orals International Association for Breast Cancer Research 2014

R-spondin-3 is an oncogene driving the development of mammary tumors distinctive from Wnt1-induced tumors (#32)

Elvira R.M. Bakker 1 , Mandy Boer 1 , Ji-Ying Song 1 , John Hilkens 1
  1. Netherlands Cancer Institute, Amsterdam, Netherlands

In an MMTV insertional mutagenesis screen, we found Rspo1, Rspo2 and Rspo3 as frequent MMTV targets. We have investigated Rspo3 as paradigm for oncogenic R-spondin signaling in more detail. We have generated a conditional Rspo3 mouse model, in which Rspo3 cDNA flanked by Lox sites was inserted into the Rosa locus in the inverse orientation (Rspo3invf). Upon Cre expression, transgenic Rspo3 switches into the sense orientation and thereby brought under control of the ubiquitous CAGGS promoter. All female MMTV-Cre; Rspo3invf mice developed mammary tumors, validating Rspo3 as a proto-oncogene. Since Rspo’s potentiate Wnt-signaling, we compared mammary tumor incidence, latency and histology in mice expressing either Wnt1 or Rspo3, and we investigated whether Rspo3 expression accelerates Wnt1 induced mammary tumorigenesis. By crossing Rspo3invf mice with MMTV-Cre;MMTV-Wnt1 transgenic mice, we obtained cohorts that express either Wnt1 or Rspo3 or both genes. Rspo3 induced mammary tumors showed a strikingly different histology than those induced by Wnt1. The former were relatively solid, invasive and poorly differentiated with abundant foci of squamous metaplasia and regions with EMT. In contrast, Wnt1 mammary tumors were more differentiated, cystic and less frequently exhibited squamous differentiation or EMT. Also expression levels and patterns of cytokeratins strikingly differed. RNAseq analysis of Rspo3 and Wnt1 tumors revealed a clearly separate clustering of Wnt1 and Rspo3 induced tumors. Among differentially expressed genes known Wnt-target genes were expressed highest in Wnt1 tumors, whereas certain genes encoding ECM degrading enzymes were expressed higher in Rspo3 tumors. Together our findings indicate that Rspo3 induces mammary tumors arise from different progenitor cells or through a mechanism that is distinct from that of Wnt1. In all tumor cohorts, the stem- and progenitor cell related Rspo receptors Lgr4-6 were expressed and we are currently investigating the in situ expression patterns of these receptors in the different tumor types.