|
| Status |
Public on Jun 24, 2013 |
| Title |
FG-3019 + gemcitabine treated tumor, biological rep2 |
| Sample type |
RNA |
| |
|
| Source name |
PDA tumor, mouse MH8259, FG-3019 + gemcitabine, day 9
|
| Organism |
Mus musculus |
| Characteristics |
tissue: pancreatic ductal adenocarcinoma
gender: female
strain: 129/SvJae/C57B6
treatment: FG-3019 + gemcitabine
|
| Treatment protocol |
All animal procedures were carried out in accordance with institutional and national guidelines. For matching purposes, LSL-KrasG12D/+;LSL-Trp53R172H/+;Pdx1-Cre (KPC) mice (Hingorani et al. 2005 Cancer Cell 7:469-83.) were screened by high-resolution ultrasound using a Visual Sonics Vevo 2100 System with a MS250, 13-24MHz scanhead. Mice with tumor diameters of 6-9mm were randomized and enrolled into one of four treatment groups: IgG, gemcitabine/IgG, FG-3019, or FG-3019/gemcitabine. Gemcitabine (100mg/kg) was administered every 3-4 days by i.p. injection. Polyclonal human IgG or human monoclonal FG-3019 (30mg/kg) was administered by i.p. injection every 3 days. Tumor volumes were measured on days 4 and 7 by reconstructed 3D ultrasound using the Vevo 2100 software package. Mice were euthanized on day 9, 2h after the final gemcitabine dose, and perfused with PBS. Half of the tumor was fixed with 10% neutral buffered formalin, and the other half was divided into 4-6 pieces, some of which were snap frozen in liquid nitrogen and stored at -80°C, while those to be used for RNA extraction were stored for at least 24hrs in RNAlater (Ambion) at 4°C then snap frozen and stored at -80°C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using the Qiagen RNeasy Universal RNA purification kit per the manufacturer's recommended methods. RNA quantity and integrity were determined using a NanoDrop ND-1000 spectrophotometer and an Agilent 2100 Bioanalyzer and Agilent RNA 6000 Nano assay kit.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared from 250ng total RNA using the Affymetrix GeneChip 3'IVT Express kit (P/N 901229) according the manufacturer's standard protocol.
|
| |
|
| Hybridization protocol |
Following fragmentation, labeled cRNA were hybridized on Affymetrix mouse genome 430A 2.0 microarrays using the Affymetrix GeneChip Hybridization, Wash, and Stain kit (P/N 901241) and standard protocol. Hybridization was performed for 16hrs at 45°C, 60rpm in an Affymetrix GeneChip Hybridization Oven 640 and staining and washing were performed using an Affymetrix GeneChip Fluidics Station 450 using the manufacturer's recommended fluidics script for 430Av2 genechips (FS450_0002).
|
| Scan protocol |
GeneChips were scanned using an Affymetrix GeneChip Scanner 7G. Scanning was performed using AGCC software and resulting CEL files were exported to GCOS v1.4.0.036 to generate CEL files compatible with Agilent GeneSpring GX 7.3.1 analysis software.
|
| Description |
Tumoral gene expression data from FG-3019 + gemcitabine treated mouse MH8259, biological replicate 2
|
| Data processing |
CEL data were preprocessed using the GC content-based Robust Multi-Array (GCRMA) algorithm and imported into GeneSpring GX 7.3.1. Between-group gene expression comparisons were performed on gene data normalized to the median of measurements in the IgG control samples V1-V7 using parametric t-tests without assuming equal variances or correcting for multiple comparisons.
|
| |
|
| Submission date |
Apr 18, 2013 |
| Last update date |
Jun 25, 2013 |
| Contact name |
Mark D. Sternlicht |
| E-mail(s) |
msternlicht@fibrogen.com
|
| Phone |
1-415-978-1496
|
| Fax |
1-415-978-1908
|
| Organization name |
FibroGen, Inc.
|
| Department |
Molecular Biology
|
| Street address |
409 Illinois Street
|
| City |
San Francisco |
| State/province |
CA |
| ZIP/Postal code |
94158 |
| Country |
USA |
| |
|
| Platform ID |
GPL8321 |
| Series (1) |
| GSE46203 |
Transcriptional effects of CTGF inhibition and gemcitabine in the KPC mouse model of pancreatic ductal adenocarcinoma |
|
