Transcription profiling of normal fibroblasts from patients with breast cancer to compare clinical radiosensitivity
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Overgaard Jens
PROVIDER: S-ECPF-MEXP-821 | biostudies-other |
REPOSITORIES: biostudies-other
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Project description:Gene expression profiles in normal fibroblasts from patients with breast cancer - comparison of clinical radiosensitivity
2007-02-01 | E-MEXP-821 | biostudies-arrayexpress
Project description:Normal-appearing epithelium of cancer patients can harbor occult genetic abnormalities. Data comprehensively comparing gene expression between histologically normal breast epithelium of breast cancer patients and cancer-free controls are limited. The present study compares global gene expression between these groups. We performed microarrays using RNA from microdissected histologically normal terminal ductal-lobular units (TDLU) from 2 groups: (i) cancer normal (CN) (TDLUs adjacent to untreated ER1 breast cancers (n = 14)) and (ii) reduction mammoplasty (RM) (TDLUs of age-matched women without breast disease (n = 15)). Cyber-T identi?ed differentially expressed genes. Quantitative RT-PCR (qRT-PCR), immunohistochemistry (IHC), and comparison to independent microarray data including 6 carcinomas in situ (CIS), validated the results. Gene ontology (GO), UniProt and published literature evaluated gene function. About 127 probesets, corresponding to 105 genes, were differentially expressed between CN and RM (p < 0.0009, corresponding to FDR <0.10). 104/127 (82%) probesets were also differentially expressed between CIS and RM, nearly always (102/104 (98%)) in the same direction as in CN vs. RM. Two-thirds of the 105 genes were implicated previously in carcinogenesis. Overrepresented functional groups included transcription, G-protein coupled and chemokine receptor activity, the MAPK cascade and immediate early genes. Most genes in these categories were under-expressed in CN vs. RM. We conclude that global gene expression abnormalities exist in normal epithelium of breast cancer patients and are also present in early cancers. Thus, cancer-related pathways may be perturbed in normal epithelium. These abnormalities could be markers of disease risk, occult disease, or the tissue’s response to an existing tumor. Experiment Overall Design: 29 samples from histologically normal microdissected breast epithelium are included in this series. 14 samples are from epithelium adjacent to a breast tumor, 15 samples were obtained from patients undergoing reduction mammoplasty without apparent breast cancer
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Project description:The epigenetic influence from microenvironment on mammary epethelial cells was investigated by coculture MCF10A cells with different breast fibroblasts that extracted from either breast cancer tissues or normal breast tissues. Experiment Overall Design: MCF10A cells were cocultured with different breast fibroblasts that extracted from either breast cancer tissues or normal breast tissues. After three weeks coculture, MCF10A cells were sorted and harvested for total RNA extraction. Affymetrix expression microarray was performed to analyse gene expression changes.
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Project description:Background and purposeAdjuvant radiotherapy for cancer can result in severe adverse side effects for normal tissues. In this respect, individuals with anomalies of the ATM (ataxia telangiectasia) protein/gene are of particular interest as they may be at risk of both breast cancer and clinical radiosensitivity. The association of specific ATM gene mutations with these pathologies has been well documented, however, there is uncertainty regarding pathological thresholds for the ATM protein.ResultsSemi-quantitative immuno-blotting provided a reliable and reproducible method to compare levels of the ATM protein for a rare cohort of 20 cancer patients selected on the basis of their severe adverse normal tissue reactions to radiotherapy. We found that 4/12 (33%) of the breast cancer patients with severe adverse normal tissue reactions following radiotherapy had ATM protein levels < 55% compared to the mean for non-reactor controls.ConclusionsATM mutations are generally considered low risk alleles for breast cancer and clinical radiosensitivity. From results reported here we propose a tentative ATM protein threshold of ~55% for high-risk of clinical radiosensitivity for breast cancer patients.
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Project description:Comparison of different culture conditions of the human breast cancer cell line SUM149. We cultured the cells adherent in standard medium in normal cell culture flasks and as mammospheres in MammoCult (stemcell technologies) in ultra low-attachment flasks
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Project description:Stromal fibroblasts influence the behavior of breast epithelial cells. Fibroblasts derived from normal breast (NAF) inhibit epithelial growth, whereas fibroblasts from breast carcinomas (CAF) have less growth inhibitory capacity and can promote epithelial growth. We sought to identify molecules that are differentially expressed in NAF versus CAF and potentially responsible for their different growth regulatory abilities. To determine the contribution of soluble molecules to fibroblast-epithelial interactions, NAF were grown in 3D, transwell or direct contact co-cultures with MCF10AT epithelial cells. NAF suppressed proliferation of MCF10AT in both direct contact and transwell co-cultures, but this suppression was significantly greater in direct co-cultures, indicating involvement of both soluble and contact factors. Gene expression profiling of early passage fibroblast cultures identified 420 genes that were differentially expressed in NAF versus CAF. Of the eight genes selected for validation by real-time PCR, FIBULIN 1, was overexpressed in NAF, and DICKKOPF 1, NEUREGULIN 1, PLASMINOGEN ACTIVATOR INHIBITOR 2, and TISSUE PLASMINOGEN ACTIVATOR were overexpressed in CAF. A higher expression of FIBULIN 1 in normal- than cancer-associated fibroblastic stroma was confirmed by immunohistochemistry of breast tissues. Among breast cancers, stromal expression of Fibulin 1 protein was higher in estrogen receptor alpha-positive cancers and low stromal expression of Fibulin 1 correlated with a higher proliferation of cancer epithelial cells. In conclusion, expression profiling of NAF and CAF cultures identified many genes with potential relevance to fibroblast-epithelial interactions in breast cancer. Furthermore, these early passage fibroblast cultures can be representative of gene expression in stromal fibroblasts in vivo.
| S-EPMC2787925 | biostudies-literature
Project description:Nearly half of all cancers are treated with radiotherapy alone or in combination with other treatments, where damage to normal tissues is a limiting factor for the treatment. Radiotherapy-induced adverse health effects, mostly of importance for cancer patients with long-term survival, may appear during or long time after finishing radiotherapy and depend on the patient's radiosensitivity. Currently, there is no assay available that can reliably predict the individual's response to radiotherapy. We profiled two study sets from breast (n = 29) and head-and-neck cancer patients (n = 74) that included radiosensitive patients and matched radioresistant controls.. We studied 55 single nucleotide polymorphisms (SNPs) in 33 genes by DNA genotyping and 130 circulating proteins by affinity-based plasma proteomics. In both study sets, we discovered several plasma proteins with the predictive power to find radiosensitive patients (adjusted p < 0.05) and validated the two most predictive proteins (THPO and STIM1) by sandwich immunoassays. By integrating genotypic and proteomic data into an analysis model, it was found that the proteins CHIT1, PDGFB, PNKD, RP2, SERPINC1, SLC4A, STIM1, and THPO, as well as the VEGFA gene variant rs69947, predicted radiosensitivity of our breast cancer (AUC = 0.76) and head-and-neck cancer (AUC = 0.89) patients. In conclusion, circulating proteins and a SNP variant of VEGFA suggest that processes such as vascular growth capacity, immune response, DNA repair and oxidative stress/hypoxia may be involved in an individual's risk of experiencing radiation-induced toxicity.
| S-EPMC7140105 | biostudies-literature
Project description:The molecular basis of breast cancer invasion and metastasis is not well understood. Our objective was to analyze transcriptome differences between stromal and epithelial cells in normal breast tissue and invasive breast cancer to define the role stroma plays in invasion. Total RNA was isolated from epithelial and stromal cells that were laser captured from normal breast tissue (n=5) and invasive breast cancer (n=28). Gene expression was measured using Affymetrix U133A 2.0 GeneChips. Differential gene expression was evaluated and compared within a model that accounted for cell type (epithelial [E] versus stromal [S]), diagnosis (cancer [C] versus normal [N]) as well as cell type-diagnosis interactions. Compared to NE, the CE transcriptome was highly enriched with genes in proliferative, motility and ECM ontologies. Differences in CS and NS transcriptomes suggested that the ECM was being remodeled in invasive breast cancer, as genes were over-represented in ECM and proteolysis ontologies. Genes more highly expressed in CS compared to CE were primarily ECM components or were involved in the remodeling of ECM, suggesting that ECM biosynthesis and remodeling were initiated in the tumor stromal compartment. Experiment Overall Design: Breast tissue was collected at surgery from patients undergoing surgical resection for invasive breast cancer (n=28) and reduction mammoplasty (n=5). Tissue was cryopreserved in OCT and serial sections were cut using a cryostat. Sections were stained and epithelial and stromal cells were isolated by laser capture microdissection. Total RNA was isolated and target was prepared using NuGen Ribo-SPIA Ovation Technology. Gene expression was measured using Affymetrix U133A 2.0 GeneChips. Gene expression differences were evaluated using a linear model of the Robust Multichip Average gene expression statistic, a model that accounted for cell type (E and S), patient diagnosis (C and N), as well as cell type-diagnosis interactions. Data were examined to determine false discovery rate and were filtered for fold changes in gene expression and p-value.
| S-ECPF-GEOD-10797 | biostudies-other
Project description:BackgroundCancer-associated fibroblasts (CAFs) constitute the primary constituents of the tumor microenvironment (TME) and exert significant influences on cancer progression. However, adequate comprehension of CAF profiles in breast cancer, as well as the precise mechanisms underlying their promotion of cancer, remains lacking.ObjectivesTo discerns the biological differences between normal fibroblasts (NFs) and CAFs in breast cancer and explore the underlying mechanism.MethodsThree pairs of CAFs and NFs were isolated from breast cancer patients of diverse subtypes who had not undergone prior radiotherapy or chemotherapy. Morphological characteristics of CAFs and NFs were assessed through optical and electron microscopy, their biological attributes were examined using cell counting kits and transwell assays, and their impact on breast cancer cells was simulated using a coculture system. Furthermore, the miRNA profiles of CAFs and NFs were sequenced via an Illumina HiSeq 2500 platform.ResultsCAFs exhibited higher growth rate and motility than NFs and a stronger potential to promote the malignancy of breast cancer cells. RNA sequencing of both NFs and CAFs revealed differentially expressed miRNAs with notable variability among distinct patients within their NFs and CAFs, while the enrichment of the target genes of differentially expressed miRNAs within both GO terms and KEGG pathways demonstrated significant similarity across patients with different profiles.ConclusionCAFs have greater malignancy and higher potential to influence the growth, migration, invasion and chemoresistance of cocultured breast cancer cells than NFs. In addition, the miRNAs that are differentially expressed in CAFs when compared to NFs display substantial variability across patients with distinct breast cancer subtypes, while the enrichment of target genes regulated by these miRNAs, within GO terms and KEGG pathways, remains remarkably consistent among patients with varying profiles.
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Project description:These patients were sensitive to docetaxel treatment, exhibiting less than 25% residual tumor.
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