Abstract 352: Murine model of chordoma: Sonic Hedgehog promoter-driven Cre activation of Brachyury (T) expression induces spinal disk abnormalities and perinatal lethal developmental defects.
Alcorta, DA; Maier, JA; Harfe, BD; Kelley, MJ
Published in: Cancer Research
Introduction: Chordoma is a rare malignancy arising in the axial skeleton from notochord remnants. The T-box transcription factor, Brachyury (T), is associated with familial chordoma, is expressed in over 90% of chordomas, and is a driver of cell growth in cultured chordoma cells. Suppression of T expression or function in vitro results in down-regulation of cell growth pathways and reduction in matrix formation. To further understand the mechanisms of T's oncogenic properties, we sought to generate a murine model of constitutive, notochord-directed T expression.Methods: To produce elevated expression of T in the notochord-derived (ND) cells, we generated a murine transgenic animal in which a cassette (T/GFP) containing the β-actin promoter, a transcriptional stop sequence flanked by LoxP sites (FS), the murine T cDNA, an intragenic ribosomal entry sequence and a GFP cDNA was homologously recombined into the ROSA26 locus. We termed these animals FS-T/GFP mice. FS-T/GFP mice were mated to mice with Cre expression regulated by the developmentally-timed and tissue-specific control of the Sonic Hedgehog promoter (Shh-Cre mice).Results: Excision of the FS sequence by Cre was confirmed to result in constitutive expression of T and GFP in 293 cells prior to transgenic creation, and subsequently in cultured fibroblasts from ear punches of the transgenic animals. Mice containing homozygous FS-T/GFP at ROSA26 were healthy with no apparent physiologic or developmental defects and with the expected Mendelian ratios of sex and genotype. A cross of FS-T/GFP with Shh-Cre mice resulted in T/Cre pups that died perinatally with grossly evident heart, lung and kidney defects. In addition, these animals showed incomplete fusion of the vertebral bodies, and dysmorphic spinal disks. Elevated T protein levels and GFP co-expression in disk cells was demonstrated by immunofluorescent detection in tissue sections.Summary: Constitutive expression of T in ND lineage cells results in alteration of the morphology of intervertebral disks possibly indicative of a proliferative effect of Brachyury on ND cells. Expression of T in “off-target” tissues that express Sonic Hedgehog likely results in perinatal death.Citation Format: David A. Alcorta, Jennifer A. Maier, Brian D. Harfe, Michael J. Kelley. Murine model of chordoma: Sonic Hedgehog promoter-driven Cre activation of Brachyury (T) expression induces spinal disk abnormalities and perinatal lethal developmental defects. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 352. doi:10.1158/1538-7445.AM2013-352Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
