Project description:The environment inside even a small tumor is characterized by total (anoxia) or partial oxygen deprivation, hypoxia. It has been shown that radiotherapy and some conventional chemotherapies may be less effective in hypoxia, and therefore it is important to investigate how different drugs act in different microenvironments. In the associated study we performed a large screening of the effects of 19 clinically used or experimental chemotherapeutic drugs on four different cell lines in conditions of normoxia, hypoxia and anoxia. A panel of 19 commercially available drugs: 5-fluorouracil, acriflavine, bortezomib, cisplatin, digitoxin, digoxin, docetaxel, doxorubicin, etoposide, gemcitabine, irinotecan, melphalan, mitomycin c, rapamycin, sorafenib, thalidomide, tirapazamine, topotecan and vincristine were tested for cytotoxic activity on the cancer cell lines A2780 (ovarian), ACHN (renal), MCF-7 (breast), H69 (SCLC) and U-937 (lymphoma). Parallel aliquots of the cells were grown at different oxygen pressures and after 72 hours of drug exposure viability was measured with the fluorometric microculture cytotoxicity assay (FMCA). Sorafenib, irinotecan and docetaxel were in general more effective in an oxygenated environment, while cisplatin, mitomycin c and tirapazamine were more effective in a low oxygen environment. Surprisingly, hypoxia in H69 and MCF-7 cells mostly rendered higher drug sensitivity. In contrast ACHN appeared more sensitive to hypoxia, giving slower proliferating cells, and consequently, was more resistant to most drugs. Gene expression analysis was performed on MCF-7 cells after 90 hours in either anoxic or hypoxic conditions, and compared to cells grown in a regular cell incubator. The gene expression analysis was performed to validate that the cells were hypoxic/anoxic and showed the characteristic hypoxia response. Microarray based mRNA profiling was used to charactarize cells grown in hypoxia and anoxia. In the associated study we performed a large screening of the effects of 19 clinically used or experimental chemotherapeutic drugs on four different cell lines in conditions of normoxia, hypoxia and anoxia. We fin that hypoxia/anoxia render cancer cells both more resistant and more sensistive, depending of the type of drug used. The gene expression analysis was performed to validate that the cells really were hypoxic/anoxic and showed the characteristic hypoxia response. The cell line used for the gene expression analysis was MCF-7.
Project description:Transcriptional expression data for a bioactive small molecule for mechanism identification. MCF7 cells were treated with either the compound NSC620358 or DMSO (control) for 6h prior to RNA isolation, to identify drug specific gene expression changes.
Project description:Microarray based mRNA profiling was used to charactarize the response to the compound VLX600 in cells grown as spheroids. Cells used was colon cancer cells HCT116 and HCT116HIF1a knock-out. We identify the small molecule VLX600 as a drug that is preferentially active against quiescent cells in colon cancer 3-D microtissues. The anticancer activity is associated with reduced mitochondrial respiration, leading to a bioenergetic catastrophe and tumor cell death. VLX600 shows enhanced cytotoxic activity under conditions of nutrient starvation. Microarray based mRNA profiling was used to charactarize the response to the compound VLX600 in cells grown as spheroids. Cells used was colon cancer cells HCT116 and HCT116HIF1a knock-out cells.
Project description:Microarray based mRNA profiling was used to charactarize and compare the gene expression in cells grown as monolayer or spheroids. Keywords: Expression profiling by array Microarray based mRNA profiling was used to charactarize and compare the gene expression in cells grown as monolayer or spheroids.
Project description:The transcription factor Zinc finger protein 148 (Zfp148) interacts physically with the tumor suppressor p53, but the siginficance of this interaction is not known. We recently showed that knockout of Zfp148 in mice leads to ectopic activation of p53 in tissues and cultured fibroblasts, suggesting that Zfp148 represses p53 activity. Here we hypothesized that targeting Zfp148 would unleash p53 activity and protect against cancer development, and test this idea in the APCMin/+ mouse model of intestinal adenomas. Crypt-enriched tissues were isolated by laser microdissection (PALM) from the small intestines (proximal) of Zfp148gt/+APCMin/+ and Zfp148+/+APCMin/+ mice for RNA extraction and hybridization to Affymetrix microarrays.
Project description:Serglycin proteoglycans contribute to proper storage and secretion of inflammatory mediators in hematopoietic cells. Serglycin is also expressed in cancer cells where increased expression has been linked to poor prognosis. In the present study we report that serglycin proteoglycan is absolutely required for metastasis in the MMTV-PyMT-driven mouse breast cancer model. Serglycin seems to play a role in promoting epithelial to mesenchymal transition, cancer-related inflammation and extravasation. Our results suggest that serglycin and serglycin-dependent mediators are potential drug targets to prevent metastatic disease/dissemination of cancer. 6 total breast tumor samples were analyzed. 3 of SG+/- and 3 of SG-/- tumour tissue. Raw data was normalized using the robust multi-array average (RMA) method. To identify potential serglycin-regulated mediators of metastasis, we performed a microarray expression analysis of RNA isolated from SG+/- and SG-/- breast tumor tissue. The expression analysis identified 672 genes with a significantly altered expression level, at log2 fold >±1,2. Strikingly, only six genes were up-regulated in the SG-/- PyMT+ tumor cells compared to SG+/- PyMT+ tumor cells while 666 were significantly down-regulated.
Project description:Primary cultures of patient tumor cells (PCPTC) were used in a cell-based cytotoxicity screen. Microarray-based mRNA profiling was used to identify the mechanism-of-action for the small molecule VLX 50. MCF7 cells were treated with the PCPTC screening hit VLX 50 or DMSO control for 6h prior to RNA isolation. One sample per treatment. Data were analyzed using both MAS5.0 and RMA.
Project description:The delicate interaction between cancer cells and the surrounding stroma plays an essential role in all stages of tumourigenesis. Despite the significance of this interplay, alterations in protein composition underlying tumour-stroma interactions are largely unknown. The aim of this study was to identify stromal proteins with clinical relevance in non-small cell lung cancer. A list encompassing 203 stromal candidate genes was compiled based on gene expression array data and available literature. The protein expression of these genes in human NSCLC was screened utilising the Human Protein Atlas. Twelve proteins were selected that showed a differential stromal staining pattern. The corresponding antibodies were applied on a tissue microarray, including 190 NSCLC samples, and stromal staining was correlated with clinical parameters. Higher stromal expression of CD99was associated with good prognosis in the univariate (p=0.037) and multivariate (p=0.039) analyses. The association was independent from the total proportion of tumour stroma, the fraction of inflammatory cells, and clinical and pathological parameters like stage, performance status and tumour histology. The prognostic impact of stromal CD99 protein expression was confirmed in an independent cohort of 240 NSCLC patients (p=0.008). Furthermore, double-staining confocal fluorescence microscopy showed that CD99 was expressed in stromal lymphocytes as well as in cancer associated fibroblasts. Based on a comprehensive screening strategy the membrane protein CD99 was identified as a novel stromal factor with clinical relevance. The result supports the concept that stromal properties have a significant impact on tumourigenesis. Tissue from five tumors were compared to corressponding microdissected tissue from the same tissue sample
Project description:Microarray based mRNA profiling was used to charactarize and compare the gene expression in cells grown as monolayer or spheroids. Microarray based mRNA profiling was used to charactarize and compare the gene expression in cells grown as monolayer or spheroids.
Project description:Transcriptional expression data for bioactive small molecules for mechanism identification. Keywords: Expression profiling by array MCF7 cells were treated with either NSC76022 (thaspine) or DMSO control for six hours, to identify compound specific gene regulation.