Transcription profiling of Mitotane induced expression changes in adrenocortical carcinoma cell-line NCI-H295
ABSTRACT: Human ACC cell-line NCI-H295 was treated in biological triplicate for 6 hours with 50åµM (16 mg/l) and 100åµM (32 mg/l) mitotane or diluent as control.Genome-wide gene expression analyses were performed using the Affymetrix HG_133_plus2 array to identify the molecular mechanisms underlying mitotane action in an unsupervised fashion.
INSTRUMENT(S): in a humid atmosphere at 37å¡C and 5% CO2
Project description:The actual drug treatment options for adrenocortical carcinoma (ACC) are rather narrow and intensive efforts are going on to find novel effective agents. In our previous functional genomics study, retinoid signaling via the retinoid X receptor (RXR) was detected as a major pathogenic pathway in ACC and we have demonstrated the in vitro activity of 9-cis retinoic acid (9-cisRA) acting via the RXR on NCI-H295R cells and also found that 9-cisRA has antitumoral effects in a small pilot xenograft study. In the present study our aim was to confirm the antitumoral effect of 9-cisRA on adrenocortical cancer in a large xenograft study involving both mitotane and 9-cisRA and their combination. 43 male SCID mice inoculated with NCI-H295R cells were treated in four groups (i. control – corn oil vehicle, ii. 5 mg/kg 9-cisRA, iii. 200 mg/kg mitotane, iv. 5 mg/kg 9-cisRA + 200 mg/kg mitotane) for 28 days. Regular tumor size follow-up, histological and immunohistochemical (Ki-67) analysis, tissue gene expression microarray have been performed. Quantitative real-time-PCR was used for the validation of the microarray results and to detect circulating and tissue microRNAs. To examine the proteome proteomics and Western-blot analysis were executed. We have found that both 9-cisRA and mitotane reduced tumor growth relative to control, but only the combination of the two agents resulted in significant tumor size reduction. The Ki-67 index was significantly reduced in both 9-cisRA and 9-cisRA+mitotane groups. Gene expression analysis revealed 483 genes with significant differences in expression, but only without Benjamini-Hochberg correction (APOA4 and PDE4A were validated). Overall design: 43 SCID mice in four groups: 1) control, 2)mitotane, 3)9-cisRA, 4)combined treated. Treatment time: 28 days. Per os treatment
Project description:NCI-60 cancer cell lines were profiled with their genome-wide gene expression patterns using Affymetrix HG-U133A chips. Experiment Overall Design: Total RNA sample of each of the NCI-60 cell lines was obtained before the treatment of any anticancer compound.
Project description:Genome variation profiling of lung adenocarcinoma cells comparing untreated NCI-H1975 cells with CNX-2006-resistant untreated cells. Goal was to determine the potential mechanism of resistance to mutant EGFR-TKIs and rationally design novel strategies for the treatment of EGFR-mutant lung cancer patients. Two-condition experiment: NCI-H1975 parental cells vs CNX-2006-resistant cells. Pooled DNA from healthy volunteers was used as reference.
Project description:Current approaches to the preclinical investigation for novel cancer therapies rely heavily on the use of traditional cancer cell lines maintained in serum-containing conditions. The discrepancy between promising preclinical efficacy and clinical outcome of most novel anticancer agents emphasizes a need for developing predictive preclinical models that preserve the integrity of original patient tumors, including cancer stem cell (CSC) compartment. In this study, we isolate and characterize CSCs from a non-small cell lung cancer cell line, NCI-H1299, by selectively propagating the cells in a stem-cell culture condition. Isolated CSCs proliferated as nonadherent spheroids, displayed capacity to differentiate and self-renew and exhibited higher tumorigenic potential compared to the parental cells. The gene expression profiles of NCI-H1299 parental cells (serum-containing medium), CSCs (stem-cell medium), and xenograft tumor derived from both cell types were studied by Affymetrix array analysis
Project description:To clarify the downstream signal pathway of EML4-ALK in NSCLC, we performed Affymetrix GeneChip analysis using ALK inhibitor CH5424802-treated NCI-H2228 xenograft tumors, and comprehensively characterized the gene expression regulated by inhibition of activated ALK. Mice bearing NCI-H2228 cells were orally administered at dose of 0 (vehicle), 4 or 20 mg/kg of CH5424802, and the tumors were collected and lysed at 6 hours post-dose. Total RNA were extracted from xenograft tumors.
Project description:To clarify the downstream signal pathway of EML4-ALK in NSCLC, we performed Affymetrix GeneChip analysis using ALK inhibitor CH5424802-treated NCI-H2228 xenograft tumors, and comprehensively characterized the gene expression regulated by inhibition of activated ALK. Overall design: Mice bearing NCI-H2228 cells were orally administered at dose of 0 (vehicle), 4 or 20 mg/kg of CH5424802, and the tumors were collected and lysed at 6 hours post-dose. Total RNA were extracted from xenograft tumors.
Project description:Background: Adrenal myelolipoma (AML) is a relatively common and invariably benign tumor composed of adipose tissue and hematopoietic elements. Due to the variable proportion of fat and hematopoietic elements, it is sometimes challenging to differentiate AML from adrenocortical carcinoma (ACC). MicroRNAs have been identified as promising biomarkers in many tumors, including adrenocortical neoplasms, but the microRNA expression of adrenal myelolipoma has not been investigated, yet. Aims: To perform a large scale microRNA expression profiling in adrenal myelolipoma, benign and malignant adrenocortical tumors to identify potential microRNA biomarkers. Methods: Next-generation sequencing (NGS) on 30 formalin-fixed paraffin-embedded archived tissue samples (discovery cohort: 10 adrenocortical adenoma (ACA), 10 ACC and 10 myelolipoma) was performed by Illumina MiSeq. Significantly differentially expressed microRNAs were validated by real-time RT-qPCR in an independent validation cohort comprised of 10 ACA, 10 myelolipoma and 9 ACC samples. Results: We have found relative overexpression of miR- 451a, miR-486-5p, miR-363-3p and miR-150-5p in myelolipoma compared to the other two tumor groups by NGS. For ACC, we have found up-regulation of miR-184, miR-483-5p, miR-431-5p and miR-183-5p compared to myelolipoma and ACA. Validation by RT-qPCR, confirmed significant overexpression of miR-451a, miR-486-5p and miR-150-5p in myelolipomas relative to ACA and ACC, whereas significant overexpression of miR-184 and miR-183-5p was confirmed in ACC relative to the other 2 groups. The overexpression of miR-483-5p has not turned out to be significant in ACC relative to myelolipomas in the validation cohort. Conclusions: Overexpressed miR-451a, miR-486-5p and miR-150-5p might be potential tissue markers of adrenal myelolipoma. The lack of significance in the expression of miR-483-5p between ACC and myelolipoma is remarkable, as miR-483-5p has been considered to be the best marker of adrenal malignancy to date. Overall design: Altogether 30 samples were investigated by high-throughput microRNA expression profiling
Project description:Further to our previous study (E-MTAB-5997), here we performed transcriptome profiling on Anlotinib-resistant NCI-H1975 and Anlotinib-treated Anlotinib-resistant NCI-H1975, and would like to understand the effects of Anlotinib on Anlotinib-resistant NCI-H1975 cell, compare the different transcriptome profiling on NCI-H1975 cells and Anlotinib-resistant NCI-H1975 cells, sought to find the biomarker for explaining Anlotinib resistance.