In:
Cancer Research, American Association for Cancer Research (AACR), Vol. 76, No. 14_Supplement ( 2016-07-15), p. 653-653
Abstract:
The Myc oncogene is overexpressed in approximately 30% of human breast cancers. Mice carrying the mammary tumor virus LTR/c-myc (MMTV-c-Myc) develop mammary tumors, but display a relatively low rate of metastases to the lung. However, not all cells that express the transgene develop neoplasms indicating that Myc-induced tumorigenesis requires additional cooperating events for full transformation. To identify genes that cooperate with Myc-induced tumorigenesis leading to the acceleration of tumor progression or an increase in metastases in MMTV-Myc transgenic mice, we have employed the PiggyBac (PB) transposon system. The PiggyBac system used in our studies was (developed by Rad et al, Science, 2010) is composed of two elements, the ATP-1 PiggyBac transposon and transposase. ATP-1 transposons are mobile DNA elements that can randomly integrate throughout the genome in the presence of transposase resulting in the activation or loss of gene function. The insertion sites can be identified through sequencing. It has been used successfully for genomic modification of cells in vivo leading to cancer gene discovery. In order to better adapt the PB system for studing mammary gland biology, we have designed a system to express a hyperactive transposase and luciferase specifically in the mammary gland using the MMTV LTR (MMTV-hyTPase-luc). The efficiency of MMTV-hyTPase-luc expression was first examined in vitro by transfection with an ATP-1 containing plasmid using the HC11 mouse mammary epithelial cell line. Selection for integration of ATP1 in the cells using puromycin selection and crystal violet staining, RT-QPCR and luciferase assays confirmed that MMTV-hyTPase-luc led to efficient integration of ATP-1 into the HC11 cells. HC11 cell proliferation further increased in the presence of dexamethasone, a hormone that stimulates MMTV promotor function. The MMTV-HyPB-Luc construct was subsequently used to generate transgenic mice and multiple founder lines are being analyzed for mammary specific transposase expression. The best expressing line will be crossed with MMTV-cMyc mice and mice carrying multiple copies of the ATP1 transposon. Transposon integration sites will be determined from tumors or metastases in mice with accelerated tumor development and/or increased metastases compared to controls. Further molecular characterization and validation of candidate genes will identify novel genes that cooperate with Myc to augment tumorigenesis and metastases. Citation Format: Qiu N. Tinghu, Laura Vera Ramirez, Roland Rad, Jeffrey E. Green. Identification of metastasis promoting genes using the PiggyBac insertional mutagenesis system. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 653.
Type of Medium:
Online Resource
ISSN:
0008-5472
,
1538-7445
DOI:
10.1158/1538-7445.AM2016-653
Language:
English
Publisher:
American Association for Cancer Research (AACR)
Publication Date:
2016
detail.hit.zdb_id:
2036785-5
detail.hit.zdb_id:
1432-1
detail.hit.zdb_id:
410466-3
