title: Int6 reduction activates stromal fibroblasts to enhance transforming activity in breast epithelial cells.
abstract: BACKGROUND
  The INT6 gene was first discovered as a site of integration in mouse mammary tumors by the mouse mammary tumor virus; however, INT6's role in the development of human breast cancer remains largely unknown. By gene silencing, we have previously shown that repressing INT6 promotes transforming activity in untransformed human mammary epithelial cells. In the present study, guided by microarray data of human tumors, we have discovered a role of Int6 in stromal fibroblasts.
  
  
  RESULTS
  We searched microarray databases of human tumors to assess Int6's role in breast cancer. While INT6 expression levels, as expected, were lower in breast tumors than in adjacent normal breast tissue samples, INT6 expression levels were also substantially lower in tumor stroma. By immunohistochemistry, we determined that the low levels of INT6 mRNA observed in the microarray databases most likely occurs in stromal fibroblasts, because far fewer fibroblasts in the tumor tissue showed detectable levels of the Int6 protein. To directly investigate the effects of Int6 repression on fibroblasts, we silenced INT6 expression in immortalized human mammary fibroblasts (HMFs). When these INT6-repressed HMFs were co-cultured with breast cancer cells, the abilities of the latter to form colonies in soft agar and to invade were enhanced. We analyzed INT6-repressed HMFs and found an increase in the levels of a key carcinoma-associated fibroblast (CAF) marker, smooth muscle actin. Furthermore, like CAFs, these INT6-repressed HMFs secreted more stromal cell-derived factor 1 (SDF-1), and the addition of an SDF-1 antagonist attenuated the INT6-repressed HMFs' ability to enhance soft agar colony formation when co-cultured with cancer cells. These INT6-repressed HMFs also expressed high levels of mesenchymal markers such as vimentin and N-cadherin. Intriguingly, when mesenchymal stem cells (MSCs) were induced to form CAFs, Int6 levels were reduced.
  
  
  CONCLUSIONS
  These data suggest that besides enhancing transforming activity in epithelial cells, INT6 repression can also induce fibroblasts, and possibly MSCs as well, via mesenchymal-mesenchymal transitions to promote the formation of CAFs, leading to a proinvasive microenvironment for tumorigenesis.
authors.affiliation: Department of Clinical Cancer Prevention, The University of Texas MD Anderson Cancer Center, Houston, 77030 TX USA ; Department of Molecular and Cellular Biology and Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, 77030 TX USA.
authors.forename: Jinfeng
authors.initials: J
authors.lastname: Suo
authors.affiliation: Department of Molecular and Cellular Biology and Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, 77030 TX USA.
authors.forename: Daniel
authors.initials: D
authors.lastname: Medina
authors.affiliation: Department of Molecular and Cellular Biology and Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, 77030 TX USA.
authors.forename: Sabrina
authors.initials: S
authors.lastname: Herrera
authors.affiliation: Department of Molecular and Cellular Biology and Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, 77030 TX USA.
authors.forename: Ze-Yi
authors.initials: ZY
authors.lastname: Zheng
authors.affiliation: Department of Molecular and Cellular Biology and Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, 77030 TX USA.
authors.forename: Lei
authors.initials: L
authors.lastname: Jin
authors.affiliation: Department of Molecular and Cellular Biology and Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, 77030 TX USA.
authors.forename: Gary C
authors.initials: GC
authors.lastname: Chamness
authors.affiliation: Department of Molecular and Cellular Biology and Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, 77030 TX USA.
authors.forename: Alejandro
authors.initials: A
authors.lastname: Contreras
authors.affiliation: Department of Molecular and Cellular Biology and Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, 77030 TX USA.
authors.forename: Carolina
authors.initials: C
authors.lastname: Gutierrez
authors.affiliation: Department of Molecular and Cellular Biology and Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, 77030 TX USA.
authors.forename: Susan
authors.initials: S
authors.lastname: Hilsenbeck
authors.affiliation: Department of Medical Oncology, Erasmus MC Cancer Institute, Erasmus University Medical Center, Rotterdam, Netherlands.
authors.forename: Arzu
authors.initials: A
authors.lastname: Umar
authors.affiliation: Department of Medical Oncology, Erasmus MC Cancer Institute, Erasmus University Medical Center, Rotterdam, Netherlands.
authors.forename: John A
authors.initials: JA
authors.lastname: Foekens
authors.affiliation: Department of Clinical Cancer Prevention, The University of Texas MD Anderson Cancer Center, Houston, 77030 TX USA.
authors.forename: Samir
authors.initials: S
authors.lastname: Hanash
authors.affiliation: Department of Molecular and Cellular Biology and Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, 77030 TX USA.
authors.forename: Rachel
authors.initials: R
authors.lastname: Schiff
authors.affiliation: Department of Molecular and Cellular Biology and Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, 77030 TX USA.
authors.forename: Xiang H-F
authors.initials: XH
authors.lastname: Zhang
authors.affiliation: Department of Molecular and Cellular Biology and Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, 77030 TX USA.
authors.forename: Eric C
authors.initials: EC
authors.lastname: Chang
keywords: CAF; CXCR4; HMFs; SDF-1; SMA; Stroma; Translation initiation factor; Tumor microenvironment; eIF3e