplatin (CIS) and two paclitaxel (PAC)-resistant variants of the A2780 drug-sensitive ovarian cancer cell line-by miRNA microarray. The next goal was to identify miRNAs responsible for the regulation of drug-resistance genes. We observed changes in the expression of 46 miRNA that may be related to drug resistance. The overexpression of miR-125b-5p, miR-99a-5p, miR-296-3p, and miR-887-3p and downregulation of miR-218-5p, miR-221-3p, and miR-222-3p was observed in both CIS-resistant cell lines. In both PAC-resistant cell lines, we observed the upregulation of miR-221-3p, miR-222-3p, and miR-4485, and decreased expression of miR-551b-3p, miR-551b-5p, and miR-218-5p. Analysis of targets suggest that expression of important drug-resistant genes like protein Tyrosine Phosphatase Receptor Type K (PTPRK), receptor tyrosine kinase-EPHA7, Semaphorin 3A (SEMA3A), or the ATP-binding cassette subfamily B member 1 gene (ABCB1) can be regulated by miRNA.
Asunto(s)
Biomarcadores de Tumor , Cisplatino/farmacología , Resistencia a Antineoplásicos/genética , MicroARNs/genética , Paclitaxel/farmacología , Línea Celular Tumoral , Biología Computacional/métodos , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Redes Reguladoras de Genes , Humanos , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Neoplasias Ováricas/patología , Interferencia de ARNRESUMEN
Ovarian cancer is the most common cause of gynecological cancer death. Cancer Stem Cells (CSCs) characterized by drug transporters and extracellular matrix (ECM) molecules expression are responsible for drug resistance development. The goal of our study was to examine the role of aldehyde dehydrogenase 1A1 (ALDH1A1) expression in paclitaxel (PAC) and topotecan (TOP) resistant ovarian cancer cell lines. In both cell lines, we knocked out the ALDH1A1 gene using the CRISPR/Cas9 technique. Additionally, we derived an ALDH1A1 positive TOP-resistant cell line with ALDH1A1 expression in all cells via clonal selection. The effect of ALDH1A1 gene knockout or clonal selection on the expression of ALDH1A1, drug transporters (P-gp and BCRP), and ECM (COL3A1) was determined by Q-PCR, Western blot and immunofluorescence. Using MTT assay, we compared drug resistance in two-dimensional (2D) and three-dimensional (3D) cell culture conditions. We did not observe any effect of ALDH1A1 gene knockout on MDR1/P-gp expression and drug resistance in the PAC-resistant cell line. The knockout of ALDH1A1 in the TOP-resistant cell line resulted in a moderate decrease of BCRP and COL3A1 expression and weakened TOP resistance. The clonal selection of ALDH1A1 cells resulted in very strong downregulation of BCPR and COL3A1 expression and overexpression of MDR1/P-gp. This finally resulted in decreased resistance to TOP but increased resistance to PAC. All spheroids were more resistant than cells growing as monolayers, but the resistance mechanism differs. The spheroids' resistance may result from the presence of cell zones with different proliferation paces, the density of the spheroid, ECM expression, and drug capacity to diffuse into the spheroid.
Asunto(s)
Neoplasias Ováricas , Topotecan , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/genética , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/metabolismo , Aldehído Deshidrogenasa/genética , Aldehído Deshidrogenasa/metabolismo , Familia de Aldehído Deshidrogenasa 1 , Carcinoma Epitelial de Ovario/genética , Línea Celular Tumoral , Resistencia a Antineoplásicos/genética , Femenino , Regulación Neoplásica de la Expresión Génica , Técnicas de Inactivación de Genes , Humanos , Proteínas de Neoplasias/metabolismo , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Paclitaxel/farmacología , Retinal-Deshidrogenasa/genética , Retinal-Deshidrogenasa/metabolismo , Topotecan/farmacologíaRESUMEN
Our goal was to examine the anticancer effects of piperine against the resistant human ovarian cancer cells and to explore the molecular mechanisms responsible for its anticancer effects. Our study used drug-sensitive ovarian cancer cell line W1 and its sublines resistant to paclitaxel (PAC) and topotecan (TOP). We analyzed the cytotoxic effect of piperine and cytostatic drugs using an MTT assay. The impact of piperine on protein expression was determined by immunofluorescence and Western blot. We also examined its effect on cell proliferation and migration. We noticed a different level of piperine resistance between cell lines. Piperine increases the cytotoxic effect of PAC and TOP in drug-resistant cells. We observed an increase in PTPRK expression correlated with decreased pTYR level after piperine treatment and downregulation of P-gp and BCRP expression. We also noted a decrease in COL3A1 and TGFBI expression in investigated cell lines and increased COL3A1 expression in media from W1PR2 cells. The expression of Ki67 protein and cell proliferation rate decreased after piperine treatment. Piperine markedly inhibited W1TR cell migration. Piperine can be considered a potential anticancer agent that can increase chemotherapy effectiveness in cancer patients.
Asunto(s)
Alcaloides/farmacología , Antineoplásicos/farmacología , Benzodioxoles/farmacología , Piperidinas/farmacología , Alcamidas Poliinsaturadas/farmacología , Familia de Aldehído Deshidrogenasa 1/genética , Familia de Aldehído Deshidrogenasa 1/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Colágeno Tipo III/genética , Resistencia a Antineoplásicos/genética , Proteínas de la Matriz Extracelular/genética , Femenino , Humanos , Neoplasias Ováricas/genética , Paclitaxel/farmacología , Fosforilación , Retinal-Deshidrogenasa/genética , Retinal-Deshidrogenasa/metabolismo , Factor de Crecimiento Transformador beta/genéticaRESUMEN
In recent years, peroxisome proliferator-activated receptor-γ (PPARγ) has been intensively studied. Because its activation is often associated with changes in the expression level of various apoptotic genes, many studies have emphasized the role of PPARγ as an important anticancer agent. However, in different types of cancer, different genes are influenced by PPARγ action. Previous studies showed that conjugated linoleic acid (CLA) was able to induce apoptosis, upregulate PPARG gene expression and activate PPARγ protein in certain human cancer cell lines. Moreover, some PPARγ agonists inhibited the growth of human lung cancer cells through the induction of apoptosis. Nevertheless, the impact of CLA on PPARγ mRNA and protein levels in non-small cell lung cancer (NSCLC) cell lines has not been investigated thus far. Therefore, in our study, we analysed the influence of the c9,t11 linoleic acid isomer on the expression of PPARG and other genes involved in the apoptotic response (BCL-2, BAX, and CDKN1A) in two NSCLC cell lines of different histological origin (A549 and Calu-1) and in normal human bronchial epithelial Beas-2B cells. Cells were treated with several doses of c9,t11 CLA, followed by RNA and protein isolation, cDNA synthesis, real-time quantitative PCR (RT-qPCR) and Western blot analysis. We showed that the investigated CLA isomer was able to enhance the expression of PPARγ in the examined cell lines and alter the mRNA and protein levels of genes involved in apoptosis. Fluorescent staining and MMT assay revealed the antiproliferative potential of CLA as well as its ability to activate pathways that lead to cell death.
Asunto(s)
Apoptosis/efectos de los fármacos , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Ácidos Linoleicos Conjugados/farmacología , Neoplasias Pulmonares/metabolismo , PPAR gamma/biosíntesis , Células A549 , Proteínas Reguladoras de la Apoptosis/biosíntesis , Proteínas Reguladoras de la Apoptosis/genética , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , PPAR gamma/genéticaRESUMEN
Ovarian cancer rates the highest mortality among all gynecological malignancies. The main reason for high mortality is the development of drug resistance. It can be related to increased expression of drug transporters and increased expression of extracellular matrix (ECM) proteins. Our foremost aim was to exhibit alterations in the miRNA expression levels in cisplatin (CIS), paclitaxel (PAC), doxorubicin (DOX), and topotecan (TOP)-resistant variants of the W1 sensitive ovarian cancer cell line-using miRNA microarray. The second goal was to identify miRNAs responsible for the regulation of drug-resistant genes. According to our observation, alterations in the expression of 40 miRNAs were present. We could observe that, in at least one drug-resistant cell line, the expression of 21 miRNAs was upregulated and that of 19 miRNAs was downregulated. We identified target genes for 22 miRNAs. Target analysis showed that miRNA regulates key genes responsible for drug resistance. Among others, we observed regulation of the ATP-binding cassette subfamily B member 1 gene (ABCB1) in the paclitaxel-resistant cell line by miR-363 and regulation of the collagen type III alpha 1 chain gene (COL3A1) in the topotekan-resistant cell line by miR-29a.
Asunto(s)
Resistencia a Antineoplásicos/genética , Matriz Extracelular/metabolismo , Regulación Neoplásica de la Expresión Génica , MicroARNs/genética , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Subfamilia B de Transportador de Casetes de Unión a ATP/genética , Línea Celular Tumoral , Colágeno Tipo III/genética , Femenino , Perfilación de la Expresión Génica , Redes Reguladoras de Genes , Humanos , Neoplasias Ováricas/patología , TranscriptomaRESUMEN
Background: Ovarian cancer is the 7th most common cancer and 8th most mortal cancer among woman. The standard treatment includes cytoreduction surgery followed by chemotherapy. Unfortunately, in most cases, after treatment, cancer develops drug resistance. Decreased expression and/or activity of protein phosphatases leads to increased signal transduction and development of drug resistance in cancer cells. Methods: Using sensitive (W1, A2780) and resistant ovarian cancer cell lines, the expression of Protein Tyrosine Phosphatase Receptor Type K (PTPRK) was performed at the mRNA (real-time PCR analysis) and protein level (Western blot, immunofluorescence analysis). The protein expression in ovarian cancer tissues was determined by immunohistochemistry. Results: The results showed a decreased level of PTPRK expression in ovarian cancer cell lines resistant to cisplatin (CIS), paclitaxel (PAC), doxorubicin (DOX), topotecan (TOP), vincristine (VIN) and methotrexate (MTX). Additionally, the lower PTPRK expression was observed in Aldehyde Dehydrogenase 1 Family Member A1 (ALDH1A1) positive cancer stem cells (CSCs) population, suggesting the role of PTPRK downregulation in primary as well as acquired resistance to cytotoxic drugs. Conclusions: These results provide important insights into the role of PTPRK in mechanism leading to drug resistance in ovarian cancer and has raised important questions about the role of imbalance in processes of phosphorylation and dephosphorylation.
Asunto(s)
Aldehído Deshidrogenasa/metabolismo , Regulación hacia Abajo/genética , Resistencia a Antineoplásicos/genética , Regulación Neoplásica de la Expresión Génica , Células Madre Neoplásicas/metabolismo , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Proteínas Tirosina Fosfatasas Clase 2 Similares a Receptores/genética , Familia de Aldehído Deshidrogenasa 1 , Línea Celular Tumoral , Femenino , Humanos , Células Madre Neoplásicas/patología , Neoplasias Ováricas/tratamiento farmacológico , Fosfotirosina/metabolismo , Proteínas Tirosina Fosfatasas Clase 2 Similares a Receptores/metabolismo , Retinal-Deshidrogenasa , Topotecan/farmacología , Topotecan/uso terapéuticoRESUMEN
One of the main obstacles to the effective treatment of ovarian cancer patients continues to be the drug resistance of cancer cells. Osteoblast-Specific Factor 2 (OSF-2, Periostin) is a secreted extracellular matrix protein (ECM) expressed in fibroblasts during bone and teeth development. Expression of OSF-2 has been also related to the progression and drug resistance of different tumors. The present study investigated the role of OSF-2 by evaluating its expression in the primary serous ovarian cancer cell line, sensitive (W1) and resistant to doxorubicin (DOX) (W1DR) and methotrexate (MTX) (W1MR). The OSF-2 transcript (real-time PCR analysis), protein expression in cell lysates and cell culture medium (western blot), and expression of the OSF-2 protein in cell lines (immunofluorescence) were investigated in this study. Increased expression of OSF-2 mRNA was observed in drug-resistant cells and followed by increased protein expression in cell culture media of drug-resistant cell lines. A subpopulation of ALDH1A1-positive cells was noted for W1DR and W1MR cell lines; however, no direct co-expression with OSF-2 was demonstrated. Both drugs induced OSF-2 expression after a short period of exposure of the drug-sensitive cell line to DOX and MTX. The obtained results indicate that OSF-2 expression might be associated with the development of DOX and MTX resistance in the primary serous W1 ovarian cancer cell line.
Asunto(s)
Antineoplásicos/farmacología , Moléculas de Adhesión Celular/genética , Resistencia a Antineoplásicos/genética , Neoplasias Ováricas/genética , Familia de Aldehído Deshidrogenasa 1/genética , Familia de Aldehído Deshidrogenasa 1/metabolismo , Moléculas de Adhesión Celular/metabolismo , Línea Celular Tumoral , Proteínas de la Matriz Extracelular/genética , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Neoplasias Ováricas/metabolismo , Retinal-Deshidrogenasa/genética , Retinal-Deshidrogenasa/metabolismoRESUMEN
A major contributor leading to treatment failure of ovarian cancer patients is the drug resistance of cancer cell. CSCs- (cancer stem cells) and ECM (extracellular matrix)-related models of drug resistance are described as independently occurring in cancer cells. Lysyl oxidase (LOX) is another extracellular protein involved in collagen cross-linking and remodeling of extracellular matrix and has been correlated with tumor progression. The expression of LOX, COL1A2, COL3A1, and ALDH1A1 was performed in sensitive (A2780, W1) and resistant to paclitaxel (PAC) (A2780PR1 and W1PR2) and topotecan (TOP) (W1TR) cell lines at the mRNA (real-time PCR analysis) and protein level (Western blot and immunofluorescence analysis). The ALDH1A1 activity was measured with the ALDEFLUOR test and flow cytometry analysis. The protein expression in ovarian cancer tissues was determined by immunohistochemistry. We observed an increased expression of LOX and collagens in PAC and TOP resistant cell lines. Subpopulations of ALDH1A1 positive and negative cells were also noted for examined cell lines. Additionally, the coexpression of LOX with ALDH1A1 and COL1A2 with ALDH1A1 was observed. The expression of LOX, collagens, and ALDH1A1 was also detected in ovarian cancer lesions. In our study LOX, ALDH1A1 and collagens were found to be coordinately expressed by cells resistant to PAC (LOX, ALDH1A1, and COL1A2) or to TOP (LOX and ALDH1A1). This represents the study where molecules related with CSCs (ALDH1A1) and ECM (LOX, collagens) models of drug resistance are described as occurring simultaneously in ovarian cancer cells treated with PAC and TOP.
Asunto(s)
Aldehído Deshidrogenasa/metabolismo , Colágeno Tipo I/metabolismo , Células Madre Neoplásicas/efectos de los fármacos , Neoplasias Ováricas/metabolismo , Proteína-Lisina 6-Oxidasa/metabolismo , Aldehído Deshidrogenasa/genética , Familia de Aldehído Deshidrogenasa 1 , Línea Celular Tumoral , Colágeno Tipo I/genética , Colágeno Tipo III/genética , Colágeno Tipo III/metabolismo , Resistencia a Antineoplásicos/efectos de los fármacos , Matriz Extracelular/metabolismo , Femenino , Humanos , Neoplasias Ováricas/tratamiento farmacológico , Paclitaxel/farmacología , Cultivo Primario de Células , Proteína-Lisina 6-Oxidasa/genética , Retinal-Deshidrogenasa , Topotecan/farmacologíaRESUMEN
The major cause of ovarian cancer treatment failure in cancer patients is inherent or acquired during treatment drug resistance of cancer. Matrix Gla protein (MGP) is a secreted, non-collagenous extracellular matrix protein involved in inhibition of tissue calcification. Recently, MGP expression was related to cellular differentiation and tumor progression. A detailed MGP expression analysis in sensitive (A2780) and resistant to paclitaxel (PAC) (A2780PR) and topotecan (TOP) (A2780TR) ovarian cancer cell lines and their corresponding media was performed. MGP mRNA level (real time PCR analysis) and protein expression in cell lysates and cell culture medium (Western blot analysis) and protein expression in cancer cells (immunofluorescence analysis) and cancer patient lesions (immunohistochemistry) were determined in this study. We observed increased expression of MGP in PAC and TOP resistant cell lines at both mRNA and protein level. MGP protein was also detected in the corresponding culture media. Finally, we detected expression of MGP protein in ovarian cancer lesions from different histological type of cancer. MGP is an important factor that might contribute to cancer resistance mechanism by augmenting the interaction of cells with ECM components leading to increased resistance of ovarian cancer cells to paclitaxel and topotecan. Expression found in ovarian cancer tissue suggests its possible role in ovarian cancer pathogenesis.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Proteínas de Unión al Calcio/genética , Resistencia a Antineoplásicos , Proteínas de la Matriz Extracelular/genética , Regulación Neoplásica de la Expresión Génica , Neoplasias Ováricas/tratamiento farmacológico , Paclitaxel/farmacología , Inhibidores de Topoisomerasa I/farmacología , Topotecan/farmacología , Proteínas de Unión al Calcio/análisis , Línea Celular Tumoral , Proteínas de la Matriz Extracelular/análisis , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Proteína Gla de la MatrizRESUMEN
BACKGROUND: Nephrotic-range proteinuria as a paraneoplastic syndrome (PNS) is an exceptional presentation, especially in children. It is usually associated with hematologic malignancies. Solid tumors are very rare causes of proteinuria. CASE-DIAGNOSIS/TREATMENT: We present the case of a 7-year-old boy with an extremely rare atypical thymic carcinoid accompanied by nephrotic-range proteinuria as PNS. The kidney biopsy was consistent with minimal change disease (MCD). Tests for a neuroendocrine tumor were performed due to symptoms of hypercortisolemia and an elevated concentration of chromogranin A in the serum. The chest computed tomography revealed a tumor in the anterior mediastinum, which was diagnosed as an atypical thymic carcinoid. A complete resolution of the nephrotic-range proteinuria was observed within 1 week after the first thoracoscopic surgery, with almost complete reduction of the tumor mass. CONCLUSIONS: This extremely rare case shows that MCD can occur as a PNS even in children. Nephrotic-range proteinuria can be a symptom of malignant solid tumor. This case highlights the possibility of secondary causes of MCD in children.
Asunto(s)
Tumor Carcinoide/complicaciones , Síndromes Paraneoplásicos/orina , Proteinuria/etiología , Enfermedades Raras/complicaciones , Neoplasias del Timo/complicaciones , Hormona Adrenocorticotrópica/sangre , Biopsia , Tumor Carcinoide/diagnóstico , Tumor Carcinoide/cirugía , Tumor Carcinoide/orina , Niño , Cromogranina A/orina , Síndrome de Cushing/diagnóstico , Diagnóstico Diferencial , Humanos , Ácido Hidroxiindolacético/orina , Hiperglucemia/etiología , Hipernatremia/etiología , Hipertensión/etiología , Hipopotasemia/etiología , Riñón/patología , Riñón/ultraestructura , Imagen por Resonancia Magnética , Masculino , Microscopía Electrónica , Síndrome Nefrótico/diagnóstico , Síndromes Paraneoplásicos/diagnóstico , Proteinuria/orina , Enfermedades Raras/diagnóstico , Enfermedades Raras/cirugía , Enfermedades Raras/orina , Toracoscopía , Neoplasias del Timo/diagnóstico , Neoplasias del Timo/cirugía , Neoplasias del Timo/orina , Tomografía Computarizada por Rayos XRESUMEN
Ovarian cancer is the most common type of gynecologic cancer. One of the leading causes of high mortality is chemoresistance, developed primarily or during treatment. Different mechanisms of drug resistance appear at the cellular and cancer tissue organization levels. We examined the differences in response to the cytotoxic drugs CIS, MTX, DOX, VIN, PAC, and TOP using 2D (two-dimensional) and 3D (three-dimensional) culture methods. We tested the drug-sensitive ovarian cancer cell line W1 and established resistant cell lines to appropriate cytotoxic drugs. The following qualitative and quantitative methods were used to assess: 1) morphology - inverted microscope and hematoxylin & eosin staining; 2) viability - MTT assay; 3) gene expression - a quantitative polymerase chain reaction; 4) identification of proteins - immunohistochemistry, and immunofluorescence. Our results indicate that the drug-sensitive and drug-resistant cells cultured in 3D conditions exhibit stronger resistance than the cells cultured in 2D conditions. A traditional 2D model shows that drug resistance of cancer cells is caused mainly by changes in the expression of genes encoding ATP-binding cassette transporter proteins, components of the extracellular matrix, "new" established genes related to drug resistance in ovarian cancer cell lines, and universal marker of cancer stem cells. Whereas in a 3D model, the drug resistance in spheroids can be related to other mechanisms such as the structure of the spheroid (dense or loose), the cell type (necrotic, quiescent, proliferating cells), drug concentrations or drug diffusion into the dense cellular/ECM structure.
Asunto(s)
Antineoplásicos , Resistencia a Antineoplásicos , Neoplasias Ováricas , Neoplasias Ováricas/química , Neoplasias Ováricas/metabolismo , Neoplasias Ováricas/patología , Línea Celular Tumoral , Humanos , Femenino , Esferoides Celulares , Técnicas de Cultivo Tridimensional de Células , Antineoplásicos/farmacologíaRESUMEN
BACKGROUND: Inherent or developed during treatment drug resistance is the main reason for the low effectiveness of chemotherapy in ovarian cancer. IFI16 is a cytoplasmic/nuclear protein involved in response to virus's infection and cell cycle arrest associated with the cellular senescence. METHODS: Here we performed a detailed IFI16 expression analysis in ovarian cancer cell lines sensitive (A2780) and resistant to doxorubicin (DOX) (A2780DR1 and A2780DR2) and paclitaxel (PAC) (A2780PR1). IFI16 mRNA level, protein level in the nuclear and cytoplasmic fraction (Western blot analysis), the protein expression in cancer cells and nuclei (immunofluorescence analysis) and cancer patient lesions (immunohistochemistry) were performed in this study. RESULTS: We observed upregulation of IFI16 expression in drug resistant cell lines with dominant cytoplasmic localization in DOX-resistant cell lines and nuclear one in the PAC-resistant cell line. The most abundantly overexpressed isoforms of IFI16 were IFI16A and IFI16C. Finally, an analysis of a histological type of ovarian cancer (immunohistochemistry) showed expression in serous ovarian cancer. CONCLUSIONS: Expression of IFI16 in drug-resistant cell lines suggests its role in drug resistance development in ovarian cancer. Expression in serous ovarian cancer suggests its role in the pathogenesis of this histological type.
Asunto(s)
Neoplasias Ováricas , Carcinoma Epitelial de Ovario , Línea Celular Tumoral , Doxorrubicina/farmacología , Doxorrubicina/uso terapéutico , Resistencia a Antineoplásicos/genética , Femenino , Humanos , Interferón gamma , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Paclitaxel/farmacología , Paclitaxel/uso terapéutico , Fosfoproteínas/metabolismoRESUMEN
Histological lesions of gallbladder were described mainly in older patients with cholelithiasis (CH). The aim of the study was to analyse morphological alterations in gallbladder mucosa and selected clinical data of young patients with CH. The studies were conducted on 57 patients with CH, subjected to cholecystectomy in the years of 2003-2007. In course of the years, 37 respective young patients (below 25 years of age) were operated. The comparative group included twenty 50-year-old patients with gallstones. The inflammatory activity (grading) was evaluated using a semiquantitative scale on HE-stained gallbladders. In either group, women with chronic cholecystitis and multiple gallstones prevailed. Histological alterations in young patients involved absence of evident epithelial metaplasia traits, low number of foamy cells and prevalence of eosinophils in gallbladder mucosa. Even if a similar grading in gallbladder walls was noted in young and older patients, only in the former ones, a higher grading was detected in patients with an acute clinical course of the gallstone disease. The results point also to a potential role of local accumulation of eosinophils in gallbladder mucosa in pathogenesis of CH in young patients.
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Vesícula Biliar/patología , Cálculos Biliares/patología , Membrana Mucosa/patología , Adolescente , Adulto , Colecistectomía , Eosinófilos/patología , Femenino , Vesícula Biliar/cirugía , Cálculos Biliares/cirugía , Humanos , Masculino , Persona de Mediana Edad , Adulto JovenRESUMEN
Ovarian cancer rates the highest mortality among all gynecological malignancies. The main reason for high mortality is the development of drug resistance. It can be related to changes in the expression of many drug resistance genes as well as expression of extracellular matrix proteins and cell density in the tumor. We developed a simple two-dimensional and three-dimensional model of drug sensitive A2780 and resistant to cisplatin and paclitaxel variants of ovarian cancer cell line. Using MTT assay, we compared drug resistance in two-dimensional and three-dimensional cell culture conditions. Real-time polymerase chain reaction analysis was used to compare the expression of drug resistance genes. The expression of proteins in spheroids was determined by immunohistochemistry. We observed a moderate increase in cisplatin resistance and a significant increase in paclitaxel resistance between two-dimensional and three-dimensional cell culture conditions. Our findings show that changes in the expression of drug resistance genes may play a crucial role in the drug resistance of cancer cells in traditional cell culture. On the other hand, the drug resistance in spheroids may result from different mechanisms such as cell density in the spheroid, extracellular matrix proteins expression and drug capacity to diffuse into the spheroid.
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Antineoplásicos/farmacología , Comunicación Celular , Técnicas de Cultivo de Célula , Cisplatino/farmacología , Resistencia a Antineoplásicos/genética , Regulación Neoplásica de la Expresión Génica , Neoplasias Ováricas/tratamiento farmacológico , Paclitaxel/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Femenino , Humanos , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Neoplasias Ováricas/patologíaRESUMEN
Ovarian cancer is the fifth leading cause of cancer-related deaths in women. Its high mortality rate results from lack of adequate and sensitive methods allowing for the detection of the early stages of the disease, as well as low efficiency of the treatment, caused by the cytotoxic drug resistance of cancer cells. Unfortunately, tumours are able to develop new pathways and protective mechanisms that allow them to survive toxic conditions of chemotherapy. Therefore, intensive search for new genes and proteins involved in resistance to cytotoxic drugs is still needed, especially from a clinical point of view. The article presents an overview of the available literature on the role of semaphorin 3A (SEMA3A), protocadherin 9 (PCDH9), and S100 calcium binding protein A3 (S100A3) in carcinogenesis and chemoresistance of various tumors including ovarian cancer. As it turns out, the role of described genes/proteins is not limited only to their native biological activity but they function also as an oncogenic or suppressor factors in the tumor development. Moreover, they can also play an important role in development of drug resistance, as it was shown in ovarian cancer cell lines.
Asunto(s)
Biomarcadores de Tumor , Cadherinas , Resistencia a Antineoplásicos , Neoplasias Ováricas , Proteínas S100 , Semaforina-3A , Antineoplásicos , Biomarcadores de Tumor/análisis , Biomarcadores de Tumor/metabolismo , Cadherinas/análisis , Cadherinas/metabolismo , Femenino , Humanos , Neoplasias/diagnóstico por imagen , Neoplasias/metabolismo , Neoplasias Ováricas/diagnóstico , Neoplasias Ováricas/metabolismo , Protocadherinas , Proteínas S100/análisis , Proteínas S100/metabolismo , Semaforina-3A/análisis , Semaforina-3A/metabolismoRESUMEN
Background: Low effectiveness of chemotherapy in ovarian cancer results from development of drug resistance during treatment. Topotecan (TOP) is a chemotherapeutic drug used in second-line chemotherapy of this cancer. Unfortunately, during treatment cancer can develop diverse cellular and tissue specific mechanisms of resistance to cytotoxic drugs. Methods: We analyzed development of TOP resistance in ovarian cancer cell lines (A2780 and W1). On the base of our previous results where a set of "new genes" with different functions that can be related to TOP-resistance was described hereby we performed detailed analysis of MYOT expression. MYOT mRNA level (real time PCR analysis), protein expression in cell lysates and cell culture medium (western blot analysis) and protein expression in cancer cells (immunofluorescence analysis) were determined in this study. Results: We observed increased expression of MYOT in TOP resistant cell lines at both mRNA and protein level. MYOT, together with extracellular matrix molecules like COL1A2 and COL15A1 were also secreted to corresponding cell culture media. Conclusion: Our results suggest that upregulation of MYOT can be related to TOP resistance in ovarian cancer cell lines.
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INTRODUCTION: The role of Wnt/ ß -catenin signaling pathway in HCV-associated hepatocellular carcinogenesis is still unknown. MATERIAL AND METHODS: This study aimed to perform quantitative analysis of immuno- and hybridocytochemical expression of ß -catenin, E- and N-cadherins and HCV proteins (C, NS3, NS5A) in long-lasting (≥ 20 years) chronic hepatitis C (CH-C) (n = 54), hepatocellular carcinoma (HCC) (n = 61), and control liver samples (n = 8). RESULTS: Typical membranous expression of ß -catenin in the control liver was higher than in the CH-C and HCC (p = 0.06). The mean ß -catenin tissue expression in CH-C was similar to controls, and significantly higher than that of HCC (p = 0.005). E-cadherin expression was lower in CH-C than in the control (p = 0.045) and HCC (p < 0.001). In HCC both ß -catenin and E-cadherin expressions were significantly lower in comparison to controls (p = 0.02, p = 0.001, respectively). Positive correlations were found between ß -catenin and E-cadherin (in CH-C and HCC), ß -catenin and N-cadherin (HCC), E- and N-cadherins expressions (HCC) (p < 0.05 in all cases). In CH-C the positive correlation was demonstrated between NS5A protein and ß -catenin, and between the all HCV proteins (C, NS3, NS5A) and E-cadherin expression (p < 0.05 in all cases). CONCLUSIONS: Alterations in cellular locations of ß -catenin and E-cadherin in CH-C and HCC pointed to structural disturbances in intercellular junctions in the livers and presence of the transcriptionally inactive form of ß -catenin. The reduced expression of E-cadherin in long-lasting CH-C may represent an early indicator of the epithelial-mesenchymal transition. The most important role in modulation of the Wnt/ ß -catenin pathway in vivo is probably played by the NS5A viral protein.
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PURPOSE: The aim of the present study is to determine the expression of LUM in drug-resistant ovarian cancer cell lines. METHODS: Doxorubicin- (DOX), topotecan- (TOP) and vincristine- (VIN) resistant variants of the W1 ovarian cancer cell line were used in this study. We used quantitative real-time polymerase chain reactions to determine LUM mRNA levels. Protein expression was detected using Western blot and immunocytochemistry assays. Protein glycosylation was investigated using PGNase F digestion. Immunohistochemistry assays were used to determine protein expression in ovarian cancer patients. RESULTS: We observed increased expression of LUM in drug-resistant cell lines at both the mRNA and the protein level. The most abundant LUM expression was observed in TOP-resistant cell line. We observed LUM bands that corresponded to different molecular masses, and the most abundant LUM form was the secreted form, which had a mass of 50 kDa. Double immunofluorescence analysis showed co-expression of LUM and COL3A1 as well as the presence of extracellular COL3A1 in the TOP-resistant cell line. Finally, we detected the LUM protein in ovarian cancer tissue. CONCLUSION: The expression of LUM in cytostatic-resistant cell lines suggests its role in drug resistance. The co-expression of LUM and COL3A1 indicates the significance of LUM in collagen fibre assembly. Expression in ovarian cancer tissue suggests that LUM can play a role in ovarian cancer pathogenesis in ways similar to other cancers.